Sample records for aeruginosa pao1 strain

  1. Emergence of the P2 Phenotype in Pseudomonas aeruginosa PAO1 Strains Involves Various Mutations in mexT or mexF

    PubMed Central

    Luong, Preston M.; Shogan, Benjamin D.; Zaborin, Alexander; Belogortseva, Natalia; Shrout, Joshua D.

    2014-01-01

    We recently demonstrated that Pseudomonas aeruginosa PAO1 undergoes a pronounced phenotypic change when introduced into the intestines of rats during surgical injury. Recovered strains displayed a specific phenotype (termed the P2 phenotype) characterized by altered pyocyanin production, high collagenase activity, high swarming motility, low resistance to chloramphenicol, and increased killing of Caenorhabditis elegans compared to the inoculating strain (termed the P1 phenotype). The aims of this study were to characterize the differences between the P. aeruginosa P1 and P2 phenotypes in quorum sensing and competitiveness. We then determined the presence of the P2 phenotype among PAO1 strains from various laboratories. Results demonstrated that P2 cells display accelerated growth during early exponential phase and early activation of quorum-sensing systems and overcome the growth of P1 cells in a mixed population. Among eight PAO1 strains obtained from different laboratories, four exhibited the P2 phenotype. Of 27 mutants analyzed from the P. aeruginosa MPAO1 transposon library, 25 displayed P2 phenotypes. The P2 phenotype in both cases correlated with a lack of expression of mexE or mexF due to mutations in mexT and mexF genes. In summary, strains possessing the P2 phenotype are distributed among PAO1 strains commonly used across a variety of research laboratories. Genetically, they are characterized by various mutations in mexT or mexF. PMID:24244000

  2. Cross-reactive and strain-specific antipeptide antibodies to Pseudomonas aeruginosa PAK and PAO pili.

    PubMed Central

    Lee, K K; Paranchych, W; Hodges, R S

    1990-01-01

    Antipeptide antibodies were raised against synthetic peptides corresponding to the amino acid sequences of eight surface predicted regions of the pilin proteins from Pseudomonas aeruginosa PAK and PAO. Four of the anti-PAK peptide antisera cross-reacted with strain PAO pili, while five anti-PAO peptide antisera cross-reacted with strain PAK pili. Only one region of the two pilin proteins (region 88-97) provided strain-specific antibodies when either strain PAK or strain PAO region 88-97 peptides were used to generate antipeptide antibodies. Our results clearly showed that cross-reactive and strain-specific antibodies cannot be based solely on the degree of homology in the aligned protein sequences. The majority of synthetic peptides bound to their homologous antipilus antiserum, suggesting that linear sequences play a significant role in the immunogenic response of native pili. PMID:1974884

  3. Analyses of Short-Term Antagonistic Evolution of Pseudomonas aeruginosa Strain PAO1 and Phage KPP22 (Myoviridae Family, PB1-Like Virus Genus).

    PubMed

    Uchiyama, Jumpei; Suzuki, Masato; Nishifuji, Koji; Kato, Shin-Ichiro; Miyata, Reina; Nasukawa, Tadahiro; Yamaguchi, Kotoe; Takemura-Uchiyama, Iyo; Ujihara, Takako; Shimakura, Hidekatsu; Murakami, Hironobu; Okamoto, Noriaki; Sakaguchi, Yoshihiko; Shibayama, Keigo; Sakaguchi, Masahiro; Matsuzaki, Shigenobu

    2016-08-01

    Pseudomonas aeruginosa causes serious intractable infections in humans and animals. Bacteriophage (phage) therapy has been applied to treat P. aeruginosa infections, and phages belonging to the PB1-like virus genus in the Myoviridae family have been used as therapeutic phages. To achieve safer and more effective phage therapy, the use of preadapted phages is proposed. To understand in detail such phage preadaptation, the short-term antagonistic evolution of bacteria and phages should be studied. In this study, the short-term antagonistic evolution of bacteria and PB1-like phage was examined by studying phage-resistant clones of P. aeruginosa strain PAO1 and mutant PB1-like phages that had recovered their infectivity. First, phage KPP22 was isolated and characterized; it was classified as belonging to the PB1-like virus genus in the Myoviridae family. Subsequently, three KPP22-resistant PAO1 clones and three KPP22 mutant phages capable of infecting these clones were isolated in three sets of in vitro experiments. It was shown that the bacterial resistance to phage KPP22 was caused by significant decreases in phage adsorption and that the improved infectivity of KPP22 mutant phages was caused by significant increases in phage adsorption. The KPP22-resistant PAO1 clones and the KPP22 mutant phages were then analyzed genetically. All three KPP22-resistant PAO1 clones, which were deficient for the O5 antigen, had a common nonsense mutation in the wzy gene. All the KPP22 mutant phage genomes showed the same four missense mutations in the open reading frames orf060, orf065, and orf086 The information obtained in this study should be useful for further development of safe and efficient phage therapy. Pseudomonas aeruginosa causes serious intractable infections in humans and animals; bacteriophage (phage) therapy has been utilized to treat P. aeruginosa infections, and phages that belong to the PB1-like virus genus in the family Myoviridae have been used as therapeutic

  4. Leaf Extracts of Selected Gardening Trees Can Attenuate Quorum Sensing and Pathogenicity of Pseudomonas aeruginosa PAO1.

    PubMed

    Niu, Kaimin; Kuk, Min; Jung, Haein; Chan, Kokgan; Kim, Sooki

    2017-09-01

    An increasing concern on resistance to multiple-antibiotics has led to the discovery of novel agents and the establishment of new precaution strategy. Numerous plant sources have been widely studied to reduce virulence of pathogenic bacteria by interfering cell-to-cell based communication called quorum sensing (QS). Leaf extracts of 17 gardening trees were collected and investigated for their anti-QS effects using a sensor strain Chromobacterium violaceum CV026. Methanolic extracts of K4 ( Acer palmatum ), K9 ( Acer pseudosieboldianum ) and K13 ( Cercis chinensis ) leaves were selected for further experiments based on their antagonism effect on QS without inhibiting C. violaceum CV026 growth. Subsequently, the leaf extracts on QS-mediated virulence of Pseudomonas aeruginosa PAO1 involved in biofilm formation, motility, bioluminescence, pyocyanin production, QS molecules production, and Caenorhabditis elegans killing activity were evaluated. The biofilm formation ability and swarming motility of P. aeruginosa PAO1 were decreased approximately 50% in the presence of these leaf extracts at a concentration of 1 mg/mL. The expression level of lecA::lux of P. aeruginosa PAO1 and pyocyanin production were also reduced. The three leaf extracts also decreased autoinducer (AI) production in P. aeruginosa PAO1 without direct degradation, suggesting that AI synthesis might have been suppressed by these extracts. The three leaf extracts also showed anti-infection activity in C. elegans model. Taken together, these results suggest that methanolic leaf extracts of K4, K9 and K13 have the potential to attenuate the virulence of P. aeruginosa PAO1.

  5. Functional Analysis of Genes for Biosynthesis of Pyocyanin and Phenazine-1-Carboxamide from Pseudomonas aeruginosa PAO1

    PubMed Central

    Mavrodi, Dmitri V.; Bonsall, Robert F.; Delaney, Shannon M.; Soule, Marilyn J.; Phillips, Greg; Thomashow, Linda S.

    2001-01-01

    Two seven-gene phenazine biosynthetic loci were cloned from Pseudomonas aeruginosa PAO1. The operons, designated phzA1B1C1D1E1F1G1 and phzA2B2C2D2E2F2G2, are homologous to previously studied phenazine biosynthetic operons from Pseudomonas fluorescens and Pseudomonas aureofaciens. Functional studies of phenazine-nonproducing strains of fluorescent pseudomonads indicated that each of the biosynthetic operons from P. aeruginosa is sufficient for production of a single compound, phenazine-1-carboxylic acid (PCA). Subsequent conversion of PCA to pyocyanin is mediated in P. aeruginosa by two novel phenazine-modifying genes, phzM and phzS, which encode putative phenazine-specific methyltransferase and flavin-containing monooxygenase, respectively. Expression of phzS alone in Escherichia coli or in enzymes, pyocyanin-nonproducing P. fluorescens resulted in conversion of PCA to 1-hydroxyphenazine. P. aeruginosa with insertionally inactivated phzM or phzS developed pyocyanin-deficient phenotypes. A third phenazine-modifying gene, phzH, which has a homologue in Pseudomonas chlororaphis, also was identified and was shown to control synthesis of phenazine-1-carboxamide from PCA in P. aeruginosa PAO1. Our results suggest that there is a complex pyocyanin biosynthetic pathway in P. aeruginosa consisting of two core loci responsible for synthesis of PCA and three additional genes encoding unique enzymes involved in the conversion of PCA to pyocyanin, 1-hydroxyphenazine, and phenazine-1-carboxamide. PMID:11591691

  6. Genetic and physiological characterization of ciprofloxacin resistance in Pseudomonas aeruginosa PAO.

    PubMed Central

    Robillard, N J; Scarpa, A L

    1988-01-01

    Spontaneous ciprofloxacin-resistant mutants of Pseudomonas aeruginosa PAO2 were isolated on ML agar containing 0.5 microgram of ciprofloxacin per ml (2 times the MIC). The mutants were 8- to 64-fold more resistant to ciprofloxacin and showed complete cross resistance to nalidixic acid, ofloxacin, enoxacin, and norfloxacin. Two chromosomal resistance genes, cfxA and cfxB, were mapped between eda-9001 and phe-2 and near pyrB52 distal to proC130, respectively. The cfxB mutation was identical to a nalB mutation and conferred cross resistance to novobiocin, tetracycline, carbenicillin, and chloramphenicol, suggesting that there is an effect on permeability. DNA gyrase A and B subunits were purified from strain PAO2 (wild type), PAO236 nalA2, PAO4704 cfxA2, and PAO4700 cfxA1 cfxB1. Inhibition of gyrase-mediated DNA supercoiling by ciprofloxacin or nalidixic acid was greatly reduced in preparations derived from each of the mutants. Inhibition studies on reconstituted heterologous gyrase subunits showed that decreased inhibition was dependent on the mutant gyrase A subunit. We conclude that ciprofloxacin resistance in P. aeruginosa PAO2 can occur by mutation in the nalB gene or the gene for DNA gyrase A (formerly nalA). PMID:2837141

  7. Proteomic profiling of Pseudomonas aeruginosa AES-1R, PAO1 and PA14 reveals potential virulence determinants associated with a transmissible cystic fibrosis-associated strain.

    PubMed

    Hare, Nathan J; Solis, Nestor; Harmer, Christopher; Marzook, N Bishara; Rose, Barbara; Harbour, Colin; Crossett, Ben; Manos, Jim; Cordwell, Stuart J

    2012-01-22

    Pseudomonas aeruginosa is an opportunistic pathogen that is the major cause of morbidity and mortality in patients with cystic fibrosis (CF). While most CF patients are thought to acquire P. aeruginosa from the environment, person-person transmissible strains have been identified in CF clinics worldwide. The molecular basis for transmissibility and colonization of the CF lung remains poorly understood. A dual proteomics approach consisting of gel-based and gel-free comparisons were undertaken to analyse protein profiles in a transmissible, early (acute) isolate of the Australian epidemic strain 1 (AES-1R), the virulent burns/wound isolate PA14, and the poorly virulent, laboratory-associated strain PAO1. Over 1700 P. aeruginosa proteins were confidently identified. AES-1R protein profiles revealed elevated abundance of proteins associated with virulence and siderophore biosynthesis and acquisition, antibiotic resistance and lipopolysaccharide and fatty acid biosynthesis. The most abundant protein in AES-1R was confirmed as a previously hypothetical protein with sequence similarity to carbohydrate-binding proteins and database search revealed this gene is only found in the CF-associated strain PA2192. The link with CF infection may suggest that transmissible strains have acquired an ability to rapidly interact with host mucosal glycoproteins. Our data suggest that AES-1R expresses higher levels of proteins, such as those involved in antibiotic resistance, iron acquisition and virulence that may provide a competitive advantage during early infection in the CF lung. Identification of novel proteins associated with transmissibility and acute infection may aid in deciphering new strategies for intervention to limit P. aeruginosa infections in CF patients.

  8. Proteomic profiling of Pseudomonas aeruginosa AES-1R, PAO1 and PA14 reveals potential virulence determinants associated with a transmissible cystic fibrosis-associated strain

    PubMed Central

    2012-01-01

    Background Pseudomonas aeruginosa is an opportunistic pathogen that is the major cause of morbidity and mortality in patients with cystic fibrosis (CF). While most CF patients are thought to acquire P. aeruginosa from the environment, person-person transmissible strains have been identified in CF clinics worldwide. The molecular basis for transmissibility and colonization of the CF lung remains poorly understood. Results A dual proteomics approach consisting of gel-based and gel-free comparisons were undertaken to analyse protein profiles in a transmissible, early (acute) isolate of the Australian epidemic strain 1 (AES-1R), the virulent burns/wound isolate PA14, and the poorly virulent, laboratory-associated strain PAO1. Over 1700 P. aeruginosa proteins were confidently identified. AES-1R protein profiles revealed elevated abundance of proteins associated with virulence and siderophore biosynthesis and acquisition, antibiotic resistance and lipopolysaccharide and fatty acid biosynthesis. The most abundant protein in AES-1R was confirmed as a previously hypothetical protein with sequence similarity to carbohydrate-binding proteins and database search revealed this gene is only found in the CF-associated strain PA2192. The link with CF infection may suggest that transmissible strains have acquired an ability to rapidly interact with host mucosal glycoproteins. Conclusions Our data suggest that AES-1R expresses higher levels of proteins, such as those involved in antibiotic resistance, iron acquisition and virulence that may provide a competitive advantage during early infection in the CF lung. Identification of novel proteins associated with transmissibility and acute infection may aid in deciphering new strategies for intervention to limit P. aeruginosa infections in CF patients. PMID:22264352

  9. Determination of the amino acid change responsible for the nontoxic, cross-reactive exotoxin A protein (CRM 66) of Pseudomonas aeruginosa PAO-PR1.

    PubMed Central

    Wick, M J; Iglewski, B H

    1988-01-01

    Analysis of purified exotoxin A from parental Pseudomonas aeruginosa PAO1 and mutant strain PAO-PR1, which produces enzymatically inactive exotoxin A (CRM 66), revealed that CRM 66 lost 90% of parental enzymatic activity. Nucleotide sequence analysis of cloned exotoxin A genes showed a single amino acid substitution in CRM 66. Position 426 in the mature protein of parental (PAO1) exotoxin A is histidine, whereas in CRM 66, it is tyrosine. Images PMID:3141388

  10. Efflux-Mediated Resistance to Tigecycline (GAR-936) in Pseudomonas aeruginosa PAO1

    PubMed Central

    Dean, Charles R.; Visalli, Melissa A.; Projan, Steven J.; Sum, Phaik-Eng; Bradford, Patricia A.

    2003-01-01

    Pseudomonas aeruginosa strains are less susceptible to tigecycline (previously GAR-936; MIC, 8 μg/ml) than many other bacteria (P. J. Petersen, N. V. Jacobus, W. J. Weiss, P. E. Sum, and R. T. Testa, Antimicrob. Agents Chemother. 43:738-744, 1999). To elucidate the mechanism of resistance to tigecycline, P. aeruginosa PAO1 strains defective in the MexAB-OprM and/or MexXY (OprM) efflux pumps were tested for susceptibility to tigecycline. Increased susceptibility to tigecycline (MIC, 0.5 to 1 μg/ml) was specifically associated with loss of MexXY. Transcription of mexX and mexY was also responsive to exposure of cells to tigecycline. To test for the emergence of compensatory efflux pumps in the absence of MexXY-OprM, mutants lacking MexXY-OprM were plated on medium containing tigecycline at 4 or 6 μg/ml. Resistant mutants were readily recovered, and these also had decreased susceptibility to several other antibiotics, suggesting efflux pump recruitment. One representative carbenicillin-resistant strain overexpressed OprM, the outer membrane channel component of the MexAB-OprM efflux pump. The mexAB-oprM repressor gene, mexR, from this strain contained a 15-bp in-frame deletion. Two representative chloramphenicol-resistant strains showed expression of an outer membrane protein slightly larger than OprM. The mexCD-OprJ repressor gene, nfxB, from these mutants contained a 327-bp in-frame deletion and an IS element insertion, respectively. Together, these data indicated drug efflux mediated by MexCD-OprJ. The MICs of the narrower-spectrum semisynthetic tetracyclines doxycycline and minocycline increased more substantially than did those of tigecycline and other glycylcyclines against the MexAB-OprM- and MexCD-OprJ-overexpressing mutant strains. This suggests that glycylcyclines, although they are subject to efflux from P. aeruginosa, are generally inferior substrates for P. aeruginosa efflux pumps than are narrower-spectrum tetracyclines. PMID:12604529

  11. Methanol Extract from Anogeissus leiocarpus (DC) Guill. et Perr. (Combretaceae) Stem Bark Quenches the Quorum Sensing of Pseudomonas aeruginosa PAO1

    PubMed Central

    Ouedraogo, Vincent; Kiendrebeogo, Martin

    2016-01-01

    Background: Due to its extensive arsenal of virulence factors and inherent resistance to antibiotics, Pseudomonas aeruginosa is a threat particularly in immunocompromised patients. Considering the central role of quorum sensing in the production of virulence factors, inhibition of bacterial communication mechanism constitute an opportunity to attenuate pathogenicity of bacteria resistant to available antibiotics. Our study aimed to assess the anti-quorum sensing activity of Anogeissus leiocarpus, traditionally used in Burkina Faso, for the treatment of infected burn wounds. Methods: Investigations were carried out on methanol extract from A. leiocarpus stem bark. The reporter strains Chromobacterium violaceum CV026 and P. aeruginosa PAO1 derivatives were used to evidence any interference with the bacterial quorum sensing and expression of related genes. P. aeruginosa PAO1 was used to measure the impact on pyocyanin production. Results: At a sub-inhibitory concentration (100 µg/mL), A. leiocarpus methanol extract quenched the quorum sensing mechanism of P. aeruginosa PAO1 by down-streaming the rhlR gene, with a subsequent reduction of pyocyanin production. Moreover, the antioxidant polyphenols evidenced are able to reduce the oxidative stress induced by pyocyanin. Conclusion: The antioxidant and anti-quorum sensing activities of A. leiocarpus stem bark could justify its traditional use in the treatment of infected burn wounds. PMID:28930136

  12. Cesium-induced inhibition of bacterial growth of Pseudomonas aeruginosa PAO1 and their possible potential applications for bioremediation of wastewater.

    PubMed

    Kang, Sung-Min; Jang, Sung-Chan; Heo, Nam Su; Oh, Seo Yeong; Cho, Hye-Jin; Rethinasabapathy, Muruganantham; Vilian, A T Ezhil; Han, Young-Kyu; Roh, Changhyun; Huh, Yun Suk

    2017-09-15

    Radioactive isotopes and fission products have attracted considerable attention because of their long lasting serious damage to the health of humans and other organisms. This study examined the toxicity and accumulation behavior of cesium towards P. aeruginosa PAO1 and its capacity to remove cesium from waste water. Interestingly, the programmed bacterial growth inhibition occurred according to the cesium environment. The influence of cesium was analyzed using several optical methods for quantitative evaluation. Cesium plays vital role in the growth of microorganisms and functions as an anti-microbial agent. The toxicity of Cs to P. aeruginosa PAO1 increases as the concentration of cesium is increased in concentration-dependent manner. P. aeruginosa PAO1 shows excellent Cs removal efficiency of 76.1% from the contaminated water. The toxicity of cesium on the cell wall and in the cytoplasm were studied by transmission electron microscopy and electron dispersive X-ray analysis. Finally, the removal of cesium from wastewater using P. aeruginosa PAO1 as a potential biosorbent and the blocking of competitive interactions of other monovalent cation, such as potassium, were assessed. Overall, P. aeruginosa PAO1 can be used as a high efficient biomaterial in the field of radioactive waste disposal and management. Copyright © 2017 Elsevier B.V. All rights reserved.

  13. Extracts of Cordia gilletii de wild (Boraginaceae) quench the quorum sensing of Pseudomonas aeruginosa PAO1

    PubMed Central

    Okusa, Philippe N.; Rasamiravaka, Tsiry; Vandeputte, Olivier; Stévigny, Caroline; Jaziri, Mondher El; Duez, Pierre

    2014-01-01

    Aim: The fight against infectious diseases and antimicrobial resistances needs the exploration of new active compounds with new proprieties like disrupting quorum sensing (QS) mechanisms, which is a cell-to-cell communication that regulates bacterial virulence factors. In this work, leaves and root barks extracts of a Congolese medicinal plant, Cordia gilletii, were investigated for their effect on the production of Pseudomonas aeruginosa major virulence factors regulated by QS. Materials and Methods: The effect of C. gilletii extracts on virulence factors of P. aeruginosa PAO1 was studied by the evaluation of the production of pyocyanine, elastase and biofilm; and by the measurement of the expression of QS-related genes. Results: The dichloromethane extract from root barks was found to quench the production of pyocyanin, a QS-dependent virulence factor in P. aeruginosa PAO1. Moreover, this extract specifically inhibits the expression of several QS-regulated genes (i.e. lasB, rhlA, lasI, lasR, rhlI, and rhlR) and reduces biofilm formation by PAO1. Conclusion: This study contributes to explain the efficacy of C. gilletii in the traditional treatment of infectious diseases caused by P. aeruginosa. PMID:26401363

  14. Extracts of Cordia gilletii de wild (Boraginaceae) quench the quorum sensing of Pseudomonas aeruginosa PAO1.

    PubMed

    Okusa, Philippe N; Rasamiravaka, Tsiry; Vandeputte, Olivier; Stévigny, Caroline; Jaziri, Mondher El; Duez, Pierre

    2014-01-01

    The fight against infectious diseases and antimicrobial resistances needs the exploration of new active compounds with new proprieties like disrupting quorum sensing (QS) mechanisms, which is a cell-to-cell communication that regulates bacterial virulence factors. In this work, leaves and root barks extracts of a Congolese medicinal plant, Cordia gilletii, were investigated for their effect on the production of Pseudomonas aeruginosa major virulence factors regulated by QS. The effect of C. gilletii extracts on virulence factors of P. aeruginosa PAO1 was studied by the evaluation of the production of pyocyanine, elastase and biofilm; and by the measurement of the expression of QS-related genes. The dichloromethane extract from root barks was found to quench the production of pyocyanin, a QS-dependent virulence factor in P. aeruginosa PAO1. Moreover, this extract specifically inhibits the expression of several QS-regulated genes (i.e. lasB, rhlA, lasI, lasR, rhlI, and rhlR) and reduces biofilm formation by PAO1. This study contributes to explain the efficacy of C. gilletii in the traditional treatment of infectious diseases caused by P. aeruginosa.

  15. Adhesion inhibition of F1C-fimbriated Escherichia coli and Pseudomonas aeruginosa PAK and PAO by multivalent carbohydrate ligands.

    PubMed

    Autar, Reshma; Khan, A Salam; Schad, Matthias; Hacker, Jörg; Liskamp, Rob M J; Pieters, Roland J

    2003-12-05

    In order to evaluate their inhibition of bacterial adhesion, the carbohydrate sequences GalNAcbeta1-->4Gal and GalNAcbeta1-->4Galbeta1-->4Glc were synthesized. The disaccharide was conjugated to dendrons based on the 3,5-di-(2-aminoethoxy)-benzoic acid branching unit to yield di- and tetravalent versions of these compounds. A divalent compound was also prepared that had significantly longer spacer arms. Relevant monovalent compounds were prepared for comparison. Their anti-adhesion properties against F1C-fimbriated uropathogenic Escherichia coli were evaluated in an ELISA-type assay by using a recombinant strain and also by using Pseudomonas aeruginosa strains PAO and PAK. Adhesion inhibition was observed in all cases, and multivalency effects of up to one order of magnitude were observed. The combination of spacer and multivalency effects led to a 38-fold increase in the potency of a divalent inhibitor with long spacer arms towards the PAO strain when compared with the free carbohydrate.

  16. Recalibration of the Pseudomonas aeruginosa Strain Pao Chromosome Map in Time Units Using High-Frequency-of-Recombination Donors

    PubMed Central

    O'Hoy, Kim; Krishnapillai, Viji

    1987-01-01

    High-frequency-of-recombination donors of P. aeruginosa strain PAO were generated using a temperature-sensitive, replication mutant of the IncP-1 plasmid R68, loaded with the transposon Tn2521. Fourteen donors so isolated mobilized the chromosome in a polarized manner from a number of different transfer origins. The donors were used to construct a time of entry map of the entire chromosome and this was achieved by determining the time of entry of 32 randomly dispersed markers in crosses using nalidixic acid to interrupt chromosome transfer. Analysis of the time of entry data enabled the recalibration of the chromosome map to 75 min. PMID:3108071

  17. Effect of nitrofurans and NO generators on biofilm formation by Pseudomonas aeruginosa PAO1 and Burkholderia cenocepacia 370.

    PubMed

    Zaitseva, Julia; Granik, Vladimir; Belik, Alexandr; Koksharova, Olga; Khmel, Inessa

    2009-06-01

    Antibacterial drugs in the nitrofuran series, such as nitrofurazone, furazidin, nitrofurantoin and nifuroxazide, as well as the nitric oxide generators sodium nitroprusside and isosorbide mononitrate in concentrations that do not suppress bacterial growth, were shown to increase the capacity of pathogenic bacteria Pseudomonas aeruginosa PAO1 and Burkholderia cenocepacia 370 to form biofilms. At 25-100microg/ml, nitrofurans 2-2.5-fold enhanced biofilm formation of P. aeruginosa PAO1, and NO donors 3-6-fold. For B. cenocepacia 370, the enhancement was 2-5-fold (nitrofurans) and 4.5-fold (sodium nitroprusside), respectively.

  18. Enhancing extracellular electron transfer between Pseudomonas aeruginosa PAO1 and light driven semiconducting birnessite.

    PubMed

    Ren, Guiping; Sun, Yuan; Ding, Yang; Lu, Anhuai; Li, Yan; Wang, Changqiu; Ding, Hongrui

    2018-06-02

    In recent years, considerable research effort has explored the interaction between semiconducting minerals and microorganisms, such relationship is a promising way to increase the efficiency of bioelectrochemical systems. Herein, the enhancement of electron transfer between birnessite photoanodes and Pseudomonas aeruginosa PAO1 under visible light was investigated. Under light illumination and positive bias, the light-birnessite-PAO1 electrochemical system generated a photocurrent of 279.57 μA/cm 2 , which is 322% and 170% higher than those in the abiotic control and dead culture, suggesting photoenhanced electrochemical interaction between birnessite and Pseudomonas. The I-t curves presented repeatable responses to light on/off cycles, and multi-conditions analyses indicated that the enhanced photocurrent was attributed to the additional redox species associated with P. aeruginosa PAO1 and with the biofilm on birnessite. Electroconductibility analysis was conducted on the biofilm cellularly by conductive atomic force microscope. Pyocyanin was isolated as the biosynthesized extracellular shuttle and characterized by cyclic voltammetry and surface-enhanced Raman spectroscopy. Rapid bioelectron transfer driven by light was observed. The results suggest new opportunities for designing photo-bioelectronic devices and expanding our understanding of extracellular electron transfer with semiconducting minerals under light in nature environments. Copyright © 2018. Published by Elsevier B.V.

  19. Pseudomonas aeruginosa ATCC 9027 is a non-virulent strain suitable for mono-rhamnolipids production.

    PubMed

    Grosso-Becerra, María-Victoria; González-Valdez, Abigail; Granados-Martínez, María-Jessica; Morales, Estefanía; Servín-González, Luis; Méndez, José-Luis; Delgado, Gabriela; Morales-Espinosa, Rosario; Ponce-Soto, Gabriel-Yaxal; Cocotl-Yañez, Miguel; Soberón-Chávez, Gloria

    2016-12-01

    Rhamnolipids produced by Pseudomonas aeruginosa are biosurfactants with a high biotechnological potential, but their extensive commercialization is limited by the potential virulence of P. aeruginosa and by restrictions in producing these surfactants in heterologous hosts. In this work, we report the characterization of P. aeruginosa strain ATCC 9027 in terms of its genome-sequence, virulence, antibiotic resistance, and its ability to produce mono-rhamnolipids when carrying plasmids with different cloned genes from the type strain PAO1. The genes that were expressed from the plasmids are those coding for enzymes involved in the synthesis of this biosurfactant (rhlA and rhlB), as well as the gene that codes for the RhlR transcriptional regulator. We confirm that strain ATCC 9027 forms part of the PA7 clade, but contrary to strain PA7, it is sensitive to antibiotics and is completely avirulent in a mouse model. We also report that strain ATCC 9027 mono-rhamnolipid synthesis is limited by the expression of the rhlAB-R operon. Thus, this strain carrying the rhlAB-R operon produces similar rhamnolipids levels as PAO1 strain. We determined that strain ATCC 9027 with rhlAB-R operon was not virulent to mice. These results show that strain ATCC 9027, expressing PAO1 rhlAB-R operon, has a high biotechnological potential for industrial mono-rhamnolipid production.

  20. Strain- and Substrate-Dependent Redox Mediator and Electricity Production by Pseudomonas aeruginosa

    PubMed Central

    Bosire, Erick M.; Blank, Lars M.

    2016-01-01

    ABSTRACT Pseudomonas aeruginosa is an important, thriving member of microbial communities of microbial bioelectrochemical systems (BES) through the production of versatile phenazine redox mediators. Pure culture experiments with a model strain revealed synergistic interactions of P. aeruginosa with fermenting microorganisms whereby the synergism was mediated through the shared fermentation product 2,3-butanediol. Our work here shows that the behavior and efficiency of P. aeruginosa in mediated current production is strongly dependent on the strain of P. aeruginosa. We compared levels of phenazine production by the previously investigated model strain P. aeruginosa PA14, the alternative model strain P. aeruginosa PAO1, and the BES isolate Pseudomonas sp. strain KRP1 with glucose and the fermentation products 2,3-butanediol and ethanol as carbon substrates. We found significant differences in substrate-dependent phenazine production and resulting anodic current generation for the three strains, with the BES isolate KRP1 being overall the best current producer and showing the highest electrochemical activity with glucose as a substrate (19 μA cm−2 with ∼150 μg ml−1 phenazine carboxylic acid as a redox mediator). Surprisingly, P. aeruginosa PAO1 showed very low phenazine production and electrochemical activity under all tested conditions. IMPORTANCE Microbial fuel cells and other microbial bioelectrochemical systems hold great promise for environmental technologies such as wastewater treatment and bioremediation. While there is much emphasis on the development of materials and devices to realize such systems, the investigation and a deeper understanding of the underlying microbiology and ecology are lagging behind. Physiological investigations focus on microorganisms exhibiting direct electron transfer in pure culture systems. Meanwhile, mediated electron transfer with natural redox compounds produced by, for example, Pseudomonas aeruginosa might enable an

  1. Isolation of the Autoinducer-Quenching Strain that Inhibits LasR in Pseudomonas aeruginosa

    PubMed Central

    Weng, Lixing; Zhang, Yuqian; Yang, Yuxiang; Wang, Lianhui

    2014-01-01

    Quorum sensing (QS) has been recognized as a general phenomenon in microorganisms and plays an important role in many pathogenic bacteria. In this report, we used the Agrobacterium tumefaciens biosensor strain NT1 to rapidly screen for autoinducer-quenching inhibitors from bacteria. After initial screening 5389 isolates obtained from land and beach soil, 53 putative positive strains were identified. A confirmatory bioassay was carried out after concentrating the putative positive culture supernatant, and 22 strains were confirmed to have anti-LasR activity. Finally, we determined the strain JM2, which could completely inhibit biofilm formation of Pseudomonas aeruginosa PAO1, belonged to the genus Pseudomonas by analysis of 16S rDNA. Partially purified inhibitor factor(s) F5 derived from culture supernatants specifically inhibited LasR-controlled elastase and protease in wild type P. aeruginosa PAO1 by 68% and 73%, respectively, without significantly affecting growth; the rhl-controlled pyocyanin and rhamnolipids were inhibited by 54% and 52% in the presence of 100 μg/mL of F5. The swarming motility and biofilm of PAO1 were also inhibited by F5. Real time RT-PCR on samples from 100 μg/mL F5-treated P. aeruginosa showed downregulation of autoinducer synthase (LasRI and rhlI) and cognate receptor (lasR and rhlR) genes by 50%, 28%, 48%, and 29%, respectively. These results provide compelling evidence that the F5 inhibitor(s) interferes with the las system and significantly inhibits biofilm formation. PMID:24736783

  2. Strain- and Substrate-Dependent Redox Mediator and Electricity Production by Pseudomonas aeruginosa.

    PubMed

    Bosire, Erick M; Blank, Lars M; Rosenbaum, Miriam A

    2016-08-15

    Pseudomonas aeruginosa is an important, thriving member of microbial communities of microbial bioelectrochemical systems (BES) through the production of versatile phenazine redox mediators. Pure culture experiments with a model strain revealed synergistic interactions of P. aeruginosa with fermenting microorganisms whereby the synergism was mediated through the shared fermentation product 2,3-butanediol. Our work here shows that the behavior and efficiency of P. aeruginosa in mediated current production is strongly dependent on the strain of P. aeruginosa We compared levels of phenazine production by the previously investigated model strain P. aeruginosa PA14, the alternative model strain P. aeruginosa PAO1, and the BES isolate Pseudomonas sp. strain KRP1 with glucose and the fermentation products 2,3-butanediol and ethanol as carbon substrates. We found significant differences in substrate-dependent phenazine production and resulting anodic current generation for the three strains, with the BES isolate KRP1 being overall the best current producer and showing the highest electrochemical activity with glucose as a substrate (19 μA cm(-2) with ∼150 μg ml(-1) phenazine carboxylic acid as a redox mediator). Surprisingly, P. aeruginosa PAO1 showed very low phenazine production and electrochemical activity under all tested conditions. Microbial fuel cells and other microbial bioelectrochemical systems hold great promise for environmental technologies such as wastewater treatment and bioremediation. While there is much emphasis on the development of materials and devices to realize such systems, the investigation and a deeper understanding of the underlying microbiology and ecology are lagging behind. Physiological investigations focus on microorganisms exhibiting direct electron transfer in pure culture systems. Meanwhile, mediated electron transfer with natural redox compounds produced by, for example, Pseudomonas aeruginosa might enable an entire microbial

  3. A two-component response regulator, gltR, is required for glucose transport activity in Pseudomonas aeruginosa PAO1.

    PubMed Central

    Sage, A E; Proctor, W D; Phibbs, P V

    1996-01-01

    A 729-bp open reading frame (gltR) was identified in Pseudomonas aeruginosa PAO1 that encodes a product homologous to the two-component response regulator family of proteins. Disruption of gltR caused loss of glucose transport activity. Restoration of gltR resulted in wild-type levels of glucose transport. These findings indicate that gltR is required for expression of the glucose transport system in P. aeruginosa. PMID:8830708

  4. Transcriptomic Analyses Elucidate Adaptive Differences of Closely Related Strains of Pseudomonas aeruginosa in Fuel.

    PubMed

    Gunasekera, Thusitha S; Bowen, Loryn L; Zhou, Carol E; Howard-Byerly, Susan C; Foley, William S; Striebich, Richard C; Dugan, Larry C; Ruiz, Oscar N

    2017-05-15

    Pseudomonas aeruginosa can utilize hydrocarbons, but different strains have various degrees of adaptation despite their highly conserved genome. P. aeruginosa ATCC 33988 is highly adapted to hydrocarbons, while P. aeruginosa strain PAO1, a human pathogen, is less adapted and degrades jet fuel at a lower rate than does ATCC 33988. We investigated fuel-specific transcriptomic differences between these strains in order to ascertain the underlying mechanisms utilized by the adapted strain to proliferate in fuel. During growth in fuel, the genes related to alkane degradation, heat shock response, membrane proteins, efflux pumps, and several novel genes were upregulated in ATCC 33988. Overexpression of alk genes in PAO1 provided some improvement in growth, but it was not as robust as that of ATCC 33988, suggesting the role of other genes in adaptation. Expression of the function unknown gene PA5359 from ATCC 33988 in PAO1 increased the growth in fuel. Bioinformatic analysis revealed that PA5359 is a predicted lipoprotein with a conserved Yx(FWY)xxD motif, which is shared among bacterial adhesins. Overexpression of the putative resistance-nodulation-division (RND) efflux pump PA3521 to PA3523 increased the growth of the ATCC 33988 strain, suggesting a possible role in fuel tolerance. Interestingly, the PAO1 strain cannot utilize n -C 8 and n -C 10 The expression of green fluorescent protein (GFP) under the control of alkB promoters confirmed that alk gene promoter polymorphism affects the expression of alk genes. Promoter fusion assays further confirmed that the regulation of alk genes was different in the two strains. Protein sequence analysis showed low amino acid differences for many of the upregulated genes, further supporting transcriptional control as the main mechanism for enhanced adaptation. IMPORTANCE These results support that specific signal transduction, gene regulation, and coordination of multiple biological responses are required to improve the survival

  5. Transcriptomic Analyses Elucidate Adaptive Differences of Closely Related Strains of Pseudomonas aeruginosa in Fuel

    PubMed Central

    Gunasekera, Thusitha S.; Bowen, Loryn L.; Zhou, Carol E.; Howard-Byerly, Susan C.; Foley, William S.; Striebich, Richard C.; Dugan, Larry C.

    2017-01-01

    ABSTRACT Pseudomonas aeruginosa can utilize hydrocarbons, but different strains have various degrees of adaptation despite their highly conserved genome. P. aeruginosa ATCC 33988 is highly adapted to hydrocarbons, while P. aeruginosa strain PAO1, a human pathogen, is less adapted and degrades jet fuel at a lower rate than does ATCC 33988. We investigated fuel-specific transcriptomic differences between these strains in order to ascertain the underlying mechanisms utilized by the adapted strain to proliferate in fuel. During growth in fuel, the genes related to alkane degradation, heat shock response, membrane proteins, efflux pumps, and several novel genes were upregulated in ATCC 33988. Overexpression of alk genes in PAO1 provided some improvement in growth, but it was not as robust as that of ATCC 33988, suggesting the role of other genes in adaptation. Expression of the function unknown gene PA5359 from ATCC 33988 in PAO1 increased the growth in fuel. Bioinformatic analysis revealed that PA5359 is a predicted lipoprotein with a conserved Yx(FWY)xxD motif, which is shared among bacterial adhesins. Overexpression of the putative resistance-nodulation-division (RND) efflux pump PA3521 to PA3523 increased the growth of the ATCC 33988 strain, suggesting a possible role in fuel tolerance. Interestingly, the PAO1 strain cannot utilize n-C8 and n-C10. The expression of green fluorescent protein (GFP) under the control of alkB promoters confirmed that alk gene promoter polymorphism affects the expression of alk genes. Promoter fusion assays further confirmed that the regulation of alk genes was different in the two strains. Protein sequence analysis showed low amino acid differences for many of the upregulated genes, further supporting transcriptional control as the main mechanism for enhanced adaptation. IMPORTANCE These results support that specific signal transduction, gene regulation, and coordination of multiple biological responses are required to improve the

  6. Determining Multiple Responses of Pseudomonas aeruginosa PAO1 to an Antimicrobial Agent, Free Nitrous Acid.

    PubMed

    Gao, Shu-Hong; Fan, Lu; Peng, Lai; Guo, Jianhua; Agulló-Barceló, Míriam; Yuan, Zhiguo; Bond, Philip L

    2016-05-17

    Free nitrous acid (FNA) has recently been demonstrated as an antimicrobial agent on a range of micro-organisms, especially in wastewater-treatment systems. However, the antimicrobial mechanism of FNA is largely unknown. Here, we report that the antimicrobial effects of FNA are multitargeted. The response of a model denitrifier, Pseudomnas aeruginosa PAO1 (PAO1), common in wastewater treatment, was investigated in the absence and presence of inhibitory level of FNA (0.1 mg N/L) under anaerobic denitrifying conditions. This was achieved through coupling gene expression analysis, by RNA sequencing, and with a suite of physiological analyses. Various transcripts exhibited significant changes in abundance in the presence of FNA. Respiration was likely inhibited because denitrification activity was severely depleted, and decreased transcript levels of most denitrification genes occurred. As a consequence, the tricarboxylic acid (TCA) cycle was inhibited due to the lowered cellular redox state in the FNA-exposed cultures. Meanwhile, during FNA exposure, PAO1 rerouted its carbon metabolic pathway from the TCA cycle to pyruvate fermentation with acetate as the end product as a possible survival mechanism. Additionally, protein synthesis was significantly decreased, and ribosome preservation was evident. These findings improve our understanding of PAO1 in response to FNA and contribute toward the potential application for use of FNA as an antimicrobial agent.

  7. Quorum quenching activity in cell-free lysate of endophytic bacteria isolated from Pterocarpus santalinus Linn., and its effect on quorum sensing regulated biofilm in Pseudomonas aeruginosa PAO1.

    PubMed

    Rajesh, P S; Ravishankar Rai, V

    2014-01-01

    Quorum sensing mechanism allows the microorganisms to resist the antibiotic treatment by forming biofilms. Quorum quenching is one of the mechanisms to control the development of drug resistance in microbes. Endophyte bacteria are beneficial to plant growth as they support the immune system against the pathogen attack. The endophytic bacteria present in Pterocarpus santalinus were screened for the presence of N-acyl homoserine lactones (AHLs) degrading bacteria using biosensor strains and further confirmed by quantifying the violacein production. Cell-free lysate of endophytic bacteria, Bacillus firmus PT18 and Enterobacter asburiae PT39 exhibited potent AHL degrading ability by inhibiting about 80% violacein production in biosensor strain. Furthermore, when the cell-free lysate was applied to Pseudomonas aeruginosa PAO1 and PAO1-JP2 biofilm it resulted in significant (p<0.01) inhibition of biofilm formation. The biofilm inhibition was confirmed by visualization of biofilm slides under fluorescence microscopy, which showed decrease in total biomass formation in treated slides. Isolation and amplification of the gene (aiiA) indicated that the presence of AHL lactonase in cell-free lysate and sequence alignment indicated that AiiA contains a "HXHXDH" zinc-binding motif that is being conserved in several groups of metallohydrolases. Therefore, the study shows the potential of AHLs degradation by AHL lactonase present in cell-free lysate of isolated endophytic bacteria and inhibition of quorum sensing regulated biofilm formation in P. aeruginosa PAO1. Copyright © 2013 Elsevier GmbH. All rights reserved.

  8. Gene PA2449 Is Essential for Glycine Metabolism and Pyocyanin Biosynthesis in Pseudomonas aeruginosa PAO1

    PubMed Central

    Lundgren, Benjamin R.; Thornton, William; Dornan, Mark H.; Villegas-Peñaranda, Luis Roberto; Boddy, Christopher N.

    2013-01-01

    Many pseudomonads produce redox active compounds called phenazines that function in a variety of biological processes. Phenazines are well known for their toxicity against non-phenazine-producing organisms, which allows them to serve as crucial biocontrol agents and virulence factors during infection. As for other secondary metabolites, conditions of nutritional stress or limitation stimulate the production of phenazines, but little is known of the molecular details underlying this phenomenon. Using a combination of microarray and metabolite analyses, we demonstrate that the assimilation of glycine as a carbon source and the biosynthesis of pyocyanin in Pseudomonas aeruginosa PAO1 are both dependent on the PA2449 gene. The inactivation of the PA2449 gene was found to influence the transcription of a core set of genes encoding a glycine cleavage system, serine hydroxymethyltransferase, and serine dehydratase. PA2449 also affected the transcription of several genes that are integral in cell signaling and pyocyanin biosynthesis in P. aeruginosa PAO1. This study sheds light on the unexpected relationship between the utilization of an unfavorable carbon source and the production of pyocyanin. PA2449 is conserved among pseudomonads and might be universally involved in the assimilation of glycine among this metabolically diverse group of bacteria. PMID:23457254

  9. Synergy of Silver Nanoparticles and Aztreonam against Pseudomonas aeruginosa PAO1 Biofilms

    PubMed Central

    Park, Amber J.; Vis, Emily C.; Harris, Robert J.

    2014-01-01

    Pathogenic bacterial biofilms, such as those found in the lungs of patients with cystic fibrosis (CF), exhibit increased antimicrobial resistance, due in part to the inherent architecture of the biofilm community. The protection provided by the biofilm limits antimicrobial dispersion and penetration and reduces the efficacy of antibiotics that normally inhibit planktonic cell growth. Thus, alternative antimicrobial strategies are required to combat persistent infections. The antimicrobial properties of silver have been known for decades, but silver and silver-containing compounds have recently seen renewed interest as antimicrobial agents for treating bacterial infections. The goal of this study was to assess the efficacy of citrate-capped silver nanoparticles (AgNPs) of various sizes, alone and in combination with the monobactam antibiotic aztreonam, to inhibit Pseudomonas aeruginosa PAO1 biofilms. Among the different sizes of AgNPs examined, 10-nm nanoparticles were most effective in inhibiting the recovery of P. aeruginosa biofilm cultures and showed synergy of inhibition when combined with sub-MIC levels of aztreonam. Visualization of biofilms treated with combinations of 10-nm AgNPs and aztreonam indicated that the synergistic bactericidal effects are likely caused by better penetration of the small AgNPs into the biofilm matrix, which enhances the deleterious effects of aztreonam against the cell envelope of P. aeruginosa within the biofilms. These data suggest that small AgNPs synergistically enhance the antimicrobial effects of aztreonam against P. aeruginosa in vitro, and they reveal a potential role for combinations of small AgNPs and antibiotics in treating patients with chronic infections. PMID:25049240

  10. Regulation of Nicotine Tolerance by Quorum Sensing and High Efficiency of Quorum Quenching Under Nicotine Stress in Pseudomonas aeruginosa PAO1.

    PubMed

    Tang, Huiming; Zhang, Yunyun; Ma, Yifan; Tang, Mengmeng; Shen, Dongsheng; Wang, Meizhen

    2018-01-01

    Quorum sensing (QS) regulates the behavior of bacterial populations and promotes their adaptation and survival under stress. As QS is responsible for the virulence of vast majority of bacteria, quorum quenching (QQ), the interruption of QS, has become an attractive therapeutic strategy. However, the role of QS in stress tolerance and the efficiency of QQ under stress in bacteria are seldom explored. In this study, we demonstrated that QS-regulated catalase (CAT) expression and biofilm formation help Pseudomonas aeruginosa PAO1 resist nicotine stress. CAT activity and biofilm formation in wild type (WT) and Δ rhlR strains are significantly higher than those in the Δ lasR strain. Supplementation of Δ lasI strain with 3OC12-HSL showed similar CAT activity and biofilm formation as those of the WT strain. LasIR circuit rather than RhlIR circuit is vital to nicotine tolerance. Acylase I significantly decreased the production of virulence factors, namely elastase, pyocyanin, and pyoverdine under nicotine stress compared to the levels observed in the absence of nicotine stress. Thus, QQ is more efficient under stress. To our knowledge, this is the first study to report that QS contributes to nicotine tolerance in P. aeruginosa . This work facilitates a better application of QQ for the treatment of bacterial infections, especially under stress.

  11. Factors influencing the accumulation of ciprofloxacin in Pseudomonas aeruginosa.

    PubMed Central

    Celesk, R A; Robillard, N J

    1989-01-01

    Ciprofloxacin accumulation in Pseudomonas aeruginosa was measured by a bioassay. Drug accumulation in strain PAO2 was compared with that of three spontaneous ciprofloxacin-resistant mutants selected with 0.5 micrograms of ciprofloxacin per ml. PAO4701 cfxA2 contains a mutation in the gyrA gene, PAO4742 cfxB5 may represent a permeability mutant based on pleiotropic drug resistance, and PAO4700 cfxA1 cfxB1 contains both types of mutations. In all strains, drug accumulation was similar, reaching steady state during the first minute of exposure. Drug accumulation was unsaturable over a range of 5 to 80 micrograms/ml, suggesting that ciprofloxacin accumulates by diffusion in P. aeruginosa. Although all four strains accumulated two- to sevenfold more ciprofloxacin in the presence of the inhibitor carbonyl cyanide m-chlorophenylhydrazone, the cfxB mutants accumulated two- to fourfold less drug than either PAO2 or the cfxA2 mutant. Polyacrylamide gel analysis revealed a protein common to cfxB mutants only, while all strains had similar lipopolysaccharide profiles. The results suggest that ciprofloxacin accumulation in P. aeruginosa is a complex phenomenon that may be affected by both an energy-dependent drug efflux process and outer envelope composition. Images PMID:2514623

  12. The Effects of Chinese Herbal Medicines on the Quorum Sensing-Regulated Virulence in Pseudomonas aeruginosa PAO1.

    PubMed

    Chong, Yee Meng; How, Kah Yan; Yin, Wai Fong; Chan, Kok Gan

    2018-04-21

    The quorum sensing (QS) system has been used by many opportunistic pathogenic bacteria to coordinate their virulence determinants in relation to cell-population density. As antibiotic-resistant bacteria are on the rise, interference with QS has been regarded as a novel way to control bacterial infections. As such, many plant-based natural products have been widely explored for their therapeutic roles. These natural products may contain anti-QS compounds that could block QS signals generation or transmission to combat QS pathogens. In this study, we report the anti-QS activities of four different Chinese herbal plant extracts: Poria cum Radix pini , Angelica dahurica , Rhizoma cibotii and Schizonepeta tenuifolia , on Pseudomonas aeruginosa PAO1. All the plants extracted using hexane, chloroform and methanol were tested and found to impair swarming motility and pyocyanin production in P. aeruginosa PAO1, particularly by Poria cum Radix pini . In addition, all the plant extracts also inhibited violacein production in C. violaceum CV026 up to 50% while bioluminescence activities were reduced in lux -based E. coli biosensors, pSB401 and pSB1075, up to about 57%. These anti-QS properties of the four medicinal plants are the first documentation that demonstrates a potential approach to attenuate pathogens’ virulence determinants.

  13. [Formation of the Pseudomonas aeruginosa PAO1 biofilms in the presence of hydrogen peroxide; the effect of the AiiA gene].

    PubMed

    Pliuta, V A; Andreenko, Iu V; Kuznetsov, A E; Khmel', I A

    2013-01-01

    In the natural ecosystems, most bacteria exist as specifically organized biofilms attached to various surfaces; the biofilms have a complex architecture and are surrounded by an exopolymeric matrix. The bacteria in the biofilms are extremely resistant to antibacterial agents. The ability of the pathogenic bacteria to produce biofilms causes serious problems in medicine. Therefore, the study of the action of different compounds with antibacterial activity is of great interest. In this work, we studied the effect of the hydrogen peroxide (H2O2) on the formation of biofilms by Pseudomonas aeruginosa PAO1. It was shown that H2O2 in concentrations that do not suppress bacterial growth (or suppress it only weakly) stimulates the formation of the biofilms. At higher concentrations, H2O2 inhibits the formation of the biofilms. In order to determine if the stimulation of the biofilm formation depends on Quorum Sensing (QS) regulation, the plasmid pME6863 containing the heterologous gene aiiA encoding the N-acyl-homoserine lactonase AiiA was introduced into P. aeruginosa PAO1. The synthesis by cells of this enzyme degrading N-acyl-homoserine lactones (AHL), signaling molecules of the QS systems, led to the absence of the stimulation of the biofilm formation by the action of H2O2. This fact indicates that the stimulation of the biofilm formation in the presence of H2O2 depends on the functioning of the QS systems of the gene expression regulation of P. aeruginosa PAO1.

  14. A diagnostic PCR assay for the detection of an Australian epidemic strain of Pseudomonas aeruginosa

    PubMed Central

    2010-01-01

    Background Chronic lung infection with the bacterium Pseudomonas aeruginosa is one of the hallmarks of cystic fibrosis (CF) and is associated with worsening lung function, increased hospitalisation and reduced life expectancy. A virulent clonal strain of P. aeruginosa (Australian epidemic strain I; AES-I) has been found to be widespread in CF patients in eastern Australia. Methods Suppression subtractive hybridization (SSH) was employed to identify genetic sequences that are present in the AES-I strain but absent from the sequenced reference strain PAO1. We used PCR to evaluate the distribution of several of the AES-I loci amongst a collection of 188 P. aeruginosa isolates which was comprised of 35 AES-I isolates (as determined by PFGE), 78 non-AES-I CF isolates including other epidemic CF strains as well as 69 P. aeruginosa isolates from other clinical and environmental sources. Results We have identified a unique AES-I genetic locus that is present in all 35 AES-I isolates tested and not present in any of the other 153 P. aeruginosa strains examined. We have used this unique AES-I locus to develop a diagnostic PCR and a real-time PCR assay to detect the presence of P. aeruginosa and AES-I in patient sputum samples. Conclusions We have developed diagnostic PCR assays that are 100% sensitive and 100% specific for the P. aeruginosa strain AES-I. We have also shown that Whatman FTA® Elute cards may be used with PCR-based assays to rapidly detect the presence of P. aeruginosa strains in CF sputum. PMID:20637114

  15. Piper betle leaf extract affects the quorum sensing and hence virulence of Pseudomonas aeruginosa PAO1.

    PubMed

    Datta, Siraj; Jana, Debanjan; Maity, Tilak Raj; Samanta, Aveek; Banerjee, Rajarshi

    2016-06-01

    Quorum sensing (QS) plays an important role in virulence of Pseudomonas aeruginosa, blocking of QS ability are viewed as viable antimicrobial chemotherapy and which may prove to be a safe anti-virulent drug. Bioactive components from Piper betle have been reported to possess antimicrobial ability. This study envisages on the anti-QS properties of ethanolic extract of P. betle leaf (PbLE) using P. aeruginosa PAO1 as a model organism. A marked reduction in swarming, swimming, and twitching ability of the bacteria is demonstrated in presence of PbLE. The biofilm and pyocyanin production also shows a marked reduction in presence of PbLE, though it does not affect the bacterial growth. Thus, the studies hint on the possible effect of the bioactive components of PbLE on reducing the virulent ability of the bacteria; identification of bioactive compounds should be investigated further.

  16. Whole-Genome Sequence Variation among Multiple Isolates of Pseudomonas aeruginosa

    PubMed Central

    Spencer, David H.; Kas, Arnold; Smith, Eric E.; Raymond, Christopher K.; Sims, Elizabeth H.; Hastings, Michele; Burns, Jane L.; Kaul, Rajinder; Olson, Maynard V.

    2003-01-01

    Whole-genome shotgun sequencing was used to study the sequence variation of three Pseudomonas aeruginosa isolates, two from clonal infections of cystic fibrosis patients and one from an aquatic environment, relative to the genomic sequence of reference strain PAO1. The majority of the PAO1 genome is represented in these strains; however, at least three prominent islands of PAO1-specific sequence are apparent. Conversely, ∼10% of the sequencing reads derived from each isolate fail to align with the PAO1 backbone. While average sequence variation among all strains is roughly 0.5%, regions of pronounced differences were evident in whole-genome scans of nucleotide diversity. We analyzed two such divergent loci, the pyoverdine and O-antigen biosynthesis regions, by complete resequencing. A thorough analysis of isolates collected over time from one of the cystic fibrosis patients revealed independent mutations resulting in the loss of O-antigen synthesis alternating with a mucoid phenotype. Overall, we conclude that most of the PAO1 genome represents a core P. aeruginosa backbone sequence while the strains addressed in this study possess additional genetic material that accounts for at least 10% of their genomes. Approximately half of these additional sequences are novel. PMID:12562802

  17. Homogentisate 1-2-Dioxygenase Downregulation in the Chronic Persistence of Pseudomonas aeruginosa Australian Epidemic Strain-1 in the CF Lung

    PubMed Central

    Harmer, Christopher J.; Wynn, Matthew; Pinto, Rachel; Cordwell, Stuart; Rose, Barbara R.; Harbour, Colin; Triccas, James A.; Manos, Jim

    2015-01-01

    Some Pseudomonas aeruginosa strains including Australian Epidemic Strain-1 (AES-1 or AUS-01) cause persistent chronic infection in cystic fibrosis (CF) patients, with greater morbidity and mortality. Factors conferring persistence are largely unknown. Previously we analysed the transcriptomes of AES-1 grown in Luria broth, nematode growth medium for Caenorhabditis elegans assay (both aerobic) and artificial sputum medium (mainly hypoxic). Transcriptional comparisons included chronic AES-1 strains against PAO1 and acute AES-1 (AES-1R) against its chronic isogen (AES-1M), isolated 10.5 years apart from a CF patient and not eradicated in the meantime. Prominent amongst genes downregulated in AES-1M in all comparisons was homogentisate-1-2-dioxygenase (hmgA); an oxygen-dependent gene known to be mutationally deactivated in many chronic infection strains of P. aeruginosa. To investigate if hmgA downregulation and deactivation gave similar virulence persistence profiles, a hmgA mutant made in UCBPP-PA14 utilising RedS-recombinase and AES-1M were assessed in the C. elegans virulence assay, and the C57BL/6 mouse for pulmonary colonisation and TNF-α response. In C. elegans, hmgA deactivation resulted in significantly increased PA14 virulence while hmgA downregulation reduced AES-1M virulence. AES-1M was significantly more persistent in mouse lung and showed a significant increase in TNF-α (p<0.0001), sustained even with no detectable bacteria. PA14ΔhmgA did not show increased TNF-α. This study suggests that hmgA may have a role in P. aeruginosa persistence in chronic infection and the results provide a starting point for clarifying the role of hmgA in chronic AES-1. PMID:26252386

  18. Proteomics of Pseudomonas aeruginosa Australian epidemic strain 1 (AES-1) cultured under conditions mimicking the cystic fibrosis lung reveals increased iron acquisition via the siderophore pyochelin.

    PubMed

    Hare, Nathan J; Soe, Cho Zin; Rose, Barbara; Harbour, Colin; Codd, Rachel; Manos, Jim; Cordwell, Stuart J

    2012-02-03

    Pseudomonas aeruginosa is an opportunistic pathogen that is the major cause of morbidity and mortality in patients with cystic fibrosis (CF). While most CF patients are thought to acquire P. aeruginosa from the environment, person-to-person transmissible strains have been identified in CF clinics worldwide, and the molecular basis for transmissibility remains poorly understood. We undertook a complementary proteomics approach to characterize protein profiles from a transmissible, acute isolate of the Australian epidemic strain 1 (AES-1R), the virulent burns/wound isolate PA14, and the poorly virulent, laboratory-associated strain PAO1 when grown in an artificial medium that mimics the CF lung environment compared to growth in standard laboratory medium. Proteins elevated in abundance in AES-1R included those involved in methionine and S-adenosylmethionine biosynthesis and in the synthesis of phenazines. Proteomic data were validated by measuring culture supernatant levels of the virulence factor pyocyanin, which is the final product of the phenazine pathway. AES-1R and PAO1 released higher extracellular levels of pyocyanin compared to PA14 when grown in conditions that mimic the CF lung. Proteins associated with biosynthesis of the iron-scavenging siderophore pyochelin (PchDEFGH and FptA) were also present at elevated abundance in AES-1R and at much higher levels than in PAO1, whereas they were reduced in PA14. These protein changes resulted phenotypically in increased extracellular iron acquisition potential and, specifically, elevated pyochelin levels in AES-1R culture supernatants as detected by chrome azurol-S assay and fluorometry, respectively. Transcript analysis of pyochelin genes (pchDFG and fptA) showed they were highly expressed during the early stage of growth in artificial sputum medium (18 h) but returned to basal levels following the establishment of microcolony growth (72 h) consistent with that observed in the CF lung. This provides further

  19. The Antiproliferative Effect of Cyclodipeptides from Pseudomonas aeruginosa PAO1 on HeLa Cells Involves Inhibition of Phosphorylation of Akt and S6k Kinases.

    PubMed

    Hernández-Padilla, Laura; Vázquez-Rivera, Dolores; Sánchez-Briones, Luis A; Díaz-Pérez, Alma L; Moreno-Rodríguez, José; Moreno-Eutimio, Mario A; Meza-Carmen, Victor; Cruz, Homero Reyes-De la; Campos-García, Jesús

    2017-06-20

    Pseudomonas aeruginosa PAO1, a potential pathogen of plants and animals, produces the cyclodipeptides cyclo(l-Pro-l-Tyr), cyclo(l-Pro-l-Phe), and cyclo(l-Pro-l-Val) (PAO1-CDPs), whose effects have been implicated in inhibition of human tumor cell line proliferation. Our purpose was to investigate in depth in the mechanisms of HeLa cell proliferation inhibition by the PAO1-CDPs. The results indicate that PAO1-CDPs, both purified individually and in mixtures, inhibited HeLa cell proliferation by arresting the cell cycle at the G0-G1 transition. The crude PAO1-CDPs mixture promoted cell death in HeLa cells in a dose-dependent manner, showing efficacy similar to that of isolated PAO1-CDPs (LD 50 of 60-250 µM) and inducing apoptosis with EC 50 between 0.6 and 3.0 µM. Moreover, PAO1-CDPs showed a higher proapoptotic activity (~10³-10⁵ fold) than their synthetic analogs did. Subsequently, the PAO1-CDPs affected mitochondrial membrane potential and induced apoptosis by caspase-9-dependent pathway. The mechanism of inhibition of cells proliferation in HeLa cells involves inhibition of phosphorylation of both Akt-S473 and S6k-T389 protein kinases, showing a cyclic behavior of their expression and phosphorylation in a time and concentration-dependent fashion. Taken together our findings indicate that PI3K-Akt-mTOR-S6k signaling pathway blockage is involved in the antiproliferative effect of the PAO1-CDPs.

  20. Nitrogen Source Stabilization of Quorum Sensing in the Pseudomonas aeruginosa Bioaugmentation Strain SD-1.

    PubMed

    Wang, Mei-Zhen; Lai, Bai-Min; Dandekar, Ajai A; Yang, Yu-Sheng; Li, Na; Yin, Jun; Shen, Dong-Sheng

    2017-08-15

    Pseudomonas aeruginosa SD-1 is efficient at degrading aromatic compounds and can therefore contribute to the bioremediation of wastewater. P. aeruginosa uses quorum sensing (QS) to regulate the production of numerous secreted "public goods." In wastewater bioaugmentation applications, there are myriad nitrogen sources, and we queried whether various nitrogen sources impact the stabilities of both QS and the bacterial populations. In a laboratory strain of P. aeruginosa , PAO1, the absence of a nitrogen source has been shown to destabilize these populations through the emergence of QS mutant "cheaters." We tested the ability of SD-1 to grow in casein broth, a condition that requires QS for growth, when the nitrogen source with either NH 4 Cl, NaNO 3 , or NaNO 2 or with no added nitrogen source. There was great variability in susceptibility to invasion by QS mutant cheaters and, by extension, the stability of the SD-1 population. When grown with NH 4 Cl as an extra nitrogen source, no population collapse was observed; by contrast, two-thirds of cultures grown in the presence of NaNO 2 collapsed. In the populations that collapsed, the frequency of social cheaters exceeded 40%. NaNO 3 and NaNO 2 directly favor QS mutants of P. aeruginosa SD-1. Although the mechanism by which these nitrogen sources act is not clear, these data indicate that the metabolism of nitrogen can affect the stability of bacterial populations, an important observation for continuing industrial applications with this species. IMPORTANCE Bioaugmentation as a method to help remediate wastewater pollutant streams holds significant potential to enhance traditional methods of treatment. Addition of microbes that can catabolize organic pollutants can be an effective method to remove several toxic compounds. Such bioaugmented strains of bacteria have been shown to be susceptible to competition from the microbiota that are present in wastewater streams, limiting their potential effectiveness. Here, we

  1. Identification of CtpL as a chromosomally encoded chemoreceptor for 4-chloroaniline and catechol in Pseudomonas aeruginosa PAO1.

    PubMed

    Vangnai, Alisa S; Takeuchi, Kazuki; Oku, Shota; Kataoka, Naoya; Nitisakulkan, Tisana; Tajima, Takahisa; Kato, Junichi

    2013-12-01

    Bacterial chemotaxis influences the ability of bacteria to survive and thrive in most environments, including polluted ones. Despite numerous reports of the phenotypic characterization of chemotactic bacteria, only a few molecular details of chemoreceptors for aromatic pollutants have been described. In this study, the molecular basis of chemotaxis toward an environmentally toxic chlorinated aromatic pollutant, 4-chloroaniline (4CA), was evaluated. Among the three Pseudomonas spp. tested, Pseudomonas aeruginosa PAO1 exhibited positive chemotaxis both to the nonmetabolizable 4CA, where 4-chloroacetanilide was formed as a dead-end transformation product, and to the metabolizable catechol. Molecular analysis of all 26 mutants with a disrupted methyl-accepting chemotaxis gene revealed that CtpL, a chromosomally encoded chemoreceptor, was responsible for the positive chemotactic response toward 4CA. Since CtpL has previously been described to be a major chemoreceptor for inorganic phosphate at low concentrations in PAO1, this report describes a fortuitous ability of CtpL to function toward aromatic pollutants. In addition, its regulation not only was dependent on the presence of the chemoattractant inducer but also was regulated by conditions of phosphate starvation. These results expand the range of known chemotactic transducers and their function in the environmental bacterium PAO1.

  2. 1H NMR-Based Global Metabolic Studies of Pseudomonas aeruginosa upon Exposure of the Quorum Sensing Inhibitor Resveratrol.

    PubMed

    Chen, Tongtong; Sheng, Jiyang; Fu, Yonghong; Li, Minghui; Wang, Junsong; Jia, Ai-Qun

    2017-02-03

    Quorum sensing (QS) is a process of bacterial communication that has been a novel target for drug discovery. Pyocyanin quantification assay confirmed that resveratrol was an effective quorum sensing inhibitor (QSI) against Pseudomonas aeruginosa PAO1. In this study, the global metabolite changes of P. aeruginosa PAO1 exposed to QSI resveratrol were investigated by 1 H NMR spectroscopy. A total of 40 metabolites containing amino acids, organic acid, organic amine, and energy storage compounds were identified. The changed metabolic profile indicated that resveratrol influenced pathways including oxidative stress, protein synthesis, and energy metabolism. Oxidative stress could upregulate the expression of genes related to QS in P. aeruginosa. It suggested that resveratrol could inhibit the QS systems in P. aeruginosa PAO1 by relieving oxidative stress due to its antioxidant activity. On the other hand, resveratrol could attenuate the pathogenicity of P. aeruginosa PAO1 by disturbing the TCA cycle so that anaerobic respiration could suppress the virulence because anaerobiosis could induce the loss of cytotoxicity regulated by QS in P. aeruginosa. These findings deepened our comprehending of the metabolic responses of P. aeruginosa PAO1 to resveratrol and pinpointed the possible underlying mechanism of resveratrol's inhibition effect on QS in P. aeruginosa PAO1.

  3. Anti-quorum sensing activity of flavonoid-rich fraction from Centella asiatica L. against Pseudomonas aeruginosa PAO1.

    PubMed

    Vasavi, H S; Arun, A B; Rekha, P D

    2016-02-01

    Inhibition of quorum sensing (QS), a cell-density dependent regulation of gene expression in bacteria by autoinducers is an attractive strategy for the development of antipathogenic agents. In this study, the anti-QS activity of the ethanolic extract of the traditional herb Centella asiatica was investigated by the biosensor bioassay using Chromobacterium violaceum CV026. The effect of ethyl acetate fraction (CEA) from the bioassay-guided fractionation of ethanol extract on QS-regulated violacein production in C. violaceum ATCC12472 and pyocyanin production, proteolytic and elastolytic activities, swarming motility, and biofilm formation in Pseudomonas aeruginosa PAO1 were evaluated. Possible mechanism of QS-inhibitory action on autoinducer activity was determined by measuring the acyl homoserine lactone using C. violaceum ATCC31532. Anti-QS compounds in the CEA fraction were identified using thin layer chromatography biosensor overlay assay. Ethanol extract of C. asiatica showed QS inhibition in C. violaceum CV026. Bioassay-guided fractionation of ethanol extract revealed that CEA was four times more active than the ethanol extract. CEA, at 400 μg/mL, completely inhibited violacein production in C. violaceum ATCC12472 without significantly affecting growth. CEA also showed inhibition of QS-regulated phenotypes, namely, pyocyanin production, elastolytic and proteolytic activities, swarming motility, and biofilm formation in P. aeruginosa PAO1 in a concentration-dependent manner. Thin layer chromatography of CEA with biosensor overlay showed anti-QS spot with an Rf value that corresponded with that of standard kaempferol. The anti-QS nature of C. asiatica herb can be further exploited for the formulation of drugs targeting bacterial infections where pathogenicity is mediated through QS. Copyright © 2014. Published by Elsevier B.V.

  4. Strain identification and quorum sensing inhibition characterization of marine-derived Rhizobium sp. NAO1

    NASA Astrophysics Data System (ADS)

    Chang, Hong; Zhou, Jin; Zhu, Xiaoshan; Yu, Shenchen; Chen, Lu; Jin, Hui; Cai, Zhonghua

    2017-03-01

    A novel strategy for combating pathogens is through the ongoing development and use of anti-quorum sensing (QS) treatments such as therapeutic bacteria or their anti-QS substances. Relatively little is known about the bacteria that inhabit the open ocean and of their potential anti-pathogenic attributes; thus, in an initiative to identify these types of therapeutic bacteria, planktonic microbes from the North Atlantic Ocean were collected, isolated, cultured and screened for anti-QS activity. Screening analysis identified one such strain, Rhizobium sp. NAO1. Extracts of Rhizobium sp. NAO1 were identified via ultra-performance liquid chromatography (UPLC) analysis. They were shown to contain N-acyl homoserine lactone (AHL)-based QS analogues (in particular, the N-butyryl homoserine lactone (C4-AHL) analogue) and could disrupt biofilm formation by Pseudomonas aeruginosa PAO1. QS inhibition was confirmed using confocal scanning laser microscopy and growth curves, and it was shown to occur in a dose-dependent manner without affecting bacterial growth. Secondary metabolites of Rhizobium sp. NAO1 inhibited PAO1 pathogenicity by downregulating AHL-mediated virulence factors such as elastase activity and siderophore production. Furthermore, as a result of biofilm structure damage, the secondary metabolite products of Rhizobium sp. NAO1 significantly increased the sensitivity of PAO1 to aminoglycoside antibiotics. Our results demonstrated that Rhizobium sp. strain NAO1 has the ability to disrupt P. aeruginosa PAO1 biofilm architecture, in addition to attenuating P. aeruginosa PAO1 virulence factor production and pathogenicity. Therefore, the newly identified ocean-derived Rhizobium sp. NAO1 has the potential to serve as a QS inhibitor and may be a new microbial resource for drug development.

  5. Cross-Regulation between the phz1 and phz2 Operons Maintain a Balanced Level of Phenazine Biosynthesis in Pseudomonas aeruginosa PAO1

    PubMed Central

    Jiang, Bei; Xiao, Bo; Liu, Linde; Ge, Yihe; Hu, Xiaomei

    2016-01-01

    Gene duplication often provides selective advantages for the survival of microorganisms in adapting to varying environmental conditions. P. aeruginosa PAO1 possesses two seven-gene operons [phz1 (phzA1B1C1D1E1F1G1) and phz2 (phzA2B2C2D2E2F2G2)] that are involved in the biosynthesis of phenazine-1-carboxylic acid and its derivatives. Although the two operons are highly homologous and their functions are well known, it is unclear how the two phz operons coordinate their expressions to maintain the phenazine biosynthesis. By constructing single and double deletion mutants of the two phz operons, we found that the phz1-deletion mutant produced the same or less amount of phenazine-1-carboxylic acid and pyocyanin in GA medium than the phz2-knockout mutant while the phz1-phz2 double knockout mutant did not produce any phenazines. By generating phzA1 and phzA2 translational and transcriptional fusions with a truncated lacZ reporter, we found that the expression of the phz1 operon increased significantly at the post-transcriptional level and did not alter at the transcriptional level in the absence of the phz2 operon. Surprisingly, the expression the phz2 operon increased significantly at the post-transcriptional level and only moderately at the transcriptional level in the absence of the phz1 operon. Our findings suggested that a complex cross-regulation existed between the phz1 and phz2 operons. By mediating the upregulation of one phz operon expression while the other was deleted, this crosstalk would maintain the homeostatic balance of phenazine biosynthesis in P. aeruginosa PAO1. PMID:26735915

  6. Bioproduction of D-Tagatose from D-Galactose Using Phosphoglucose Isomerase from Pseudomonas aeruginosa PAO1.

    PubMed

    Patel, Manisha J; Patel, Arti T; Akhani, Rekha; Dedania, Samir; Patel, Darshan H

    2016-07-01

    Pseudomonas aeruginosa PAO1 phosphoglucose isomerase was purified as an active soluble form by a single-step purification using Ni-NTA chromatography that showed homogeneity on SDS-PAGE with molecular mass ∼62 kDa. The optimum temperature and pH for the maximum isomerization activity with D-galactose were 60 °C and 7.0, respectively. Generally, sugar phosphate isomerases show metal-independent activity but PA-PGI exhibited metal-dependent isomerization activity with aldosugars and optimally catalyzed the D-galactose isomerization in the presence of 1.0 mM MnCl2. The apparent Km and Vmax for D-galactose under standardized conditions were calculated to be 1029 mM (±31.30 with S.E.) and 5.95 U/mg (±0.9 with S.E.), respectively. Equilibrium reached after 180 min with production of 567.51 μM D-tagatose from 1000 mM of D-galactose. Though, the bioconversion ratio is low but it can be increased by immobilization and enzyme engineering. Although various L-arabinose isomerases have been characterized for bioproduction of D-tagatose, P. aeruginosa glucose phosphate isomerase is distinguished from the other L-arabinose isomerases by its optimal temperature (60 °C) for D-tagatose production being mesophilic bacteria, making it an alternate choice for bulk production.

  7. A new approach to study attached biofilms and floating communities from Pseudomonas aeruginosa strains of various origins reveals diverse effects of divalent ions.

    PubMed

    Gagné-Thivierge, Cynthia; Barbeau, Jean; Levesque, Roger C; Charette, Steve J

    2018-06-25

    Pseudomonas aeruginosa is an opportunistic pathogen associated with nosocomial infections and disease complications. In the lungs of cystic fibrosis (CF) individuals, biofilm growth plays a crucial role in the persistence and antibiotic resistance of P. aeruginosa. Some strains, adapted to the CF lung microenvironment, show distinguishable phenotypes linked to biofilm production when compared to other strains. Using a novel image analysis quantification approach with crystal violet-stained biofilms, we compared the biofilm formation of four different P. aeruginosa isolates in 24-well plates: PAO1, the reference strain, LESB58 from CF patients' lungs, and PPF-1 and Urg-7, two environmental isolates from dental unit waterlines. We also observed the formation of biofilm-like structures (BLSs) floating in the medium and investigated growth inhibition of the attached biofilm and BLS with Mg2+ or Zn2+. Urg-7 produced the most attached biofilms, but not the most BLSs. Attached biofilms had different responses to cations than BLSs did, but the effect of the cations was similar for all strains. These results demonstrate some diversity of biofilm formation in P. aeruginosa and indicate that chemical inhibition of attached biofilm formation for a specific strain or isolate cannot be predicative of a result on other P. aeruginosa strains or on BLSs.

  8. Genetically and Phenotypically Distinct Pseudomonas aeruginosa Cystic Fibrosis Isolates Share a Core Proteomic Signature

    PubMed Central

    Penesyan, Anahit; Kumar, Sheemal S.; Kamath, Karthik; Shathili, Abdulrahman M.; Venkatakrishnan, Vignesh; Krisp, Christoph; Packer, Nicolle H.; Molloy, Mark P.; Paulsen, Ian T.

    2015-01-01

    The opportunistic pathogen Pseudomonas aeruginosa is among the main colonizers of the lungs of cystic fibrosis (CF) patients. We have isolated and sequenced several P. aeruginosa isolates from the sputum of CF patients and compared them with each other and with the model strain PAO1. Phenotypic analysis of CF isolates showed significant variability in colonization and virulence-related traits suggesting different strategies for adaptation to the CF lung. Genomic analysis indicated these strains shared a large set of core genes with the standard laboratory strain PAO1, and identified the genetic basis for some of the observed phenotypic differences. Proteomics revealed that in a conventional laboratory medium PAO1 expressed 827 proteins that were absent in the CF isolates while the CF isolates shared a distinctive signature set of 703 proteins not detected in PAO1. PAO1 expressed many transporters for the uptake of organic nutrients and relatively few biosynthetic pathways. Conversely, the CF isolates expressed a narrower range of transporters and a broader set of metabolic pathways for the biosynthesis of amino acids, carbohydrates, nucleotides and polyamines. The proteomic data suggests that in a common laboratory medium PAO1 may transport a diverse set of “ready-made” nutrients from the rich medium, whereas the CF isolates may only utilize a limited number of nutrients from the medium relying mainly on their own metabolism for synthesis of essential nutrients. These variations indicate significant differences between the metabolism and physiology of P. aeruginosa CF isolates and PAO1 that cannot be detected at the genome level alone. The widening gap between the increasing genomic data and the lack of phenotypic data means that researchers are increasingly reliant on extrapolating from genomic comparisons using experimentally characterized model organisms such as PAO1. While comparative genomics can provide valuable information, our data suggests that such

  9. Anti-quorum sensing activity of Psidium guajava L. flavonoids against Chromobacterium violaceum and Pseudomonas aeruginosa PAO1.

    PubMed

    Vasavi, Halkare Suryanarayana; Arun, Ananthapadmanabha Bhagwath; Rekha, Punchapady-Devasya

    2014-05-01

    Psidium guajava L., which has been used traditionally as a medicinal plant, was explored for anti-quorum sensing (QS) activity. The anti-QS activity of the flavonoid (FL) fraction of P. guajava leaves was determined using a biosensor bioassay with Chromobacterium violaceum CV026. Detailed investigation of the effects of the FL-fraction on QS-regulated violacein production in C. violaceum ATCC12472 and pyocyanin production, proteolytic, elastolytic activities, swarming motility and biofilm formation in Pseudomonas aeruginosa PAO1 was performed using standard methods. Possible mechanisms of QS-inhibition were studied by assessing violacein production in response to N-acyl homoserine lactone (AHL) synthesis in the presence of the FL-fraction in C. violaceum ATCC31532 and by evaluating the induction of violacein in the mutant C. violaceum CV026 by AHL extracted from the culture supernatants of C. violaceum 31532. Active compounds in the FL-fraction were identified by liquid chromatography-mass spectrometry (LC-MS). Inhibition of violacein production by the FL-fraction in a C. violaceum CV026 biosensor bioassay indicated possible anti-QS activity. The FL-fraction showed concentration-dependent decreases in violacein production in C. violaceum 12472 and inhibited pyocyanin production, proteolytic and elastolytic activities, swarming motility and biofilm formation in P. aeruginosa PAO1. Interestingly, the FL-fraction did not inhibit AHL synthesis; AHL extracted from cultures of C. violaceum 31532 grown in the presence of the FL-fraction induced violacein in the mutant C. violaceum CV026. LC-MS analysis revealed the presence of quercetin and quercetin-3-O-arabinoside in the FL-fraction. Both quercetin and quercetin-3-O-arabinoside inhibited violacein production in C. violaceum 12472, at 50 and 100 μg/mL, respectively. Results of this study provide scope for further research to exploit these active molecules as anti-QS agents. © 2014 The Societies and Wiley Publishing

  10. Activities of Antibiotic Combinations against Resistant Strains of Pseudomonas aeruginosa in a Model of Infected THP-1 Monocytes

    PubMed Central

    Buyck, Julien M.

    2014-01-01

    Antibiotic combinations are often used for treating Pseudomonas aeruginosa infections but their efficacy toward intracellular bacteria has not been investigated so far. We have studied combinations of representatives of the main antipseudomonal classes (ciprofloxacin, meropenem, tobramycin, and colistin) against intracellular P. aeruginosa in a model of THP-1 monocytes in comparison with bacteria growing in broth, using the reference strain PAO1 and two clinical isolates (resistant to ciprofloxacin and meropenem, respectively). Interaction between drugs was assessed by checkerboard titration (extracellular model only), by kill curves, and by using the fractional maximal effect (FME) method, which allows studying the effects of combinations when dose-effect relationships are not linear. For drugs used alone, simple sigmoidal functions could be fitted to all concentration-effect relationships (extracellular and intracellular bacteria), with static concentrations close to (ciprofloxacin, colistin, and meropenem) or slightly higher than (tobramycin) the MIC and with maximal efficacy reaching the limit of detection in broth but only a 1 to 1.5 (colistin, meropenem, and tobramycin) to 2 to 3 (ciprofloxacin) log10 CFU decrease intracellularly. Extracellularly, all combinations proved additive by checkerboard titration but synergistic using the FME method and more bactericidal in kill curve assays. Intracellularly, all combinations proved additive only based on both FME and kill curve assays. Thus, although combinations appeared to modestly improve antibiotic activity against intracellular P. aeruginosa, they do not allow eradication of these persistent forms of infections. Combinations including ciprofloxacin were the most active (even against the ciprofloxacin-resistant strain), which is probably related to the fact this drug was the most effective alone intracellularly. PMID:25348528

  11. Role of TonB1 in pyoverdine-mediated signaling in Pseudomonas aeruginosa.

    PubMed

    Shirley, Matt; Lamont, Iain L

    2009-09-01

    Pyoverdines are siderophores secreted by Pseudomonas aeruginosa. Uptake of ferripyoverdine in P. aeruginosa PAO1 occurs via the FpvA receptor protein and requires the energy-transducing protein TonB1. Interaction of (ferri)pyoverdine with FpvA activates pyoverdine gene expression in a signaling process involving the cytoplasmic-membrane-spanning anti-sigma factor FpvR and the sigma factor PvdS. Here, we show that mutation of a region of FpvA that interacts with TonB1 (the TonB box) prevents this signaling process, as well as inhibiting bacterial growth in the presence of the iron-chelating compound ethylenediamine-di(o-hydroxy-phenylacetic acid). Signaling via wild-type FpvA was also eliminated in strains lacking TonB1 but was unaffected in strains lacking either (or both) of two other TonB proteins in P. aeruginosa, TonB2 and TonB3. An absence of pyoverdine-mediated signaling corresponded with proteolysis of PvdS. These data show that interactions between FpvA and TonB1 are required for (ferri)pyoverdine signal transduction, as well as for ferripyoverdine transport, consistent with a mechanistic link between the signaling and transport functions of FpvA.

  12. Resistance of Pseudomonas aeruginosa PAO to nalidixic acid and low levels of beta-lactam antibiotics: mapping of chromosomal genes.

    PubMed Central

    Rella, M; Haas, D

    1982-01-01

    Resistance to high concentrations of nalidixic acid in Pseudomonas aeruginosa PAO was due to mutations in one locus designated nalA, which was mapped by transduction between hex-9001 and leu-10. The nalA mutants were cross-resistant to pipemidic acid, a nalidixic acid analog, at relatively low concentrations. Replicative DNA synthesis was resistant to both drugs in permeabilized cells of nalA mutants. A locus coding for low-level resistance to nalidixic acid, nalB, was cotransducible with pyrB, proC, and met-28. The nalB mutants were also resistant to low levels of pipemidic acid, novobiocin, and beta-lactam antibiotics (e.g., carbenicillin, azlocillin, and cefsulodin), but not to other drugs, such as gentamicin, rifampin, kanamycin, or tetracycline. In nalB mutants, DNA replication showed wild-type sensitivity to nalidixic acid, whereas carbenicillin-induced filamentation required higher drug levels than in the wild-type strain. Thus, nalB mutations appear to decrease cell permeability to some antibiotics. The sensitivity of replicative DNA synthesis to nalidixic acid and novobiocin was very similar in P. aeruginosa and Escherichia coli; by contrast, the concentrations of these drugs needed to inhibit growth of P. aeruginosa were higher than those reported for E. coli by one or two orders of magnitude. PMID:6821455

  13. Multiple virulence factors regulated by quorum sensing may help in establishment and colonisation of urinary tract by Pseudomonas aeruginosa during experimental urinary tract infection.

    PubMed

    Gupta, P; Gupta, R K; Harjai, K

    2013-01-01

    Damage caused by an organism during infection is attributed to production of virulence factors. Different virulence factors produced by the organism contribute to its pathogenicity, individually. During infectious conditions, role of virulence factors produced by the pathogen is different, depending upon the site of involvement. Pseudomonas aeruginosa is an opportunistic nosocomial pathogen known to cause infections of the respiratory tract, burn wound, urinary tract and eye. Importance of virulence factors produced by P. Aeruginosa during infections such as keratitis, burn wound and respiratory tract is known. The present study was designed to understand the importance of different virulence factors of P. aeruginosa in urinary tract infection in vivo. An ascending urinary tract infection model was established in mice using standard parent strain PAO1 and its isogenic mutant, JP2. Mice were sacrificed at different time intervals and renal tissue homogenates were used for estimation of renal bacterial load and virulence factors. Both parent and mutant strains were able to reach the renal tissue. PAO 1 PAO1 was isolated from renal tissue till day 5 post-infection. However, the mutant strain was unable to colonise the renal tissue. Failure of mutant strain to colonise was attributed to its inability to produce protease, elastase and rhamnolipid. This study suggests that protease, elastase and rhamnolipid contribute to pathogenesis and survival of P. aeruginosa during urinary tract infection.

  14. Terpenoids from Platostoma rotundifolium (Briq.) A. J. Paton Alter the Expression of Quorum Sensing-Related Virulence Factors and the Formation of Biofilm in Pseudomonas aeruginosa PAO1

    PubMed Central

    Rasamiravaka, Tsiry; Ngezahayo, Jérémie; Pottier, Laurent; Oliveira Ribeiro, Sofia; Souard, Florence; Hari, Léonard; Stévigny, Caroline; El Jaziri, Mondher; Duez, Pierre

    2017-01-01

    Platostoma rotundifolium (Briq.) A. J. Paton aerial parts are widely used in Burundi traditional medicine to treat infectious diseases. In order to investigate their probable antibacterial activities, crude extracts from P. rotundifolium were assessed for their bactericidal and anti-virulence properties against an opportunistic bacterial model, Pseudomonas aeruginosa PAO1. Whereas none of the tested extracts exert bacteriostatic and/or bactericidal proprieties, the ethyl acetate and dichloromethane extracts exhibit anti-virulence properties against Pseudomonas aeruginosa PAO1 characterized by an alteration in quorum sensing gene expression and biofilm formation without affecting bacterial viability. Bioguided fractionation of the ethyl acetate extract led to the isolation of major anti-virulence compounds that were identified from nuclear magnetic resonance and high-resolution molecular spectroscopy spectra as cassipourol, β-sitosterol and α-amyrin. Globally, cassipourol and β-sitosterol inhibit quorum sensing-regulated and -regulatory genes expression in las and rhl systems without affecting the global regulators gacA and vfr, whereas α-amyrin had no effect on the expression of these genes. These terpenoids disrupt the formation of biofilms at concentrations down to 12.5, 50 and 50 µM for cassipourol, β-sitosterol and α-amyrin, respectively. Moreover, these terpenoids reduce the production of total exopolysaccharides and promote flagella-dependent motilities (swimming and swarming). The isolated terpenoids exert a wide range of inhibition processes, suggesting a complex mechanism of action targeting P. aeruginosa virulence mechanisms which support the wide anti-infectious use of this plant species in traditional Burundian medicine. PMID:28613253

  15. A Drug-Repositioning Screening Identifies Pentetic Acid as a Potential Therapeutic Agent for Suppressing the Elastase-Mediated Virulence of Pseudomonas aeruginosa

    PubMed Central

    Gi, Mia; Jeong, Junhui; Lee, Keehoon; Lee, Kang-Mu; Toyofuku, Masanori; Yong, Dong Eun

    2014-01-01

    Pseudomonas aeruginosa, a Gram-negative bacterium of clinical significance, produces elastase as a predominant exoprotease. Here, we screened a library of chemical compounds currently used for human medication and identified diethylene triamine penta-acetic acid (DTPA, pentetic acid) as an agent that suppresses the production of elastase. Elastase activity found in the prototype P. aeruginosa strain PAO1 was significantly decreased when grown with a concentration as low as 20 μM DTPA. Supplementation with Zn2+ or Mn2+ ions restored the suppressive effect of DTPA, suggesting that the DTPA-mediated decrease in elastase activity is associated with ion-chelating activity. In DTPA-treated PAO1 cells, transcription of the elastase-encoding lasB gene and levels of the Pseudomonas quinolone signal (PQS), a molecule that mediates P. aeruginosa quorum sensing (QS), were significantly downregulated, reflecting the potential involvement of the PQS QS system in DTPA-mediated elastase suppression. Biofilm formation was also decreased by DTPA treatment. When A549 alveolar type II-like adenocarcinoma cells were infected with PAO1 cells in the presence of DTPA, A549 cell viability was substantially increased. Furthermore, the intranasal delivery of DTPA to PAO1-infected mice alleviated the pathogenic effects of PAO1 cells in the animals. Together, our results revealed a novel function for a known molecule that may help treat P. aeruginosa airway infection. PMID:25246397

  16. Quorum Sensing Down-Regulation Counteracts the Negative Impact of Pseudomonas aeruginosa on CFTR Channel Expression, Function and Rescue in Human Airway Epithelial Cells

    PubMed Central

    Maillé, Émilie; Ruffin, Manon; Adam, Damien; Messaoud, Hatem; Lafayette, Shantelle L.; McKay, Geoffrey; Nguyen, Dao; Brochiero, Emmanuelle

    2017-01-01

    The function of cystic fibrosis transmembrane conductance regulator (CFTR) channels is crucial in human airways. However unfortunately, chronic Pseudomonas aeruginosa infection has been shown to impair CFTR proteins in non-CF airway epithelial cells (AEC) and to alter the efficiency of new treatments with CFTR modulators designed to correct the basic CFTR default in AEC from cystic fibrosis (CF) patients carrying the F508del mutation. Our aim was first to compare the effect of laboratory strains, clinical isolates, engineered and natural mutants to determine the role of the LasR quorum sensing system in CFTR impairment, and second, to test the efficiency of a quorum sensing inhibitor to counteract the deleterious impact of P. aeruginosa both on wt-CFTR and on the rescue of F508del-CFTR by correctors. We first report that exoproducts from either the laboratory PAO1 strain or a clinical ≪Early≫ isolate (from an early stage of infection) altered CFTR expression, localization and function in AEC expressing wt-CFTR. Genetic inactivation of the quorum-sensing LasR in PAO1 (PAO1ΔlasR) or in a natural clinical mutant (≪Late≫ CF-adapted clinical isolate) abolished wt-CFTR impairment. PAO1 exoproducts also dampened F508del-CFTR rescue by VRT-325 or Vx-809 correctors in CF cells, whereas PAO1ΔlasR had no impact. Importantly, treatment of P. aeruginosa cultures with a quorum sensing inhibitor (HDMF) prevented the negative effect of P. aeruginosa exoproducts on wt-CFTR and preserved CFTR rescue by correctors in CF AEC. These findings indicate that LasR-interfering strategies could be of benefits to counteract the deleterious effect of P. aeruginosa in infected patients. PMID:29177135

  17. Multiple roles of Pseudomonas aeruginosa TBCF10839 PilY1 in motility, transport and infection

    PubMed Central

    Bohn, Yu-Sing Tammy; Brandes, Gudrun; Rakhimova, Elza; Horatzek, Sonja; Salunkhe, Prabhakar; Munder, Antje; van Barneveld, Andrea; Jordan, Doris; Bredenbruch, Florian; Häußler, Susanne; Riedel, Kathrin; Eberl, Leo; Jensen, Peter Østrup; Bjarnsholt, Thomas; Moser, Claus; Hoiby, Niels; Tümmler, Burkhard; Wiehlmann, Lutz

    2008-01-01

    Polymorphonuclear neutrophils are the most important mammalian host defence cells against infections with Pseudomonas aeruginosa. Screening of a signature tagged mutagenesis library of the non-piliated P. aeruginosa strain TBCF10839 uncovered that transposon inactivation of its pilY1 gene rendered the bacterium more resistant against killing by neutrophils than the wild type and any other of the more than 3000 tested mutants. Inactivation of pilY1 led to the loss of twitching motility in twitching-proficient wild-type PA14 and PAO1 strains, predisposed to autolysis and impaired the secretion of quinolones and pyocyanin, but on the other hand promoted growth in stationary phase and bacterial survival in murine airway infection models. The PilY1 population consisted of a major full-length and a minor shorter PilY1* isoform. PilY1* was detectable in small extracellular quinolone-positive aggregates, but not in the pilus. P. aeruginosa PilY1 is not an adhesin on the pilus tip, but assists in pilus biogenesis, twitching motility, secretion of secondary metabolites and in the control of cell density in the bacterial population. PMID:19054330

  18. Secretome of transmissible Pseudomonas aeruginosa AES-1R grown in a cystic fibrosis lung-like environment.

    PubMed

    Scott, Nichollas E; Hare, Nathan J; White, Melanie Y; Manos, Jim; Cordwell, Stuart J

    2013-12-06

    Pseudomonas aeruginosa is the predominant cause of mortality in patients with cystic fibrosis (CF). We examined the secretome of an acute, transmissible CF P. aeruginosa (Australian epidemic strain 1-R; AES-1R) compared with laboratory-adapted PAO1. Culture supernatant proteins from rich (LB) and minimal (M9) media were compared using 2-DE and 2DLC-MS/MS, which revealed elevated abundance of PasP protease and absence of AprA protease in AES-1R. CF lung-like artificial sputum medium (ASMDM) contains serum and mucin that generally preclude proteomics of secreted proteins. ASMDM culture supernatants were subjected to 2DLC-MS/MS, which allowed the identification of 57 P. aeruginosa proteins, and qualitative spectral counting was used to estimate relative abundance. AES-1R-specific AES_7139 and PasP were more abundant in AES-1R ASMDM culture supernatants, while AprA could only be identified in PAO1. Relative quantitation was performed using selected reaction monitoring. Significantly elevated levels of PasP, LasB, chitin-binding protein (CbpD), and PA4495 were identified in AES-1R ASMDM supernatants. Quantitative PCR showed elevated pasP in AES-1R during early (18 h) ASMDM growth, while no evidence of aprA expression could be observed. Genomic screening of CF isolates revealed aes_7139 was present in all AES-1 and one pair of sequential nonepidemic isolates. Secreted proteins may be crucial in aiding CF-associated P. aeruginosa to establish infection and for adaptation to the CF lung.

  19. Anti-quorum sensing activity of Pistacia atlantica against Pseudomonas aeruginosa PAO1 and identification of its bioactive compounds.

    PubMed

    Kordbacheh, H; Eftekhar, F; Ebrahimi, S N

    2017-09-01

    Pseudomonas aeruginosa is a multidrug resistant opportunistic pathogen and an important cause of nosocomial infections. Quorum-sensing (QS) is a process in which bacterial cell-cell communication can regulates production of many virulence factors including pigment formation and the ability to form biofilm which is essential for establishment of chronic infections. We examined the inhibitory effect of Pistacia atlantica (Anacardiaceae) methanolic leaf extract and its bioactive components on biofilm formation and pigment production by P. aeruginosa PAO1. Fractionation of the methanolic leaf extract was carried out using HPLC based activity profiling. Identification of the active compounds was carried out by the integrated approach of HPLC-DAD and LC-MS followed by molecular docking analysis. Pistacia atlantica crude extract at 2 and 1 mg/mL, inhibited 92% and 79% biofilm formation, respectively. Minimum biofilm inhibitory concentration (MBIC) determined by microbroth dilution was 0.25 mg/mL with 39% inhibition. Pyocyanin production measured by spectrophotometry showed 100% and 83% inhibition at 2 and 1 mg/mL and minimum inhibitory concentration (MIC) was 0.5 mg/mL with 40% inhibition. Four active HPLC fractions (11, 15, 16 and 19) showed MBIC values of 0.06, 0.16, 0.10, 0.15 mg/mL, and MICs for pyocyanin production of 0.49, 0.31, 0.76, >0.30 mg/mL, respectively. The active compounds were identified as rutin (1), myricetin, 3-O-rutinoside (2) and kaempferol-3-O-rutinoside (4), all belonging to the flavonoid family. Molecular docking simulation of the active compounds showed that all had high affinity for LasR protein which is an important quorum-sensing signal receptor. The results of this study suggest that the active components of P. atlantica have high anti-QS activities and may have the potential for treatment of chronic infections caused by Pseudomonas aeruginosa. Copyright © 2017 Elsevier Ltd. All rights reserved.

  20. High β-Lactamase Levels Change the Pharmacodynamics of β-Lactam Antibiotics in Pseudomonas aeruginosa Biofilms

    PubMed Central

    Ciofu, Oana; Yang, Liang; Wu, Hong; Song, Zhijun; Oliver, Antonio; Høiby, Niels

    2013-01-01

    Resistance to β-lactam antibiotics is a frequent problem in Pseudomonas aeruginosa lung infection of cystic fibrosis (CF) patients. This resistance is mainly due to the hyperproduction of chromosomally encoded β-lactamase and biofilm formation. The purpose of this study was to investigate the role of β-lactamase in the pharmacokinetics (PK) and pharmacodynamics (PD) of ceftazidime and imipenem on P. aeruginosa biofilms. P. aeruginosa PAO1 and its corresponding β-lactamase-overproducing mutant, PAΔDDh2Dh3, were used in this study. Biofilms of these two strains in flow chambers, microtiter plates, and on alginate beads were treated with different concentrations of ceftazidime and imipenem. The kinetics of antibiotics on the biofilms was investigated in vitro by time-kill methods. Time-dependent killing of ceftazidime was observed in PAO1 biofilms, but concentration-dependent killing activity of ceftazidime was observed for β-lactamase-overproducing biofilms of P. aeruginosa in all three models. Ceftazidime showed time-dependent killing on planktonic PAO1 and PAΔDDh2Dh3. This difference is probably due to the special distribution and accumulation in the biofilm matrix of β-lactamase, which can hydrolyze the β-lactam antibiotics. The PK/PD indices of the AUC/MBIC and Cmax/MBIC (AUC is the area under concentration-time curve, MBIC is the minimal biofilm-inhibitory concentration, and Cmax is the maximum concentration of drug in serum) are probably the best parameters to describe the effect of ceftazidime in β-lactamase-overproducing P. aeruginosa biofilms. Meanwhile, imipenem showed time-dependent killing on both PAO1 and PAΔDDh2Dh3 biofilms. An inoculum effect of β-lactams was found for both planktonic and biofilm P. aeruginosa cells. The inoculum effect of ceftazidime for the β-lactamase-overproducing mutant PAΔDDh2Dh3 biofilms was more obvious than for PAO1 biofilms, with a requirement of higher antibiotic concentration and a longer period of treatment

  1. Pharmacodynamic Evaluation of the Intracellular Activity of Antibiotics towards Pseudomonas aeruginosa PAO1 in a Model of THP-1 Human Monocytes

    PubMed Central

    Buyck, Julien M.; Tulkens, Paul M.

    2013-01-01

    Pseudomonas aeruginosa invades epithelial and phagocytic cells, which may play an important role in the persistence of infection. We have developed a 24-h model of THP-1 monocyte infection with P. aeruginosa PAO1 in which bacteria are seen multiplying in vacuoles by electron microscopy. The model has been used to quantitatively assess antibiotic activity against intracellular and extracellular bacteria by using a pharmacodynamic approach (concentration-dependent experiments over a wide range of extracellular concentrations to calculate bacteriostatic concentrations [Cs] and maximal relative efficacies [Emax]; Hill-Langmuir equation). Using 16 antipseudomonal antibiotics (three aminoglycosides, nine β-lactams, three fluoroquinolones, and colistin), dose-response curves were found to be undistinguishable for antibiotics of the same pharmacological class if data were expressed as a function of the corresponding MICs. Extracellularly, all of the antibiotics reached a bacteriostatic effect at their MIC, and their Emax exceeded the limit of detection (−4.5 log10 CFU compared to the initial inoculum). Intracellularly, Cs values remained unchanged for β-lactams, fluoroquinolones, and colistin but were approximately 10 times higher for aminoglycosides, whereas Emax values were markedly reduced (less negative), reaching −3 log10 CFU for fluoroquinolones and only −1 to −1.5 log10 CFU for all other antibiotics. The decrease in intracellular aminoglycoside potency (higher Cs) can be ascribed to the acid pH to which bacteria are exposed in vacuoles. The decrease in the Emax may reflect a reversible alteration of bacterial responsiveness to antibiotics in the intracellular milieu. The model may prove useful for comparison of antipseudomonal antibiotics to reduce the risk of persistence or relapse of pseudomonal infections. PMID:23478951

  2. Pseudomonas aeruginosa AES-1 exhibits increased virulence gene expression during chronic infection of cystic fibrosis lung.

    PubMed

    Naughton, Sharna; Parker, Dane; Seemann, Torsten; Thomas, Torsten; Turnbull, Lynne; Rose, Barbara; Bye, Peter; Cordwell, Stuart; Whitchurch, Cynthia; Manos, Jim

    2011-01-01

    Pseudomonas aeruginosa, the leading cause of morbidity and mortality in people with cystic fibrosis (CF), adapts for survival in the CF lung through both mutation and gene expression changes. Frequent clonal strains such as the Australian Epidemic Strain-1 (AES-1), have increased ability to establish infection in the CF lung and to superimpose and replace infrequent clonal strains. Little is known about the factors underpinning these properties. Analysis has been hampered by lack of expression array templates containing CF-strain specific genes. We sequenced the genome of an acute infection AES-1 isolate from a CF infant (AES-1R) and constructed a non-redundant micro-array (PANarray) comprising AES-1R and seven other sequenced P. aeruginosa genomes. The unclosed AES-1R genome comprised 6.254Mbp and contained 6957 putative genes, including 338 not found in the other seven genomes. The PANarray contained 12,543 gene probe spots; comprising 12,147 P. aeruginosa gene probes, 326 quality-control probes and 70 probes for non-P. aeruginosa genes, including phage and plant genes. We grew AES-1R and its isogenic pair AES-1M, taken from the same patient 10.5 years later and not eradicated in the intervening period, in our validated artificial sputum medium (ASMDM) and used the PANarray to compare gene expression of both in duplicate. 675 genes were differentially expressed between the isogenic pairs, including upregulation of alginate, biofilm, persistence genes and virulence-related genes such as dihydroorotase, uridylate kinase and cardiolipin synthase, in AES-1M. Non-PAO1 genes upregulated in AES-1M included pathogenesis-related (PAGI-5) genes present in strains PACS2 and PA7, and numerous phage genes. Elucidation of these genes' roles could lead to targeted treatment strategies for chronically infected CF patients.

  3. Pseudomonas aeruginosa AES-1 Exhibits Increased Virulence Gene Expression during Chronic Infection of Cystic Fibrosis Lung

    PubMed Central

    Naughton, Sharna; Parker, Dane; Seemann, Torsten; Thomas, Torsten; Turnbull, Lynne; Rose, Barbara; Bye, Peter; Cordwell, Stuart; Whitchurch, Cynthia; Manos, Jim

    2011-01-01

    Pseudomonas aeruginosa, the leading cause of morbidity and mortality in people with cystic fibrosis (CF), adapts for survival in the CF lung through both mutation and gene expression changes. Frequent clonal strains such as the Australian Epidemic Strain-1 (AES-1), have increased ability to establish infection in the CF lung and to superimpose and replace infrequent clonal strains. Little is known about the factors underpinning these properties. Analysis has been hampered by lack of expression array templates containing CF-strain specific genes. We sequenced the genome of an acute infection AES-1 isolate from a CF infant (AES-1R) and constructed a non-redundant micro-array (PANarray) comprising AES-1R and seven other sequenced P. aeruginosa genomes. The unclosed AES-1R genome comprised 6.254Mbp and contained 6957 putative genes, including 338 not found in the other seven genomes. The PANarray contained 12,543 gene probe spots; comprising 12,147 P. aeruginosa gene probes, 326 quality-control probes and 70 probes for non-P. aeruginosa genes, including phage and plant genes. We grew AES-1R and its isogenic pair AES-1M, taken from the same patient 10.5 years later and not eradicated in the intervening period, in our validated artificial sputum medium (ASMDM) and used the PANarray to compare gene expression of both in duplicate. 675 genes were differentially expressed between the isogenic pairs, including upregulation of alginate, biofilm, persistence genes and virulence-related genes such as dihydroorotase, uridylate kinase and cardiolipin synthase, in AES-1M. Non-PAO1 genes upregulated in AES-1M included pathogenesis-related (PAGI-5) genes present in strains PACS2 and PA7, and numerous phage genes. Elucidation of these genes' roles could lead to targeted treatment strategies for chronically infected CF patients. PMID:21935417

  4. [TL, the new bacteriophage of Pseudomonas aeruginosa and its application for the search of halo-producing bacteriophages].

    PubMed

    Pleteneva, E A; Burkal'tseva, M V; Shaburova, O V; Krylov, S V; Pechnikova, E V; Sokolova, O S; Krylov, V N

    2011-01-01

    The properties of new virulent bacteriophage TL of Pseudomonas aeruginosa belonging to the family Podoviridae (genome size of 46 kb) were investigated. This bacteriophage is capable of lysogenizing the bacterial lawn in halo zones around negative colonies (NC) of other bacteriophages. TL forms large NC, that are hardly distinguishable on the lawn of P. aeruginisa PAO1. At the same time, on the lawns of some phage-resistant PAO1 mutants, as well as on those produced by a number of clinical isolates, TL forms more transparent NC. It is suggested that more effective growth of the bacteriophage TL NC is associated with the differences in outer lipopolysaccharide (LPS) layer of the cell walls of different bacterial strains, as well as of the bacteria inside and outside of the halos. This TL property was used to optimize selection of bacteriophages producing halos around NC on the lawn of P. aeruginosa PAO1. As a result, a group of bacteriophages differing in the patterns of interaction between their halos and TL bacteriophage, as well as in some characters was identified. Taking into consideration the importance of cell-surfaced structures of P. aeruginosa in manifestation of virulence and pathogenicity, possible utilization of specific phage enzymes, polysacchadide depolymerases, for more effective treatment of P. aeruginosa infections is discussed.

  5. l-Methionine anti-biofilm activity against Pseudomonas aeruginosa is enhanced by the cystic fibrosis transmembrane conductance regulator potentiator, ivacaftor.

    PubMed

    Cho, Do-Yeon; Lim, Dong-Jin; Mackey, Calvin; Weeks, Christopher G; Peña Garcia, Jaime A; Skinner, Daniel; Grayson, Jessica W; Hill, Harrison S; Alexander, David K; Zhang, Shaoyan; Woodworth, Bradford A

    2018-05-01

    Biofilms may contribute to refractory chronic rhinosinusitis (CRS), as they lead to antibiotic resistance and failure of effective clinical treatment. l-Methionine is an amino acid with reported biofilm-inhibiting properties. Ivacaftor is a cystic fibrosis transmembrane conductance regulator (CFTR) potentiator with mild antimicrobial activity via inhibition of bacterial DNA gyrase and topoisomerase IV. The objective of this study was to evaluate whether co-treatment with ivacaftor and l-methionine can reduce the formation of Pseudomonas aeruginosa biofilms. P aeruginosa (PAO-1 strain) biofilms were studied in the presence of l-methionine and/or ivacaftor. For static biofilm assays, PAO-1 was cultured in a 48-well plate for 72 hours with stepwise combinations of these agents. Relative biofilm inhibitions were measured according to optical density of crystal violet stain at 590 nm. Live/dead assays (BacTiter-Glo™ assay, Promega) were imaged with laser scanning confocal microscopy. An agar diffusion test was used to confirm antibacterial effects of the drugs. l-Methionine (0.5 μM) significantly reduced PAO-1 biofilm mass (32.4 ± 18.0%; n = 4; p < 0.001) compared with controls. Low doses of ivacaftor alone (4, 8, and 12 μg/mL) had no effect on biofilm formation. When combined with ivacaftor (4 μg/mL), a synergistic anti-biofilm effect was noted at 0.05 μM and 0.5 μM of l-methionine (two-way analysis of variane, p = 0.0415) compared with corresponding concentrations of l-methionine alone. Ivacaftor enhanced the anti-biofilm activity of l-methionine against the PAO-1 strain of P aeruginosa. Further studies evaluating the efficacy of ivacaftor/l-methionine combinations for P aeruginosa sinusitis are planned. © 2018 ARS-AAOA, LLC.

  6. Temperature-Dependent Expression of phzM and Its Regulatory Genes lasI and ptsP in Rhizosphere Isolate Pseudomonas sp. Strain M18▿

    PubMed Central

    Huang, Jiaofang; Xu, Yuquan; Zhang, Hongyan; Li, Yaqian; Huang, Xianqing; Ren, Bin; Zhang, Xuehong

    2009-01-01

    Pseudomonas sp. strain M18, an effective biological control agent isolated from the melon rhizosphere, has a genetic background similar to that of the opportunistic human pathogen Pseudomonas aeruginosa PAO1. However, the predominant phenazine produced by strain M18 is phenazine-1-carboxylic acid (PCA) rather than pyocyanin (PYO); the quantitative ratio of PCA to PYO is 105 to 1 at 28°C in strain M18, while the ratio is 1 to 2 at 37°C in strain PAO1. We first provided evidence that the differential production of the two phenazines in strains M18 and PAO1 is related to the temperature-dependent and strain-specific expression patterns of phzM, a gene involved in the conversion of PCA to PYO. Transcriptional levels of phzM were measured by quantitative real-time PCR, and the activities of both transcriptional and translational phzM′-′lacZ fusions were determined in strains M18 and PAO1, respectively. Using lasI::Gm and ptsP::Gm inactivation M18 mutants, we further show that expression of the phzM gene is positively regulated by the quorum-sensing protein LasI and negatively regulated by the phosphoenolpyruvate phosphotransferase protein PtsP. Surprisingly, the lasI and ptsP regulatory genes were also expressed in a temperature-dependent and strain-specific manner. The differential production of the phenazines PCA and PYO by strains M18 and PAO1 may be a consequence of selective pressure imposed on P. aeruginosa PAO1 and its relative M18 in the two different niches over a long evolutionary process. PMID:19717631

  7. A plasmid-encoded UmuD homologue regulates expression of Pseudomonas aeruginosa SOS genes.

    PubMed

    Díaz-Magaña, Amada; Alva-Murillo, Nayeli; Chávez-Moctezuma, Martha P; López-Meza, Joel E; Ramírez-Díaz, Martha I; Cervantes, Carlos

    2015-07-01

    The Pseudomonas aeruginosa plasmid pUM505 contains the umuDC operon that encodes proteins similar to error-prone repair DNA polymerase V. The umuC gene appears to be truncated and its product is probably not functional. The umuD gene, renamed umuDpR, possesses an SOS box overlapped with a Sigma factor 70 type promoter; accordingly, transcriptional fusions revealed that the umuDpR gene promoter is activated by mitomycin C. The predicted sequence of the UmuDpR protein displays 23 % identity with the Ps. aeruginosa SOS-response LexA repressor. The umuDpR gene caused increased MMC sensitivity when transferred to the Ps. aeruginosa PAO1 strain. As expected, PAO1-derived knockout lexA-  mutant PW6037 showed resistance to MMC; however, when the umuDpR gene was transferred to PW6037, MMC resistance level was reduced. These data suggested that UmuDpR represses the expression of SOS genes, as LexA does. To test whether UmuDpR exerts regulatory functions, expression of PAO1 SOS genes was evaluated by reverse transcription quantitative PCR assays in the lexA-  mutant with or without the pUC_umuD recombinant plasmid. Expression of lexA, imuA and recA genes increased 3.4-5.3 times in the lexA-  mutant, relative to transcription of the corresponding genes in the lexA+ strain, but decreased significantly in the lexA- /umuDpR transformant. These results confirmed that the UmuDpR protein is a repressor of Ps. aeruginosa SOS genes controlled by LexA. Electrophoretic mobility shift assays, however, did not show binding of UmuDpR to 5' regions of SOS genes, suggesting an indirect mechanism of regulation.

  8. A Pseudomonas aeruginosa EF-Hand Protein, EfhP (PA4107), Modulates Stress Responses and Virulence at High Calcium Concentration

    PubMed Central

    Sarkisova, Svetlana A.; Lotlikar, Shalaka R.; Guragain, Manita; Kubat, Ryan; Cloud, John

    2014-01-01

    Pseudomonas aeruginosa is a facultative human pathogen, and a major cause of nosocomial infections and severe chronic infections in endocarditis and in cystic fibrosis (CF) patients. Calcium (Ca2+) accumulates in pulmonary fluids of CF patients, and plays a role in the hyperinflamatory response to bacterial infection. Earlier we showed that P. aeruginosa responds to increased Ca2+ levels, primarily through the increased production of secreted virulence factors. Here we describe the role of putative Ca2+-binding protein, with an EF-hand domain, PA4107 (EfhP), in this response. Deletion mutations of efhP were generated in P. aeruginosa strain PAO1 and CF pulmonary isolate, strain FRD1. The lack of EfhP abolished the ability of P. aeruginosa PAO1 to maintain intracellular Ca2+ homeostasis. Quantitative high-resolution 2D-PAGE showed that the efhP deletion also affected the proteomes of both strains during growth with added Ca2+. The greatest proteome effects occurred when the pulmonary isolate was cultured in biofilms. Among the proteins that were significantly less abundant or absent in the mutant strains were proteins involved in iron acquisition, biosynthesis of pyocyanin, proteases, and stress response proteins. In support, the phenotypic responses of FRD1 ΔefhP showed that the mutant strain lost its ability to produce pyocyanin, developed less biofilm, and had decreased resistance to oxidative stress (H2O2) when cultured at high [Ca2+]. Furthermore, the mutant strain was unable to produce alginate when grown at high [Ca2+] and no iron. The effect of the ΔefhP mutations on virulence was determined in a lettuce model of infection. Growth of wild-type P. aeruginosa strains at high [Ca2+] causes an increased area of disease. In contrast, the lack of efhP prevented this Ca2+-induced increase in the diseased zone. The results indicate that EfhP is important for Ca2+ homeostasis and virulence of P. aeruginosa when it encounters host environments with high [Ca2+]. PMID

  9. Identification of essential genes of Pseudomonas aeruginosa for its growth in airway mucus.

    PubMed

    Alrahman, Mohammed Abd; Yoon, Sang Sun

    2017-01-01

    Pseudomonas aeruginosa has been identified as an important causative agent of airway infection, mainly in cystic fibrosis. This disease is characterized by defective mucociliary clearance induced in part by mucus hyper-production. Mucin is a major component of airway mucus and is heavily O-glycosylated, with a protein backbone. Airway infection is known to be established with bacterial adhesion to mucin. However, the genes involved in mucin degradation or utilization remain elusive. In this study, we sought to provide a genetic basis of P. aeruginosa airway growth by identifying those genes. First, using RNASeq analyses, we compared genome-wide expression profiles of PAO1, a prototype P. aeruginosa laboratory strain, grown in M9-mucin (M9M) and M9-glucose (M9G) media. Additionally, a PAO1 transposon (Tn) insertion mutants library was screened for mutants defective in growth in M9M medium. One mutant with a Tn insertion in the xcpU gene (PA3100) was determined to exhibit faulty growth in M9M medium. This gene contributes to the type II secretion system, suggesting that P. aeruginosa uses this secretion system to produce a number of proteins to break down and assimilate the mucin molecule. Furthermore, we screened the PAO1 genome for genes with protease activity. Of 13 mutants, one with mutation in PA3247 gene exhibited defective growth in M9M, suggesting that the PA3247-encoded protease plays a role in mucin utilization. Further mechanistic dissection of this particular process will reveal new drug targets, the inhibition of which could control recalcitrant P. aeruginosa infections.

  10. Contribution of Cell Elongation to the Biofilm Formation of Pseudomonas aeruginosa during Anaerobic Respiration

    PubMed Central

    Park, Yongjin; Yoon, Sang Sun

    2011-01-01

    Pseudomonas aeruginosa, a gram-negative bacterium of clinical importance, forms more robust biofilm during anaerobic respiration, a mode of growth presumed to occur in abnormally thickened mucus layer lining the cystic fibrosis (CF) patient airway. However, molecular basis behind this anaerobiosis-triggered robust biofilm formation is not clearly defined yet. Here, we identified a morphological change naturally accompanied by anaerobic respiration in P. aeruginosa and investigated its effect on the biofilm formation in vitro. A standard laboratory strain, PAO1 was highly elongated during anaerobic respiration compared with bacteria grown aerobically. Microscopic analysis demonstrated that cell elongation likely occurred as a consequence of defective cell division. Cell elongation was dependent on the presence of nitrite reductase (NIR) that reduces nitrite (NO2 −) to nitric oxide (NO) and was repressed in PAO1 in the presence of carboxy-PTIO, a NO antagonist, demonstrating that cell elongation involves a process to respond to NO, a spontaneous byproduct of the anaerobic respiration. Importantly, the non-elongated NIR-deficient mutant failed to form biofilm, while a mutant of nitrate reductase (NAR) and wild type PAO1, both of which were highly elongated, formed robust biofilm. Taken together, our data reveal a role of previously undescribed cell biological event in P. aeruginosa biofilm formation and suggest NIR as a key player involved in such process. PMID:21267455

  11. Characterization of mechanisms of quinolone resistance in Pseudomonas aeruginosa strains isolated in vitro and in vivo during experimental endocarditis.

    PubMed Central

    Chamberland, S; Bayer, A S; Schollaardt, T; Wong, S A; Bryan, L E

    1989-01-01

    Mechanisms of resistance to quinolones were characterized in Pseudomonas aeruginosa strains isolated after Tn5 insertional mutagenesis and in resistant strains that emerged during pefloxacin therapy of experimental aortic endocarditis. Quinolone resistance achieved in in vitro-selected mutants Qr-1 and Qr-2 was associated with cross-resistance to several groups of antimicrobial agents, including beta-lactams, tetracycline, and chloramphenicol. A significant reduction of norfloxacin uptake was also observed. After ether permeabilization of the cells, DNA synthesis of these two isolates was as susceptible to norfloxacin as DNA synthesis of the parent strain (PAO1). These results indicate that alteration of outer membrane permeability is the primary determinant of resistance in these isolates. This altered cell permeability was correlated with reduction of outer membrane protein G (25.5 kilodaltons) and loss of a 40-kilodalton outer membrane protein in strain Qr-1. Resistance to quinolones that emerged during experimental endocarditis therapy was associated with both modification of outer membrane permeability (decreased uptake of norfloxacin) and decreased susceptibility of DNA synthesis to norfloxacin. Resistance was limited to quinolones and chloramphenicol. For these strains, norfloxacin inhibitory doses (50%) for DNA synthesis were identical to the drug MICs, suggesting that despite the identification of a permeability change, perhaps due to changes of lipopolysaccharide, the alteration of the quinolone intracellular target(s) susceptibility constitutes the primary determinant of resistance. Also, two distinct levels of norfloxacin resistance of DNA synthesis were found in these isolates, indicating that at least two distinct alterations of the drug target(s) are possible in P. aeruginosa. Images PMID:2502066

  12. TonB-Dependent Receptor Repertoire of Pseudomonas aeruginosa for Uptake of Siderophore-Drug Conjugates.

    PubMed

    Luscher, Alexandre; Moynié, Lucile; Auguste, Pamela Saint; Bumann, Dirk; Mazza, Lena; Pletzer, Daniel; Naismith, James H; Köhler, Thilo

    2018-06-01

    The conjugation of siderophores to antimicrobial molecules is an attractive strategy to overcome the low outer membrane permeability of Gram-negative bacteria. In this Trojan horse approach, the transport of drug conjugates is redirected via TonB-dependent receptors (TBDR), which are involved in the uptake of essential nutrients, including iron. Previous reports have demonstrated the involvement of the TBDRs PiuA and PirA from Pseudomonas aeruginosa and their orthologues in Acinetobacter baumannii in the uptake of siderophore-beta-lactam drug conjugates. By in silico screening, we further identified a PiuA orthologue, termed PiuD, present in clinical isolates, including strain LESB58. The piuD gene in LESB58 is located at the same genetic locus as piuA in strain PAO1. PiuD has a similar crystal structure as PiuA and is involved in the transport of the siderophore-drug conjugates BAL30072, MC-1, and cefiderocol in strain LESB58. To screen for additional siderophore-drug uptake systems, we overexpressed 28 of the 34 TBDRs of strain PAO1 and identified PfuA, OptE, OptJ, and the pyochelin receptor FptA as novel TBDRs conferring increased susceptibility to siderophore-drug conjugates. The existence of a TBDR repertoire in P. aeruginosa able to transport siderophore-drug molecules potentially decreases the likelihood of resistance emergence during therapy. Copyright © 2018 Luscher et al.

  13. Carbapenem-resistant Pseudomonas aeruginosa strains from a Spanish hospital: characterization of metallo-beta-lactamases, porin OprD and integrons.

    PubMed

    Rojo-Bezares, Beatriz; Estepa, Vanesa; Cebollada, Rocío; de Toro, María; Somalo, Sergio; Seral, Cristina; Castillo, Francisco Javier; Torres, Carmen; Sáenz, Yolanda

    2014-05-01

    Molecular typing and mechanisms of carbapenem resistance such as alterations in porin OprD and presence of metallo-beta-lactamases (MBLs), as well as integrons have been studied in a collection of carbapenem-resistant Pseudomonas aeruginosa (CRPA) isolates from a Spanish hospital. One hundred and twenty-three CRPA isolates were recovered from different samples of 80 patients. Clonal relationship among CRPA was analyzed by SpeI-PFGE. Susceptibility testing to 11 antibiotics and MBL phenotype was determined by microdilution, IP/IPI E-test and double disc method. The oprD gene was studied by PCR and sequencing, and mutations were determined comparing with P. aeruginosa PAO1 sequence. Characterization of MBLs, and class 1 and 2 integrons were studied by PCR and sequencing. SDS-PAGE analysis of outer membrane proteins of selected strains was performed. Seventy-four-per-cent of patients with CRPA were hospitalised in the ICU setting and 50% had long hospitalization stays. Sixty-four different PFGE patterns were detected, and 87 CRPA strains were further analyzed. MBL phenotype was detected in 43 of 87 strains (49.4%), which contained blaVIM-2 gene inside class 1 integrons. VIM-2-producing strains belonged to lineages ST175, ST235, and ST973. A great diversity of nucleotide insertions, deletions, and mutations in oprD gene, and the presence of a new insertion sequence (ISPa45) truncating oprD were identified among CRPA strains. Class 1 integrons were detected in 75% of CRPA strains, blaVIM-2 and the new arrangement aac(3)-Ia+ISPa34+aadA1 (named as In661) being the most frequent gene-cassette arrays detected. Other gene cassettes detected in integrons were: aadB, aadA6, aadA7, aac(6')-Ib', and blaOXA-46. Copyright © 2014 Elsevier GmbH. All rights reserved.

  14. Network-assisted investigation of virulence and antibiotic-resistance systems in Pseudomonas aeruginosa

    NASA Astrophysics Data System (ADS)

    Hwang, Sohyun; Kim, Chan Yeong; Ji, Sun-Gou; Go, Junhyeok; Kim, Hanhae; Yang, Sunmo; Kim, Hye Jin; Cho, Ara; Yoon, Sang Sun; Lee, Insuk

    2016-05-01

    Pseudomonas aeruginosa is a Gram-negative bacterium of clinical significance. Although the genome of PAO1, a prototype strain of P. aeruginosa, has been extensively studied, approximately one-third of the functional genome remains unknown. With the emergence of antibiotic-resistant strains of P. aeruginosa, there is an urgent need to develop novel antibiotic and anti-virulence strategies, which may be facilitated by an approach that explores P. aeruginosa gene function in systems-level models. Here, we present a genome-wide functional network of P. aeruginosa genes, PseudomonasNet, which covers 98% of the coding genome, and a companion web server to generate functional hypotheses using various network-search algorithms. We demonstrate that PseudomonasNet-assisted predictions can effectively identify novel genes involved in virulence and antibiotic resistance. Moreover, an antibiotic-resistance network based on PseudomonasNet reveals that P. aeruginosa has common modular genetic organisations that confer increased or decreased resistance to diverse antibiotics, which accounts for the pervasiveness of cross-resistance across multiple drugs. The same network also suggests that P. aeruginosa has developed mechanism of trade-off in resistance across drugs by altering genetic interactions. Taken together, these results clearly demonstrate the usefulness of a genome-scale functional network to investigate pathogenic systems in P. aeruginosa.

  15. Pseudomonas aeruginosa reveals high intrinsic resistance to penem antibiotics: penem resistance mechanisms and their interplay.

    PubMed

    Okamoto, K; Gotoh, N; Nishino, T

    2001-07-01

    Pseudomonas aeruginosa exhibits high intrinsic resistance to penem antibiotics such as faropenem, ritipenem, AMA3176, sulopenem, Sch29482, and Sch34343. To investigate the mechanisms contributing to penem resistance, we used the laboratory strain PAO1 to construct a series of isogenic mutants with an impaired multidrug efflux system MexAB-OprM and/or impaired chromosomal AmpC beta-lactamase. The outer membrane barrier of PAO1 was partially eliminated by inducing the expression of the plasmid-encoded Escherichia coli major porin OmpF. Susceptibility tests using the mutants and the OmpF expression plasmid showed that MexAB-OprM and the outer membrane barrier, but not AmpC beta-lactamase, are the main mechanisms involved in the high intrinsic penem resistance of PAO1. However, reducing the high intrinsic penem resistance of PAO1 to the same level as that of penem-susceptible gram-negative bacteria such as E. coli required the loss of either both MexAB-OprM and AmpC beta-lactamase or both MexAB-OprM and the outer membrane barrier. Competition experiments for penicillin-binding proteins (PBPs) revealed that the affinity of PBP 1b and PBP 2 for faropenem were about 1.8- and 1.5-fold lower, than the respective affinity for imipenem. Loss of the outer membrane barrier, MexAB, and AmpC beta-lactamase increased the susceptibility of PAO1 to almost all penems tested compared to the susceptibility of the AmpC-deficient PAO1 mutants to imipenem. Thus, it is suggested that the high intrinsic penem resistance of P. aeruginosa is generated from the interplay among the outer membrane barrier, the active efflux system, and AmpC beta-lactamase but not from the lower affinity of PBPs for penems.

  16. Pseudomonas aeruginosa Reveals High Intrinsic Resistance to Penem Antibiotics: Penem Resistance Mechanisms and Their Interplay

    PubMed Central

    Okamoto, Kiyomi; Gotoh, Naomasa; Nishino, Takeshi

    2001-01-01

    Pseudomonas aeruginosa exhibits high intrinsic resistance to penem antibiotics such as faropenem, ritipenem, AMA3176, sulopenem, Sch29482, and Sch34343. To investigate the mechanisms contributing to penem resistance, we used the laboratory strain PAO1 to construct a series of isogenic mutants with an impaired multidrug efflux system MexAB-OprM and/or impaired chromosomal AmpC β-lactamase. The outer membrane barrier of PAO1 was partially eliminated by inducing the expression of the plasmid-encoded Escherichia coli major porin OmpF. Susceptibility tests using the mutants and the OmpF expression plasmid showed that MexAB-OprM and the outer membrane barrier, but not AmpC β-lactamase, are the main mechanisms involved in the high intrinsic penem resistance of PAO1. However, reducing the high intrinsic penem resistance of PAO1 to the same level as that of penem-susceptible gram-negative bacteria such as E. coli required the loss of either both MexAB-OprM and AmpC β-lactamase or both MexAB-OprM and the outer membrane barrier. Competition experiments for penicillin-binding proteins (PBPs) revealed that the affinity of PBP 1b and PBP 2 for faropenem were about 1.8- and 1.5-fold lower, than the respective affinity for imipenem. Loss of the outer membrane barrier, MexAB, and AmpC β-lactamase increased the susceptibility of PAO1 to almost all penems tested compared to the susceptibility of the AmpC-deficient PAO1 mutants to imipenem. Thus, it is suggested that the high intrinsic penem resistance of P. aeruginosa is generated from the interplay among the outer membrane barrier, the active efflux system, and AmpC β-lactamase but not from the lower affinity of PBPs for penems. PMID:11408209

  17. Promotion of acute-phase skin wound healing by Pseudomonas aeruginosa C4 -HSL.

    PubMed

    Kanno, Emi; Kawakami, Kazuyoshi; Miyairi, Shinichi; Tanno, Hiromasa; Suzuki, Aiko; Kamimatsuno, Rina; Takagi, Naoyuki; Miyasaka, Tomomitsu; Ishii, Keiko; Gotoh, Naomasa; Maruyama, Ryoko; Tachi, Masahiro

    2016-12-01

    A Pseudomonas aeruginosa quorum-sensing system, which produces N-(3-oxododecanoyl)-l-homoserine lactone (3-oxo-C 12 -HSL) and N-butanoyl-l-homoserine lactone (C 4 -HSL), regulates the virulence factors. In our previous study, 3-oxo-C 12 -HSL, encoded by lasI gene, was shown to promote wound healing. However, the effect of C 4 -HSL, encoded by rhlI gene, remains to be elucidated. We addressed the effect of C 4 -HSL on wounds in P. aeruginosa infection. Wounds were created on the backs of Sprague-Dawley SD rats, and P. aeruginosa PAO1 (PAO1) or its rhlI deletion mutant (ΔrhlI) or lasI deletion mutant (ΔlasI) was inoculated onto the wound. Rats were injected intraperitoneally with anti-C 4 -HSL antiserum or treated with C 4 -HSL at the wound surface. PAO1 inoculation led to significant acceleration of wound healing, which was associated with neutrophil infiltration and TNF-α synthesis. These responses were reversed, except for TNF-α production, when ΔrhlI was inoculated instead of PAO1 or when rats were co-treated with PAO1 and anti-C 4 -HSL antiserum. In contrast, the healing process and neutrophil infiltration, but not TNF-α synthesis, were accelerated when C 4 -HSL was administered in the absence of PAO1. This acceleration was not affected by anti-TNF-α antibody. These results suggest that C 4 -HSL may be involved in the acceleration of acute wound healing in P. aeruginosa infection by modifying the neutrophilic inflammation. © 2015 Medicalhelplines.com Inc and John Wiley & Sons Ltd.

  18. Insertional inactivation of oprD in carbapenem-resistant Pseudomonas aeruginosa strains isolated from burn patients in Tehran, Iran.

    PubMed

    Shariati, A; Azimi, T; Ardebili, A; Chirani, A S; Bahramian, A; Pormohammad, A; Sadredinamin, M; Erfanimanesh, S; Bostanghadiri, N; Shams, S; Hashemi, A

    2018-01-01

    In this study, we report the insertion sequence IS Ppu 21 in the opr D porin gene of carbapenem-resistant Pseudomonas aeruginosa isolates from burn patients in Tehran, Iran. Antibiotic susceptibility tests for P. aeruginosa isolates were determined. Production of metallo-β-lactamases (MBLs) and carbapenemase was evaluated and the β-lactamase-encoding and aminoglycoside-modifying enzyme genes were investigated by PCR and sequencing methods. The mRNA transcription level of oprD and mex efflux pump genes were evaluated by real-time PCR. The outer membrane protein profile was determined by SDS-PAGE. The genetic relationship between the P. aeruginosa isolates was assessed by random amplified polymorphic DNA PCR. In all, 10.52% (10/95) of clinical isolates of P. aeruginosa harboured the IS Ppu 21 insertion element in the opr D gene. The extended-spectrum β-lactamase-encoding gene in IS Ppu 21-carrying isolates was bla TEM . PCR assays targeting MBL and carbapenemase-encoding genes were also negative in all ten isolates. The rmt A, aad A, aad B and arm A genes were positive in all IS Ppu 21 harbouring isolates. The relative expression levels of the mex X, mex B, mex T and mex D genes in ten isolates ranged from 0.1- to 1.4-fold, 1.1- to 3.68-fold, 0.3- to 8.22-fold and 1.7- to 35.17-fold, respectively. The relative expression levels of the oprD in ten isolates ranged from 0.57- to 35.01-fold, which was much higher than those in the control strain P. aeruginosa PAO1. Evaluation of the outer membrane protein by SDS-PAGE suggested that opr D was produced at very low levels by all isolates. Using random amplified polymorphic DNA PCR genotyping, eight of the ten isolates containing IS Ppu 21 were shown to be clonally related. The present study describes a novel molecular mechanism, IS Ppu 21 insertion of the opr D gene, associated with carbapenem resistance in clinical P. aeruginosa isolates.

  19. Deletions in the tetracycline resistance determinant reduce the thermosensitivity of a trfA(Ts) derivative of plasmid RP1 in Pseudomonas aeruginosa.

    PubMed

    Rella, M; Watson, J M; Thomas, C M; Haas, D

    1987-01-01

    A derivative of the broad-host-range plasmid RP1, pME301, was temperature-sensitive (Ts) at 43 degrees C for maintenance in Pseudomonas aeruginosa, P. mendocina, Klebsiella aerogenes and Escherichia coli. In E. coli, the Ts defect of pME301 could be complemented in trans by the cloned trfA gene, which is known to be essential for RP1 replication in E. coli and P. aeruginosa. Because pME301 expressed a Ts phenotype in P. mendocina and K. aerogenes, we assume that the trfA function is also vital in these organisms. When plasmid-encoded carbenicillin resistance (on transposon Tn801) was selected at non-permissive temperatures in P. aeruginosa strain PAO carrying pME301, we obtained either Tn801 insertions into the chromosome or pME301 derivatives having a deletion (or point mutation) in their tet genes, which determine resistance to tetracycline and are not transposable. From cloning experiments, we infer that the tet gene product(s) destabilize the pME301 replicon in P. aeruginosa at 40-43 degrees C.

  20. A demonstration of athermal effects of continuous microwave irradiation on the growth and antibiotic sensitivity of Pseudomonas aeruginosa PAO1.

    PubMed

    Nakouti, Ismini; Hobbs, Glyn; Teethaisong, Yothin; Phipps, David

    2017-01-01

    Stress, caused by exposure to microwaves (2.45 GHz) at constant temperature (37 ± 0.5°C), alters the growth profile of Pseudomonas aeruginosa PAO1. In the absence of microwave treatment a simple, highly reproducible growth curve was observed over 24 h or more. Microwave treatment caused no reduction in growth during the first 6 h, but at a later stage (>12 h) the growth was markedly different to the controls. Secondary growth, typical of the presence of persisters clearly became apparent, as judged by both the dissolved oxygen and the cell density profiles. These treated cells showed distinct morphological changes, but on regrowth these cells reverted to normal. The microwave induced persisters were subject to antibiotic challenge (tobramycin) and showed increased sensitivity when compared to the unstressed planktonic cells. This is in marked contrast to antibiotic induced persisters which show increased resistance. This provides evidence for both a nonthermal effect of microwaves and a previously undescribed route to a novel form of antibiotic susceptible persister cells. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 33:37-44, 2017. © 2016 American Institute of Chemical Engineers.

  1. Prioritization of potential drug targets against P. aeruginosa by core proteomic analysis using computational subtractive genomics and Protein-Protein interaction network.

    PubMed

    Uddin, Reaz; Jamil, Faiza

    2018-06-01

    Pseudomonas aeruginosa is an opportunistic gram-negative bacterium that has the capability to acquire resistance under hostile conditions and become a threat worldwide. It is involved in nosocomial infections. In the current study, potential novel drug targets against P. aeruginosa have been identified using core proteomic analysis and Protein-Protein Interactions (PPIs) studies. The non-redundant reference proteome of 68 strains having complete genome and latest assembly version of P. aeruginosa were downloaded from ftp NCBI RefSeq server in October 2016. The standalone CD-HIT tool was used to cluster ortholog proteins (having >=80% amino acid identity) present in all strains. The pan-proteome was clustered in 12,380 Clusters of Orthologous Proteins (COPs). By using in-house shell scripts, 3252 common COPs were extracted out and designated as clusters of core proteome. The core proteome of PAO1 strain was selected by fetching PAO1's proteome from common COPs. As a result, 1212 proteins were shortlisted that are non-homologous to the human but essential for the survival of the pathogen. Among these 1212 proteins, 321 proteins are conserved hypothetical proteins. Considering their potential as drug target, those 321 hypothetical proteins were selected and their probable functions were characterized. Based on the druggability criteria, 18 proteins were shortlisted. The interacting partners were identified by investigating the PPIs network using STRING v10 database. Subsequently, 8 proteins were shortlisted as 'hub proteins' and proposed as potential novel drug targets against P. aeruginosa. The study is interesting for the scientific community working to identify novel drug targets against MDR pathogens particularly P. aeruginosa. Copyright © 2018 Elsevier Ltd. All rights reserved.

  2. Contribution of permeability and sensitivity to inhibition of DNA synthesis in determining susceptibilities of Escherichia coli, Pseudomonas aeruginosa, and Alcaligenes faecalis to ciprofloxacin.

    PubMed Central

    Bedard, J; Chamberland, S; Wong, S; Schollaardt, T; Bryan, L E

    1989-01-01

    To examine the correlation between bacterial cell susceptibility to ciprofloxacin and the magnitude of uptake and cell target sensitivity, the relative contribution of ciprofloxacin accumulation in intact cells and its ability to inhibit DNA synthesis were investigated among strains of Escherichia coli, Pseudomonas aeruginosa, and Alcaligenes faecalis. Uptake studies of [14C]ciprofloxacin demonstrated diffusion kinetics for P. aeruginosa and E. coli. Ciprofloxacin was more readily removed from E. coli J53 and A. faecalis ATCC 19018 by washing than from P. aeruginosa PAO503. These results indicate that the process of cell accumulation is different for P. aeruginosa in that the drug is firmly bound at an extracellular site. Whatever the washing conditions, A. faecalis accumulated less drug than either of the other two bacteria. Magnesium chloride (10 mM) caused a substantial decrease of ciprofloxacin accumulated and an increase in the MIC, depending upon the nature of the medium. The addition of carbonyl cyanide m-chlorophenylhydrazone caused a variable increase in drug accumulated, depending on the medium and the bacterial strain. The concentration of ciprofloxacin required to obtain 50% inhibition (ID50) of DNA synthesis for P. aeruginosa PAO503 and A. faecalis ATCC 19018 did not correlate with their corresponding MICs but did for E. coli J53. Treatment with EDTA decreased the ID50 of ciprofloxacin for P. aeruginosa PAO503 and its gyrA derivative by 5- and 2-fold, respectively, and decreased the ID50 for E. coli JB5R, a strain with a known decrease in OmpF, by 1.4-fold but did not decrease the ID50 for the normally susceptible E. coli J53. The ID(50) for P. aeruginosa obtained after EDTA treatment or in ether-permeabilized cells was higher than that obtained for the other two strains. The protonophore carbonyl cyanide m-chlorophenylhydrazone prevented killing by low ciprofloxacin concentrtaions, but sodium azide did not. The latter compound did not enhance killing

  3. Protective role of extracellular catalase (KatA) against UVA radiation in Pseudomonas aeruginosa biofilms.

    PubMed

    Pezzoni, Magdalena; Pizarro, Ramón A; Costa, Cristina S

    2014-02-05

    One of the more stressful factors that Pseudomonas aeruginosa must face in nature is solar UVA radiation. In this study, the protective role of KatA catalase in both planktonic cells and biofilms of P. aeruginosa against UVA radiation was determined by using the wild-type (PAO1) and an isogenic catalase deficient strain (katA). The katA strain was more sensitive than the wild-type, especially in the case of biofilms. Moreover, the wild-type biofilm was more resistant than its planktonic counterpart, but this was not observed in the katA strain. Striking KatA activity was detected in the matrix of katA(+) strains, and to our knowledge, this is the first report of this activity in the matrix of P. aeruginosa biofilms. Provision of bovine catalase or KatA to the matrix of a katA biofilm significantly increased its UVA tolerance, demonstrating that extracellular KatA is essential to optimal defense against UVA in P. aeruginosa biofilms. Efficiency of photocatalytic treatments using TiO2 and UVA was lower in biofilms than in planktonic cells, but KatA and KatB catalases seem not to be responsible for the higher resistance of the sessile cells to this treatment. Copyright © 2014 Elsevier B.V. All rights reserved.

  4. Inhibition of Pseudomonas aeruginosa biofilm formation by 2,2'-bipyridyl, lipoic, kojic and picolinic acids.

    PubMed

    Çevik, Kübra; Ulusoy, Seyhan

    2015-08-01

    The inhibitory effects of iron chelators, and FeCl3 chelation on biofilm formation and swarming motility were investigated against an opportunistic human pathogen Pseudomonas aeruginosa. The inhibitory activity of 2,2'-bipyridyl, lipoic acid, kojic acid and picolinic acid on biofilm formation of P. aeruginosa strain PAO1 and three clinical isolates (P. aeruginosa PAK01, P. aeruginosa PAK02 and P. aeruginosa PAK03) were investigated, based on crystal violet assay, and swarming motility test. The kojic, lipoic and picolinic acid inhibited biofilm formation by 5-33% in all tested P. aeruginosa isolates. When chelated iron was added, biofilm inhibition rates were determined to be 39-57%. Among the tested chelators against P. aeruginosa, lipoic acid (84%) and kojic acid (68%) presented the highest inhibition of swarming motility. This is the first study to report the inhibitory effect of lipoic acid on biofilm formation and swarming motility of P. aeruginosa. It is considered that lipoic and picolinic acids can serve as alternatives for the treatment of the P. aeruginosa infections by inhibiting biofilm formation.

  5. Pseudomonas aeruginosa inhibits the growth of Cryptococcus species.

    PubMed

    Rella, Antonella; Yang, Mo Wei; Gruber, Jordon; Montagna, Maria Teresa; Luberto, Chiara; Zhang, Yong-Mei; Del Poeta, Maurizio

    2012-06-01

    Pseudomonas aeruginosa is a ubiquitous and opportunistic bacterium that inhibits the growth of different microorganisms, including Gram-positive bacteria and fungi such as Candida spp. and Aspergillus fumigatus. In this study, we investigated the interaction between P. aeruginosa and Cryptococcus spp. We found that P. aeruginosa PA14 and, to a lesser extent, PAO1 significantly inhibited the growth of Cryptococcus spp. The inhibition of growth was observed on solid medium by the visualization of a zone of inhibition of yeast growth and in liquid culture by viable cell counting. Interestingly, such inhibition was only observed when P. aeruginosa and Cryptococcus were co-cultured. Minimal inhibition was observed when cell-cell contact was prevented using a separation membrane, suggesting that cell contact is required for inhibition. Using mutant strains of Pseudomonas quinoline signaling, we showed that P. aeruginosa inhibited the growth of Cryptococcus spp. by producing antifungal molecules pyocyanin, a redox-active phenazine, and 2-heptyl-3,4-dihydroxyquinoline (PQS), an extracellular quorum-sensing signal. Because both P. aeruginosa and Cryptococcus neoformans are commonly found in lung infections of immunocompromised patients, this study may have important implication for the interaction of these microbes in both an ecological and a clinical point of view.

  6. Genome-scale metabolic modeling of responses to polymyxins in Pseudomonas aeruginosa.

    PubMed

    Zhu, Yan; Czauderna, Tobias; Zhao, Jinxin; Klapperstueck, Matthias; Maifiah, Mohd Hafidz Mahamad; Han, Mei-Ling; Lu, Jing; Sommer, Björn; Velkov, Tony; Lithgow, Trevor; Song, Jiangning; Schreiber, Falk; Li, Jian

    2018-04-01

    Pseudomonas aeruginosa often causes multidrug-resistant infections in immunocompromised patients, and polymyxins are often used as the last-line therapy. Alarmingly, resistance to polymyxins has been increasingly reported worldwide recently. To rescue this last-resort class of antibiotics, it is necessary to systematically understand how P. aeruginosa alters its metabolism in response to polymyxin treatment, thereby facilitating the development of effective therapies. To this end, a genome-scale metabolic model (GSMM) was used to analyze bacterial metabolic changes at the systems level. A high-quality GSMM iPAO1 was constructed for P. aeruginosa PAO1 for antimicrobial pharmacological research. Model iPAO1 encompasses an additional periplasmic compartment and contains 3022 metabolites, 4265 reactions, and 1458 genes in total. Growth prediction on 190 carbon and 95 nitrogen sources achieved an accuracy of 89.1%, outperforming all reported P. aeruginosa models. Notably, prediction of the essential genes for growth achieved a high accuracy of 87.9%. Metabolic simulation showed that lipid A modifications associated with polymyxin resistance exert a limited impact on bacterial growth and metabolism but remarkably change the physiochemical properties of the outer membrane. Modeling with transcriptomics constraints revealed a broad range of metabolic responses to polymyxin treatment, including reduced biomass synthesis, upregulated amino acid catabolism, induced flux through the tricarboxylic acid cycle, and increased redox turnover. Overall, iPAO1 represents the most comprehensive GSMM constructed to date for Pseudomonas. It provides a powerful systems pharmacology platform for the elucidation of complex killing mechanisms of antibiotics.

  7. Intraclonal Genome Stability of the Metallo-β-lactamase SPM-1-producing Pseudomonas aeruginosa ST277, an Endemic Clone Disseminated in Brazilian Hospitals.

    PubMed

    Nascimento, Ana P B; Ortiz, Mauro F; Martins, Willames M B S; Morais, Guilherme L; Fehlberg, Lorena C C; Almeida, Luiz G P; Ciapina, Luciane P; Gales, Ana C; Vasconcelos, Ana T R

    2016-01-01

    Carbapenems represent the mainstay therapy for the treatment of serious P. aeruginosa infections. However, the emergence of carbapenem resistance has jeopardized the clinical use of this important class of compounds. The production of SPM-1 metallo-β-lactamase has been the most common mechanism of carbapenem resistance identified in P. aeruginosa isolated from Brazilian medical centers. Interestingly, a single SPM-1-producing P. aeruginosa clone belonging to the ST277 has been widely spread within the Brazilian territory. In the current study, we performed a next-generation sequencing of six SPM-1-producing P. aeruginosa ST277 isolates. The core genome contains 5899 coding genes relative to the reference strain P. aeruginos a PAO1. A total of 26 genomic islands were detected in these isolates. We identified remarkable elements inside these genomic islands, such as copies of the bla SPM-1 gene conferring resistance to carbapenems and a type I-C CRISPR-Cas system, which is involved in protection of the chromosome against foreign DNA. In addition, we identified single nucleotide polymorphisms causing amino acid changes in antimicrobial resistance and virulence-related genes. Together, these factors could contribute to the marked resistance and persistence of the SPM-1-producing P. aeruginosa ST277 clone. A comparison of the SPM-1-producing P. aeruginosa ST277 genomes showed that their core genome has a high level nucleotide similarity and synteny conservation. The variability observed was mainly due to acquisition of genomic islands carrying several antibiotic resistance genes.

  8. Bioproduction of L-Aspartic Acid and Cinnamic Acid by L-Aspartate Ammonia Lyase from Pseudomonas aeruginosa PAO1.

    PubMed

    Patel, Arti T; Akhani, Rekha C; Patel, Manisha J; Dedania, Samir R; Patel, Darshan H

    2017-06-01

    Aspartase (L-aspartate ammonia lyase, EC 4.3.1.1) catalyses the reversible amination and deamination of L-aspartic acid to fumaric acid which can be used to produce important biochemical. In this study, we have explored the characteristics of aspartase from Pseudomonas aeruginosa PAO1 (PA-AspA). To overproduce PA-AspA, the 1425-bp gene was introduced in Escherichia coli BL21 and purified. A 51.0-kDa protein was observed as a homogenous purified protein on SDS-PAGE. The enzyme was optimally active at pH 8.0 and 35 °C. PA-AspA has retained 56% activity after 7 days of incubation at 35 °C, which displays the hyperthermostablility characteristics of the enzyme. PA-AspA is activated in the presence of metal ions and Mg2+ is found to be most effective. Among the substrates tested for specificity of PA-AspA, L-phenylalanine (38.35 ± 2.68) showed the highest specific activity followed by L-aspartic acid (31.21 ± 3.31) and fumarate (5.42 ± 2.94). K m values for L-phenylalanine, L-aspartic acid and fumarate were 1.71 mM, 0.346 μM and 2 M, respectively. The catalytic efficiency (k cat /K m ) for L-aspartic acid (14.18 s -1  mM -1 ) was higher than that for L-phenylalanine (4.65 s -1  mM -1 ). For bioconversion, from an initial concentration of 1000 mM of fumarate and 30 mM of L-phenylalanine, PA-AspA was found to convert 395.31 μM L-aspartic acid and 3.47 mM cinnamic acid, respectively.

  9. Impact of higher alginate expression on deposition of Pseudomonas aeruginosa in radial stagnation point flow and reverse osmosis systems.

    PubMed

    Herzberg, Moshe; Rezene, Tesfalem Zere; Ziemba, Christopher; Gillor, Osnat; Mathee, Kalai

    2009-10-01

    Extracellular polymeric substances (EPS) have major impact on biofouling of reverse osmosis (RO) membranes. On one hand, EPS can reduce membrane permeability and on the other, EPS production by the primary colonizers may influence their deposition and attachment rate and subsequently affect the biofouling propensity of the membrane. The role of bacterial exopolysaccharides in bacterial deposition followed by the biofouling potential of an RO membrane was evaluated using an alginate overproducing (mucoid) Pseudomonas aeruginosa. The mucoid P. aeruginosa PAOmucA22 was compared with its isogenic nonmucoid prototypic parent PAO1 microscopically in a radial stagnation point flow (RSPF) system for their bacterial deposition characteristics. Then, biofouling potential of PAO1 and PAOmucA22 was determined in a crossflow rectangular plate-and-frame membrane cell, in which the strains were cultivated on a thin-film composite, polyamide, flat RO membrane coupon (LFC-1) under laminar flow conditions. In the RSPF system, the observed deposition rate of the mucoid strain was between 5- and 10-fold lower than of the wild type using either synthetic wastewater medium (with ionic strength of 14.7 mM and pH 7.4) or 15 mM KCl solution (pH of 6.2). The slower deposition rate of the mucoid strain is explained by 5- to 25-fold increased hydrophilicity of the mucoid strain as compared to the isogenic wild type, PAO1. Corroborating with these results, a significant delay in the onset of biofouling of the RO membrane was observed when the mucoid strain was used as the membrane colonizer, in which the observed time for the induced permeate flux decline was delayed (ca. 2-fold). In conclusion, the lower initial cell attachment of the mucoid strain decelerated biofouling of the RO membrane. Bacterial deposition and attachment is a critical step in biofilm formation and governed by intimate interactions between outer membrane proteins of the bacteria and the surface. Shielding these

  10. [Prevalence of cytotoxicity effectors in nosocomial Pseudomonas Aeruginosa strains].

    PubMed

    Kuznetsova, M V; Maksimova, A V; Karpunina, T I; Demakov, V A

    2014-01-01

    Analysis of occurrence of the third type secretory system (TTSS) effectors in clinical P. aeruginosa strains. Intra-hospital (n = 164) and extra-hospital (n = 30) strains of P. aeruginosa were studied. Detection of exoS and exoU genes was carried out by PCR in DNA Engine Dyad Thermal Cycler ("Bio-Rad", USA). Metallo-beta-lactamase (MBL) producers were detected by the presence of blaVIM-2 gene. Screening of intra- and extra-hospital strains for the presence of genes coding ExoS and ExoU showed, that exoS is detected in genome of clinical isolates in 59.8% and exoU--31.1% of cases. At the same time, strains with exoS-/exoU+ genotype predominated in lCU (Φ = 0.466; p = 0.0000). A significant association between the presence of the respective effectors and material of strain isolation was not detected. exoU gene was more frequently detected in genome of MBL producers (Φ = 0.784; p = 0.0004). A significant association between exoU and blaVIM-2 could be explained by clonal prevalence of P. aeruginosa ST235 VIM-2, circulation of those is noted on all the territory of Russia. As a rule, ExoU is produced by highly virulent poly-antibiotic resistant hospital isolates that determine unfavorable outcomes of pseudomonas infection.

  11. Formation of hydroxyl radicals contributes to the bactericidal activity of ciprofloxacin against Pseudomonas aeruginosa biofilms.

    PubMed

    Jensen, Peter Ø; Briales, Alejandra; Brochmann, Rikke P; Wang, Hengzhuang; Kragh, Kasper N; Kolpen, Mette; Hempel, Casper; Bjarnsholt, Thomas; Høiby, Niels; Ciofu, Oana

    2014-04-01

    Antibiotic-tolerant, biofilm-forming Pseudomonas aeruginosa has long been recognized as a major cause of chronic lung infections of cystic fibrosis patients. The mechanisms involved in the activity of antibiotics on biofilm are not completely clear. We have investigated whether the proposed induction of cytotoxic hydroxyl radicals (OH˙) during antibiotic treatment of planktonically grown cells may contribute to action of the commonly used antibiotic ciprofloxacin on P. aeruginosa biofilms. For this purpose, WT PAO1, a catalase deficient ΔkatA and a ciprofloxacin resistant mutant of PAO1 (gyrA), were grown as biofilms in microtiter plates and treated with ciprofloxacin. Formation of OH˙ and total amount of reactive oxygen species (ROS) was measured and viability was estimated. Formation of OH˙ and total ROS in PAO1 biofilms treated with ciprofloxacin was shown but higher levels were measured in ΔkatA biofilms, and no ROS production was seen in the gyrA biofilms. Treatment with ciprofloxacin decreased the viability of PAO1 and ΔkatA biofilms but not of gyrA biofilms. Addition of thiourea, a OH˙ scavenger, decreased the OH˙ levels and killing of PAO1 biofilm. Our study shows that OH˙ is produced by P. aeruginosa biofilms treated with ciprofloxacin, which may contribute to the killing of biofilm subpopulations. © 2013 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.

  12. Transferable Drug Resistance in Pseudomonas aeruginosa1

    PubMed Central

    Bryan, L. E.; Elzen, H. M. Van Den; Tseng, Jui Teng

    1972-01-01

    Three strains of Pseudomonas aeruginosa were demonstrated to transfer double-drug resistance by conjugation to a P. aeruginosa recipient at frequencies of 10−4 to 10−2 per recipient cell. Two of the three strains also transferred to Escherichia coli at frequencies which were 103- to 105-fold lower, but the third strain could not be demonstrated to do so. The latter strain, however, conferred maleness on the Pseudomonas recipient. The transfer of streptomycin resistance was associated with the acquisition of streptomycin phosphorylase by both P. aeruginosa and E. coli recipients. Maximal broth mating frequencies were obtained with nonagitated cultures less than 1 mm in depth. A pyocine selection system based on donor sensitivity and recipient resistance is described and appears to have future value as a generalized selective device for use after matings. PMID:4207756

  13. Prediction of vaccine candidates against Pseudomonas aeruginosa: An integrated genomics and proteomics approach.

    PubMed

    Rashid, Muhammad Ibrahim; Naz, Anam; Ali, Amjad; Andleeb, Saadia

    2017-07-01

    Pseudomonas aeruginosa is among top critical nosocomial infectious agents due to its persistent infections and tendency for acquiring drug resistance mechanisms. To date, there is no vaccine available for this pathogen. We attempted to exploit the genomic and proteomic information of P. aeruginosa though reverse-vaccinology approaches to unveil the prospective vaccine candidates. P. aeruginosa strain PAO1 genome was subjected to sequential prioritization approach following genomic, proteomics and structural analyses. Among, the predicted vaccine candidates: surface components of antibiotic efflux pumps (Q9HY88, PA2837), chaperone-usher pathway components (CupC2, CupB3), penicillin binding protein of bacterial cell wall (PBP1a/mrcA), extracellular component of Type 3 secretory system (PscC) and three uncharacterized secretory proteins (PA0629, PA2822, PA0978) were identified as potential candidates qualifying all the set criteria. These proteins were then analyzed for potential immunogenic surface exposed epitopes. These predicted epitopes may provide a basis for development of a reliable subunit vaccine against P. aeruginosa. Copyright © 2017 Elsevier Inc. All rights reserved.

  14. Combination of hypothiocyanite and lactoferrin (ALX-109) enhances the ability of tobramycin and aztreonam to eliminate Pseudomonas aeruginosa biofilms growing on cystic fibrosis airway epithelial cells

    PubMed Central

    Moreau-Marquis, Sophie; Coutermarsh, Bonita; Stanton, Bruce A.

    2015-01-01

    Objectives Chelating iron may be a promising new therapy to eliminate Pseudomonas aeruginosa biofilms in the lungs of cystic fibrosis (CF) patients. Here, we investigate whether ALX-109 [a defined combination of an investigational drug containing lactoferrin (an iron-binding glycoprotein) and hypothiocyanite (a bactericidal agent)], alone and in combination with tobramycin or aztreonam, reduces P. aeruginosa biofilms grown on human CF airway epithelial cells. Methods P. aeruginosa (PAO1 and six clinical isolates of Pseudomonas) biofilms grown at the apical surface of confluent monolayers of CF airway epithelial cells were treated with ALX-109, either alone or in combination with tobramycin or aztreonam. Bacterial cfu remaining after treatment were determined by plate counting. Results ALX-109 alone reduced PAO1 biofilm formation, but had no effect on established biofilms. ALX-109 enhanced the ability of tobramycin and aztreonam to inhibit PAO1 biofilm formation and to reduce established PAO1 biofilms. ALX-109 and tobramycin were additive in disrupting established biofilms formed by six clinical isolates of P. aeruginosa obtained from the sputum of CF patients. Mucoid P. aeruginosa isolates were most susceptible to the combination of ALX-109 and tobramycin. In addition, ALX-109 also enhanced the ability of aztreonam to reduce established PAO1 biofilms. Conclusions Inhalation therapy combining hypothiocyanite and lactoferrin with TOBI® (tobramycin) or Cayston® (aztreonam) may be beneficial to CF patients by decreasing the airway bacterial burden of P. aeruginosa. PMID:25213272

  15. Inhibition of Pseudomonas aeruginosa biofilm formation by 2,2’-bipyridyl, lipoic, kojic and picolinic acids

    PubMed Central

    Çevik, Kübra; Ulusoy, Seyhan

    2015-01-01

    Objective(s): The inhibitory effects of iron chelators, and FeCl3 chelation on biofilm formation and swarming motility were investigated against an opportunistic human pathogen Pseudomonas aeruginosa. Materials and Methods: The inhibitory activity of 2,2’-bipyridyl, lipoic acid, kojic acid and picolinic acid on biofilm formation of P. aeruginosa strain PAO1 and three clinical isolates (P. aeruginosa PAK01, P. aeruginosa PAK02 and P. aeruginosa PAK03) were investigated, based on crystal violet assay, and swarming motility test. Results: The kojic, lipoic and picolinic acid inhibited biofilm formation by 5-33% in all tested P. aeruginosa isolates. When chelated iron was added, biofilm inhibition rates were determined to be 39-57%. Among the tested chelators against P. aeruginosa, lipoic acid (84%) and kojic acid (68%) presented the highest inhibition of swarming motility. This is the first study to report the inhibitory effect of lipoic acid on biofilm formation and swarming motility of P. aeruginosa. Conclusion: It is considered that lipoic and picolinic acids can serve as alternatives for the treatment of the P. aeruginosa infections by inhibiting biofilm formation. PMID:26557964

  16. Induced Formation of Chelating Agents by Pseudomonas aeruginosa Grown in Presence of Thorium and Uranium

    DTIC Science & Technology

    1985-07-01

    aerugiaosa PAO-l, Saccharomyces cerevisiae, Aspergillus niger , P. fluorescens, Escherichia coli, and Thiobacillus ferroxidans. Interaction of these...shown that P. aeruginosa CSU has..a-••reference for uranium while P. aeruginosa PAO-l, Aspergillus niger and-P. fluorescens exhibits a preference for...exhibits a preference for chromium. Aspergillus niger under identical conditions is chromium and manganese selective. P. aeruginosa when grown in th

  17. Pseudomonas aeruginosa biofilm aggravates skin inflammatory response in BALB/c mice in a novel chronic wound model.

    PubMed

    Trøstrup, Hannah; Thomsen, Kim; Christophersen, Lars J; Hougen, Hans P; Bjarnsholt, Thomas; Jensen, Peter Ø; Kirkby, Nikolai; Calum, Henrik; Høiby, Niels; Moser, Claus

    2013-01-01

    Chronic wounds are presumed to persist in the inflammatory state, preventing healing. Emerging evidence indicates a clinical impact of bacterial biofilms in soft tissues, including Pseudomonas aeruginosa (PA) biofilms. To further investigate this, we developed a chronic PA biofilm wound infection model in C3H/HeN and BALB/c mice. The chronic wound was established by an injection of seaweed alginate-embedded P. aeruginosa PAO1 beneath a third-degree thermal lesion providing full thickness skin necrosis, as in human chronic wounds. Cultures revealed growth of PA, and both alginate with or without PAO1 generated a polymorphonuclear-dominated inflammation early after infection. However, both at days 4 and 7, there were a more acute polymorphonuclear-dominated and higher degree of inflammation in the PAO1 containing group (p < 0.05). Furthermore, PNA-FISH and supplemented DAPI staining showed bacteria organized in clusters, resembling biofilms, and inflammation located adjacent to the PA. The chronic wound infection showed a higher number of PAO1 in the BALB/c mice at day 4 after infection as compared to C3H/HeN mice (p < 0.006). In addition, a higher concentration of interleukin-1beta in the chronic wounds of BALB/c mice was observed at day 7 (p < 0.02), despite a similar number of bacteria in the two mouse strains. The present study succeeded in establishing a chronic PA biofilm infection in mice. The results showed an aggravating impact of local inflammation induced by PA biofilms. In conclusion, our findings indicate that improved infection control of chronic wounds reduces the inflammatory response and may improve healing. © 2013 by the Wound Healing Society.

  18. Reconstruction of the metabolic network of Pseudomonas aeruginosa to interrogate virulence factor synthesis

    NASA Astrophysics Data System (ADS)

    Bartell, Jennifer A.; Blazier, Anna S.; Yen, Phillip; Thøgersen, Juliane C.; Jelsbak, Lars; Goldberg, Joanna B.; Papin, Jason A.

    2017-03-01

    Virulence-linked pathways in opportunistic pathogens are putative therapeutic targets that may be associated with less potential for resistance than targets in growth-essential pathways. However, efficacy of virulence-linked targets may be affected by the contribution of virulence-related genes to metabolism. We evaluate the complex interrelationships between growth and virulence-linked pathways using a genome-scale metabolic network reconstruction of Pseudomonas aeruginosa strain PA14 and an updated, expanded reconstruction of P. aeruginosa strain PAO1. The PA14 reconstruction accounts for the activity of 112 virulence-linked genes and virulence factor synthesis pathways that produce 17 unique compounds. We integrate eight published genome-scale mutant screens to validate gene essentiality predictions in rich media, contextualize intra-screen discrepancies and evaluate virulence-linked gene distribution across essentiality datasets. Computational screening further elucidates interconnectivity between inhibition of virulence factor synthesis and growth. Successful validation of selected gene perturbations using PA14 transposon mutants demonstrates the utility of model-driven screening of therapeutic targets.

  19. A medicinal herb Cassia alata attenuates quorum sensing in Chromobacterium violaceum and Pseudomonas aeruginosa.

    PubMed

    Rekha, P D; Vasavi, H S; Vipin, C; Saptami, K; Arun, A B

    2017-03-01

    Quorum sensing (QS) has been shown to play a crucial role in the pathogenesis in many bacteria, and attenuation of QS is one of the targets of antimicrobial therapy with particular interest in combating drug resistance. This study reports the QS inhibitory activity of metabolites from Cassia alata L. (Ca. alata), an important medicinal herb widely used in the treatment of microbial infections. For investigating the QS inhibition (QSI), the potential of Ca. alata L., initially, metabolites of the leaves extracted using ethanol was tested against biosensor strain Chromobacterium violaceum CV026 and C. violaceum wild-type strains. Furthermore, a purified fraction rich in flavonoids (F-AF) was used for establishing QSI activity by studying the inhibition of violacein production in C. violaceum, and QS controlled virulence and biofilm formation in Pseudomonas aeruginosa PAO1. The study results showed 50% inhibition of violacein production in C. violaceum at 0·05 mg ml -1 concentration of F-AF. In P. aeruginosa PAO1, it inhibited the tested virulence factors and biofilm formation significantly. The F-AF contained major flavonoids namely, quercetin, quercetrin and kaempferol displaying QSI activity individually against the test organisms. Present study demonstrates the quorum sensing inhibitory activity of metabolites from Cassia alata, an important medicinal herb which is commonly used worldwide in the treatment of infections caused by microorganisms. An extract prepared from the leaves of the plant showed activity against quorum sensing in Chromobacterium violaceum and was also effective against attenuating the quorum sensing controlled virulence factors in Pseudomonas aeruginosa. Activity is attributed to the rich flavonoid composition of the plant. Results of the present investigation throw an insight into the possibility of developing drug formulations using the isolated compounds against infections caused by quorum sensing-mediated pathogenicity of bacteria

  20. Molecular Characterization and Regulation of the aguBA Operon, Responsible for Agmatine Utilization in Pseudomonas aeruginosa PAO1

    PubMed Central

    Nakada, Yuji; Jiang, Ying; Nishijyo, Takayuki; Itoh, Yoshifumi; Lu, Chung-Dar

    2001-01-01

    Pseudomonas aeruginosa PAO1 utilizes agmatine as the sole carbon and nitrogen source via two reactions catalyzed successively by agmatine deiminase (encoded by aguA; also called agmatine iminohydrolase) and N-carbamoylputrescine amidohydrolase (encoded by aguB). The aguBA and adjacent aguR genes were cloned and characterized. The predicted AguB protein (Mr 32,759; 292 amino acids) displayed sequence similarity (≤60% identity) to enzymes of the β-alanine synthase/nitrilase family. While the deduced AguA protein (Mr 41,190; 368 amino acids) showed no significant similarity to any protein of known function, assignment of agmatine deiminase to AguA in this report discovered a new family of carbon-nitrogen hydrolases widely distributed in organisms ranging from bacteria to Arabidopsis. The aguR gene encoded a putative regulatory protein (Mr 24,424; 221 amino acids) of the TetR protein family. Measurements of agmatine deiminase and N-carbamoylputrescine amidohydrolase activities indicated the induction effect of agmatine and N-carbamoylputrescine on expression of the aguBA operon. The presence of an inducible promoter for the aguBA operon in the aguR-aguB intergenic region was demonstrated by lacZ fusion experiments, and the transcription start of this promoter was localized 99 bp upstream from the initiation codon of aguB by S1 nuclease mapping. Experiments with knockout mutants of aguR established that expression of the aguBA operon became constitutive in the aguR background. Interaction of AguR overproduced in Escherichia coli with the aguBA regulatory region was demonstrated by gel retardation assays, supporting the hypothesis that AguR serves as the negative regulator of the aguBA operon, and binding of agmatine and N-carbamoylputrescine to AguR would antagonize its repressor function. PMID:11673419

  1. Exploring the In Vitro Thrombolytic Activity of Nattokinase From a New Strain Pseudomonas aeruginosa CMSS.

    PubMed

    Chandrasekaran, Subathra Devi; Vaithilingam, Mohanasrinivasan; Shanker, Ravi; Kumar, Sanjeev; Thiyur, Swathi; Babu, Vaishnavi; Selvakumar, Jemimah Naine; Prakash, Suyash

    2015-10-01

    Thrombolytic therapy has become a conventional treatment for acute myocardial infarction (AMI), yet currently, clinically prescribed thrombolytic drugs have problems such as delayed action and other side effects. Fibrinolytic enzymes have attracted interest as thrombolytic agents because of their efficiency in the fibrinolytic process, including plasmin activation. Nattokinase (NK) is a potent fibrinolytic agent for thrombosis therapy. The aim of this study was to enhance the production of NK from Pseudomonas aeruginosa CMSS by media optimization and strain improvement. In the present study, a potent NK-producing strain was isolated from cow milk and identified. To enhance the yield of NK, effect of various parameters such as pH, temperature, carbon source, nitrogen source and inoculum size were optimized. Strain improvement of P. aeruginosa CMSS was done by random UV-mutagenesis. Nattokinase was partially purified and the activity was determined by the casein digestion method, blood clot lysis and fibrin degradation assay. Based on morphological, biochemical and molecular characterization, the strain was confirmed as P. aeruginosa (GenBank accession number: JX112657), designated as P. aeruginosa CMSS. The optimum condition at pH 7 and temperature at 25˚C showed activity of NK as 1514 U mL(-1) and 1532 U mL(-1), respectively. Sucrose as the carbon source and shrimp shell powder (SSP) as the nitrogen source expressed NK activity of 1721 U mL(-1) and 2524 U mL(-1), respectively. At 1% inoculum size, the maximum rate of enzyme production was achieved with 2581 U mL(-1). The NK activity of the mutant strain UV60 was 4263 U mL(-1), indicating a two-fold increase in activity compared to the wild strain (2581 UmL(-1)). Nattokinase produced from mutant strain P. aeruginosa CMSS UV60 showed 94% blood clot lysis at ten minutes. The degradation of fibrin clot by the produced NK was observed after two hours of incubation. Sodium dodecyl sulfate polyacrylamide gel

  2. Exploring the In Vitro Thrombolytic Activity of Nattokinase From a New Strain Pseudomonas aeruginosa CMSS

    PubMed Central

    Chandrasekaran, Subathra Devi; Vaithilingam, Mohanasrinivasan; Shanker, Ravi; Kumar, Sanjeev; Thiyur, Swathi; Babu, Vaishnavi; Selvakumar, Jemimah Naine; Prakash, Suyash

    2015-01-01

    Background: Thrombolytic therapy has become a conventional treatment for acute myocardial infarction (AMI), yet currently, clinically prescribed thrombolytic drugs have problems such as delayed action and other side effects. Fibrinolytic enzymes have attracted interest as thrombolytic agents because of their efficiency in the fibrinolytic process, including plasmin activation. Nattokinase (NK) is a potent fibrinolytic agent for thrombosis therapy. Objectives: The aim of this study was to enhance the production of NK from Pseudomonas aeruginosa CMSS by media optimization and strain improvement. Materials and Methods: In the present study, a potent NK-producing strain was isolated from cow milk and identified. To enhance the yield of NK, effect of various parameters such as pH, temperature, carbon source, nitrogen source and inoculum size were optimized. Strain improvement of P. aeruginosa CMSS was done by random UV-mutagenesis. Nattokinase was partially purified and the activity was determined by the casein digestion method, blood clot lysis and fibrin degradation assay. Results: Based on morphological, biochemical and molecular characterization, the strain was confirmed as P. aeruginosa (GenBank accession number: JX112657), designated as P. aeruginosa CMSS. The optimum condition at pH 7 and temperature at 25˚C showed activity of NK as 1514 U mL-1 and 1532 U mL-1, respectively. Sucrose as the carbon source and shrimp shell powder (SSP) as the nitrogen source expressed NK activity of 1721 U mL-1 and 2524 U mL-1, respectively. At 1% inoculum size, the maximum rate of enzyme production was achieved with 2581 U mL-1. The NK activity of the mutant strain UV60 was 4263 U mL-1, indicating a two-fold increase in activity compared to the wild strain (2581 UmL-1). Nattokinase produced from mutant strain P. aeruginosa CMSS UV60 showed 94% blood clot lysis at ten minutes. The degradation of fibrin clot by the produced NK was observed after two hours of incubation. Sodium

  3. Serological Typing of 31 Achromogenic and 40 Melanogenic Pseudomonas aeruginosa Strains

    PubMed Central

    Yabuuchi, Eiko; Miyajima, Noriko; Hotta, Hisako; Furu, Youichi

    1971-01-01

    Thirty-one achromogenic and 40 melanogenic Pseudomonas aeruginosa strains were studied with 10 monovalent typing sera (3). Twenty-one of the achromogenic (67.7%) and seven of the melanogenic (17.5%) strains were agglutinated by one of the 10 typing sera. Ten achromogenic and 33 melanogenic strains were not agglutinated by any of the 10 typing sera. As far as this set of antisera is concerned, the typability of achromogenic and melanogenic P. aeruginosa strains appears to be much lower than that of the chromogenic, nonmelanogenic strains of the species reported previously. PMID:5002137

  4. [Antiseptic sensitivity of clinical strains of Pseudomonas aeruginosa].

    PubMed

    Adarchenko, A A; Krasil'nikov, A P; Sobeshchuk, O P

    1989-12-01

    MICs, the frequency of clinical and statistic resistance and the antiseptic activity index were studied in complex on out-of-hospital and hospital ecovars of P. aeruginosa. The forms resistant to a number of antiseptics, i.e. chloramine B, chlorhexidine, decamethoxine and dioxidine whose frequency eventually increased were shown to be widely distributed. The antiseptic sensitivity spectrum was more narrow and more heterogeneous than that of other bacteria, the heterogeneity level being dependent on the antiseptic type and bacterial ecovar. The activity of pervomur, phenol, resorcin and boric acid was higher against the clinical strains of P. aeruginosa while iodopyrin, sulfacetamide sodium and dioxidine were less active. The P. aeruginosa strains had natural resistance to cetylpyridinium chloride, rokkal, ethonium, sodium laurate and laurylsulfate and rivanol. It was recommended to assay antiseptic sensitivity of agents causing purulent inflammatory infections and to control circulation of antiseptic resistant variants of bacteria in hospitals.

  5. Detection of VIM-2-, IMP-1- and NDM-1-producing multidrug resistant Pseudomonas aeruginosa in Malaysia.

    PubMed

    Liew, Siew Mun; Rajasekaram, Ganeswrei; Puthucheary, Savithri D; Chua, Kek Heng

    2018-02-09

    The increasing incidence of carbapenem-resistant Pseudomonas aeruginosa along with the discovery of novel metallo-β-lactamases (MBLs) is of concern. In this study, the isolation of Malaysian MBL-producing P. aeruginosa clinical strains was investigated. Fifty-three P. aeruginosa clinical strains were isolated from different patients in Sultanah Aminah Hospital, Johor Bahru, Malaysia in 2015. Antimicrobial susceptibility test was conducted. Minimum inhibitory concentrations (MICs) of imipenem and meropenem were determined by Etest. The carbapenem-resistant strains were screened for MBL production by IMP-EDTA double disk synergy test (DDST), MBL imipenem/imipenem-inhibitor (IP/IPI) Etest and polymerase chain reaction (PCR). Genotyping was performed by multilocus sequence typing (MLST) analysis. Three (5.7%) clinical strains were identified as MBL producers. Multidrug resistance was observed in the three strains, and two were resistant to all the antimicrobials tested. Sequencing analysis confirmed the three strains to harbour carbapenemase genes: one with bla IMP-1 , one with bla VIM-2 and the other with bla NDM-1 genes. These multidrug resistant strains were identified as sequence type (ST) 235 and ST308. None of the bla IMP-1 and bla NDM-1 genes have been reported in Malaysian P. aeruginosa. The emergence of imipenemase 1 (IMP-1)- and New Delhi metallo-β-lactamase 1 (NDM-1)-producing P. aeruginosa in Malaysia maybe travel-associated. Copyright © 2018. Published by Elsevier Ltd.

  6. Quorum sensing signal molecules produced by Pseudomonas aeruginosa cause inflammation and escape host factors in murine model of urinary tract infection.

    PubMed

    Gupta, Parul; Gupta, Ravi Kumar; Harjai, Kusum

    2013-10-01

    Quorum sensing (QS) is well established for its role in pathogenesis of various infections of Pseudomonas aeruginosa. However, its role in local tissue damage during urinary tract infection (UTI) is not yet fully established. To have insight in this, the present study was planned. UTI was established in mice using standard strain PAO1 and its isogenic QS mutant JP2. One group was challenged only with QS signals. Damage was assessed in terms of histopathology and pathology markers, malondialdehyde (MDA) and reactive nitrogen intermediates (RNI). Effect on pathogen motility, uroepithelial adhesion, and host serum sensitivity was also ascertained. PAO1-infected mice showed severe inflammation and tissue destruction, while mice infected with JP2 showed no significant destruction. JP2 was also unable to mount any tissue pathology markers, MDA and RNI, whereas PAO1 showed significantly higher levels of these two. Presence of only QS signals also showed considerable renal pathology. Strain JP2 also showed less motility, reduced uroepithelial cell adhesion, and increased serum sensitivity. Result highlights that QS signals induce local tissue pathology along with interference of host protective mechanisms during UTI.

  7. Anaerobic Corrosion of 304 Stainless Steel Caused by the Pseudomonas aeruginosa Biofilm

    PubMed Central

    Jia, Ru; Yang, Dongqing; Xu, Dake; Gu, Tingyue

    2017-01-01

    Pseudomonas aeruginosa is a ubiquitous bacterium capable of forming problematic biofilms in many environments. They cause biocorrosion of medical implants and industrial equipment and infrastructure. Aerobic corrosion of P. aeruginosa against stainless steels has been reported by some researchers while there is a lack of reports on anaerobic P. aeruginosa corrosion in the literature. In this work, the corrosion by a wild-type P. aeruginosa (strain PAO1) biofilm against 304 stainless steel (304 SS) was investigated under strictly anaerobic condition for up to 14 days. The anaerobic corrosion of 304 SS by P. aeruginosa was reported for the first time. Results showed that the average sessile cell counts on 304 SS coupons after 7- and 14-day incubations were 4.8 × 107 and 6.2 × 107 cells/cm2, respectively. Scanning electron microscopy and confocal laser scanning microscopy corroborated the sessile cell counts. The X-ray diffraction analysis identified the corrosion product as iron nitride, confirming that the corrosion was caused by the nitrate reducing biofilm. The largest pit depths on 304 SS surfaces after the 7- and 14-day incubations with P. aeruginosa were 3.9 and 7.4 μm, respectively. Electrochemical tests corroborated the pitting data. PMID:29230206

  8. Photodynamic inactivation of antibiotic resistant strain of Pseudomonas aeruginosa in vivo

    NASA Astrophysics Data System (ADS)

    Hashimoto, M. C. E.; Toffoli, D. J.; Prates, R. A.; Courrol, Lilia C.; Ribeiro, M. S.

    2009-06-01

    Burns are frequently contamined by pathogenic microorganisms and the widespread occurrence of antibiotic resistant strains of Pseudomonas aeruginosa in hospitals is a matter of growing concern. Hypocrellin B (HB) is a new generation photosensitizer extracted from the fungus Hypocrella bambusae with absorption bands at 460, 546 and 584 nm. Lanthanide ions change the HB molecular structure and a red shift in the absorption band is observed as well as an increase in the singlet oxygen quantum yield. In this study, we report the use of HB:La+3 to kill resistant strain of P. aeruginosa infected burns. Burns were produced on the back of mice and wounds were infected subcutaneously with 1x109 cfu/mL of P. aeruginosa. Three-hours after inoculation, the animals were divided into 4 groups: control, HB:La+3, blue LED and HB:La+3+blue LED. PDT was performed using 10μM HB:La+3 and 500mW light-emitting diode (LED) emitting at λ=470nm+/-20nm during 120s. The animals of all groups were killed and the infected skin was removed for bacterial counting. Mice with photosensitizer alone, light alone or untreated infected wounds presented 1x108 cfu/g while mice PDT-treated showed a reduction of 2 logs compared to untreated control. These results suggest that HB:La+3 associated to blue LED is effective in diminishing antibiotic resistant strain P. aeruginosa in infected burns.

  9. Honey-sensitive Pseudomonas aeruginosa mutants are impaired in catalase A.

    PubMed

    Bolognese, Fabrizio; Bistoletti, Michela; Barbieri, Paola; Orlandi, Viviana Teresa

    2016-09-01

    The antimicrobial power of honey seems to be ascribable to several factors, including oxidative and osmotic stress. The aim of this study was to find genetic determinants involved in the response to honey stress in the opportunistic pathogen Pseudomonas aeruginosa, chosen as model micro-organism. A library of transposon mutants of P. aeruginosa PAO1 was constructed and only four mutants unable to grow in presence of fir honeydew honey were selected. All four mutants were impaired in the major H2O2-scavenging enzyme catalase A (KatA). The knockout of katA gene caused sensitivity, as expected, not only to hydrogen peroxide but also to different types of honey including Manuka GMO 220 honey. Genetic complementation, as well as the addition of PAO1 supernatant containing extracellular catalase, restored tolerance to honey stress in all the mutants. As P. aeruginosa PAO1 catalase KatA copes with H2O2 stress, it is conceivable that the antimicrobial activity of honey is, at least partially, due to the presence of hydrogen peroxide in honey or the ability of honey to induce production of hydrogen peroxide. The katA-deficient mutants could be used as tester micro-organisms to compare the power of different types of natural and curative honeys in eliciting oxidative stress mediated by hydrogen peroxide.

  10. Prevalence and spread of pseudomonas aeruginosa and Klebsiella pneumoniae strains in patients with hematological malignancies.

    PubMed

    Kolar, Milan; Sauer, Pavel; Faber, Edgar; Kohoutova, Jarmila; Stosová, Tatana; Sedlackova, Michaela; Chroma, Magdalena; Koukalova, Dagmar; Indrak, Karel

    2009-01-01

    The aim of the study was to determine the prevalence of Pseudomonas aeruginosa and Klebsiella pneumoniae strains in patients with acute leukemias, to assess their clinical significance, and to define the sources and ways of their spread using genetic analysis. Thirty-four patients were investigated during the observed period. Twenty-one strains of Pseudomonas aeruginosa and 35 strains of Klebsiella pneumoniae were isolated from patient samples. In the case of Pseudomonas aeruginosa, 47.6% of strains were identified as pathogens and caused infection. By contrast, only 4 isolates (11.4%) of Klebsiella pneumoniae could be regarded as etiological agents of bacterial infection. Based on the obtained results, Klebsiella pneumoniae strains are assumed to be of mostly endogenous origin. In the case of Pseudomonas aeruginosa strains, the proportion of identical strains detected in various patients was higher and exogenous sources were more significant. In addition, our results confirmed the ability of Pseudomonas aeruginosa strains to survive on a particular site in the hospital for a longer time.

  11. Anaerobically controlled expression system derived from the arcDABC operon of Pseudomonas aeruginosa: application to lipase production.

    PubMed Central

    Winteler, H V; Schneidinger, B; Jaeger, K E; Haas, D

    1996-01-01

    The anaerobically inducible arcDABC operon encodes the enzymes of the arginine deiminase pathway in Pseudomonas aeruginosa. Upon induction, the arcAB mRNAs and proteins reach high intracellular levels, because of a strong anaerobically controlled promoter and mRNA processing in arcD, leading to stable downstream transcripts. We explored the usefulness of this system for the construction of expression vectors. The lacZ gene of Escherichia coli was expressed to the highest levels when fused close to the arc promoter. Insertion of lacZ further downstream into arcA or arcB did not stabilize the intrinsically unstable lacZ mRNA. On the contrary, lacZ mRNA appeared to be a vulnerable endonuclease target destabilizing arcAB mRNAs in the 5'-to-3' direction in P. aeruginosa. The native arc promoter was modified for optional expression in the -10 sequence and in the -40 region, which is a binding site for the anaerobic regulator ANR. In P. aeruginosa grown either anaerobically or with oxygen limitation in unshaken cultures, this promoter was stronger than the induced tac promoter. The P. aeruginosa lipAH genes, which encode extracellular lipase and lipase foldase, respectively, were fused directly to the modified arc promoter in an IncQ vector plasmid. Semianaerobic static cultures of P. aeruginosa PAO1 carrying this recombinant plasmid overproduced extracellular lipase 30-fold during stationary phase compared with the production by strain PAO1 without the plasmid. Severe oxygen limitation, in contrast, resulted in poor lipase productivity despite effective induction of the ANR-dependent promoter, suggesting that secretion of active lipase is blocked by the absence of oxygen. In conclusion, the modified arc promoter is useful for driving the expression of cloned genes in P. aeruginosa during oxygen-limited growth and stationary phase. PMID:8795231

  12. Combination of hypothiocyanite and lactoferrin (ALX-109) enhances the ability of tobramycin and aztreonam to eliminate Pseudomonas aeruginosa biofilms growing on cystic fibrosis airway epithelial cells.

    PubMed

    Moreau-Marquis, Sophie; Coutermarsh, Bonita; Stanton, Bruce A

    2015-01-01

    Chelating iron may be a promising new therapy to eliminate Pseudomonas aeruginosa biofilms in the lungs of cystic fibrosis (CF) patients. Here, we investigate whether ALX-109 [a defined combination of an investigational drug containing lactoferrin (an iron-binding glycoprotein) and hypothiocyanite (a bactericidal agent)], alone and in combination with tobramycin or aztreonam, reduces P. aeruginosa biofilms grown on human CF airway epithelial cells. P. aeruginosa (PAO1 and six clinical isolates of Pseudomonas) biofilms grown at the apical surface of confluent monolayers of CF airway epithelial cells were treated with ALX-109, either alone or in combination with tobramycin or aztreonam. Bacterial cfu remaining after treatment were determined by plate counting. ALX-109 alone reduced PAO1 biofilm formation, but had no effect on established biofilms. ALX-109 enhanced the ability of tobramycin and aztreonam to inhibit PAO1 biofilm formation and to reduce established PAO1 biofilms. ALX-109 and tobramycin were additive in disrupting established biofilms formed by six clinical isolates of P. aeruginosa obtained from the sputum of CF patients. Mucoid P. aeruginosa isolates were most susceptible to the combination of ALX-109 and tobramycin. In addition, ALX-109 also enhanced the ability of aztreonam to reduce established PAO1 biofilms. Inhalation therapy combining hypothiocyanite and lactoferrin with TOBI(®) (tobramycin) or Cayston(®) (aztreonam) may be beneficial to CF patients by decreasing the airway bacterial burden of P. aeruginosa. © The Author 2014. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  13. Characteristics of carbapenem-resistant Pseudomonas aeruginosa strains in patients with ventilator-associated pneumonia in intensive care units.

    PubMed

    Vitkauskienė, Astra; Skrodenienė, Erika; Dambrauskienė, Asta; Bakšytė, Giedrė; Macas, Andrius; Sakalauskas, Raimundas

    2011-01-01

    The aim of this study was to determine the characteristics of carbapenem-resistant Pseudomonas aeruginosa (P. aeruginosa) strains and 5-year changes in resistance in a tertiary university hospital. The study included 90 and 101 randomly selected P. aeruginosa strains serotyped in 2003 and 2008, respectively. The standardized disk diffusion test and E-test were used to determine resistance to antibiotics. P. aeruginosa strains were considered to have high-level resistance if a minimum inhibitory concentration (MIC) for imipenem or meropenem was >32 µg/mL. To identify serogroups, sera containing specific antibodies against O group antigens of P. aeruginosa were used. P. aeruginosa isolates resistant to imipenem or/and meropenem were screened for metallo-β-lactamase (MBL) production by using the MBL E-test. Comparison of the changes in resistance of P. aeruginosa strains to carbapenems within the 5-year period revealed that the level of resistance to imipenem increased. In 2003, 53.3% of P. aeruginosa strains were found to be highly resistant to imipenem, while in 2008, this percentage increased to 87.8% (P=0.01). The prevalence of MBL-producing strains increased from 15.8% in 2003 to 61.9% in 2008 (P<0.001). In 2003 and 2008, carbapenem-resistant P. aeruginosa strains were more often resistant to ciprofloxacin and gentamicin than carbapenem-sensitive strains. In 2008, carbapenem-resistant strains additionally were more often resistant to ceftazidime, cefepime, aztreonam, piperacillin, and amikacin than carbapenem-sensitive strains. MBL-producing P. aeruginosa strains belonged more often to the O:11 serogroup than MBL-non-producing strains (51.7% vs. 34.3%, P<0.05). A greater percentage of non-MBL-producing strains had low MICs against ciprofloxacin and amikacin as compared with MBL-producing strains. The results of our study emphasize the need to restrict the spread of O:11 serogroup P. aeruginosa strains and usage of carbapenems to treat infections with P

  14. Cinnamic acid attenuates quorum sensing associated virulence factors and biofilm formation in Pseudomonas aeruginosa PAO1.

    PubMed

    Rajkumari, Jobina; Borkotoky, Subhomoi; Murali, Ayaluru; Suchiang, Kitlangki; Mohanty, Saswat Kumar; Busi, Siddhardha

    2018-04-21

    Anti-quorum sensing and anti-biofilm efficacy of Cinnamic acid against Pseudomonas aeruginosa was comparatively assessed with respect to potent quorum sensing inhibitor, Baicalein. At sub-lethal concentration, Cinnamic acid effectively inhibited both the production of the QS-dependent virulence factors and biofilm formation in P. aeruginosa without affecting the viability of the bacterium. The phytocompound interfered with the initial attachment of planktonic cells to the substratum thereby causing reduction in biofilm development. In addition, the in vivo study indicated that the test compound protected Caenorhabditis elegans from the virulence factors of P. aeruginosa leading to reduced mortality. The in silico analysis revealed that Cinnamic acid can act as a competitive inhibitor for the natural ligands towards the ligand binding domain of the transcriptional activators of the quorum sensing circuit in P. aeruginosa, LasR and RhlR. The findings suggest that Cinnamic acid may serve as a novel quorum sensing based anti-infective in controlling P. aeruginosa infections.

  15. Gallium-Protoporphyrin IX Inhibits Pseudomonas aeruginosa Growth by Targeting Cytochromes.

    PubMed

    Hijazi, Sarah; Visca, Paolo; Frangipani, Emanuela

    2017-01-01

    Pseudomonas aeruginosa is a challenging pathogen due to both innate and acquired resistance to antibiotics. It is capable of causing a variety of infections, including chronic lung infection in cystic fibrosis (CF) patients. Given the importance of iron in bacterial physiology and pathogenicity, iron-uptake and metabolism have become attractive targets for the development of new antibacterial compounds. P. aeruginosa can acquire iron from a variety of sources to fulfill its nutritional requirements both in the environment and in the infected host. The adaptation of P. aeruginosa to heme iron acquisition in the CF lung makes heme utilization pathways a promising target for the development of new anti- Pseudomonas drugs. Gallium [Ga(III)] is an iron mimetic metal which inhibits P. aeruginosa growth by interfering with iron-dependent metabolism. The Ga(III) complex of the heme precursor protoporphyrin IX (GaPPIX) showed enhanced antibacterial activity against several bacterial species, although no inhibitory effect has been reported on P. aeruginosa . Here, we demonstrate that GaPPIX is indeed capable of inhibiting the growth of clinical P. aeruginosa strains under iron-deplete conditions, as those encountered by bacteria during infection, and that GaPPIX inhibition is reversed by iron. Using P. aeruginosa PAO1 as model organism, we show that GaPPIX enters cells through both the heme-uptake systems has and phu , primarily via the PhuR receptor which plays a crucial role in P. aeruginosa adaptation to the CF lung. We also demonstrate that intracellular GaPPIX inhibits the aerobic growth of P. aeruginosa by targeting cytochromes, thus interfering with cellular respiration.

  16. The Impact of ExoS on Pseudomonas aeruginosa Internalization by Epithelial Cells Is Independent of fleQ and Correlates with Bistability of Type Three Secretion System Gene Expression

    PubMed Central

    Kroken, Abby R.; Chen, Camille K.; Evans, David J.; Yahr, Timothy L.

    2018-01-01

    ABSTRACT Pseudomonas aeruginosa is internalized into multiple types of epithelial cell in vitro and in vivo and yet is often regarded as an exclusively extracellular pathogen. Paradoxically, ExoS, a type three secretion system (T3SS) effector, has antiphagocytic activities but is required for intracellular survival of P. aeruginosa and its occupation of bleb niches in epithelial cells. Here, we addressed mechanisms for this dichotomy using invasive (ExoS-expressing) P. aeruginosa and corresponding effector-null isogenic T3SS mutants, effector-null mutants of cytotoxic P. aeruginosa with and without ExoS transformation, antibiotic exclusion assays, and imaging using a T3SS-GFP reporter. Except for effector-null PA103, all strains were internalized while encoding ExoS. Intracellular bacteria showed T3SS activation that continued in replicating daughter cells. Correcting the fleQ mutation in effector-null PA103 promoted internalization by >10-fold with or without ExoS. Conversely, mutating fleQ in PAO1 reduced internalization by >10-fold, also with or without ExoS. Effector-null PA103 remained less well internalized than PAO1 matched for fleQ status, but only with ExoS expression, suggesting additional differences between these strains. Quantifying T3SS activation using GFP fluorescence and quantitative reverse transcription-PCR (qRT-PCR) showed that T3SS expression was hyperinducible for strain PA103ΔexoUT versus other isolates and was unrelated to fleQ status. These findings support the principle that P. aeruginosa is not exclusively an extracellular pathogen, with internalization influenced by the relative proportions of T3SS-positive and T3SS-negative bacteria in the population during host cell interaction. These data also challenge current thinking about T3SS effector delivery into host cells and suggest that T3SS bistability is an important consideration in studying P. aeruginosa pathogenesis. PMID:29717012

  17. Light and phosphate competition between Cylindrospermopsis raciborskii and Microcystis aeruginosa is strain dependent.

    PubMed

    Marinho, Marcelo Manzi; Souza, Maria Betânia Gonçalves; Lürling, Miquel

    2013-10-01

    The hypothesis that outcomes of phosphorus and light competition between Cylindrospermopsis raciborskii and Microcystis aeruginosa are strain dependent was tested experimentally. Critical requirements of phosphorus (P*) and of light (I*) of two strains of each species were determined through monoculture experiments, which indicated a trade-off between species and also between Microcystis strains. Competition experiments between species were performed using the weakest predicted competitors (with the highest values of P* and of I*) and with the strongest predicted competitors (with the lowest values of P* and of I*). Under light limitation, competition between the weakest competitors led C. raciborskii to dominate. Between the strongest competitors, the opposite was observed, M. aeruginosa displaced C. raciborskii, but both strains co-existed in equilibrium. Under phosphate limitation, competition between the weakest competitors led C. raciborskii to exclude M. aeruginosa, and between the strongest competitors, the opposite was observed, M. aeruginosa displaced C. raciborskii, but the system did not reach an equilibrium and both strains were washed out. Hence, outcomes of the competition depended on the pair of competing strains and not only on species or on type of limitation. We concluded that existence of different trade-offs among strains and between species underlie our results showing that C. raciborskii can either dominate or be displaced by M. aeruginosa when exposed to different conditions of light or phosphate limitation.

  18. Sublethal Ciprofloxacin Treatment Leads to Rapid Development of High-Level Ciprofloxacin Resistance during Long-Term Experimental Evolution of Pseudomonas aeruginosa

    PubMed Central

    Jørgensen, Karin Meinike; Wassermann, Tina; Jensen, Peter Østrup; Hengzuang, Wang; Molin, Søren; Høiby, Niels

    2013-01-01

    The dynamics of occurrence and the genetic basis of ciprofloxacin resistance were studied in a long-term evolution experiment (940 generations) in wild-type, reference strain (PAO1) and hypermutable (PAOΔmutS and PAOMY-Mgm) P. aeruginosa populations continuously exposed to sub-MICs (1/4) of ciprofloxacin. A rapid occurrence of ciprofloxacin-resistant mutants (MIC of ≥12 μg/ml, representing 100 times the MIC of the original population) were observed in all ciprofloxacin-exposed lineages of PAOΔmutS and PAOMY-Mgm populations after 100 and 170 generations, respectively, and in one of the PAO1 lineages after 240 generations. The genetic basis of resistance was mutations in gyrA (C248T and G259T) and gyrB (C1397A). Cross-resistance to beta-lactam antibiotics was observed in the bacterial populations that evolved during exposure to sublethal concentrations of ciprofloxacin. Our study shows that mutants with high-level ciprofloxacin resistance are selected in P. aeruginosa bacterial populations exposed to sub-MICs of ciprofloxacin. This can have implications for the long-term persistence of resistant bacteria and spread of antibiotic resistance by exposure of commensal bacterial flora to low antibiotic concentrations. PMID:23774442

  19. Computational discovery of putative quorum sensing inhibitors against LasR and RhlR receptor proteins of Pseudomonas aeruginosa

    NASA Astrophysics Data System (ADS)

    Annapoorani, Angusamy; Umamageswaran, Venugopal; Parameswari, Radhakrishnan; Pandian, Shunmugiah Karutha; Ravi, Arumugam Veera

    2012-09-01

    Drugs have been discovered in the past mainly either by identification of active components from traditional remedies or by unpredicted discovery. A key motivation for the study of structure based virtual screening is the exploitation of such information to design targeted drugs. In this study, structure based virtual screening was used in search for putative quorum sensing inhibitors (QSI) of Pseudomonas aeruginosa. The virtual screening programme Glide version 5.5 was applied to screen 1,920 natural compounds/drugs against LasR and RhlR receptor proteins of P. aeruginosa. Based on the results of in silico docking analysis, five top ranking compounds namely rosmarinic acid, naringin, chlorogenic acid, morin and mangiferin were subjected to in vitro bioassays against laboratory strain PAO1 and two more antibiotic resistant clinical isolates, P. aeruginosa AS1 (GU447237) and P. aeruginosa AS2 (GU447238). Among the five compounds studied, except mangiferin other four compounds showed significant inhibition in the production of protease, elastase and hemolysin. Further, all the five compounds potentially inhibited the biofilm related behaviours. This interaction study provided promising ligands to inhibit the quorum sensing (QS) mediated virulence factors production in P. aeruginosa.

  20. Investigating the link between imipenem resistance and biofilm formation by Pseudomonas aeruginosa.

    PubMed

    Musafer, Hadeel K; Kuchma, Sherry L; Naimie, Amanda A; Schwartzman, Joseph D; Al-Mathkhury, Harith J Fahad; O'Toole, George A

    2014-07-01

    Pseudomonas aeruginosa, a ubiquitous environmental organism, is a difficult-to-treat opportunistic pathogen due to its broad-spectrum antibiotic resistance and its ability to form biofilms. In this study, we investigate the link between resistance to a clinically important antibiotic, imipenem, and biofilm formation. First, we observed that the laboratory strain P. aeruginosa PAO1 carrying a mutation in the oprD gene, which confers resistance to imipenem, showed a modest reduction in biofilm formation. We also observed an inverse relationship between imipenem resistance and biofilm formation for imipenem-resistant strains selected in vitro, as well as for clinical isolates. We identified two clinical isolates of P. aeruginosa from the sputum of cystic fibrosis patients that formed robust biofilms, but were sensitive to imipenem (MIC ≤ 2 μg/ml). To test the hypothesis that there is a general link between imipenem resistance and biofilm formation, we performed transposon mutagenesis of these two clinical strains to identify mutants defective in biofilm formation, and then tested these mutants for imipenem resistance. Analysis of the transposon mutants revealed a role for previously described biofilm factors in these clinical isolates of P. aeruginosa, including mutations in the pilY1, pilX, pilW, algC, and pslI genes, but none of the biofilm-deficient mutants became imipenem resistant (MIC ≥ 8 μg/ml), arguing against a general link between biofilm formation and resistance to imipenem. Thus, assessing biofilm formation capabilities of environmental isolates is unlikely to serve as a good predictor of imipenem resistance. We also discuss our findings in light of the limited literature addressing planktonic antibiotic resistance factors that impact biofilm formation.

  1. Antiphagocytic Effect of Slime from a Mucoid Strain of Pseudomonas aeruginosa

    PubMed Central

    Schwarzmann, Stephen; Boring, John R.

    1971-01-01

    Mucoid strains of Pseudomonas aeruginosa produce a viscid slime when grown on the surface of agar media. These strains are known to colonize persistently the tracheobronchial tree of children with cystic fibrosis. Colonization may result from inhibition of phagocytosis due to slime produced by the organism. Slime separated from one mucoid strain was examined to determine whether it possessed antiphagocytic activity in vitro. Cells of P. aeruginosa, Escherichia coli, and Staphylococcus aureus were rapidly phagocytized by rabbit polymorphonuclear leukocytes when mixtures were rotated for 2 hr at 37 C in the absence of slime. The addition of relatively small amounts of slime to bacteria and leukocytes inhibited phagocytosis as measured by phagocytic killing of the organisms. Inhibition was found to be most complete with P. aeruginosa. PMID:16558051

  2. Effect of impaired twitching motility and biofilm dispersion on performance of Pseudomonas aeruginosa-powered microbial fuel cells.

    PubMed

    Shreeram, Devesh D; Panmanee, Warunya; McDaniel, Cameron T; Daniel, Susan; Schaefer, Dale W; Hassett, Daniel J

    2018-02-01

    Pseudomonas aeruginosa is a metabolically voracious bacterium that is easily manipulated genetically. We have previously shown that the organism is also highly electrogenic in microbial fuel cells (MFCs). Polarization studies were performed in MFCs with wild-type strain PAO1 and three mutant strains (pilT, bdlA and pilT bdlA). The pilT mutant was hyperpiliated, while the bdlA mutant was suppressed in biofilm dispersion chemotaxis. The double pilT bdlA mutant was expected to have properties of both mutations. Polarization data indicate that the pilT mutant showed 5.0- and 3.2-fold increases in peak power compared to the wild type and the pilT bdlA mutant, respectively. The performance of the bdlA mutant was surprisingly the lowest, while the pilT bdlA electrogenic performance fell between the pilT mutant and wild-type bacteria. Measurements of biofilm thickness and bacterial viability showed equal viability among the different strains. The thickness of the bdlA mutant, however, was twice that of wild-type strain PAO1. This observation implicates the presence of dead or dormant bacteria in the bdlA mutant MFCs, which increases biofilm internal resistance as confirmed by electrochemical measurements.

  3. Ferritin and ferrihydrite nanoparticles as iron sources for Pseudomonas aeruginosa

    PubMed Central

    Dehner, Carolyn; Morales-Soto, Nydia; Behera, Rabindra K.; Shrout, Joshua; Theil, Elizabeth C.; Maurice, Patricia A.

    2013-01-01

    Metabolism of iron derived from insoluble and/ or scarce sources is essential for pathogenic and environmental microbes. The ability of Pseudomonas aeruginosa to acquire iron from exogenous ferritin was assessed; ferritin is an iron-concentrating and antioxidant protein complex composed of a catalytic protein and caged ferrihydrite nanomineral synthesized from Fe(II) and O2 or H2O2. Ferritin and free ferrihydrite supported growth of P. aeruginosa with indistinguishable kinetics and final culture densities. The P. aeruginosa PAO1 mutant (ΔpvdDΔpchEF), which is incapable of siderophore production, grew as well as the wild type when ferritin was the iron source. Such data suggest that P. aeruginosa can acquire iron by siderophore-independent mechanisms, including secretion of small-molecule reductant(s). Protease inhibitors abolished the growth of the siderophore-free strain on ferritins, with only a small effect on growth of the wild type; predictably, protease inhibitors had no effect on growth with free ferrihydrite as the iron source. Proteolytic activity was higher with the siderophore-free strain, suggesting that the role of proteases in the degradation of ferritin is particularly important for iron acquisition in the absence of siderophores. The combined results demonstrate the importance of both free ferrihydrite, a natural environmental form of iron and a model for an insoluble form of partly denatured ferritin called hemosiderin, and caged ferritin iron minerals as bacterial iron sources. Ferritin is also revealed as a growth promoter of opportunistic, pathogenic bacteria such a P. aeruginosa in diseased tissues such as the cystic fibrotic lung, where ferritin concentrations are abnormally high. PMID:23417538

  4. Gallium-Protoporphyrin IX Inhibits Pseudomonas aeruginosa Growth by Targeting Cytochromes

    PubMed Central

    Hijazi, Sarah; Visca, Paolo; Frangipani, Emanuela

    2017-01-01

    Pseudomonas aeruginosa is a challenging pathogen due to both innate and acquired resistance to antibiotics. It is capable of causing a variety of infections, including chronic lung infection in cystic fibrosis (CF) patients. Given the importance of iron in bacterial physiology and pathogenicity, iron-uptake and metabolism have become attractive targets for the development of new antibacterial compounds. P. aeruginosa can acquire iron from a variety of sources to fulfill its nutritional requirements both in the environment and in the infected host. The adaptation of P. aeruginosa to heme iron acquisition in the CF lung makes heme utilization pathways a promising target for the development of new anti-Pseudomonas drugs. Gallium [Ga(III)] is an iron mimetic metal which inhibits P. aeruginosa growth by interfering with iron-dependent metabolism. The Ga(III) complex of the heme precursor protoporphyrin IX (GaPPIX) showed enhanced antibacterial activity against several bacterial species, although no inhibitory effect has been reported on P. aeruginosa. Here, we demonstrate that GaPPIX is indeed capable of inhibiting the growth of clinical P. aeruginosa strains under iron-deplete conditions, as those encountered by bacteria during infection, and that GaPPIX inhibition is reversed by iron. Using P. aeruginosa PAO1 as model organism, we show that GaPPIX enters cells through both the heme-uptake systems has and phu, primarily via the PhuR receptor which plays a crucial role in P. aeruginosa adaptation to the CF lung. We also demonstrate that intracellular GaPPIX inhibits the aerobic growth of P. aeruginosa by targeting cytochromes, thus interfering with cellular respiration. PMID:28184354

  5. Complexity of resistance mechanisms to imipenem in intensive care unit strains of Pseudomonas aeruginosa.

    PubMed

    Fournier, Damien; Richardot, Charlotte; Müller, Emeline; Robert-Nicoud, Marjorie; Llanes, Catherine; Plésiat, Patrick; Jeannot, Katy

    2013-08-01

    Pseudomonas aeruginosa can become resistant to carbapenems by both intrinsic (mutation-driven) and transferable (β-lactamase-based) mechanisms. Knowledge of the prevalence of these various mechanisms is important in intensive care units (ICUs) in order to define optimal prevention and therapeutic strategies. A total of 109 imipenem-non-susceptible (MIC >4 mg/L) strains of P. aeruginosa were collected in June 2010 from the ICUs of 26 French public hospitals. Their resistance mechanisms were characterized by phenotypic, enzymatic, western blotting and molecular methods. Single or associated imipenem resistance mechanisms were identified among the 109 strains. Seven isolates (6.4%) were found to produce a metallo-β-lactamase (one VIM-1, four VIM-2, one VIM-4 and one IMP-29). Porin OprD was lost in 94 (86.2%) strains as a result of mutations or gene disruption by various insertion sequences (ISPa1635, ISPa1328, IS911, ISPs1, IS51, IS222 and ISPa41). Thirteen other strains were shown to be regulatory mutants in which down-regulation of oprD was coupled with overexpressed efflux pumps CzcCBA (n = 1), MexXY (n = 9) and MexEF-OprN (n = 3). The lack of OprD was due to disruption of the oprD promoter by ISPsy2 in one strain and alteration of the porin signal sequence in another. Imipenem resistance in ICU P. aeruginosa strains may result from multiple mechanisms involving metallo-β-lactamase gene acquisition and genetic events (mutations and ISs) inactivating oprD, turning down its expression while increasing efflux activities or preventing insertion of porin OprD in the outer membrane. This diversity of mechanisms allows P. aeruginosa, more than any other nosocomial pathogen, to rapidly adapt to carbapenems in ICUs.

  6. The Impact of ExoS on Pseudomonas aeruginosa Internalization by Epithelial Cells Is Independent of fleQ and Correlates with Bistability of Type Three Secretion System Gene Expression.

    PubMed

    Kroken, Abby R; Chen, Camille K; Evans, David J; Yahr, Timothy L; Fleiszig, Suzanne M J

    2018-05-01

    Pseudomonas aeruginosa is internalized into multiple types of epithelial cell in vitro and in vivo and yet is often regarded as an exclusively extracellular pathogen. Paradoxically, ExoS, a type three secretion system (T3SS) effector, has antiphagocytic activities but is required for intracellular survival of P. aeruginosa and its occupation of bleb niches in epithelial cells. Here, we addressed mechanisms for this dichotomy using invasive (ExoS-expressing) P. aeruginosa and corresponding effector-null isogenic T3SS mutants, effector-null mutants of cytotoxic P. aeruginosa with and without ExoS transformation, antibiotic exclusion assays, and imaging using a T3SS-GFP reporter. Except for effector-null PA103, all strains were internalized while encoding ExoS. Intracellular bacteria showed T3SS activation that continued in replicating daughter cells. Correcting the fleQ mutation in effector-null PA103 promoted internalization by >10-fold with or without ExoS. Conversely, mutating fleQ in PAO1 reduced internalization by >10-fold, also with or without ExoS. Effector-null PA103 remained less well internalized than PAO1 matched for fleQ status, but only with ExoS expression, suggesting additional differences between these strains. Quantifying T3SS activation using GFP fluorescence and quantitative reverse transcription-PCR (qRT-PCR) showed that T3SS expression was hyperinducible for strain PA103Δ exoUT versus other isolates and was unrelated to fleQ status. These findings support the principle that P. aeruginosa is not exclusively an extracellular pathogen, with internalization influenced by the relative proportions of T3SS-positive and T3SS-negative bacteria in the population during host cell interaction. These data also challenge current thinking about T3SS effector delivery into host cells and suggest that T3SS bistability is an important consideration in studying P. aeruginosa pathogenesis. IMPORTANCE P. aeruginosa is often referred to as an extracellular

  7. C-type natriuretic peptide modulates quorum sensing molecule and toxin production in Pseudomonas aeruginosa

    PubMed Central

    Blier, Anne-Sophie; Veron, Wilfried; Bazire, Alexis; Gerault, Eloïse; Taupin, Laure; Vieillard, Julien; Rehel, Karine; Dufour, Alain; Le Derf, Franck; Orange, Nicole; Hulen, Christian; Feuilloley, Marc G. J.

    2011-01-01

    Pseudomonas aeruginosa coordinates its virulence expression and establishment in the host in response to modification of its environment. During the infectious process, bacteria are exposed to and can detect eukaryotic products including hormones. It has been shown that P. aeruginosa is sensitive to natriuretic peptides, a family of eukaryotic hormones, through a cyclic nucleotide-dependent sensor system that modulates its cytotoxicity. We observed that pre-treatment of P. aeruginosa PAO1 with C-type natriuretic peptide (CNP) increases the capacity of the bacteria to kill Caenorhabditis elegans through diffusive toxin production. In contrast, brain natriuretic peptide (BNP) did not affect the capacity of the bacteria to kill C. elegans. The bacterial production of hydrogen cyanide (HCN) was enhanced by both BNP and CNP whereas the production of phenazine pyocyanin was strongly inhibited by CNP. The amount of 2-heptyl-4-quinolone (HHQ), a precursor to 2-heptyl-3-hydroxyl-4-quinolone (Pseudomonas quinolone signal; PQS), decreased after CNP treatment. The quantity of 2-nonyl-4-quinolone (HNQ), another quinolone which is synthesized from HHQ, was also reduced after CNP treatment. Conversely, both BNP and CNP significantly enhanced bacterial production of acylhomoserine lactone (AHL) [e.g. 3-oxo-dodecanoyl-homoserine lactone (3OC12-HSL) and butanoylhomoserine lactone (C4-HSL)]. These results correlate with an induction of lasI transcription 1 h after bacterial exposure to BNP or CNP. Concurrently, pre-treatment of P. aeruginosa PAO1 with either BNP or CNP enhanced PAO1 exotoxin A production, via a higher toxA mRNA level. At the same time, CNP led to elevated amounts of algC mRNA, indicating that algC is involved in C. elegans killing. Finally, we observed that in PAO1, Vfr protein is essential to the pro-virulent effect of CNP whereas the regulator PtxR supports only a part of the CNP pro-virulent activity. Taken together, these data reinforce the hypothesis that during

  8. A Pseudomonas aeruginosa strain isolated from a contact lens-induced acute red eye (CLARE) is protease-deficient.

    PubMed

    Estrellas, P S; Alionte, L G; Hobden, J A

    2000-03-01

    Pseudomonas aeruginosa proteases are thought to be important virulence factors in the pathogenesis of corneal disease. This study examined protease production from two strains of P. aeruginosa responsible for two very distinct clinical diseases: strain Paer1, isolated from a Contact Lens-induced Acute Red Eye (CLARE), and strain KEI 1025, isolated from a corneal ulcer. Strains were compared to a laboratory strain (ATCC 19660) known to produce severe keratitis in experimentally infected mice for protease production and for ocular virulence. Protease production was examined with colorimetric assays, gelatin zymography and western blots. Elastase A activity was quantitated with a staphylolytic assay. Ocular virulence was examined using a mouse scratch model of keratitis. In contrast to strains KEI 1025 or ATCC 19660, Paer1 was unable to produce enzymatically active elastase A, elastase, and protease IV. All three strains produced active alkaline protease. Strains KEI 1025 and ATCC 19660 produced a fulminant keratitis in mice whereas Paer1 produced a mild transient infection. Restoration of elastase activity in Paer1 via genetic complementation did not result in a virulent phenotype. Co-infection of mouse eyes with strains Paer1 and ATCC 19660 resulted in the eventual loss of Paer1 from corneal tissue. These studies suggest that P. aeruginosa elastase A and/or protease IV, but not alkaline protease or elastase, contribute to the ocular virulence of this organism.

  9. Label-free molecular imaging of bacterial communities of the opportunistic pathogen Pseudomonas aeruginosa

    NASA Astrophysics Data System (ADS)

    Baig, Nameera; Polisetti, Sneha; Morales-Soto, Nydia; Dunham, Sage J. B.; Sweedler, Jonathan V.; Shrout, Joshua D.; Bohn, Paul W.

    2016-09-01

    Biofilms, such as those formed by the opportunistic human pathogen Pseudomonas aeruginosa are complex, matrix enclosed, and surface-associated communities of cells. Bacteria that are part of a biofilm community are much more resistant to antibiotics and the host immune response than their free-floating counterparts. P. aeruginosa biofilms are associated with persistent and chronic infections in diseases such as cystic fibrosis and HIV-AIDS. P. aeruginosa synthesizes and secretes signaling molecules such as the Pseudomonas quinolone signal (PQS) which are implicated in quorum sensing (QS), where bacteria regulate gene expression based on population density. Processes such as biofilms formation and virulence are regulated by QS. This manuscript describes the powerful molecular imaging capabilities of confocal Raman microscopy (CRM) and surface enhanced Raman spectroscopy (SERS) in conjunction with multivariate statistical tools such as principal component analysis (PCA) for studying the spatiotemporal distribution of signaling molecules, secondary metabolites and virulence factors in biofilm communities of P. aeruginosa. Our observations reveal that the laboratory strain PAO1C synthesizes and secretes 2-alkyl-4-hydroxyquinoline N-oxides and 2-alkyl-4-hydroxyquinolones in high abundance, while the isogenic acyl homoserine lactone QS-deficient mutant (ΔlasIΔrhlI) strain produces predominantly 2-alkyl-quinolones during biofilm formation. This study underscores the use of CRM, along with traditional biological tools such as genetics, for studying the behavior of microbial communities at the molecular level.

  10. Royal Jelly Inhibits Pseudomonas aeruginosa Adherence and Reduces Excessive Inflammatory Responses in Human Epithelial Cells

    PubMed Central

    Susilowati, Heni; Amoh, Takashi; Hirao, Kouji; Hirota, Katsuhiko; Matsuo, Takashi; Miyake, Yoichiro

    2017-01-01

    Pseudomonas aeruginosa is a Gram-negative bacterium and causes respiratory infection especially in elderly patients. Royal jelly has been used worldwide as a traditional remedy and as a nutrient; however, the effect against P. aeruginosa is unclear. The aim of this study was to analyze antibacterial, antiadherent, and anti-inflammatory effects of royal jelly against P. aeruginosa. Wild-type strain PAO1 and clinical isolates of P. aeruginosa were used for antibacterial assay and antiadherent assay to abiotic surface and epithelial cells, which are pharynx (Detroit 562) and lung (NCI-H292) epithelial cells. In anti-inflammatory assay, epithelial cells were pretreated with royal jelly before bacterial exposure to investigate its inhibitory effect on interleukin (IL-8) and macrophage inflammatory protein-3α/CCL20 overproduction. Although royal jelly did not have antibacterial activity at concentration of 50% w/v, antiadherent activity was confirmed on the abiotic surface and epithelial cells under concentration of 25%. Pretreatment with royal jelly significantly inhibited overproduction of IL-8 and CCL20 from both cells. These results demonstrated that royal jelly inhibits P. aeruginosa adherence and protects epithelial cells from excessive inflammatory responses against P. aeruginosa infection. Our findings suggested that royal jelly may be a useful supplement as complementary and alternative medicine for preventing respiratory infection caused by P. aeruginosa. PMID:29075644

  11. Royal Jelly Inhibits Pseudomonas aeruginosa Adherence and Reduces Excessive Inflammatory Responses in Human Epithelial Cells.

    PubMed

    Susilowati, Heni; Murakami, Keiji; Yumoto, Hiromichi; Amoh, Takashi; Hirao, Kouji; Hirota, Katsuhiko; Matsuo, Takashi; Miyake, Yoichiro

    2017-01-01

    Pseudomonas aeruginosa is a Gram-negative bacterium and causes respiratory infection especially in elderly patients. Royal jelly has been used worldwide as a traditional remedy and as a nutrient; however, the effect against P. aeruginosa is unclear. The aim of this study was to analyze antibacterial, antiadherent, and anti-inflammatory effects of royal jelly against P. aeruginosa . Wild-type strain PAO1 and clinical isolates of P. aeruginosa were used for antibacterial assay and antiadherent assay to abiotic surface and epithelial cells, which are pharynx (Detroit 562) and lung (NCI-H292) epithelial cells. In anti-inflammatory assay, epithelial cells were pretreated with royal jelly before bacterial exposure to investigate its inhibitory effect on interleukin (IL-8) and macrophage inflammatory protein-3 α /CCL20 overproduction. Although royal jelly did not have antibacterial activity at concentration of 50% w/v, antiadherent activity was confirmed on the abiotic surface and epithelial cells under concentration of 25%. Pretreatment with royal jelly significantly inhibited overproduction of IL-8 and CCL20 from both cells. These results demonstrated that royal jelly inhibits P. aeruginosa adherence and protects epithelial cells from excessive inflammatory responses against P. aeruginosa infection. Our findings suggested that royal jelly may be a useful supplement as complementary and alternative medicine for preventing respiratory infection caused by P. aeruginosa .

  12. Pseudomonas aeruginosa LasB protease impairs innate immunity in mice and humans by targeting a lung epithelial cystic fibrosis transmembrane regulator–IL-6–antimicrobial–repair pathway

    PubMed Central

    Saint-Criq, Vinciane; Villeret, Bérengère; Bastaert, Fabien; Kheir, Saadé; Hatton, Aurélie; Cazes, Aurélie; Xing, Zhou; Sermet-Gaudelus, Isabelle; Garcia-Verdugo, Ignacio; Edelman, Aleksander

    2018-01-01

    Background Pseudomonas aeruginosa lung infections are a huge problem in ventilator-associated pneumonia, cystic fibrosis (CF) and in chronic obstructive pulmonary disease (COPD) exacerbations. This bacterium secretes virulence factors that may subvert host innate immunity. Objective We evaluated the effect of P. aeruginosa elastase LasB, an important virulence factor secreted by the type II secretion system, on ion transport, innate immune responses and epithelial repair, both in vitro and in vivo. Methods Wild-type (WT) or cystic fibrosis transmembrane conductance regulator (CFTR)-mutated epithelial cells (cell lines and primary cells from patients) were treated with WT or ΔLasB pseudomonas aeruginosa O1 (PAO1) secretomes. The effect of LasB and PAO1 infection was also assessed in vivo in murine models. Results We showed that LasB was the most abundant protein in WT PAO1 secretomes and that it decreased epithelial CFTR expression and activity. In airway epithelial cell lines and primary bronchial epithelial cells, LasB degraded the immune mediators interleukin (IL)-6 and trappin-2, an important epithelial-derived antimicrobial molecule. We further showed that an IL-6/STAT3 signalling pathway was downregulated by LasB, resulting in inhibition of epithelial cell repair. In mice, intranasally instillated LasB induced significant weight loss, inflammation, injury and death. By contrast, we showed that overexpression of IL-6 and trappin-2 protected mice against WT-PAO1-induced death, by upregulating IL-17/IL-22 antimicrobial and repair pathways. Conclusions Our data demonstrate that PAO1 LasB is a major P. aeruginosa secreted factor that modulates ion transport, immune response and tissue repair. Targeting this virulence factor or upregulating protective factors such as IL-6 or antimicrobial molecules such as trappin-2 could be beneficial in P. aeruginosa-infected individuals. PMID:28790180

  13. Detection of drug-resistance mechanism of Pseudomonas aeruginosa developing from a sensitive strain to a persister during carbapenem treatment.

    PubMed

    Shen, J L; Fang, Y P

    2015-06-18

    We explored the mechanism of the development from sensitivity to resistance to carbapenem in Pseudomonas aeruginosa. Two P. aeruginosa strains were collected during treatment with carbapenem. Strain homology was investigated using pulsed-field gel electrophoresis. Porin oprD2 expression was analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The minimum inhibitory concentrations (MICs) of imipenem and meropenem with or without MC207110 were determined using the agar dilution method. The expression level of efflux pump mRNA was tested using real-time polymerase chain reaction. Metallo-lactamases (MBLs) were screened using the EDTA-disk synergy test. Genes encoding MBLs were amplified and then analyzed by DNA sequencing. The two treated strains belonged to the same pulsed-field gel electrophoresis type. The SDS-PAGE profile of the P. aeruginosa strains revealed that the 46-kDa membrane protein OprD2 of IMP(R)MEM(R) type strains was lost, whereas OprD2 of 1 IMP(S)MEM(S) strain was normal. With or without MC207110 treatment, the MIC of carbapenem-resistant P. aeruginosa decreased by 4-fold, while the MIC of carbapenem-sensitive P. aeruginosa did not. Compared with the carbapenem-sensitive strain, MexX mRNA expression in the carbapenem-resistant strain increased by 102.5-fold, while the mRNA expression of other efflux pumps did not markedly increase. Neither carbapenem-resistant nor carbapenem-sensitive P. aeruginosa produced MBL. The mechanism of development from sensitivity to resistance of P. aeruginosa to carbapenem during carbapenem treatment is due to porin oprD2 loss and an increased expression level of MexXY-OprM.

  14. NirN Protein from Pseudomonas aeruginosa is a Novel Electron-bifurcating Dehydrogenase Catalyzing the Last Step of Heme d1 Biosynthesis*

    PubMed Central

    Adamczack, Julia; Hoffmann, Martin; Papke, Ulrich; Haufschildt, Kristin; Nicke, Tristan; Bröring, Martin; Sezer, Murat; Weimar, Rebecca; Kuhlmann, Uwe; Hildebrandt, Peter; Layer, Gunhild

    2014-01-01

    Heme d1 plays an important role in denitrification as the essential cofactor of the cytochrome cd1 nitrite reductase NirS. At present, the biosynthesis of heme d1 is only partially understood. The last step of heme d1 biosynthesis requires a so far unknown enzyme that catalyzes the introduction of a double bond into one of the propionate side chains of the tetrapyrrole yielding the corresponding acrylate side chain. In this study, we show that a Pseudomonas aeruginosa PAO1 strain lacking the NirN protein does not produce heme d1. Instead, the NirS purified from this strain contains the heme d1 precursor dihydro-heme d1 lacking the acrylic double bond, as indicated by UV-visible absorption spectroscopy and resonance Raman spectroscopy. Furthermore, the dihydro-heme d1 was extracted from purified NirS and characterized by UV-visible absorption spectroscopy and finally identified by high-resolution electrospray ionization mass spectrometry. Moreover, we show that purified NirN from P. aeruginosa binds the dihydro-heme d1 and catalyzes the introduction of the acrylic double bond in vitro. Strikingly, NirN uses an electron bifurcation mechanism for the two-electron oxidation reaction, during which one electron ends up on its heme c cofactor and the second electron reduces the substrate/product from the ferric to the ferrous state. On the basis of our results, we propose novel roles for the proteins NirN and NirF during the biosynthesis of heme d1. PMID:25204657

  15. Development of a Novel Method for Analyzing Pseudomonas aeruginosa Twitching Motility and Its Application to Define the AmrZ Regulon

    PubMed Central

    Xu, Binjie; Wozniak, Daniel J.

    2015-01-01

    Twitching motility is an important migration mechanism for the Gram-negative bacterium Pseudomonas aeruginosa. In the commonly used subsurface twitching assay, the sub-population of P. aeruginosa with active twitching motility is difficult to harvest for high-throughput studies. Here we describe the development of a novel method that allows efficient isolation of bacterial sub-populations conducting highly active twitching motility. The transcription factor AmrZ regulates multiple P. aeruginosa virulence factors including twitching motility, yet the mechanism of this activation remains unclear. We therefore set out to understand this mechanism by defining the AmrZ regulon using DNA microarrays in combination with the newly developed twitching motility method. We discovered 112 genes in the AmrZ regulon and many encode virulence factors. One gene of interest and the subsequent focus was lecB, which encodes a fucose-binding lectin. DNA binding assays revealed that AmrZ activates lecB transcription by directly binding to its promoter. The lecB gene was previously shown to be required for twitching motility in P. aeruginosa strain PAK; however, our lecB deletion had no effect on twitching motility in strain PAO1. Collectively, in this study a novel condition was developed for quantitative studies of twitching motility, under which the AmrZ regulon was defined. PMID:26309248

  16. A comprehensive multi-omics approach uncovers adaptations for growth and survival of Pseudomonas aeruginosa on n-alkanes.

    PubMed

    Grady, Sarah L; Malfatti, Stephanie A; Gunasekera, Thusitha S; Dalley, Brian K; Lyman, Matt G; Striebich, Richard C; Mayhew, Michael B; Zhou, Carol L; Ruiz, Oscar N; Dugan, Larry C

    2017-04-28

    Examination of complex biological systems has long been achieved through methodical investigation of the system's individual components. While informative, this strategy often leads to inappropriate conclusions about the system as a whole. With the advent of high-throughput "omic" technologies, however, researchers can now simultaneously analyze an entire system at the level of molecule (DNA, RNA, protein, metabolite) and process (transcription, translation, enzyme catalysis). This strategy reduces the likelihood of improper conclusions, provides a framework for elucidation of genotype-phenotype relationships, and brings finer resolution to comparative genomic experiments. Here, we apply a multi-omic approach to analyze the gene expression profiles of two closely related Pseudomonas aeruginosa strains grown in n-alkanes or glycerol. The environmental P. aeruginosa isolate ATCC 33988 consumed medium-length (C 10 -C 16 ) n-alkanes more rapidly than the laboratory strain PAO1, despite high genome sequence identity (average nucleotide identity >99%). Our data shows that ATCC 33988 induces a characteristic set of genes at the transcriptional, translational and post-translational levels during growth on alkanes, many of which differ from those expressed by PAO1. Of particular interest was the lack of expression from the rhl operon of the quorum sensing (QS) system, resulting in no measurable rhamnolipid production by ATCC 33988. Further examination showed that ATCC 33988 lacked the entire lasI/lasR arm of the QS response. Instead of promoting expression of QS genes, ATCC 33988 up-regulates a small subset of its genome, including operons responsible for specific alkaline proteases and sphingosine metabolism. This work represents the first time results from RNA-seq, microarray, ribosome footprinting, proteomics, and small molecule LC-MS experiments have been integrated to compare gene expression in bacteria. Together, these data provide insights as to why strain ATCC 33988

  17. [Persistence of Pseudomonas aeruginosa strains in patients of Federal Scientific Center of Transplantology and Artificial Organs].

    PubMed

    Avetisian, L R; Voronina, O L; Chernukha, M Iu; Kunda, M S; Gabrielian, N I; Lunin, V G; Shaginian, I A

    2012-01-01

    Study genetic diversity of P. aeruginosa strains persisting in patients of Federal Scientific Center of Transplantology and Artificial Organs, and main factors facilitating persistence of strains in the hospital. 136 P. aeruginosa strains isolated from patients of the center for 3 years 6 months were genotyped by RAPD-PCR and MLST methods and studied for antibiotics resistance and presence of integrons. Genetic diversity of strains persisting in hospital was established. Strains of main genotypes ST235, ST446, ST598 were isolated from patients of various surgical departments. Patients were shown to be colonized by these strains during stay in reanimation and intensive therapy department (RITD) of the hospital. Strains of dominant genotype 235 were isolated from 47% of examined patients during more than 3 years. Only genotype 235 strains contained integron with cassettes of antibiotics resistance genes blaGES5 and aadA6 in the genome. The data obtained show that over the period of observation in the center 1 clone of P. aeruginosa that belonged to genotype 235 dominated. This clone was endemic for this hospital and in the process of prolonged persistence became more resistant to antibiotics. Colonization of patients with these strains occurs in RITD. This confirms the necessity of constant monitoring of hospital microflora for advance detection of potentially dangerous epidemic hospital strains able to cause hospital infections.

  18. Iron-Regulated Expression of Alginate Production, Mucoid Phenotype, and Biofilm Formation by Pseudomonas aeruginosa

    PubMed Central

    Wiens, Jacinta R.; Vasil, Adriana I.; Schurr, Michael J.; Vasil, Michael L.

    2014-01-01

    ABSTRACT Pseudomonas aeruginosa strains of non-cystic fibrosis (non-CF) origin do not produce significant amounts of extracellular alginate and are nonmucoid. In CF, such isolates can become mucoid through mutation of one of the genes (mucA, mucB, mucC, or mucD) that produce regulatory factors that sequester AlgU, required for increased expression of alginate genes. Mutation of the muc genes in the nonmucoid PAO1, PA14, PAKS-1, and Ps388 strains led to increased levels of extracellular alginate and an obvious mucoid phenotype, but only under iron-limiting growth conditions (≤5 µM), not under iron-replete conditions (≥10 µM). In contrast, >50% of P. aeruginosa isolates from chronic CF pulmonary infections expressed increased levels of alginate and mucoidy both under iron-limiting and iron-replete conditions (i.e., iron-constitutive phenotype). No single iron regulatory factor (e.g., Fur, PvdS) was associated with this loss of iron-regulated alginate expression and mucoidy in these CF isolates. However, the loss of only pyoverdine production, or its uptake, abrogated the ability of P. aeruginosa to produce a robust biofilm that represents the Psl-type of biofilm. In contrast, we show that mutation of the pyoverdine and pyochelin biosynthesis genes and the pyoverdine receptor (FpvA) lead to iron-constitutive expression of the key alginate biosynthesis gene, algD, and an explicitly mucoid phenotype in both iron-limiting and iron-replete conditions. These data indicate that alginate production and mucoidy, in contrast to other types of biofilms produced by P. aeruginosa, are substantially enhanced under iron limitation. These results also have compelling implications in relation to the use of iron chelators in the treatment of P. aeruginosa CF infections. PMID:24496793

  19. A Bacillus sp. strain with antagonistic activity against Fusarium graminearum kills Microcystis aeruginosa selectively.

    PubMed

    Xuan, Huanling; Dai, Xianzhu; Li, Jing; Zhang, Xiaohui; Yang, Caiyun; Luo, Feng

    2017-04-01

    Cyanobacterial harmful algal blooms (CyanoHABs) cause severe environmental problems, economic losses and threaten human health seriously. In the present study, a Bacillus sp. strain, designated as AF-1, with strong antagonistic activity against plant pathogenic fungus Fusarium graminearum was isolated from purple soil. Bacillus sp. AF-1 selectively killed Microcystis aeruginosa at low cell density (1.6×10 3 cfu/mL), and showed the strongest bactericidal activity against M. aeruginosa NIES-843 (A e =93%, t=6d). The algicidal substances originated from strain AF-1 were stable in the temperature range of 35-100°C, and pH range of 3-11. Cell-free filtrate of AF-1 culture caused excessive accumulation of intracellular reactive oxygen species (ROS), cell death and the efflux of intracellular components of M. aeruginosa NIES-843 cells. The expression of genes recA, psbA1, psbD1, rbcL and mcyB, involved in DNA repair, photosynthesis and microcystin synthesis of NIES 843, were significantly influenced by the cell-free filtrate of AF-1 culture. Bacillus sp. AF-1 has the potential to be developed as a bifunctional biocontrol agent to control CyanoHABs and F. graminearum caused plant disease. Copyright © 2017 Elsevier B.V. All rights reserved.

  20. In-Vivo Expression Profiling of Pseudomonas aeruginosa Infections Reveals Niche-Specific and Strain-Independent Transcriptional Programs

    PubMed Central

    Bielecki, Piotr; Puchałka, Jacek; Wos-Oxley, Melissa L.; Loessner, Holger; Glik, Justyna; Kawecki, Marek; Nowak, Mariusz; Tümmler, Burkhard; Weiss, Siegfried; dos Santos, Vítor A. P. Martins

    2011-01-01

    Pseudomonas aeruginosa is a threatening, opportunistic pathogen causing disease in immunocompromised individuals. The hallmark of P. aeruginosa virulence is its multi-factorial and combinatorial nature. It renders such bacteria infectious for many organisms and it is often resistant to antibiotics. To gain insights into the physiology of P. aeruginosa during infection, we assessed the transcriptional programs of three different P. aeruginosa strains directly after isolation from burn wounds of humans. We compared the programs to those of the same strains using two infection models: a plant model, which consisted of the infection of the midrib of lettuce leaves, and a murine tumor model, which was obtained by infection of mice with an induced tumor in the abdomen. All control conditions of P. aeruginosa cells growing in suspension and as a biofilm were added to the analysis. We found that these different P. aeruginosa strains express a pool of distinct genetic traits that are activated under particular infection conditions regardless of their genetic variability. The knowledge herein generated will advance our understanding of P. aeruginosa virulence and provide valuable cues for the definition of prospective targets to develop novel intervention strategies. PMID:21931663

  1. In-vivo expression profiling of Pseudomonas aeruginosa infections reveals niche-specific and strain-independent transcriptional programs.

    PubMed

    Bielecki, Piotr; Puchałka, Jacek; Wos-Oxley, Melissa L; Loessner, Holger; Glik, Justyna; Kawecki, Marek; Nowak, Mariusz; Tümmler, Burkhard; Weiss, Siegfried; dos Santos, Vítor A P Martins

    2011-01-01

    Pseudomonas aeruginosa is a threatening, opportunistic pathogen causing disease in immunocompromised individuals. The hallmark of P. aeruginosa virulence is its multi-factorial and combinatorial nature. It renders such bacteria infectious for many organisms and it is often resistant to antibiotics. To gain insights into the physiology of P. aeruginosa during infection, we assessed the transcriptional programs of three different P. aeruginosa strains directly after isolation from burn wounds of humans. We compared the programs to those of the same strains using two infection models: a plant model, which consisted of the infection of the midrib of lettuce leaves, and a murine tumor model, which was obtained by infection of mice with an induced tumor in the abdomen. All control conditions of P. aeruginosa cells growing in suspension and as a biofilm were added to the analysis. We found that these different P. aeruginosa strains express a pool of distinct genetic traits that are activated under particular infection conditions regardless of their genetic variability. The knowledge herein generated will advance our understanding of P. aeruginosa virulence and provide valuable cues for the definition of prospective targets to develop novel intervention strategies.

  2. Pleiotropic effects of temperature-regulated 2-OH-lauroytransferase (PA0011) on Pseudomonas aeruginosa antibiotic resistance, virulence and type III secretion system.

    PubMed

    Wang, Bobo; Li, Bo; Liang, Ying; Li, Jing; Gao, Lang; Chen, Lin; Duan, Kangmin; Shen, Lixin

    2016-02-01

    Pseudomonas aeruginosa is an important human pathogen which adapts to changing environment, such as temperature variations and entering host by regulating their gene expression. Here, we report that gene PA0011 in P. aeruginosa PAO1, which encodes a 2-OH-lauroytransferase participating in lipid A biosynthesis, is involved in carbapenem resistance and virulence in a temperature-regulated manner in PAO1. The expression of PA0011 was higher at an environment temperature (21 °C) than that at a body temperature (37 °C). The inactivation of PA0011 rendered increased antibiotic susceptibility and decreased virulence both in vivo and in vitro. The impaired integrity and the decreased stability of the outer membrane were the cause of the increased susceptibility of PAO1(Δ0011) to carbapenem and many other common antibiotics. The reduced endotoxic activity of lipopolysaccharide (LPS) contributed to the decreased virulence both at 21 °C and 37 °C in PAO1 (Δ0011). In addition, we have found that PA0011 repressed the expression of TTSS virulence factors both at transcriptional and translational levels, similar to the effect of O antigen of LPS but unlike any effect of its homologue reported in other bacteria. The effect of PA0011 on resistance to many antibiotics including carbapenem and virulence in P. aeruginosa makes it a target for novel antimicrobial therapies. Copyright © 2015 Elsevier Ltd. All rights reserved.

  3. Novel drug targets in cell wall biosynthesis exploited by gene disruption in Pseudomonas aeruginosa.

    PubMed

    Elamin, Ayssar A; Steinicke, Susanne; Oehlmann, Wulf; Braun, Yvonne; Wanas, Hanaa; Shuralev, Eduard A; Huck, Carmen; Maringer, Marko; Rohde, Manfred; Singh, Mahavir

    2017-01-01

    For clinicians, Pseudomonas aeruginosa is a nightmare pathogen that is one of the top three causes of opportunistic human infections. Therapy of P. aeruginosa infections is complicated due to its natural high intrinsic resistance to antibiotics. Active efflux and decreased uptake of drugs due to cell wall/membrane permeability appear to be important issues in the acquired antibiotic tolerance mechanisms. Bacterial cell wall biosynthesis enzymes have been shown to be essential for pathogenicity of Gram-negative bacteria. However, the role of these targets in virulence has not been identified in P. aeruginosa. Here, we report knockout (k.o) mutants of six cell wall biosynthesis targets (murA, PA4450; murD, PA4414; murF, PA4416; ppiB, PA1793; rmlA, PA5163; waaA, PA4988) in P. aeruginosa PAO1, and characterized these in order to find out whether these genes and their products contribute to pathogenicity and virulence of P. aeruginosa. Except waaA k.o, deletion of cell wall biosynthesis targets significantly reduced growth rate in minimal medium compared to the parent strain. The k.o mutants showed exciting changes in cell morphology and colonial architectures. Remarkably, ΔmurF cells became grossly enlarged. Moreover, the mutants were also attenuated in vivo in a mouse infection model except ΔmurF and ΔwaaA and proved to be more sensitive to macrophage-mediated killing than the wild-type strain. Interestingly, the deletion of the murA gene resulted in loss of virulence activity in mice, and the virulence was restored in a plant model by unknown mechanism. This study demonstrates that cell wall targets contribute significantly to intracellular survival, in vivo growth, and pathogenesis of P. aeruginosa. In conclusion, these findings establish a link between cell wall targets and virulence of P. aeruginosa and thus may lead to development of novel drugs for the treatment of P. aeruginosa infection.

  4. Lipotoxin F of Pseudomonas aeruginosa is an AlgU-dependent and alginate-independent outer membrane protein involved in resistance to oxidative stress and adhesion to A549 human lung epithelia.

    PubMed

    Damron, F Heath; Napper, Jennifer; Teter, M Allison; Yu, Hongwei D

    2009-04-01

    Chronic lung infection with P. aeruginosa and excessive neutrophil-associated inflammation are major causes of morbidity and mortality in patients with cystic fibrosis (CF). Overproduction of an exopolysaccharide known as alginate leads to the formation of mucoid biofilms that are resistant to antibiotics and host defences. Alginate overproduction or mucoidy is controlled by a stress-related ECF sigma factor AlgU/T. Mutation in the anti-sigma factor MucA is a known mechanism for conversion to mucoidy. Recently, we showed that inactivation of a kinase (KinB) in nonmucoid strain PAO1 results in overproduction of alginate. Here, we report the initial characterization of lipotoxin F (LptF, PA3692), an OmpA-like outer membrane protein that exhibited increased expression in the mucoid PAO1kinB mutant. The lipotoxin family of proteins has been previously shown to induce inflammation in lung epithelia, which may play a role in CF disease progression. Expression of LptF was observed to be AlgU-dependent and upregulated in CF isolates. Deletion of lptF from the kinB mutant had no effect on alginate production. Deletion of lptF from PAO1 caused a differential susceptibility to oxidants that can be generated by phagocytes. The lptF and algU mutants were more sensitive to hypochlorite than PAO1. However, the lptF mutant displayed increased resistance to hydrogen peroxide. LptF also contributed to adhesion to A549 human lung epithelial cells. Our data suggest that LptF is an outer membrane protein that may be important for P. aeruginosa survival in harsh environments, including lung colonization in CF.

  5. Lipotoxin F of Pseudomonas aeruginosa is an AlgU-dependent and alginate-independent outer membrane protein involved in resistance to oxidative stress and adhesion to A549 human lung epithelia

    PubMed Central

    Damron, F. Heath; Napper, Jennifer; Teter, M. Allison; Yu, Hongwei D.

    2009-01-01

    Chronic lung infection with P. aeruginosa and excessive neutrophil-associated inflammation are major causes of morbidity and mortality in patients with cystic fibrosis (CF). Overproduction of an exopolysaccharide known as alginate leads to the formation of mucoid biofilms that are resistant to antibiotics and host defences. Alginate overproduction or mucoidy is controlled by a stress-related ECF sigma factor AlgU/T. Mutation in the anti-sigma factor MucA is a known mechanism for conversion to mucoidy. Recently, we showed that inactivation of a kinase (KinB) in nonmucoid strain PAO1 results in overproduction of alginate. Here, we report the initial characterization of lipotoxin F (LptF, PA3692), an OmpA-like outer membrane protein that exhibited increased expression in the mucoid PAO1kinB mutant. The lipotoxin family of proteins has been previously shown to induce inflammation in lung epithelia, which may play a role in CF disease progression. Expression of LptF was observed to be AlgU-dependent and upregulated in CF isolates. Deletion of lptF from the kinB mutant had no effect on alginate production. Deletion of lptF from PAO1 caused a differential susceptibility to oxidants that can be generated by phagocytes. The lptF and algU mutants were more sensitive to hypochlorite than PAO1. However, the lptF mutant displayed increased resistance to hydrogen peroxide. LptF also contributed to adhesion to A549 human lung epithelial cells. Our data suggest that LptF is an outer membrane protein that may be important for P. aeruginosa survival in harsh environments, including lung colonization in CF. PMID:19332805

  6. The host control of a clinical isolate strain of P. aeruginosa infection is independent of Nod-1 but depends on MyD88.

    PubMed

    Sônego, Fabiane; Castanheira, Fernanda V S; Horta, Catarina V; Kanashiro, Alexandre; Czaikoski, Paula G; Zamboni, Dario S; Alves-Filho, José Carlos; Cunha, Fernando Q

    2018-05-01

    The objective of this study was to investigate the role of Nod1 in the recruitment of neutrophils into the infection site and in the establishment of the inflammatory response elicited by a clinical isolate strain of P. aeruginosa in vivo, while comparing it to the well-established role of MyD88 in this process. Wild-type, Nod1 -/- and MyD88 -/- mice, all with a C57Bl/6 background. Mice were intranasally infected with Pseudomonas aeruginosa DZ605. Bronchoalveolar lavage and blood were harvested 6 or 20 h post-infection for evaluating bacterial load, chemokine levels and neutrophil migration. Survival post-infection was also observed. We show here that wild-type and Nod1 -/- mice induce similar lung chemokine levels, neutrophil recruitment, and bacterial load, thus leading to equal survival rates upon P. aeruginosa pulmonary infection. Furthermore, we confirmed the essential role of MyD88-dependent signalling in recruiting neutrophils and controlling P. aeruginosa-induced pulmonary infection. The results suggest that in contrast to MyD88, under our experimental conditions, the absence of Nod1 does not impair the recruitment of neutrophils in response to P. aeruginosa DZ605.

  7. Anti-Quorum Sensing Activity of Forsythia suspense on Chromobacterium violaceum and Pseudomonas aeruginosa.

    PubMed

    Zhang, An; Chu, Wei-Hua

    2017-01-01

    Quorum sensing (QS) plays an important role in the production of virulence factors and pathogenicity in Pseudomonas aeruginosa , and the interruption of QS will be a hopeful pathway to combat bacterial infection. In this study, we selected Forsythia suspense (Thunb.) Vahl from traditional Chinese herbal medicines for its anti-QS activity. Anti-QS of F. suspense extracts (FSE) was monitored using the Chromobacterium violaceum 12472 bioassay. Standard methods were used to investigate the effects of FSE on QS-controlled virulence factors production, swimming motility, and biofilm establishment in P. aeruginosa PAO1. FSE could obviously inhibit the violacein production in C. violaceum 12472 and also could inhibit quorum sensing-regulated virulence factors production and biofilm formation in P. aeruginosa in a concentration-dependent manner. The elastase activity and pyocyanin production were inhibited at a maximum of 40.97 and 47.58% when P. aeruginosa was grown in the presence of 0.25 g/mL FSE, which can also inhibit swimming motility of P. aeruginosa . The biofilm formation ability was decreased about 72.45% when in PAO1 cultured with the 0.25 g/mL FSE. The results suggested that FSE may be used as an alternative drug to control and handle harmful infections caused by bacterial pathogens based on QS inhibition. Forsythia suspense water extract could obviously inhibit the purple pigment production in C. violaceum 12472 Forsythia suspense water extract could inhibit QS-regulated virulence factors production and biofilm formation in P. aeruginosa . Abbreviations used: QS: Quorum sensing, Pseudomonas aeruginosa P. aeruginosa , Forsythia suspense F. suspense , FSE: F. suspense extracts, Chromobacterium violaceum 12472 C. violaceum 12472, AIs: autoinducers, AHLs: N -acyl-homoserinelactones, LB: Luria-Bertani, MICs: Minimum inhibitory concentrations, CFU: Colony-Forming Units, ATCC: American Type Culture Collection, PBS: phosphate buffered saline.

  8. Anti-Quorum Sensing Activity of Forsythia suspense on Chromobacterium violaceum and Pseudomonas aeruginosa

    PubMed Central

    Zhang, An; Chu, Wei-Hua

    2017-01-01

    Background: Quorum sensing (QS) plays an important role in the production of virulence factors and pathogenicity in Pseudomonas aeruginosa, and the interruption of QS will be a hopeful pathway to combat bacterial infection. Objective: In this study, we selected Forsythia suspense (Thunb.) Vahl from traditional Chinese herbal medicines for its anti-QS activity. Materials and Methods: Anti-QS of F. suspense extracts (FSE) was monitored using the Chromobacterium violaceum 12472 bioassay. Standard methods were used to investigate the effects of FSE on QS-controlled virulence factors production, swimming motility, and biofilm establishment in P. aeruginosa PAO1. Results: FSE could obviously inhibit the violacein production in C. violaceum 12472 and also could inhibit quorum sensing–regulated virulence factors production and biofilm formation in P. aeruginosa in a concentration-dependent manner. The elastase activity and pyocyanin production were inhibited at a maximum of 40.97 and 47.58% when P. aeruginosa was grown in the presence of 0.25 g/mL FSE, which can also inhibit swimming motility of P. aeruginosa. The biofilm formation ability was decreased about 72.45% when in PAO1 cultured with the 0.25 g/mL FSE. The results suggested that FSE may be used as an alternative drug to control and handle harmful infections caused by bacterial pathogens based on QS inhibition. SUMMARY Forsythia suspense water extract could obviously inhibit the purple pigment production in C. violaceum 12472Forsythia suspense water extract could inhibit QS-regulated virulence factors production and biofilm formation in P. aeruginosa. Abbreviations used: QS: Quorum sensing, Pseudomonas aeruginosa P. aeruginosa, Forsythia suspense F. suspense, FSE: F. suspense extracts, Chromobacterium violaceum 12472 C. violaceum 12472, AIs: autoinducers, AHLs: N-acyl-homoserinelactones, LB: Luria-Bertani, MICs: Minimum inhibitory concentrations, CFU: Colony-Forming Units, ATCC: American Type Culture Collection

  9. CrpP Is a Novel Ciprofloxacin-Modifying Enzyme Encoded by the Pseudomonas aeruginosa pUM505 Plasmid.

    PubMed

    Chávez-Jacobo, Víctor M; Hernández-Ramírez, Karen C; Romo-Rodríguez, Pamela; Pérez-Gallardo, Rocío Viridiana; Campos-García, Jesús; Gutiérrez-Corona, J Félix; García-Merinos, Juan Pablo; Meza-Carmen, Víctor; Silva-Sánchez, Jesús; Ramírez-Díaz, Martha I

    2018-06-01

    The pUM505 plasmid, isolated from a clinical Pseudomonas aeruginosa isolate, confers resistance to ciprofloxacin (CIP) when transferred into the standard P. aeruginosa strain PAO1. CIP is an antibiotic of the quinolone family that is used to treat P. aeruginosa infections. In silico analysis, performed to identify CIP resistance genes, revealed that the 65-amino-acid product encoded by the orf131 gene in pUM505 displays 40% amino acid identity to the Mycobacterium smegmatis aminoglycoside phosphotransferase (an enzyme that phosphorylates and inactivates aminoglycoside antibiotics). We cloned orf131 (renamed crpP , for c iprofloxacin r esistance p rotein, p lasmid encoded) into the pUCP20 shuttle vector. The resulting recombinant plasmid, pUC- crpP , conferred resistance to CIP on Escherichia coli strain J53-3, suggesting that this gene encodes a protein involved in CIP resistance. Using coupled enzymatic analysis, we determined that the activity of CrpP on CIP is ATP dependent, while little activity against norfloxacin was detected, suggesting that CIP may undergo phosphorylation. Using a recombinant His-tagged CrpP protein and liquid chromatography-tandem mass spectrometry, we also showed that CIP was phosphorylated prior to its degradation. Thus, our findings demonstrate that CrpP, encoded on the pUM505 plasmid, represents a new mechanism of CIP resistance in P. aeruginosa , which involves phosphorylation of the antibiotic. Copyright © 2018 American Society for Microbiology.

  10. RESULTS OF MONITORING METALLO-BETA-LACTAMASE-PRODUCING STRAINS OF PSEUDOMONAS AERUGINOSA IN A MULTI-PROFILE HOSPITAL.

    PubMed

    Shamaeva, S K; Portnyagina, U S; Edelstein, M V; Kuzmina, A A; Maloguloval, S; Varfolomeeva, N A

    2015-01-01

    The authors present the results of long-term monitoring of metallo-beta-lactamase (MBL) producing strains of Pseudomonas aeruginosa in the Republican Hospital No 2 of Yakutsk, Russian Federation. Hospitals across Russia, as well as the rest of the world, face a rapid appearance and a virtually unchecked spread of multiresistant and panresistant nosocomial pathogens. Especially prevalent are multidrug-resistant isolates of P. aeruginosa, most often found among the patients of intensive care and intensive therapy units, as well as surgery departments. The aim of this study is to investigate the prevalence of metallo-beta-lactamase-producing strains of P. aeruginosa in a multi-profile hospital. 2,135 isolates of P. aeruginosa were studied, collected during a time span of seven years (2008-2014) from clinical specimens of hospitalised patients in acute surgery, purulent surgery, neurosurgery, otolaryngology, coloproctology departments, intensive care and intensive therapy, burn units, as well as intensive care unit for patients with acute cerebrovascular accidents and coronary care unit. Strains were identified and re-identified using established methods, NEFERMtest 24 (MICROLATEST) biochemical microtest and API (bioMerieux) test systems were used. For all carbapenem-resistant strains a phenotype screening for MBL was performed using the double-disks method with EDTA. In order to identify VIM-type and IMP-type MBL genes a real-time multiplex polymerase chain reaction was used. Among the investigated strains the largest number of P. aeruginosa - 35.6% (761 isolates) was found in patients at intensive care and intensive therapy units. Clonal expansion of extensively drug-resistant strain P. aeruginosa ST235 (VIM-2) was determined, the resistance mechanism of which is connected to MBL. Sensitivity determination of MBL-producing isolates of P. aeruginosa has shown that isolated strains have a high level of resistance (100%) to all tested antibacterial agents: piperacillin

  11. Biosynthesis of the Common Polysaccharide Antigen of Pseudomonas aeruginosa PAO1: Characterization and Role of GDP-D-Rhamnose:GlcNAc/GalNAc-Diphosphate-Lipid α1,3-D-Rhamnosyltransferase WbpZ.

    PubMed

    Wang, Shuo; Hao, Youai; Lam, Joseph S; Vlahakis, Jason Z; Szarek, Walter A; Vinnikova, Anna; Veselovsky, Vladimir V; Brockhausen, Inka

    2015-06-15

    The opportunistic pathogen Pseudomonas aeruginosa produces two major cell surface lipopolysaccharides, characterized by distinct O antigens, called common polysaccharide antigen (CPA) and O-specific antigen (OSA). CPA contains a polymer of D-rhamnose (D-Rha) in α1-2 and α1-3 linkages. Three putative glycosyltransferase genes, wbpX, wbpY, and wbpZ, are part of the CPA biosynthesis cluster. To characterize the enzymatic function of the wbpZ gene product, we chemically synthesized the donor substrate GDP-D-Rha and enzymatically synthesized GDP-D-[(3)H]Rha. Using nuclear magnetic resonance (NMR) spectroscopy, we showed that WbpZ transferred one D-Rha residue from GDP-D-Rha in α1-3 linkage to both GlcNAc- and GalNAc-diphosphate-lipid acceptor substrates. WbpZ is also capable of transferring D-mannose (D-Man) to these acceptors. Therefore, WbpZ has a relaxed specificity with respect to both acceptor and donor substrates. The diphosphate group of the acceptor, however, is required for activity. WbpZ does not require divalent metal ion for activity and exhibits an unusually high pH optimum of 9. WbpZ from PAO1 is therefore a GDP-D-Rha:GlcNAc/GalNAc-diphosphate-lipid α1,3-D-rhamnosyltransferase that has significant activity of GDP-D-Man:GlcNAc/GalNAc-diphosphate-lipid α1,3-D-mannosyltransferase. We used site-directed mutagenesis to replace the Asp residues of the two DXD motifs with Ala. Neither of the mutant constructs of wbpZ (D172A or D254A) could be used to rescue CPA biosynthesis in the ΔwbpZ knockout mutant in a complementation assay. This suggested that D172 and D254 are essential for WbpZ function. This work is the first detailed characterization study of a D-Rha-transferase and a critical step in the development of CPA synthesis inhibitors. This is the first characterization of a D-rhamnosyltransferase and shows that it is essential in Pseudomonas aeruginosa for the synthesis of the common polysaccharide antigen. Copyright © 2015, American Society for

  12. Role of Pseudomonas aeruginosa low-molecular-mass penicillin-binding proteins in AmpC expression, β-lactam resistance, and peptidoglycan structure.

    PubMed

    Ropy, Alaa; Cabot, Gabriel; Sánchez-Diener, Irina; Aguilera, Cristian; Moya, Bartolome; Ayala, Juan A; Oliver, Antonio

    2015-07-01

    This study aimed to characterize the role of Pseudomonas aeruginosa low-molecular-mass penicillin-binding proteins (LMM PBPs), namely, PBP4 (DacB), PBP5 (DacC), and PBP7 (PbpG), in peptidoglycan composition, β-lactam resistance, and ampC regulation. For this purpose, we constructed all single and multiple mutants of dacB, dacC, pbpG, and ampC from the wild-type P. aeruginosa PAO1 strain. Peptidoglycan composition was determined by high-performance liquid chromatography (HPLC), ampC expression by reverse transcription-PCR (RT-PCR), PBP patterns by a Bocillin FL-binding test, and antimicrobial susceptibility by MIC testing for a panel of β-lactams. Microscopy and growth rate analyses revealed no apparent major morphological changes for any of the mutants compared to the wild-type PAO1 strain. Of the single mutants, only dacC mutation led to significantly increased pentapeptide levels, showing that PBP5 is the major dd-carboxypeptidase in P. aeruginosa. Moreover, our results indicate that PBP4 and PBP7 play a significant role as dd-carboxypeptidase only if PBP5 is absent, and their dd-endopeptidase activity is also inferred. As expected, the inactivation of PBP4 led to a significant increase in ampC expression (around 50-fold), but, remarkably, the sequential inactivation of the three LMM PBPs produced a much greater increase (1,000-fold), which correlated with peptidoglycan pentapeptide levels. Finally, the β-lactam susceptibility profiles of the LMM PBP mutants correlated well with the ampC expression data. However, the inactivation of ampC in these mutants also evidenced a role of LMM PBPs, especially PBP5, in intrinsic β-lactam resistance. In summary, in addition to assessing the effect of P. aeruginosa LMM PBPs on peptidoglycan structure for the first time, we obtained results that represent a step forward in understanding the impact of these PBPs on β-lactam resistance, apparently driven by the interplay between their roles in AmpC induction,

  13. [Antibiotic susceptibility and occurrence of ESBL, IBL and MBL in Pseudomonas aeruginosa strains].

    PubMed

    Wolska, Katarzyna; Jakubczak, Antoni; Soszyńska, Agnieszka

    2008-01-01

    The aim of this study was to evaluate the drug susceptibility of P. aeruginosa strains and to detect strains producing inducible beta-lactamases (IBL), extended-spectrum beta-lactamases (ESBL), and metallo-beta-lactamases (MBL). During 6 month (October 2005 - March 2006), 66 strains of P. aeruginosa strains were cultured from clinical specimens obtained from patients of two of hospitals in Siedlce and from patients of outpatient clinics. All the strains were identified in the automatic ATB (bio Mérieux). The susceptibility of bacteria to antibiotics was tested by standard disc diffusion method. The majority of strains were susceptible to meropenem (89.4%), piperacillin combined with tazobactam (84.8%), ciprofloxacin (84.8%) and piperacillin (83.3%). Many of our strains were resistant to carbenicillin (69.7%), mezlocillin (45.5%), gentamicin (42.4%) and netylmicin (30.3%). 6 strains (9.1%) were multidrug-resistant (MDR). Inducible beta-lactamases were detected with the use double disc method according to Sanders and Sanders. ESBL-producing strains were detected with double disc test (DDST) according to Jarlier et al. These strains were identified as ESBL-positive on the basis of the DDST were also determined using a double disc (DD) test according to Appleton. Production of metallo-beta-lactamases (MBL) was examined with the use of Etest MBL (AB Biodisk, Sweden) and the double disc test according to Arakava et al. Sixty-five IBL-producing strains (98.5% of all strains) and three strains (4.5%) with MBL activity were detected. Strains producing extended beta-lactamases (ESBL) were not found.

  14. Identification of Novel Genomic Islands in Liverpool Epidemic Strain of Pseudomonas aeruginosa Using Segmentation and Clustering

    PubMed Central

    Jani, Mehul; Mathee, Kalai; Azad, Rajeev K.

    2016-01-01

    Pseudomonas aeruginosa is an opportunistic pathogen implicated in a myriad of infections and a leading pathogen responsible for mortality in patients with cystic fibrosis (CF). Horizontal transfers of genes among the microorganisms living within CF patients have led to highly virulent and multi-drug resistant strains such as the Liverpool epidemic strain of P. aeruginosa, namely the LESB58 strain that has the propensity to acquire virulence and antibiotic resistance genes. Often these genes are acquired in large clusters, referred to as “genomic islands (GIs).” To decipher GIs and understand their contributions to the evolution of virulence and antibiotic resistance in P. aeruginosa LESB58, we utilized a recursive segmentation and clustering procedure, presented here as a genome-mining tool, “GEMINI.” GEMINI was validated on experimentally verified islands in the LESB58 strain before examining its potential to decipher novel islands. Of the 6062 genes in P. aeruginosa LESB58, 596 genes were identified to be resident on 20 GIs of which 12 have not been previously reported. Comparative genomics provided evidence in support of our novel predictions. Furthermore, GEMINI unraveled the mosaic structure of islands that are composed of segments of likely different evolutionary origins, and demonstrated its ability to identify potential strain biomarkers. These newly found islands likely have contributed to the hyper-virulence and multidrug resistance of the Liverpool epidemic strain of P. aeruginosa. PMID:27536294

  15. A Novel indole compound that inhibits Pseudomonas aeruginosa growth by targeting MreB is a substrate for MexAB-OprM.

    PubMed

    Robertson, Gregory T; Doyle, Timothy B; Du, Qun; Duncan, Leonard; Mdluli, Khisimuzi E; Lynch, A Simon

    2007-10-01

    Drug efflux systems contribute to the intrinsic resistance of Pseudomonas aeruginosa to many antibiotics and biocides and hamper research focused on the discovery and development of new antimicrobial agents targeted against this important opportunistic pathogen. Using a P. aeruginosa PAO1 derivative bearing deletions of opmH, encoding an outer membrane channel for efflux substrates, and four efflux pumps belonging to the resistance nodulation/cell division class including mexAB-oprM, we identified a small-molecule indole-class compound (CBR-4830) that is inhibitory to growth of this efflux-compromised strain. Genetic studies established MexAB-OprM as the principal pump for CBR-4830 and revealed MreB, a prokaryotic actin homolog, as the proximal cellular target of CBR-4830. Additional studies establish MreB as an essential protein in P. aeruginosa, and efflux-compromised strains treated with CBR-4830 transition to coccoid shape, consistent with MreB inhibition or depletion. Resistance genetics further suggest that CBR-4830 interacts with the putative ATP-binding pocket in MreB and demonstrate significant cross-resistance with A22, a structurally unrelated compound that has been shown to promote rapid dispersion of MreB filaments in vivo. Interestingly, however, ATP-dependent polymerization of purified recombinant P. aeruginosa MreB is blocked in vitro in a dose-dependent manner by CBR-4830 but not by A22. Neither compound exhibits significant inhibitory activity against mutant forms of MreB protein that bear mutations identified in CBR-4830-resistant strains. Finally, employing the strains and reagents prepared and characterized during the course of these studies, we have begun to investigate the ability of analogues of CBR-4830 to inhibit the growth of both efflux-proficient and efflux-compromised P. aeruginosa through specific inhibition of MreB function.

  16. Rapid Classification and Identification of Microcystis aeruginosa Strains Using MALDI-TOF MS and Polygenetic Analysis.

    PubMed

    Sun, Li-Wei; Jiang, Wen-Jing; Sato, Hiroaki; Kawachi, Masanobu; Lu, Xi-Wu

    2016-01-01

    Matrix-assisted laser desorption-ionization-time-of-flight mass spectrometry (MALDI-TOF MS) was used to establish a rapid, simple, and accurate method to differentiate among strains of Microcystis aeruginosa, one of the most prevalent types of bloom-forming cyanobacteria. M. aeruginosa NIES-843, for which a complete genome has been sequenced, was used to characterize ribosomal proteins as biomarkers and to optimize conditions for observing ribosomal proteins as major peaks in a given mass spectrum. Thirty-one of 52 ribosomal subunit proteins were detected and identified along the mass spectrum. Fifty-five strains of M. aeruginosa from different habitats were analyzed using MALDI-TOF MS; among these samples, different ribosomal protein types were observed. A polygenetic analysis was performed using an unweighted pair-group method with arithmetic means and different ribosomal protein types to classify the strains into five major clades. Two clades primarily contained toxic strains, and the other three clades contained exclusively non-toxic strains. This is the first study to differentiate cyanobacterial strains using MALDI-TOF MS.

  17. Rapid detection of Pseudomonas aeruginosa biomarkers in biological fluids using surface-enhanced Raman scattering

    NASA Astrophysics Data System (ADS)

    Wu, Xiaomeng; Chen, Jing; Zhao, Yiping; Zughaier, Susu M.

    2014-05-01

    Pseudomonas aeruginosa (PA) is an opportunistic pathogen that causes major infection not only in Cystic Fibrosis patients but also in chronic obstructive pulmonary disease and in critically ill patients in intensive care units. Successful antibiotic treatment of the infection relies on accurate and rapid identification of the infectious agents. Conventional microbiological detection methods usually take more than 3 days to obtain accurate results. We have developed a rapid diagnostic technique based on surface-enhanced Raman scattering to directly identify PA from biological fluids. P. aeruginosa strains, PAO1 and PA14, are cultured in lysogeny broth, and the SERS spectra of the broth show the signature Raman peaks from pyocyanin and pyoverdine, two major biomarkers that P. aeruginosa secretes during its growth, as well as lipopolysaccharides. This provides the evidence that the presence of these biomarkers can be used to indicate P. aeruginosa infection. A total of 22 clinical exhaled breath condensates (EBC) samples were obtained from subjects with CF disease and from non-CF healthy donors. SERS spectra of these EBC samples were obtained and further analyzed by both principle component analysis and partial least square-discriminant analysis (PLS-DA). PLS-DA can discriminate the samples with P. aeruginosa infection and the ones without P. aeruginosa infection at 99.3% sensitivity and 99.6% specificity. In addition, this technique can also discriminate samples from subject with CF disease and healthy donor with 97.5% sensitivity and 100% specificity. These results demonstrate the potential of using SERS of EBC samples as a rapid diagnostic tool to detect PA infection.

  18. Films of Bacteria at Interfaces (FBI): Remodeling of Fluid Interfaces by Pseudomonas aeruginosa.

    PubMed

    Niepa, Tagbo H R; Vaccari, Liana; Leheny, Robert L; Goulian, Mark; Lee, Daeyeon; Stebe, Kathleen J

    2017-12-19

    Bacteria at fluid interfaces endure physical and chemical stresses unique to these highly asymmetric environments. The responses of Pseudomonas aeruginosa PAO1 and PA14 to a hexadecane-water interface are compared. PAO1 cells form elastic films of bacteria, excreted polysaccharides and proteins, whereas PA14 cells move actively without forming an elastic film. Studies of PAO1 mutants show that, unlike solid-supported biofilms, elastic interfacial film formation occurs in the absence of flagella, pili, or certain polysaccharides. Highly induced genes identified in transcriptional profiling include those for putative enzymes and a carbohydrate metabolism enzyme, alkB2; this latter gene is not upregulated in PA14 cells. Notably, PAO1 mutants lacking the alkB2 gene fail to form an elastic layer. Rather, they form an active film like that formed by PA14. These findings demonstrate that genetic expression is altered by interfacial confinement, and suggest that the ability to metabolize alkanes may play a role in elastic film formation at oil-water interfaces.

  19. Biotoxic impact of heavy metals on growth, oxidative stress and morphological changes in root structure of wheat (Triticum aestivum L.) and stress alleviation by Pseudomonas aeruginosa strain CPSB1.

    PubMed

    Rizvi, Asfa; Khan, Mohd Saghir

    2017-10-01

    Rapid industrialization and uncontrolled metal discharge into environment is a global concern for crop production. Metal tolerant bacterium isolated from chilli rhizosphere was identified as Pseudomonas aeruginosa by 16S rDNA sequence analysis. Pseudomonas aeruginosa tolerated high concentrations of Cu (1400 μg ml -1 ), Cd (1000 μg ml -1 ) and Cr (1000 μg ml -1 ). Pseudomonas aeruginosa CPSB1 produced multiple plant growth promoting biomolecules in the presence and absence of metals. Strain CPSB1 solubilized P at 400 μg ml -1 of Cd, Cr and Cu. The strain was positive for indole-3-acetic acid (IAA), siderophores, hydrogen cyanide (HCN), ammonia (NH 3 ) and 1-aminocyclopropane-1-carboxylate (ACC) deaminase when grown with/without metals. The phytotoxic effects on wheat increased with increasing Cd, Cr and Cu rates. The P. aeruginosa CPSB1 inoculated wheat in contrast had better growth and yields under Cu, Cd and Cr stress. The root dry biomass of inoculated plants was enhanced by 44, 28 and 48% at 2007 mg Cu kg -1 , 36 mg Cd kg -1 and 204 mg Cr kg -1 , respectively. The bioinoculant enhanced number of spikes, grain and straw yields by 25, 17 and 12%, respectively. Pseudomonas aeruginosa CPSB1 significantly declined the levels of catalase (CAT), glutathione reductase (GR) and superoxide dismutase SOD), proline and malondialdehyde (MDA), and reduced metal uptake by wheat. The study demonstrated that P. aeruginosa CPSB1 possessed plant growth promoting potentials, showed metal tolerance capability and had ability to counteract deleterious metal impacts. Due to these, P. aeruginosa CPSB1 could be used as bioinoculant for enhancing wheat production even in metal contaminated soils. Copyright © 2017 Elsevier Ltd. All rights reserved.

  20. Catheter-Associated Urinary Tract Infection by Pseudomonas aeruginosa Is Mediated by Exopolysaccharide-Independent Biofilms

    PubMed Central

    Cole, Stephanie J.; Records, Angela R.; Orr, Mona W.; Linden, Sara B.

    2014-01-01

    Pseudomonas aeruginosa is an opportunistic human pathogen that is especially adept at forming surface-associated biofilms. P. aeruginosa causes catheter-associated urinary tract infections (CAUTIs) through biofilm formation on the surface of indwelling catheters. P. aeruginosa encodes three extracellular polysaccharides, PEL, PSL, and alginate, and utilizes the PEL and PSL polysaccharides to form biofilms in vitro; however, the requirement of these polysaccharides during in vivo infections is not well understood. Here we show in a murine model of CAUTI that PAO1, a strain harboring pel, psl, and alg genes, and PA14, a strain harboring pel and alg genes, form biofilms on the implanted catheters. To determine the requirement of exopolysaccharide during in vivo biofilm infections, we tested isogenic mutants lacking the pel, psl, and alg operons and showed that PA14 mutants lacking these operons can successfully form biofilms on catheters in the CAUTI model. To determine the host factor(s) that induces the ΔpelD mutant to form biofilm, we tested mouse, human, and artificial urine and show that urine can induce biofilm formation by the PA14 ΔpelD mutant. By testing the major constituents of urine, we show that urea can induce a pel-, psl-, and alg-independent biofilm. These pel-, psl-, and alg-independent biofilms are mediated by the release of extracellular DNA. Treatment of biofilms formed in urea with DNase I reduced the biofilm, indicating that extracellular DNA supports biofilm formation. Our results indicate that the opportunistic pathogen P. aeruginosa utilizes a distinct program to form biofilms that are independent of exopolysaccharides during CAUTI. PMID:24595142

  1. Assessment of the Effects of Light Availability on Growth and Competition Between Strains of Planktothrix agardhii and Microcystis aeruginosa.

    PubMed

    Torres, Camila de Araujo; Lürling, Miquel; Marinho, Marcelo Manzi

    2016-05-01

    In this study, we tested the hypothesis that Planktothrix agardhii strains isolated from a tropical water body were better competitors for light than Microcystis aeruginosa strains. These cyanobacteria are common in eutrophic systems, where light is one of the main drivers of phytoplankton, and Planktothrix is considered more shade-adapted and Microcystis more high-light tolerant. First, the effect of light intensities on growth was studied in batch cultures. Next, the minimum requirement of light (I*) and the effect of light limitation on the outcome of competition was investigated in chemostats. All strains showed similar growth at 10 μmol photons m(-2) s(-1), demonstrating the ability of the two species to grow in low light. The optimum light intensity was lower for P. agardhii, but at the highest light intensity, Microcystis strains reached higher biovolume, confirming that P. agardhii has higher sensitivity to high light. Nonetheless, P. agardhii grew in light intensities considered high (500 μmol photons m(-2) s(-1)) for this species. M. aeruginosa showed a higher carrying capacity in light-limited condition, but I* was similar between all the strains. Under light competition, Microcystis strains displaced P. agardhii and dominated. In two cases, there was competitive exclusion and in the other two P. agardhii managed to remain in the system with a low biovolume (≈15%). Our findings not only show that strains of P. agardhii can grow under higher light intensities than generally assumed but also that strains of M. aeruginosa are better competitors for light than supposed. These results help to understand the co-occurrence of these species in tropical environments and the dominance of M. aeruginosa even in low-light conditions.

  2. Biosorption of uranium by Pseudomonas aeruginosa strain CSU: Characterization and comparison studies

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Hu, M.Z.C.; Norman, J.M.; Faison, B.D.

    1996-07-20

    Pseudomonas aeruginosa strain CSU, a nongenetically engineered bacterial strain known to bind dissolved hexavalent uranium (as UO{sub 2}{sup 2+} and/or its cationic hydroxo complexes) was characterized with respect to its sorptive activity. The uranium biosorption equilibrium could be described by the Langmuir isotherm. The rate of uranium adsorption increased following permeabilization of the outer and/or cytoplasmic membrane by organic solvents such as acetone. P. aeruginosa CSU biomass was significantly more sorptive toward uranium than certain novel, patented biosorbents derived from algal or fungal biomass sources. P. aeruginosa CSU biomass was also competitive with commercial cation-exchange resins, particularly in the presencemore » of dissolved transition metals. Uranium binding by P. aeruginosa CSU was clearly pH dependent. Uranium loading capacity increased with increasing pH under acidic conditions, presumably as a function of uranium speciation and due to the H{sup +} competition at some binding sites. Nevertheless, preliminary evidence suggests that this microorganism is also capable of binding anionic hexavalent uranium complexes. Ferric iron was a strong inhibitor of uranium binding to P. aeruginosa CSU biomass, and the presence of uranium also decreased the Fe{sup 3+} loading when the biomass was not saturated with Fe{sup 3+}. Thus, a two-state process in which iron and uranium are removed in consecutive steps was proposed for efficient use of the biomass as a biosorbent in uranium removal from mine wastewater, especially acidic leachates.« less

  3. A comprehensive multi-omics approach uncovers adaptations for growth and survival of Pseudomonas aeruginosa on n-alkanes

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Grady, Sarah L.; Malfatti, Stephanie A.; Gunasekera, Thusitha S.

    Examination of complex biological systems has long been achieved through methodical investigation of the system’s individual components. While informative, this strategy often leads to inappropriate conclusions about the system as a whole. With the advent of high-throughput “omic” technologies, but, researchers can now simultaneously analyze an entire system at the level of molecule (DNA, RNA, protein, metabolite) and process (transcription, translation, enzyme catalysis). This strategy reduces the likelihood of improper conclusions, provides a framework for elucidation of genotype-phenotype relationships, and brings finer resolution to comparative genomic experiments. Here in this paper, we apply a multi-omic approach to analyze the genemore » expression profiles of two closely related Pseudomonas aeruginosa strains grown in n-alkanes or glycerol. The environmental P. aeruginosa isolate ATCC 33988 consumed medium-length (C 10–C 16) n-alkanes more rapidly than the laboratory strain PAO1, despite high genome sequence identity (average nucleotide identity >99%). Our data shows that ATCC 33988 induces a characteristic set of genes at the transcriptional, translational and post-translational levels during growth on alkanes, many of which differ from those expressed by PAO1. Of particular interest was the lack of expression from the rhl operon of the quorum sensing (QS) system, resulting in no measurable rhamnolipid production by ATCC 33988. Further examination showed that ATCC 33988 lacked the entire lasI/lasR arm of the QS response. Instead of promoting expression of QS genes, ATCC 33988 up-regulates a small subset of its genome, including operons responsible for specific alkaline proteases and sphingosine metabolism. Our work represents the first time results from RNA-seq, microarray, ribosome footprinting, proteomics, and small molecule LC-MS experiments have been integrated to compare gene expression in bacteria. Altogether, these data provide insights as

  4. A comprehensive multi-omics approach uncovers adaptations for growth and survival of Pseudomonas aeruginosa on n-alkanes

    DOE PAGES

    Grady, Sarah L.; Malfatti, Stephanie A.; Gunasekera, Thusitha S.; ...

    2017-04-28

    Examination of complex biological systems has long been achieved through methodical investigation of the system’s individual components. While informative, this strategy often leads to inappropriate conclusions about the system as a whole. With the advent of high-throughput “omic” technologies, but, researchers can now simultaneously analyze an entire system at the level of molecule (DNA, RNA, protein, metabolite) and process (transcription, translation, enzyme catalysis). This strategy reduces the likelihood of improper conclusions, provides a framework for elucidation of genotype-phenotype relationships, and brings finer resolution to comparative genomic experiments. Here in this paper, we apply a multi-omic approach to analyze the genemore » expression profiles of two closely related Pseudomonas aeruginosa strains grown in n-alkanes or glycerol. The environmental P. aeruginosa isolate ATCC 33988 consumed medium-length (C 10–C 16) n-alkanes more rapidly than the laboratory strain PAO1, despite high genome sequence identity (average nucleotide identity >99%). Our data shows that ATCC 33988 induces a characteristic set of genes at the transcriptional, translational and post-translational levels during growth on alkanes, many of which differ from those expressed by PAO1. Of particular interest was the lack of expression from the rhl operon of the quorum sensing (QS) system, resulting in no measurable rhamnolipid production by ATCC 33988. Further examination showed that ATCC 33988 lacked the entire lasI/lasR arm of the QS response. Instead of promoting expression of QS genes, ATCC 33988 up-regulates a small subset of its genome, including operons responsible for specific alkaline proteases and sphingosine metabolism. Our work represents the first time results from RNA-seq, microarray, ribosome footprinting, proteomics, and small molecule LC-MS experiments have been integrated to compare gene expression in bacteria. Altogether, these data provide insights as

  5. Genomic analysis and temperature-dependent transcriptome profiles of the rhizosphere originating strain Pseudomonas aeruginosa M18

    PubMed Central

    2011-01-01

    Background Our previously published reports have described an effective biocontrol agent named Pseudomonas sp. M18 as its 16S rDNA sequence and several regulator genes share homologous sequences with those of P. aeruginosa, but there are several unusual phenotypic features. This study aims to explore its strain specific genomic features and gene expression patterns at different temperatures. Results The complete M18 genome is composed of a single chromosome of 6,327,754 base pairs containing 5684 open reading frames. Seven genomic islands, including two novel prophages and five specific non-phage islands were identified besides the conserved P. aeruginosa core genome. Each prophage contains a putative chitinase coding gene, and the prophage II contains a capB gene encoding a putative cold stress protein. The non-phage genomic islands contain genes responsible for pyoluteorin biosynthesis, environmental substance degradation and type I and III restriction-modification systems. Compared with other P. aeruginosa strains, the fewest number (3) of insertion sequences and the most number (3) of clustered regularly interspaced short palindromic repeats in M18 genome may contribute to the relative genome stability. Although the M18 genome is most closely related to that of P. aeruginosa strain LESB58, the strain M18 is more susceptible to several antimicrobial agents and easier to be erased in a mouse acute lung infection model than the strain LESB58. The whole M18 transcriptomic analysis indicated that 10.6% of the expressed genes are temperature-dependent, with 22 genes up-regulated at 28°C in three non-phage genomic islands and one prophage but none at 37°C. Conclusions The P. aeruginosa strain M18 has evolved its specific genomic structures and temperature dependent expression patterns to meet the requirement of its fitness and competitiveness under selective pressures imposed on the strain in rhizosphere niche. PMID:21884571

  6. Resistance Emergence Mechanism and Mechanism of Resistance Suppression by Tobramycin for Cefepime for Pseudomonas aeruginosa

    PubMed Central

    Bonomo, Robert A.; Bahniuk, Nadzeya; Bulitta, Juergen B.; VanScoy, Brian; DeFiglio, Holland; Fikes, Steven; Brown, David; Drawz, Sarah M.; Kulawy, Robert; Louie, Arnold

    2012-01-01

    The panoply of resistance mechanisms in Pseudomonas aeruginosa makes resistance suppression difficult. Defining optimal regimens is critical. Cefepime is a cephalosporin whose 3′ side chain provides some stability against AmpC β-lactamases. We examined the activity of cefepime against P. aeruginosa wild-type strain PAO1 and its isogenic AmpC stably derepressed mutant in our hollow-fiber infection model. Dose-ranging studies demonstrated complete failure with resistance emergence (both isolates). Inoculum range studies demonstrated ultimate failure for all inocula. Lower inocula failed last (10 days to 2 weeks). Addition of a β-lactamase inhibitor suppressed resistance even with the stably derepressed isolate. Tobramycin combination studies demonstrated resistance suppression in both the wild-type and the stably derepressed isolates. Quantitating the RNA message by quantitative PCR demonstrated that tobramycin decreased the message relative to that in cefepime-alone experiments. Western blotting with AmpC-specific antibody for P. aeruginosa demonstrated decreased expression. We concluded that suppression of β-lactamase expression by tobramycin (a protein synthesis inhibitor) was at least part of the mechanism behind resistance suppression. Monte Carlo simulation demonstrated that a regimen of 2 g of cefepime every 8 h plus 7 mg/kg of body weight of tobramycin daily would provide robust resistance suppression for Pseudomonas isolates with cefepime MIC values up to 8 mg/liter and tobramycin MIC values up to 1 mg/liter. For P. aeruginosa resistance suppression, combination therapy is critical. PMID:22005996

  7. Quantifying the adhesion and interaction forces between Pseudomonas aeruginosa and natural organic matter.

    PubMed

    Abu-Lail, Laila I; Liu, Yatao; Atabek, Arzu; Camesano, Terri A

    2007-12-01

    Atomic force microscopy (AFM) was used to characterize interactions between natural organic matter (NOM), and glass or bacteria. Poly(methacrylic acid) (PMA), soil humic Acid (SHA), and Suwannee River humic Acid (SRHA), were adsorbed to silica AFM probes. Adhesion forces (Fadh) for the interaction of organic-probes and glass slides correlated with organic molecular weight (MW), but not with radius of the organic aggregate (R), charge density (Q), or zeta potential (zeta). Two Pseudomonas aeruginosa strains with different lipopolysaccharides (LPS) were chosen: PAO1 (A+B+), whose LPS have common antigen (A-band) + O-antigen (B-band); and mutant AK1401 (A+B-). Fadh between bacteria and organics correlated with organic MW, R, and Q, but not zeta. PAO1 had lower Fadh with silica than NOM, which was attributed to negative charges from the B-band polymers causing electrostatic repulsion. AK1401 adhered stronger to silica than to the organics, perhaps because the absence of the B-band exposed underlying positively charged proteins. DLVO calculations could not explain the differences in the two bacteria or predict qualitative or quantitative trends in interaction forces in these systems. Molecular-level information from AFM studies can bring us closer to understanding the complex nature of bacterial-NOM interactions.

  8. Pseudomonas aeruginosa and their small diffusible extracellular molecules inhibit Aspergillus fumigatus biofilm formation.

    PubMed

    Mowat, Eilidh; Rajendran, Ranjith; Williams, Craig; McCulloch, Elaine; Jones, Brian; Lang, Sue; Ramage, Gordon

    2010-12-01

    Aspergillus fumigatus is often isolated from the lungs of cystic fibrosis (CF) patients, but unlike in severely immunocompromised individuals, the mortality rates are low. This suggests that competition from bacteria within the CF lung may be inhibitory. The purpose of this study was to investigate how Pseudomonas aeruginosa influences A. fumigatus conidial germination and biofilm formation. Aspergillus fumigatus biofilm formation was inhibited by direct contact with P. aeruginosa, but had no effect on preformed biofilm. A secreted heat-stable soluble factor was also shown to exhibit biofilm inhibition. Coculture of P. aeruginosa quorum-sensing mutants (PAO1:ΔLasI, PAO1:ΔLasR) did not significantly inhibit A. fumigatus biofilms (52.6-58.8%) to the same extent as that of the PA01 wild type (22.9-30.1%), both by direct and by indirect interaction (P<0.001). Planktonic and sessile inhibition assays with a series of short carbon chain molecules (decanol, decanoic acid and dodecanol) demonstrated that these molecules could both inhibit and disrupt biofilms in a concentration-dependent manner. Overall, this suggests that small diffusible and heat-stable molecules may be responsible for the competitive inhibition of filamentous fungal growth in polymicrobial environments such as the CF lung. © 2010 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.

  9. Detection of a Gentamicin-Resistant Burn Wound Strain of Pseudomonas Aeruginosa but Sensitive to Honey and Garcinia Kola (Heckel) Seed Extract

    PubMed Central

    Adeleke, O.E.; Coker, M.E.; Oke, O.B.

    2010-01-01

    Summary Studies on Staphylococcus aureus and Staphylococcus intermedius from dog and cat, and also on Staphylococcus aureus from wound and pyoderma infections, have shown a correlation between the site of microbial infection and antimicrobial susceptibility. Both the methanolic extract concentrate of Garcinia kola (Heckel) seeds and natural honey have been associated with activity on bacterial isolates from respiratory tract infections. In this study, selected bacteria belonging to genera from burn wound infection sites were treated with natural honey and methanolic extract concentrate of Garcinia kola in antimicrobial susceptibility tests separately and in combined form, and also with gentamicin and methanol as controls. The two natural products were found to be active on the bacterial isolates, excluding Klebsiella pneumoniae strains, all of which showed resistance to honey. Combination forms of the two natural products were active only on the strains of Pseudomonas aeruginosa. At 4 and 8 µg/ml, gentamicin was ineffective on the three strains of Klebsiella pneumoniae while 8 µg/ml was moderately active on only two strains of Pseudomonas aeruginosa. One strain of Pseudomonas aeruginosa, UCH002, was resistant to gentamicin beyond 1,000 µ/ml. Gentamicin at 4 µ/ml was inhibitory to one strain of Escherichia coli and two strains of Staphylococcus aureus. Though the antimicrobial activity of the two natural products tested had been previously reported against microbial agents of respiratory tract infection, it was also recorded in this study. The lack of activity of each of the three honey types used in this study against the Klebsiella pneumoniae strains tested underscores the need to exclude this organism from burn wound infections before embarking on treatment with honey. The sensitivity of one high-level gentamicin-resistant strain of Pseudomonas aeruginosa to honey and Garcinia kola seed extract was noteworthy considering the therapeutic failures of gentamicin

  10. Comparison of Epidemiological and Antibiotic Susceptibility Pattern of Metallo-Beta-Lactamase-Positive and Metallo-Beta-Lactamase-Negative Strains of Pseudomonas Aeruginosa

    PubMed Central

    Ranjan, Shikha; Banashankari, GS; Babu, PR Sreenivasa

    2014-01-01

    Background: The infections caused by metallo-beta-lactamases (MBLs) producing Pseudomonas aeruginosa are associated with higher rates of mortality, morbidity, and overall healthcare costs compared to non-MBL P. aeruginosa infections. Purpose: To compare the epidemiologic factors and antibiograms of MBL-positive and MBL-negative P. aeruginosa isolates in a tertiary care hospital. Methods: In an observational study, from January 2011 to December 2012, all non-duplicate P. aeruginosa isolates were subjected to an antimicrobial sensitivity test against 10 antibiotics of five different classes. All P. aeruginosa strains showing resistance to at least one of the carbapenems were subjected to the MBL-E test. Epidemiological features and antibiograms of MBL-positive and MBL-negative strains were compared and statistically analyzed. Results: Out of 350 isolates (total sample = 5330) of P. aeruginosa, MBL was detected in 58 isolates by the E-test, resulting in a prevalence of 16.57%. Resistance to most of the antibiotics was significantly higher in the MBL-positive strains with 100% resistance to ciprofloxacin, tobramycin, and meropenem, followed by imipenem (93.10%) and gentamicin (89.66%). The prevalence of multidrug-resistant and pandrug-resistant strains was significantly higher among the MBL group as compared to that in the non-MBL group ((55.17 vs. 7.88% (P < 0.0001) and 8.62 vs. 0.68% (P = 0.0006)), respectively. Conclusions: MBL-positive P. aeruginosa strains showed very high resistance to various antibiotics, as compared to the non-MBL strains. Increasing prevalence of MBL-producing isolates in hospital settings makes it important to perform routine detection of MBL-positive P. aeruginosa strains by in vitro testing before antibiotic use, for the purposes of infection prevention, and control, and for minimizing the adverse outcomes of infections with MBL-producing strains. PMID:25328336

  11. Comparison of epidemiological and antibiotic susceptibility pattern of metallo-Beta-lactamase-positive and metallo-Beta-lactamase-negative strains of pseudomonas aeruginosa.

    PubMed

    Ranjan, Shikha; Banashankari, Gs; Babu, Pr Sreenivasa

    2014-07-01

    The infections caused by metallo-beta-lactamases (MBLs) producing Pseudomonas aeruginosa are associated with higher rates of mortality, morbidity, and overall healthcare costs compared to non-MBL P. aeruginosa infections. To compare the epidemiologic factors and antibiograms of MBL-positive and MBL-negative P. aeruginosa isolates in a tertiary care hospital. In an observational study, from January 2011 to December 2012, all non-duplicate P. aeruginosa isolates were subjected to an antimicrobial sensitivity test against 10 antibiotics of five different classes. All P. aeruginosa strains showing resistance to at least one of the carbapenems were subjected to the MBL-E test. Epidemiological features and antibiograms of MBL-positive and MBL-negative strains were compared and statistically analyzed. Out of 350 isolates (total sample = 5330) of P. aeruginosa, MBL was detected in 58 isolates by the E-test, resulting in a prevalence of 16.57%. Resistance to most of the antibiotics was significantly higher in the MBL-positive strains with 100% resistance to ciprofloxacin, tobramycin, and meropenem, followed by imipenem (93.10%) and gentamicin (89.66%). The prevalence of multidrug-resistant and pandrug-resistant strains was significantly higher among the MBL group as compared to that in the non-MBL group ((55.17 vs. 7.88% (P < 0.0001) and 8.62 vs. 0.68% (P = 0.0006)), respectively. MBL-positive P. aeruginosa strains showed very high resistance to various antibiotics, as compared to the non-MBL strains. Increasing prevalence of MBL-producing isolates in hospital settings makes it important to perform routine detection of MBL-positive P. aeruginosa strains by in vitro testing before antibiotic use, for the purposes of infection prevention, and control, and for minimizing the adverse outcomes of infections with MBL-producing strains.

  12. An efflux pump (MexAB-OprM) of Pseudomonas aeruginosa is associated with antibacterial activity of Epigallocatechin-3-gallate (EGCG).

    PubMed

    Kanagaratnam, Rashmi; Sheikh, Rida; Alharbi, Fahad; Kwon, Dong H

    2017-12-01

    Pseudomonas aeruginosa is a notorious multidrug resistant nosocomial pathogen. An efflux pump (MexAB-OprM) is the main contributor to the multidrug resistance in clinical isolates of P. aeruginosa. Epigallocatechin-3-gallate (EGCG), a polyphenolic compound extracted from green tea, exhibits antibacterial activity. It is unclear that molecular details of the antibacterial activity of EGCG, EGCG-effect on antibiotic susceptibility, and clinical relevance of EGCG in bacteria. This study aimed to determine the roles of the efflux pump and an efflux pump inhibitor (phenylalanine-arginine β-naphthylamide; PAβN) in the antibacterial activity of EGCG and the EGCG-effect on antibiotic susceptibility. Twenty-two multidrug resistant clinical isolates of P. aeruginosa and a wild type P. aeruginosa PAO1 were used to determine antibacterial activity of EGCG and EGCG-effect on antibiotic susceptibility. An efflux pump (MexAB-OPrM) mutant strain, its complemented strain carrying an intact mexAB-oprM, and their parental strain were used to determine roles of MexAB-OprM in the antibacterial activity of EGCG and EGCG-mediated antibiotic susceptibility. PAβN was also used to evaluate EGCG as a possible efflux pump inhibitor. EGCG inhibited cellular growth and killed 100% of cells at 64-512 µg/ml and at 256-1024 µg/ml, respectively, in all tested 22 clinical isolates including the wild type strain. A subinhibitory concentration of EGCG significantly enhanced susceptibility to antibiotics, unexceptionally to chloramphenicol and tetracyclines (≥4-fold) of the clinical isolates. Both the antibacterial activity of EGCG and the EGCG-mediated antibiotic susceptibility were enhanced more in the efflux pump mutant strain (mexB::Gm) than the parental strain, suggesting additionally accumulated-EGCG produced the more antibacterial activity in the mutant strain. EGCG was synergistically interacted with PAβN with enhancing susceptibility to all tested antibiotics (up to >500-fold) at

  13. [Application of recombinase polymerase amplification in the detection of Pseudomonas aeruginosa].

    PubMed

    Jin, X J; Gong, Y L; Yang, L; Mo, B H; Peng, Y Z; He, P; Zhao, J N; Li, X L

    2018-04-20

    Objective: To establish an optimized method of recombinase polymerase amplification (RPA) to rapidly detect Pseudomonas aeruginosa in clinic. Methods: (1) The DNA templates of one standard Pseudomonas aeruginosa strain was extracted and detected by polymerase chain reaction (PCR), real-time fluorescence quantitative PCR and RPA. Time of sample loading, time of amplification, and time of detection of the three methods were recorded. (2) One standard Pseudomonas aeruginosa strain was diluted in 7 concentrations of 1×10(7,) 1×10(6,) 1×10(5,) 1×10(4,) 1×10(3,) 1×10(2,) and 1×10(1) colony forming unit (CFU)/mL after recovery and cultivation. The DNA templates of Pseudomonas aeruginosa and negative control strain Pseudomonas putida were extracted and detected by PCR, real-time fluorescence quantitative PCR, and RPA separately. The sensitivity of the three methods in detecting Pseudomonas aeruginosa was analyzed. (3) The DNA templates of one standard Pseudomonas aeruginosa strain and four negative control strains ( Staphylococcus aureus, Acinetobacter baumanii, Candida albicans, and Pseudomonas putida ) were extracted separately, and then they were detected by PCR, real-time fluorescence quantitative PCR, and RPA. The specificity of the three methods in detecting Pseudomonas aeruginosa was analyzed. (4) The DNA templates of 28 clinical strains of Pseudomonas aeruginosa preserved in glycerin, 1 clinical strain of which was taken by cotton swab, and negative control strain Pseudomonas putida were extracted separately, and then they were detected by RPA. Positive amplification signals of the clinical strains were observed, and the detection rate was calculated. All experiments were repeated for 3 times. Sensitivity results were analyzed by GraphPad Prism 5.01 statistical software. Results: (1) The loading time of RPA, PCR, and real-time fluorescence quantitative PCR for detecting Pseudomonas aeruginosa were all 20 minutes. In PCR, time of amplification was 98 minutes

  14. Gene expression in Pseudomonas aeruginosa exposed to hydroxyl-radicals.

    PubMed

    Aharoni, Noa; Mamane, Hadas; Biran, Dvora; Lakretz, Anat; Ron, Eliora Z

    2018-05-01

    Recent studies have shown the efficiency of hydroxyl radicals generated via ultraviolet (UV)-based advanced oxidation processes (AOPs) combined with hydrogen peroxide (UV/H 2 O 2 ) as a treatment process in water. The effects of AOP treatments on bacterial gene expression was examined using Pseudomonas aeruginosa strain PAO1 as a model-organism bacterium. Many bacterial genes are not expressed all the time, but their expression is regulated. The regulation is at the beginning of the gene, in a genetic region called "promoter" and affects the level of transcription (synthesis of messenger RNA) and translation (synthesis of protein). The level of expression of the regulated genes can change as a function of environmental conditions, and they can be expressed more (induced, upregulated) or less (downregulated). Exposure of strain PAO1 to UV/H 2 O 2 treatment resulted in a major change in gene expression, including elevated expression of several genes. One interesting gene is PA3237, which was significantly upregulated under UV/H 2 O 2 as compared to UV or H 2 O 2 treatments alone. The induction of this gene is probably due to formation of radicals, as it is abolished in the presence of the radical scavenger tert-butanol (TBA) and is seen even when the bacteria are added after the treatment (post-treatment exposure). Upregulation of the PA3237 promoter could also be detected using a reporter gene, suggesting the use of such genetic constructs to develop biosensors for monitoring AOPs in water-treatment plants. Currently biosensors for AOPs do not exist, consequently impairing the ability to monitor these processes on-line according to radical exposure in natural waters. Copyright © 2018 Elsevier Ltd. All rights reserved.

  15. Blue light enhances the antimicrobial activity of honey against Pseudomonas aeruginosa

    NASA Astrophysics Data System (ADS)

    Orlandi, Viviana Teresa; Bolognese, Fabrizio; Barbieri, Paola

    2018-02-01

    Pseudomonas aeruginosa may be isolated from skin wounds of burn patients, bedsore and diabetic ulcers. The healing of wounds is often impaired by the intrinsic antibiotic resistance, the tolerance to many antimicrobials and the ability to form biofilm of this opportunistic pathogen. Finding new topical treatments to combine with antibiotics is thus essential. Among natural products, the antimicrobial properties of honeys have been known for millennia. In this study honey and visible light have been combined to control the growth of P. aeruginosa PAO1. The irradiation by a broad spectrum light source of bacteria inoculated onto 2 % w/v fir and forest honeydew (HD) honeys caused a killing effect that the honeys alone or the light alone did not show. This antimicrobial activity was light energy-dose and honey-concentration dependent. Among the tested honeys, the fir and forest HD honeys were the most efficient ones. In particular, the irradiation by blue LED (λmax = 466 nm) yielded good rates of killing, that were significantly higher in comparison to irradiation alone and honey alone. Interestingly, a similar effect was obtained by plating bacteria on blue LED pre-irradiated HD honeys. The combined use of honey and blue light was also successful in inhibiting the biofilm formation of P. aeruginosa. The blue LED irradiation of PAO1 administered with 10 % w/v forest HD honey significantly enhanced the inhibition of biofilm formation in comparison to dark incubated honey.

  16. The Pseudomonas aeruginosa pirA gene encodes a second receptor for ferrienterobactin and synthetic catecholate analogues.

    PubMed

    Ghysels, Bart; Ochsner, Urs; Möllman, Ute; Heinisch, Lothar; Vasil, Michael; Cornelis, Pierre; Matthijs, Sandra

    2005-05-15

    Actively secreted iron chelating agents termed siderophores play an important role in the virulence and rhizosphere competence of fluorescent pseudomonads, including Pseudomonas aeruginosa which secretes a high affinity siderophore, pyoverdine, and the low affinity siderophore, pyochelin. Uptake of the iron-siderophore complexes is an active process that requires specific outer membrane located receptors, which are dependent of the inner membrane-associated protein TonB and two other inner membrane proteins, ExbB and ExbC. P. aeruginosa is also capable of using a remarkable variety of heterologous siderophores as sources of iron, apparently by expressing their cognate receptors. Illustrative of this feature are the 32 (of which 28 putative) siderophore receptor genes observed in the P. aeruginosa PAO1 genome. However, except for a few (pyoverdine, pyochelin, enterobactin), the vast majority of P. aeruginosa siderophore receptor genes still remain to be characterized. Ten synthetic iron chelators of catecholate type stimulated growth of a pyoverdine/pyochelin deficient P. aeruginosa PAO1 mutant under condition of severe iron limitation. Null mutants of the 32 putative TonB-dependent siderophore receptor encoding genes engineered in the same genetic background were screened for obvious deficiencies in uptake of the synthetic siderophores, but none showed decreased growth stimulation in the presence of the different siderophores. However, a double knock-out mutant of ferrienterobactin receptor encoding gene pfeA (PA 2688) and pirA (PA0931) failed to be stimulated by 4 of the tested synthetic catecholate siderophores whose chemical structures resemble enterobactin. Ferric-enterobactin also failed to stimulate growth of the double pfeA-pirA mutant although, like its synthetic analogues, it stimulated growth of the corresponding single mutants. Hence, we confirmed that pirA represents a second P. aeruginosa ferric-enterobactin receptor. The example of these two

  17. Biofilm formation, antibiotic susceptibility and RAPD genotypes in Pseudomonas aeruginosa clinical strains isolated from single centre intensive care unit patients.

    PubMed

    Vaněrková, Martina; Mališová, Barbora; Kotásková, Iva; Holá, Veronika; Růžička, Filip; Freiberger, Tomáš

    2017-11-01

    The aim of this study was to analyse genotypes, antimicrobial susceptibility patterns and serotypes in Pseudomonas aeruginosa clinical strains, including the clonal dissemination of particular strains throughout various intensive care units in one medical centre. Using random amplified polymorphic DNA (RAPD-PCR) and P. aeruginosa antisera, 22 different genotypes and 8 serotypes were defined among 103 isolates from 48 patients. No direct association between P. aeruginosa strain genotypes and serotypes was observed. RAPD typing in strains with the same serotype revealed different genotypes and, on the contrary, most strains with a different serotype displayed the same amplification pattern. The resulting banding patterns showed a high degree of genetic heterogeneity among all isolates from the patients examined, suggesting a non-clonal relationship between isolates from these patients. A higher degree of antibiotic resistance and stronger biofilm production in common genotypes compared to rare ones and genetic homogeneity of the most resistant strains indicated the role of antibiotic pressure in acquiring resistant and more virulent strains in our hospital. In conclusion, genetic characterisation of P. aeruginosa strains using RAPD method was shown to be more accurate in epidemiological analyses than phenotyping.

  18. The role of alginate in Pseudomonas aeruginosa EPS adherence, viscoelastic properties and cell attachment.

    PubMed

    Orgad, Oded; Oren, Yoram; Walker, Sharon L; Herzberg, Moshe

    2011-08-01

    Among various functions, extracellular polymeric substances (EPS) provide microbial biofilms with mechanical stability and affect initial cell attachment, the first stage in the biofilm formation process. The role of alginate, an abundant polysaccharide in Pseudomonas aeruginosa biofilms, in the viscoelastic properties and adhesion kinetics of EPS was analyzed using a quartz crystal microbalance with dissipation (QCM-D) monitoring technology. EPS was extracted from two P. aeruginosa biofilms, a wild type strain, PAO1, and a mucoid strain, PAOmucA22 that over-expresses alginate production. The higher alginate content in the EPS originating from the mucoid biofilms was clearly shown to increase both the rate and the extent of attachment of the EPS, as well as the layer's thickness. Also, the presence of calcium and elevated ionic strength increased the thickness of the EPS layer. Dynamic light scattering (DLS) showed that the presence of calcium and elevated ionic strength induced intermolecular attractive interactions in the mucoid EPS molecules. For the wild type EPS, in the presence of calcium, an elevated shift in the distribution of the diffusion coefficients was observed with DLS due to a more compacted conformation of the EPS molecules. Moreover, the alginate over-expression effect on EPS adherence was compared to the effect of alginate over-expression on P. aeruginosa cell attachment. In a parallel plate flow cell, under similar hydraulic and aquatic conditions as those applied for the EPS adsorption tests in the QCM-D flow cell, reduced adherence of the mucoid strain was clearly observed compared to the wild type isogenic bacteria. The results suggest that alginate contributes to steric hindrance and shielding of cell surface features and adhesins that are known to promote cell attachment. © 2011 Taylor & Francis

  19. Genome Sequence of Pseudomonas aeruginosa Strain LCT-PA220, Which Was Selected after Space Flight by Using Biolog's Powerful Carbon Source Utilization Technology.

    PubMed

    Xu, Guogang; Hu, Juan; Fang, Xiangqun; Zhang, Xuelin; Wang, Junfeng; Guo, Yinghua; Li, Tianzhi; Chen, Zhenghong; Dai, Wenkui; Liu, Changting

    2014-03-13

    To explore the changes of Pseudomonas aeruginosa in space flight, we present the draft genome sequence of P. aeruginosa strain LCT-PA220, which originated from a P. aeruginosa strain, ATCC 27853, that traveled on the Shenzhou-VIII spacecraft.

  20. Comparative study on the in vitro effects of Pseudomonas aeruginosa and seaweed alginates on human gut microbiota

    PubMed Central

    Bai, Shaofeng; Chen, Huahai; Zhu, Liying; Liu, Wei; Yu, Hongwei D.; Wang, Xin; Yin, Yeshi

    2017-01-01

    Alginates pertain to organic polysaccharides that have been extensively used in food- and medicine-related industries. The present study obtained alginates from an alginate overproducing Pseudomonas aeruginosa PAO1 mutant by screening transposon mutagenesis libraries. The interaction between bacterial and seaweed alginates and gut microbiota were further studied by using an in vitro batch fermentation system. Thin-layer chromatography (TLC) analysis indicated that both bacterial and seaweed alginates can be completely degraded by fecal bacteria isolated from study volunteers, indicating that a minor structural difference between bacterial and seaweed alginates (O-acetylation and lack of G-G blocks) didn’t affect the digestion of alginates by human microbiota. Although, the digestion of bacterial and seaweed alginates was attributed to different Bacteroides xylanisolvens strains, they harbored similar alginate lyase genes. Genus Bacteroides with alginate-degrading capability were enriched in growth medium containing bacterial or seaweed alginates after in vitro fermentation. Short-chain fatty acid (SCFA) production in both bacterial and seaweed alginates was also comparable, but was significantly higher than the same medium using starch. In summary, the present study has isolated an alginate-overproducing P. aeruginosa mutant strain. Both seaweed and bacterial alginates were degraded by human gut microbiota, and their regulatory function on gut microbiota was similar. PMID:28170428

  1. Comparative study on the in vitro effects of Pseudomonas aeruginosa and seaweed alginates on human gut microbiota.

    PubMed

    Bai, Shaofeng; Chen, Huahai; Zhu, Liying; Liu, Wei; Yu, Hongwei D; Wang, Xin; Yin, Yeshi

    2017-01-01

    Alginates pertain to organic polysaccharides that have been extensively used in food- and medicine-related industries. The present study obtained alginates from an alginate overproducing Pseudomonas aeruginosa PAO1 mutant by screening transposon mutagenesis libraries. The interaction between bacterial and seaweed alginates and gut microbiota were further studied by using an in vitro batch fermentation system. Thin-layer chromatography (TLC) analysis indicated that both bacterial and seaweed alginates can be completely degraded by fecal bacteria isolated from study volunteers, indicating that a minor structural difference between bacterial and seaweed alginates (O-acetylation and lack of G-G blocks) didn't affect the digestion of alginates by human microbiota. Although, the digestion of bacterial and seaweed alginates was attributed to different Bacteroides xylanisolvens strains, they harbored similar alginate lyase genes. Genus Bacteroides with alginate-degrading capability were enriched in growth medium containing bacterial or seaweed alginates after in vitro fermentation. Short-chain fatty acid (SCFA) production in both bacterial and seaweed alginates was also comparable, but was significantly higher than the same medium using starch. In summary, the present study has isolated an alginate-overproducing P. aeruginosa mutant strain. Both seaweed and bacterial alginates were degraded by human gut microbiota, and their regulatory function on gut microbiota was similar.

  2. Modular Approach to Select Bacteriophages Targeting Pseudomonas aeruginosa for Their Application to Children Suffering With Cystic Fibrosis

    PubMed Central

    Krylov, Victor; Shaburova, Olga; Pleteneva, Elena; Bourkaltseva, Maria; Krylov, Sergey; Kaplan, Alla; Chesnokova, Elena; Kulakov, Leonid; Magill, Damian; Polygach, Olga

    2016-01-01

    This review discusses the potential application of bacterial viruses (phage therapy) toward the eradication of antibiotic resistant Pseudomonas aeruginosa in children with cystic fibrosis (CF). In this regard, several potential relationships between bacteria and their bacteriophages are considered. The most important aspect that must be addressed with respect to phage therapy of bacterial infections in the lungs of CF patients is in ensuring the continuity of treatment in light of the continual occurrence of resistant bacteria. This depends on the ability to rapidly select phages exhibiting an enhanced spectrum of lytic activity among several well-studied phage groups of proven safety. We propose a modular based approach, utilizing both mono-species and hetero-species phage mixtures. With an approach involving the visual recognition of characteristics exhibited by phages of well-studied phage groups on lawns of the standard P. aeruginosa PAO1 strain, the simple and rapid enhancement of the lytic spectrum of cocktails is permitted, allowing the development of tailored preparations for patients capable of circumventing problems associated with phage resistant bacterial mutants. PMID:27790211

  3. A Rapid Phenotypic Whole Cell Screening Approach for the Identification of Small Molecule Inhibitors that Counter Beta-lactamase Resistance in Pseudomonas aeruginosa

    PubMed Central

    Collia, Deanna; Bannister, Thomas D.; Tan, Hao; Jin, Shouguang; Langaee, Taimour; Shumate, Justin; Scampavia, Louis; Spicer, Timothy P.

    2017-01-01

    Pseudomonas aeruginosa is an opportunistic human pathogen which is prevalent in hospitals and continues to develop resistance to multiple classes of antibiotics. Historically, β-lactam antibiotics have been the first line of therapeutic defense. However, the emergence of multidrug-resistant (MDR) strains of P. aeruginosa, such as AmpC β-lactamase overproducing mutants, limits the effectiveness of current antibiotics. Among AmpC hyper producing clinical isolates, inactivation of AmpG, which is essential for the expression of AmpC, increases bacterial sensitivity to β-lactam antibiotics. We hypothesize that inhibition of AmpG activity will enhance the efficacy of β-lactams against P. aeruginosa. Here, using a highly drug resistant AmpC inducible laboratory strain PAO1, we describe an ultra-high throughput whole cell turbidity assay designed to identify small molecule inhibitors of the AmpG. We screened 645K compounds to identify compounds with the ability to inhibit bacterial growth in the presence of Cefoxitin; an AmpC inducer, and identified 2,663 inhibitors which were also tested in the absence of Cefoxitin to determine AmpG specificity. The Z′ and S:B were robust at 0.87 ± 0.05 and 2.2 ± 0.2, respectively. Through a series of secondary and tertiary studies, including a novel luciferase based counterscreen, we ultimately identified 8 potential AmpG specific inhibitors. PMID:28850797

  4. A Rapid Phenotypic Whole-Cell Screening Approach for the Identification of Small-Molecule Inhibitors That Counter β-Lactamase Resistance in Pseudomonas aeruginosa.

    PubMed

    Collia, Deanna; Bannister, Thomas D; Tan, Hao; Jin, Shouguang; Langaee, Taimour; Shumate, Justin; Scampavia, Louis; Spicer, Timothy P

    2018-01-01

    Pseudomonas aeruginosa is an opportunistic human pathogen that is prevalent in hospitals and continues to develop resistance to multiple classes of antibiotics. Historically, β-lactam antibiotics have been the first line of therapeutic defense. However, the emergence of multidrug-resistant (MDR) strains of P. aeruginosa, such as AmpC β-lactamase overproducing mutants, limits the effectiveness of current antibiotics. Among AmpC hyperproducing clinical isolates, inactivation of AmpG, which is essential for the expression of AmpC, increases bacterial sensitivity to β-lactam antibiotics. We hypothesize that inhibition of AmpG activity will enhance the efficacy of β-lactams against P. aeruginosa. Here, using a highly drug-resistant AmpC-inducible laboratory strain PAO1, we describe an ultra-high-throughput whole-cell turbidity assay designed to identify small-molecule inhibitors of the AmpG. We screened 645,000 compounds to identify compounds with the ability to inhibit bacterial growth in the presence of cefoxitin, an AmpC inducer, and identified 2663 inhibitors that were also tested in the absence of cefoxitin to determine AmpG specificity. The Z' and signal-to-background ratio were robust at 0.87 ± 0.05 and 2.2 ± 0.2, respectively. Through a series of secondary and tertiary studies, including a novel luciferase-based counterscreen, we ultimately identified eight potential AmpG-specific inhibitors.

  5. Prevalence of genomic island PAPI-1 in clinical isolates of Pseudomonas aeruginosa in Iran.

    PubMed

    Sadeghifard, Nourkhoda; Rasaei, Seyedeh Zahra; Ghafourian, Sobhan; Zolfaghary, Mohammad Reza; Ranjbar, Reza; Raftari, Mohammad; Mohebi, Reza; Maleki, Abbas; Rahbar, Mohammad

    2012-03-01

    Pseudomonas aeruginosa, a gram-negative rod-shaped bacterium, is an opportunistic pathogen, which causes various serious diseases in humans and animals. The aims of this study were to evaluate of the presence of genomic island PAPI-1 in Pseudomonas aeruginosa isolated from Reference Laboratory of Ilam, Milad Hospital and Emam Khomeini Hospital, Iran and to study the frequency of extended spectrum beta-lactamases (ESBLs) among isolates. Forty-eight clinical isolates of P. aeruginosa were obtained during April to September 2010, and were evaluated for ESBLs by screening and confirmatory disk diffusion methods and PAPI-1 by PCR. Fifteen of 48 P. aeruginosa isolates were positive for ESBLs and 17 isolates positive for PAPI-1. This was first study of the prevalence of PAPI-1 in clinical isolates of P. aeruginosa in Iran, showing that most of PAPI-1 positive strains had high levels of antibiotic resistance and produced ESBLs.

  6. Antibiofilm and Anti-Infection of a Marine Bacterial Exopolysaccharide Against Pseudomonas aeruginosa

    PubMed Central

    Wu, Shimei; Liu, Ge; Jin, Weihua; Xiu, Pengyuan; Sun, Chaomin

    2016-01-01

    Pseudomonas aeruginosa is a well-known pathogenic bacterium that forms biofilms and produces virulence factors, thus leading to major problems in many fields, such as clinical infection, food contamination, and marine biofouling. In this study, we report the purification and characterization of an exopolysaccharide EPS273 from the culture supernatant of marine bacterium P. stutzeri 273. The exopolysaccharide EPS273 not only effectively inhibits biofilm formation but also disperses preformed biofilm of P. aeruginosa PAO1. High performance liquid chromatography traces of the hydrolyzed polysaccharides shows that EPS273 primarily consists of glucosamine, rhamnose, glucose and mannose. Further investigation demonstrates that EPS273 reduces the production of the virulence factors pyocyanin, exoprotease, and rhamnolipid, and the virulence of P. aeruginosa PAO1 to human lung cells A549 and zebrafish embryos is also obviously attenuated by EPS273. In addition, EPS273 also greatly reduces the production of hydrogen peroxide (H2O2) and extracellular DNA (eDNA), which are important factors for biofilm formation. Furthermore, EPS273 exhibits strong antioxidant potential by quenching hydroxyl and superoxide anion radicals. Notably, the antibiofouling activity of EPS273 is observed in the marine environment up to 2 weeks according to the amounts of bacteria and diatoms in the glass slides submerged in the ocean. Taken together, the properties of EPS273 indicate that it has a promising prospect in combating bacterial biofilm-associated infection, food-processing contamination and marine biofouling. PMID:26903981

  7. Chemical Analysis of Cellular and Extracellular Carbohydrates of a Biofilm-Forming Strain Pseudomonas aeruginosa PA14

    PubMed Central

    Coulon, Charlène; Vinogradov, Evgeny; Filloux, Alain; Sadovskaya, Irina

    2010-01-01

    Background Pseudomonas aeruginosa is a Gram-negative bacterium and an opportunistic pathogen, which causes persisting life-threatening infections in cystic fibrosis (CF) patients. Biofilm mode of growth facilitates its survival in a variety of environments. Most P. aeruginosa isolates, including the non-mucoid laboratory strain PA14, are able to form a thick pellicle, which results in a surface-associated biofilm at the air-liquid (A–L) interface in standing liquid cultures. Exopolysaccharides (EPS) are considered as key components in the formation of this biofilm pellicle. In the non-mucoid P. aeruginosa strain PA14, the “scaffolding” polysaccharides of the biofilm matrix, and the molecules responsible for the structural integrity of rigid A–L biofilm have not been identified. Moreover, the role of LPS in this process is unclear, and the chemical structure of the LPS O-antigen of PA14 has not yet been elucidated. Principal Findings In the present work we carried out a systematic analysis of cellular and extracellular (EC) carbohydrates of P. aeruginosa PA14. We also elucidated the chemical structure of the LPS O-antigen by chemical methods and 2-D NMR spectroscopy. Our results showed that it is composed of linear trisaccharide repeating units, identical to those described for P. aeruginosa Lanýi type O:2a,c (Lanýi-Bergman O-serogroup 10a, 10c; IATS serotype 19) and having the following structure: -4)-α-L-GalNAcA-(1–3)-α-D-QuiNAc-(1–3)- α-L-Rha-(1-. Furthermore, an EC O-antigen polysaccharide (EC O-PS) and the glycerol-phosphorylated cyclic β-(1,3)-glucans were identified in the culture supernatant of PA14, grown statically in minimal medium. Finally, the extracellular matrix of the thick biofilm formed at the A-L interface contained, in addition to eDNA, important quantities (at least ∼20% of dry weight) of LPS-like material. Conclusions We characterized the chemical structure of the LPS O-antigen and showed that the O-antigen polysaccharide is

  8. [The antibacterial activity of oregano essential oil (Origanum heracleoticum L.) against clinical strains of Escherichia coli and Pseudomonas aeruginosa].

    PubMed

    Sienkiewicz, Monika; Wasiela, Małgorzata; Głowacka, Anna

    2012-01-01

    The aim of this study was to investigate the antibacterial properties of oregano (Origanum heracleoticum L.) essential oil against clinical strains of Escherichia coli and Pseudomonas aeruginosa. The antibacterial activity of oregano essential oil was investigate against 2 tested and 20 clinical bacterial strains of Escherichia coli and 20 clinical strains o Pseudomonas aeruginosa come from patients with different clinical conditions. The agar dilution method was used for microbial growth inhibition at various concentrations ofoil. Susceptibility testing to antibiotics was carried out using disc-diffusion method. The results of experiments showed that the tested oil was active against all of the clinical strains from both genus of bacteria, but strains of Escherichia coli were more sensitive to tested oil. Essential oil from Origanum heracleoticum L. inhibited the growth of Escherichia coli and Pseudomonas aeruginosa clinical strains with different patters of resistance. The obtained outcomes will enable further investigations using oregano essential oil obtained from Origanum heracleoticum L. as alternative antibacterial remedies enhancing healing process in bacterial infections and as an effective means for the prevention of antibiotic-resistant strain development.

  9. Molecular detection of metallo-β-lactamase gene blaVIM-1 in imipenem-resistant Pseudomonas aeruginosa strains isolated from hospitalized patients in the hospitals of Isfahan.

    PubMed

    Sedighi, Mansour; Vaez, Hamid; Moghoofeie, Mohsen; Hadifar, Shima; Oryan, Golfam; Faghri, Jamshid

    2015-01-01

    Pseudomonas aeruginosa is an opportunistic human pathogen that causes serious problems, especially in people, who have immunodeficiency. In recent times, metallo-β-lactamase (MBLs) resistance in this bacterium has led to some difficulties in treating bacterial infections. The metallo-beta-lactamase family of genes, including blaVIM-1, is being reported with increasing frequency worldwide. The aim of this study is the detection of the metallo-β-lactamase gene blaVIM-1 in imipenem-resistant P. aeruginosa (IRPA) strains isolated from hospitalized patients. In this study, 106 P. aeruginosa samples were isolated from various nosocomial infections. The isolates were identified, tested for susceptibility to various antimicrobial agents by the Kirby-Bauer disk diffusion method, and all the imipenem-resistant isolates were screened for the presence of MBLs by using the combined disk (IMP-EDTA). The minimal inhibitory concentration (MIC) of imipenem was determined by E-test on the Mueller-Hinton agar. To detect the blaVIM-1 gene, the isolates were subjected to a polymerase chain reaction (PCR). Of all the P. aeruginosa isolates, 62 (58.5%) were found to be imipenem-resistant P. aeruginosa (MIC ≥32 μg/ml). Twenty-six (42%) of the imipenem-resistant isolates were MBL positive. None of these isolates carried the blaVIM-1 gene using the PCR assay. The results demonstrated the serious therapeutic threat of the MBL-producing P. aeruginosa populations. The rate of imipenem resistance due to MBL was increased dramatically. Early detection and infection-control practices are the best antimicrobial strategies for this organism. None of MBL-producing isolates in this study carry the blaVIM-1 gene; therefore, another gene in the MBL family should be investigated.

  10. Mucosal fluid glycoprotein DMBT1 suppresses twitching motility and virulence of the opportunistic pathogen Pseudomonas aeruginosa

    PubMed Central

    Evans, David J.; Fleiszig, Suzanne M. J.

    2017-01-01

    It is generally thought that mucosal fluids protect underlying epithelial surfaces against opportunistic infection via their antimicrobial activity. However, our published data show that human tear fluid can protect against the major opportunistic pathogen Pseudomonas aeruginosa independently of bacteriostatic activity. Here, we explored the mechanisms for tear protection, focusing on impacts of tear fluid on bacterial virulence factor expression. Results showed that tear fluid suppressed twitching motility, a type of surface-associated movement conferred by pili. Previously, we showed that twitching is critical for P. aeruginosa traversal of corneal epithelia, exit from epithelial cells after internalization, and corneal virulence. Inhibition of twitching by tear fluid was dose-dependent with dilutions to 6.25% retaining activity. Purified lactoferrin, lysozyme, and contrived tears containing these, and many other, tear components lacked the activity. Systematic protein fractionation, mass spectrometry, and immunoprecipitation identified the glycoprotein DMBT1 (Deleted in Malignant Brain Tumors 1) in tear fluid as required. DMBT1 purified from human saliva also inhibited twitching, as well as P. aeruginosa traversal of human corneal epithelial cells in vitro, and reduced disease pathology in a murine model of corneal infection. DMBT1 did not affect PilA expression, nor bacterial intracellular cyclicAMP levels, and suppressed twitching motility of P. aeruginosa chemotaxis mutants (chpB, pilK), and an adenylate cyclase mutant (cyaB). However, dot-immunoblot assays showed purified DMBT1 binding of pili extracted from PAO1 suggesting that twitching inhibition may involve a direct interaction with pili. The latter could affect extension or retraction of pili, their interactions with biotic or abiotic surfaces, or cause their aggregation. Together, the data suggest that DMBT1 inhibition of twitching motility contributes to the mechanisms by which mucosal fluids protect

  11. Algicidal activity of an actinomycete strain, Streptomyces rameus, against Microcystis aeruginosa.

    PubMed

    Phankhajon, Kanchariya; Somdee, Anchana; Somdee, Theerasak

    2016-09-01

    An actinomycete strain (KKU-A3) with algicidal activity against Microcystis aeruginosa was isolated from soil in Khon Kaen Province, Thailand. Based on its phenotypic characteristics and 16S rDNA sequence, strain KKU-A3 was identified as Streptomyces rameus. Strain KKU-A3 also exhibited algicidal activity against the cyanobacteria Synechococcus elongatus, Cylindrospermum sp. and Oscillatoria sp. A mathematical and statistical technique was used to optimize the culture conditions and maximize its anti-Microcystis activity. The single factor experiments indicated that glucose and casein were the most effective carbon and nitrogen sources, respectively, and produced the highest anti-Microcystis activity. Response surface methodology indicated that the optimum culture conditions were 19.81 g/L glucose and 2.0 g/L casein at an initial pH of 7.8 and an incubation temperature of 30 °C. The anti-Microcystis activity increased from 82% to 95% under optimum conditions. In an internal airlift loop bioreactor, the removal of M. aeruginosa KKU-13 by the bacterium was investigated in batch and continuous flow experiments. In the batch experiment, KKU-A3 displayed maximum anti-Microcystis activity of 95% at day 7, whereas in the continuous flow experiment, KKU-A3 displayed maximum anti-Microcystis activity of 95% at day 10.

  12. Levofloxacin/imipenem prevents the emergence of high-level resistance among Pseudomonas aeruginosa strains already lacking susceptibility to one or both drugs.

    PubMed

    Lister, Philip D; Wolter, Daniel J; Wickman, Paul A; Reisbig, Mark D

    2006-05-01

    Previous studies have demonstrated that a combination of levofloxacin with imipenem could prevent the emergence of resistance during the treatment of susceptible Pseudomonas aeruginosa isolates in a two-compartment pharmacodynamic model of infection. In this study, the efficacy of levofloxacin/imipenem was further evaluated against a panel of characterized P. aeruginosa strains that lacked susceptibility to one or both drugs in the combination. Five P. aeruginosa strains with characterized resistance mechanisms were evaluated. Log-phase cultures were inoculated into the peripheral compartment of the in vitro pharmacokinetic model and treated using simulated doses of 750 mg levofloxacin (dosed every 24 h) and 250 mg or 1 g doses of imipenem (dosed every 12 h). Peak levels were adjusted for protein binding. Pharmacodynamic interactions were evaluated by measuring the changes in viable counts over 30 h. To evaluate the emergence of resistance, samples removed at 30 h were plated onto agar containing the drug at 4x MIC, and potential mutants were evaluated for changes in susceptibility. Against strains overexpressing MexAB-OprM, MexCD-OprJ and MexEF-OprN efflux pumps, levofloxacin/imipenem prevented the emergence of resistance and achieved a 5 log total kill of one strain and eradication of two strains. Levofloxacin/imipenem also eradicated an imipenem-resistant strain lacking OprD. Although the combination initially killed 6-7 logs of a dual-resistant strain lacking OprD and overexpressing MexXY, it could not prevent the emergence of resistance when the 250 mg dose of imipenem was simulated in the combination. However, when the 1 g dose of imipenem was simulated with the combination, resistance was suppressed. These data suggest that levofloxacin/imipenem may be an effective combination for preventing the emergence of resistance among P. aeruginosa, even with strains already lacking susceptibility to one or both drugs in the combination. Clinical evaluation of this

  13. Antimicrobial Resistance Pattern and Their Beta-Lactamase Encoding Genes among Pseudomonas aeruginosa Strains Isolated from Cancer Patients

    PubMed Central

    Zafer, Mai M.; Al-Agamy, Mohamed H.; El-Mahallawy, Hadir A.; Amin, Magdy A.; Ashour, Mohammed Seif El-Din

    2014-01-01

    This study was designed to investigate the prevalence of metallo-β-lactamases (MBL) and extended-spectrum β-lactamases (ESBL) in P. aeruginosa isolates collected from two different hospitals in Cairo, Egypt. Antibiotic susceptibility testing and phenotypic screening for ESBLs and MBLs were performed on 122 P. aeruginosa isolates collected in the period from January 2011 to March 2012. MICs were determined. ESBLs and MBLs genes were sought by PCR. The resistant rate to imipenem was 39.34%. The resistance rates for P. aeruginosa to cefuroxime, cefoperazone, ceftazidime, aztreonam, and piperacillin/tazobactam were 87.7%, 80.3%, 60.6%, 45.1%, and 25.4%, respectively. Out of 122 P. aeruginosa, 27% and 7.4% were MBL and ESBL, respectively. The prevalence of bla VIM-2, bla OXA-10-, bla VEB-1, bla NDM-, and bla IMP-1-like genes were found in 58.3%, 41.7%, 10.4%, 4.2%, and 2.1%, respectively. GIM-, SPM-, SIM-, and OXA-2-like genes were not detected in this study. OXA-10-like gene was concomitant with VIM-2 and/or VEB. Twelve isolates harbored both OXA-10 and VIM-2; two isolates carried both OXA-10 and VEB. Only one strain contained OXA-10, VIM-2, and VEB. In conclusion, bla VIM-2- and bla OXA-10-like genes were the most prevalent genes in P. aeruginosa in Egypt. To our knowledge, this is the first report of bla VIM-2, bla IMP-1, bla NDM, and bla OXA-10 in P. aeruginosa in Egypt. PMID:24707471

  14. Mutations in β-Lactamase AmpC Increase Resistance of Pseudomonas aeruginosa Isolates to Antipseudomonal Cephalosporins

    PubMed Central

    Berrazeg, M.; Jeannot, K.; Ntsogo Enguéné, Véronique Yvette; Broutin, I.; Loeffert, S.; Fournier, D.

    2015-01-01

    Mutation-dependent overproduction of intrinsic β-lactamase AmpC is considered the main cause of resistance of clinical strains of Pseudomonas aeruginosa to antipseudomonal penicillins and cephalosporins. Analysis of 31 AmpC-overproducing clinical isolates exhibiting a greater resistance to ceftazidime than to piperacillin-tazobactam revealed the presence of 17 mutations in the β-lactamase, combined with various polymorphic amino acid substitutions. When overexpressed in AmpC-deficient P. aeruginosa 4098, the genes coding for 20/23 of these AmpC variants were found to confer a higher (2-fold to >64-fold) resistance to ceftazidime and ceftolozane-tazobactam than did the gene from reference strain PAO1. The mutations had variable effects on the MICs of ticarcillin, piperacillin-tazobactam, aztreonam, and cefepime. Depending on their location in the AmpC structure and their impact on β-lactam MICs, they could be assigned to 4 distinct groups. Most of the mutations affecting the omega loop, the R2 domain, and the C-terminal end of the protein were shared with extended-spectrum AmpCs (ESACs) from other Gram-negative species. Interestingly, two new mutations (F121L and P154L) were predicted to enlarge the substrate binding pocket by disrupting the stacking between residues F121 and P154. We also found that the reported ESACs emerged locally in a variety of clones, some of which are epidemic and did not require hypermutability. Taken together, our results show that P. aeruginosa is able to adapt to efficacious β-lactams, including the newer cephalosporin ceftolozane, through a variety of mutations affecting its intrinsic β-lactamase, AmpC. Data suggest that the rates of ESAC-producing mutants are ≥1.5% in the clinical setting. PMID:26248364

  15. WCK 5107 (Zidebactam) and WCK 5153 Are Novel Inhibitors of PBP2 Showing Potent "β-Lactam Enhancer" Activity against Pseudomonas aeruginosa, Including Multidrug-Resistant Metallo-β-Lactamase-Producing High-Risk Clones.

    PubMed

    Moya, Bartolome; Barcelo, Isabel M; Bhagwat, Sachin; Patel, Mahesh; Bou, German; Papp-Wallace, Krisztina M; Bonomo, Robert A; Oliver, Antonio

    2017-06-01

    Zidebactam and WCK 5153 are novel β-lactam enhancers that are bicyclo-acyl hydrazides (BCH), derivatives of the diazabicyclooctane (DBO) scaffold, targeted for the treatment of serious infections caused by highly drug-resistant Gram-negative pathogens. In this study, we determined the penicillin-binding protein (PBP) inhibition profiles and the antimicrobial activities of zidebactam and WCK 5153 against Pseudomonas aeruginosa , including multidrug-resistant (MDR) metallo-β-lactamase (MBL)-producing high-risk clones. MIC determinations and time-kill assays were conducted for zidebactam, WCK 5153, and antipseudomonal β-lactams using wild-type PAO1, MexAB-OprM-hyperproducing ( mexR ), porin-deficient ( oprD ), and AmpC-hyperproducing ( dacB ) derivatives of PAO1, and MBL-expressing clinical strains ST175 ( bla VIM-2 ) and ST111 ( bla VIM-1 ). Furthermore, steady-state kinetics was used to assess the inhibitory potential of these compounds against the purified VIM-2 MBL. Zidebactam and WCK 5153 showed specific PBP2 inhibition and did not inhibit VIM-2 (apparent K i [ K i app ] > 100 μM). MICs for zidebactam and WCK 5153 ranged from 2 to 32 μg/ml (amdinocillin MICs > 32 μg/ml). Time-kill assays revealed bactericidal activity of zidebactam and WCK 5153. LIVE-DEAD staining further supported the bactericidal activity of both compounds, showing spheroplast formation. Fixed concentrations (4 or 8 μg/ml) of zidebactam and WCK 5153 restored susceptibility to all of the tested β-lactams for each of the P. aeruginosa mutant strains. Likewise, antipseudomonal β-lactams (CLSI breakpoints), in combination with 4 or 8 μg/ml of zidebactam or WCK 5153, resulted in enhanced killing. Certain combinations determined full bacterial eradication, even with MDR MBL-producing high-risk clones. β-Lactam-WCK enhancer combinations represent a promising β-lactam "enhancer-based" approach to treat MDR P. aeruginosa infections, bypassing the need for MBL inhibition. Copyright © 2017

  16. [The description of an esculin-positive biovar of Pseudomonas aeruginosa].

    PubMed

    Sivolodskiĭ, E P

    2000-01-01

    In the study of 280 P. aeruginosa strains isolated in different hospitals of St. Petersburg for the first time 48 strains capable of hydrolyzing esculin have been detected. The hydrolysis of esculin is determined in plates with the use of the microvolume techniques the results were evaluated after 3-hour incubation at 37 degrees C. The data confirming the existence of the exculin-positive biovar of P. aeruginosa have been obtained; these data show the wide spread of esculin-positive strains in hospitals of different specialization (17.1 +/- 5.1% of P. aeruginosa strains), the characteristic combination of the sign of esculin hydrolysis with such signs as the absence of the smell of trimethylamine and the phenomenon of "iridescent lysis" of the colonies, the stability of the sign of esculin hydrolysis in strains, repeatedly isolated from patients, after the storage of the cultures and their treatment with plasmid-eliminating preparation. The name "esculinolytica" has been proposed for this biovar. The typing strain of biovar esculinolytica has been deposited in the culture collection of the Russian Research Institute of Agricultural Microbiology as P. aeruginosa ARRIAM 64-A. This biovar been found to be most widely spread in urological hospitals, where esculin-positive strains are isolated 3 times more frequently (32.2 +/- 5.1% of P. aeruginosa strains) than in surgical hospitals (10.7 +/- 2.2%).

  17. WCK 5107 (Zidebactam) and WCK 5153 Are Novel Inhibitors of PBP2 Showing Potent “β-Lactam Enhancer” Activity against Pseudomonas aeruginosa, Including Multidrug-Resistant Metallo-β-Lactamase-Producing High-Risk Clones

    PubMed Central

    Barcelo, Isabel M.; Bhagwat, Sachin; Patel, Mahesh; Bou, German; Papp-Wallace, Krisztina M.; Bonomo, Robert A.; Oliver, Antonio

    2017-01-01

    ABSTRACT Zidebactam and WCK 5153 are novel β-lactam enhancers that are bicyclo-acyl hydrazides (BCH), derivatives of the diazabicyclooctane (DBO) scaffold, targeted for the treatment of serious infections caused by highly drug-resistant Gram-negative pathogens. In this study, we determined the penicillin-binding protein (PBP) inhibition profiles and the antimicrobial activities of zidebactam and WCK 5153 against Pseudomonas aeruginosa, including multidrug-resistant (MDR) metallo-β-lactamase (MBL)-producing high-risk clones. MIC determinations and time-kill assays were conducted for zidebactam, WCK 5153, and antipseudomonal β-lactams using wild-type PAO1, MexAB-OprM-hyperproducing (mexR), porin-deficient (oprD), and AmpC-hyperproducing (dacB) derivatives of PAO1, and MBL-expressing clinical strains ST175 (blaVIM-2) and ST111 (blaVIM-1). Furthermore, steady-state kinetics was used to assess the inhibitory potential of these compounds against the purified VIM-2 MBL. Zidebactam and WCK 5153 showed specific PBP2 inhibition and did not inhibit VIM-2 (apparent Ki [Ki app] > 100 μM). MICs for zidebactam and WCK 5153 ranged from 2 to 32 μg/ml (amdinocillin MICs > 32 μg/ml). Time-kill assays revealed bactericidal activity of zidebactam and WCK 5153. LIVE-DEAD staining further supported the bactericidal activity of both compounds, showing spheroplast formation. Fixed concentrations (4 or 8 μg/ml) of zidebactam and WCK 5153 restored susceptibility to all of the tested β-lactams for each of the P. aeruginosa mutant strains. Likewise, antipseudomonal β-lactams (CLSI breakpoints), in combination with 4 or 8 μg/ml of zidebactam or WCK 5153, resulted in enhanced killing. Certain combinations determined full bacterial eradication, even with MDR MBL-producing high-risk clones. β-Lactam–WCK enhancer combinations represent a promising β-lactam “enhancer-based” approach to treat MDR P. aeruginosa infections, bypassing the need for MBL inhibition. PMID:28289035

  18. Molecular detection of metallo-β-lactamase genes, bla IMP-1, bla VIM-2 and bla SPM-1 in imipenem resistant Pseudomonas aeruginosa isolated from clinical specimens in teaching hospitals of Ahvaz, Iran.

    PubMed

    Moosavian, Mojtaba; Rahimzadeh, Mohammad

    2015-02-01

    Carbapenem resistant Pseudomonas aeruginosa is a serious cause of nosocomial infections. The main purpose of the study is to determine the prevalence rate of imipenem resistant Pseudomonas aeruginosa carrying metallo-ß-lactamase (MBL) genes. 236 Pseudomonas aeruginosa isolates were collected from teaching hospitals of Ahvaz University of Medical Sciences during a period of 9 months in 2012. These strains were identified using conventional microbiological tests. The susceptibility of isolates to antibiotics were assessed using disk diffusion test. The IMP-EDTA combination disk phenotypic test was performed for detection of MBL producing strains. Finally, polymerase chain reaction (PCR) was performed to detect MBL genes, bla IMP-1, bla VIM-2 and bla SPM-1 in imipenem resistant strains. Out of 236 examined isolates, 122 isolates (51.4%) were resistant to imipenem. The IMP-EDTA combination test showed that among 122 imipenem resistant strains, 110 strains (90%) were phenotipically MBL producers. Additionally, the results of PCR method showed that 2 strains (1.6%) and 67strains (55%) of imipenem resistant Pseudomonas aeruginosa isolates contained bla VIM-2 and bla IMP-1 genes respectively. No SPM-1gene was found in the examined samples. Resistance of P. aeruginosa isolates to imipenem due to MBL enzymes is increasing in Ahavaz. Because of clinical significance of this kind of resistance, rapid detection of MBL producing strains and followed by appropriate treatment is necessary to prevent the spreading of these organisms.

  19. Genomic characterisation of clinical and environmental Pseudomonas putida group strains and determination of their role in the transfer of antimicrobial resistance genes to Pseudomonas aeruginosa.

    PubMed

    Peter, Silke; Oberhettinger, Philipp; Schuele, Leonard; Dinkelacker, Ariane; Vogel, Wichard; Dörfel, Daniela; Bezdan, Daniela; Ossowski, Stephan; Marschal, Matthias; Liese, Jan; Willmann, Matthias

    2017-11-10

    Pseudomonas putida is a Gram-negative, non-fermenting bacterium frequently encountered in various environmental niches. P. putida rarely causes disease in humans, though serious infections and outbreaks have been reported from time to time. Some have suggested that P. putida functions as an exchange platform for antibiotic resistance genes (ARG), and thus represents a serious concern in the spread of ARGs to more pathogenic organisms within a hospital. Though poorly understood, the frequency of ARG exchange between P. putida and the more virulent Pseudomonas aeruginosa and its clinical relevance are particularly important for designing efficient infection control strategies, such as deciding whether high-risk patients colonized with a multidrug resistant but typically low pathogenic P. putida strain should be contact isolated or not. In this study, 21,373 screening samples (stool, rectal and throat swab) were examined to determine the presence of P. putida in a high-risk group of haemato-oncology patients during a 28-month period. A total of 89 P. putida group strains were isolated from 85 patients, with 41 of 89 (46.1%) strains harbouring the metallo-beta-lactamase gene bla VIM . These 41 clinical isolates, plus 18 bla VIM positive environmental P. putida isolates, and 17 bla VIM positive P. aeruginosa isolates, were characterized by whole genome sequencing (WGS). We constructed a maximum-likelihood tree to separate the 59 bla VIM positive P. putida group strains into eight distinct phylogenetic clusters. Bla VIM-1 was present in 6 clusters while bla VIM-2 was detected in 4 clusters. Five P. putida group strains contained both, bla VIM-1 and bla VIM-2 genes. In contrast, all P. aeruginosa strains belonged to a single genetic cluster and contained the same ARGs. Apart from bla VIM-2 and sul genes, no other ARGs were shared between P. aeruginosa and P. putida. Furthermore, the bla VIM-2 gene in P. aeruginosa was predicted to be only chromosomally located. These data

  20. Growth and Photosynthetic Characteristics of Toxic and Non-Toxic Strains of the Cyanobacteria Microcystis aeruginosa and Anabaena circinalis in Relation to Light

    PubMed Central

    Islam, M. Ashraful; Beardall, John

    2017-01-01

    Cyanobacteria are major bloom-forming organisms in freshwater ecosystems and many strains are known to produce toxins. Toxin production requires an investment in energy and resources. As light is one of the most important factors for cyanobacterial growth, any changes in light climate might affect cyanobacterial toxin production as well as their growth and physiology. To evaluate the effects of light on the growth and physiological parameters of both toxic and non-toxic strains of Microcystis aeruginosa and Anabaena circinalis, cultures were grown at a range of light intensities (10, 25, 50, 100, 150 and 200 µmol m−2 s−1). The study revealed that the toxic strains of both species (CS558 for M. aeruginosa and CS537 and CS541 for A. circinalis) showed growth (µ) saturation at a higher light intensity compared to the non-toxic strains (CS338 for M. aeruginosa and CS534 for A. circinalis). Both species showed differences in chlorophyll a, carotenoid, allophycocyanin (APC) and phycoerythrin (PE) content between strains. There were also differences in dark respiration (Rd), light saturated oxygen evolution rates (Pmax) and efficiency of light harvesting (α) between strains. All other physiological parameters showed no statistically significant differences between strains. This study suggest that the different strains respond differently to different light habitats. Thus, changes in light availability may affect bloom intensity of toxic and nontoxic strains of cyanobacteria by changing the dominance and succession patterns. PMID:28777340

  1. Role of the Pseudomonas quinolone signal (PQS) in sensitising Pseudomonas aeruginosa to UVA radiation.

    PubMed

    Pezzoni, Magdalena; Meichtry, Martín; Pizarro, Ramón A; Costa, Cristina S

    2015-01-01

    One of the main stress factors that bacteria face in the environment is solar ultraviolet-A (UVA) radiation, which leads to lethal effects through oxidative damage. The aim of this work was to investigate the role of 2-heptyl-3-hydroxi-4-quinolone (the Pseudomonas quinolone signal or PQS) in the response of Pseudomonas aeruginosa to UVA radiation. PQS is an intercellular quorum sensing signal associated to membrane vesicles which, among other functions, regulates genes related to iron acquisition, forms stable complexes with iron and participates in oxidative phenomena. UVA exposure of the wild-type PAO1 strain and a pqsA mutant unable to produce PQS revealed a sensitising role for this signal. Research into the mechanism involved in this phenomenon revealed that catalase, an essential factor in the UVA defence, is not related to PQS-mediated UVA sensitivity. Absorption of UVA by PQS produced its own photo-degradation, oxidation of the probe 2',7'- dichlorodihydrofluorescein and generation of singlet oxygen and superoxide anion, suggesting that this signal could be acting as an endogenous photosensitiser. The results presented in this study could explain the high sensitivity to UVA of P. aeruginosa when compared to enteric bacteria. Copyright © 2014 Elsevier B.V. All rights reserved.

  2. Flagellin inhibits Myoviridae phage phiCTX infection of Pseudomonas aeruginosa strain GuA18: purification and mapping of binding site.

    PubMed

    Geiben-Lynn, R; Sauber, K; Lutz, F

    2001-11-01

    PhiCTX is a double-stranded DNA phage of the Myoviridae family that converts Pseudomonas aeruginosa into a cytotoxin producer. A 42-kDa phiCTX-inhibiting protein was purified from the outer membrane fraction of P. aeruginosa strain GuA18 by octyl-beta-glucoside extraction, DEAE-chromatography, and mono-Q HPLC. This protein had an isoelectric point of 5.4 and bound specifically [125I]-labeled phiCTX. The N-terminal amino acid sequence of six out of seven Lys-C fragments was highly similar (87%) to that of the entire of type-a flagellin of P. aeruginosa strain PAK. At a concentration of 14 nM, purified flagellin protein caused a 50% decrease in the phage titer after a 20-min incubation at 37 degrees C (PhI50). The presence of ethanol was necessary to reconstitute the inhibitory activity. In contrast, no ethanol treatment was necessary for the inhibitory activity of the sheared flagellin filaments from P. aeruginosa strain GuA18, which consists of the 42-kDa flagellin subunits and the synthesized 17-mer phage-binding-peptide NGSNSDSERTALNGEAK, representing flagellin residues 100-116 of P. aeruginosa strain PAK. The PhI50 was 10 nM and 200 nM, respectively. Antisera against the flagellin filament protein as well as against the 17-mer peptide neutralized phage infection. These results indicated that the amino acid region 100-116 of the flagellin subunit of strain GuA18 is involved in phiCTX binding. This region might play a role in phage attachment.

  3. Molecular analysis of carbapenem-resistant strains of Pseudomonas aeruginosa isolated from patients hospitalized in various transplantation wards between 2008 and 2011.

    PubMed

    Kosykowska, E; Szymanek-Majchrzak, K; Walter de Walthoffen, S; Izdebski, R; Mlynarczyk, A; Ciszek, M; Chmura, A; Durlik, M; Paczek, L; Deborska-Materkowska, D; Sawicka-Grzelak, A; Mlynarczyk, G

    2014-10-01

    Recent years have seen a concerning increase in the number of carbapenem-resistant Pseudomonas aeruginosa strains. P aeruginosa is one of the most dangerous factors causing nosocomial infections, and immunosuppressed patients constitute a special risk group. The purpose of our study was to conduct a molecular analysis of 22 clinical isolates of carbapenem-resistant P aeruginosa obtained between 2008 and 2011. Metallo-beta-lactamase (MBL) phenotype tests were conducted. A polymerase chain reaction technique was used to detect VIM, IMP, NDM, and GIM carbapenemase-encoding genes. The minimum inhibitory concentrations were determined for imipenem, meropenem, and doripenem. Molecular typing was conducted with the use of restriction fragment length polymorphism/pulsed-field gel electrophoresis (RFLP-PFGE). Of the 22 strains initially resistant to at least one carbapenem, we selected 18 that exhibited the MBL phenotype. Of those 18, we identified 15 strains expressing VIM carbapenemase-encoding genes. None of the other evaluated genes were detected. VIM-positive isolates exhibited higher levels of resistance than the other ones. The RFLP technique revealed 10 different PFGE types and 6 epidemic foci. Identical strains were isolated over the period of up to 3 years. The reason for resistance to carbapenems in the majority (68%) of P aeruginosa strains isolated at the evaluated hospital was the presence of VIM carbapenemase. It is safe to say that the VIM carbapenemase is responsible for a higher level of resistance than unidentified mechanisms. Carbapenem-resistant strains of P aeruginosa spread clonally within individual wards and are likely to be of hospital origin.

  4. [Degradation characteristics of naphthalene with a Pseudomonas aeruginosa strain isolated from soil contaminated by diesel].

    PubMed

    Liu, Wen-Chao; Wu, Bin-Bin; Li, Xiao-Sen; Lu, Dian-Nan; Liu, Yong-Min

    2015-02-01

    Abstract: A naphthalene-degrading bacterium (referred as HD-5) was isolated from the diesel-contaminated soil and was assigned to Pseudomonas aeruginosa according to 16S rDNA sequences analysis. Gene nah, which encodes naphthalene dioxygenase, was identified from strain HD-5 by PCR amplification. Different bioremediation approaches, including nature attenuation, bioaugmentation with strain Pseudomonas aeruginosa, biostimulation, and an integrated degradation by bioaugmentation and biostimulation, were evaluated for their effectiveness in the remediating soil containing 5% naphthalene. The degradation rates of naphthalene in the soil were compared among the different bioremediation approaches, the FDA and dehydrogenase activity in bioremediation process were measured, and the gene copy number of 16S rRNA and nah in soil were dynamically monitored using real-time PCR. It was shown that the naphthalene removal rate reached 71.94%, 62.22% and 83.14% in approaches of bioaugmentation (B), biostimulation(S) and integrated degradation composed of bioaugmentation and biostimulation (BS), respectively. The highest removal rate of naphthalene was achieved by using BS protocol, which also gives the highest FDA and dehydrogenase activity. The gene copy number of 16S rRNA and nah in soil increased by about 2.67 x 10(11) g(-1) and 8.67 x 10(8) g(-1) after 31 days treatment using BS protocol. Above-mentioned results also demonstrated that the screened bacterium, Pseudomonas aeruginosa, could grow well in naphthalene-contaminated soil and effectively degrade naphthalene, which is of fundamental importance for bioremediation of naphthalene-contaminated soil.

  5. The atu and liu clusters are involved in the catabolic pathways for acyclic monoterpenes and leucine in Pseudomonas aeruginosa.

    PubMed

    Aguilar, J A; Zavala, A N; Díaz-Pérez, C; Cervantes, C; Díaz-Pérez, A L; Campos-García, J

    2006-03-01

    Evidence suggests that the Pseudomonas aeruginosa PAO1 gnyRDBHAL cluster, which is involved in acyclic isoprenoid degradation (A. L. Díaz-Pérez, N. A. Zavala-Hernández, C. Cervantes, and J. Campos-García, Appl. Environ. Microbiol. 70:5102-5110, 2004), corresponds to the liuRABCDE cluster (B. Hoschle, V. Gnau, and D. Jendrossek, Microbiology 151:3649-3656, 2005). A liu (leucine and isovalerate utilization) homolog cluster was found in the PAO1 genome and is related to the catabolism of acyclic monoterpenes of the citronellol family (AMTC); it was named the atu cluster (acyclic terpene utilization), consisting of the atuCDEF genes and lacking the hydroxymethyl-glutaryl-coenzyme A (CoA) lyase (HMG-CoA lyase) homolog. Mutagenesis of the atu and liu clusters showed that both are involved in AMTC and leucine catabolism by encoding the enzymes related to the geranyl-CoA and the 3-methylcrotonyl-CoA pathways, respectively. Intermediary metabolites of the acyclic monoterpene pathway, citronellic and geranic acids, were accumulated, and leucine degradation rates were affected in both atuF and liuD mutants. The alpha subunit of geranyl-CoA carboxylase and the alpha subunit of 3-methylcrotonyl-CoA carboxylase (alpha-MCCase), encoded by the atuF and liuD genes, respectively, were both induced by citronellol, whereas only the alpha-MCCase subunit was induced by leucine. Both citronellol and leucine also induced a LacZ transcriptional fusion at the liuB gene. The liuE gene encodes a probable hydroxy-acyl-CoA lyase (probably HMG-CoA lyase), an enzyme with bifunctional activity that is essential for both AMTC and leucine degradation. P. aeruginosa PAO1 products encoded by the liuABCD cluster showed a higher sequence similarity (77.2 to 79.5%) with the probable products of liu clusters from several Pseudomonas species than with the atuCDEF cluster from PAO1 (41.5%). Phylogenetic studies suggest that the atu cluster from P. aeruginosa could be the result of horizontal transfer

  6. Characterization of carbapenem resistance mechanisms and integrons in Pseudomonas aeruginosa strains from blood samples in a French hospital.

    PubMed

    Rojo-Bezares, Beatriz; Cavalié, Laurent; Dubois, Damien; Oswald, Eric; Torres, Carmen; Sáenz, Yolanda

    2016-04-01

    Metallo-β-lactamases (MBLs), porin OprD, integrons, virulence factors and the clonal relationships were characterized in imipenem-resistant Pseudomonas aeruginosa (IRPA) isolates. Fifty-six IRPA strains were recovered from blood samples of different patients at a Toulouse teaching hospital from 2011 to 2013. Susceptibility testing of 14 antibiotics was performed by the disc diffusion method. Detection and characterization of MBLs, the oprD gene and integrons were studied by PCR and sequencing. Thirteen genes involved in the virulence of P. aeruginosa were analysed. Molecular typing of IRPA strains was performed by PFGE and multilocus sequence typing. In this study, 61 % of the IRPA isolates showed a multi-resistance phenotype. The MBL phenotype, detected in three isolates (5.4 %), was linked to the blaVIM-2 gene. The oprD gene was amplified in 55 (98.2 %) IRPA strains, and variations were observed in 54 of them. Insertion sequences (IS) truncating oprD were detected in eight IRPA strains, with the novel ISPa56 identified in two strains. Class 1 integrons were detected in 24 (42.9 %) IRPA strains. The blaVIM-2 gene was found inside the class 1 integron arrangements. The new integrons In1054 (intI1-aacA56-qacEΔ1-sul1) and In1160 (intI1-aacA4-aacC1d-ISKpn4-gcuE-qacEΔ1-sul1) have been described for the first time, to the best of our knowledge, in this study. A high clonal diversity was found in our strains. Among the variety of sequence types (STs) found, ST175, ST233, ST235, ST244 and ST654 were noteworthy as epidemic clones. In conclusion, 5.4 % of IRPA strains showed an MBL phenotype linked to the blaVIM-2 gene. The identified oprD high polymorphism could be implicated in the variable resistance to carbapenems in IRPA strains. The dissemination of high-risk clones is a cause of concern.

  7. Population-based epidemiological study of infections caused by carbapenem-resistant Pseudomonas aeruginosa in the Calgary Health Region: importance of metallo-beta-lactamase (MBL)-producing strains.

    PubMed

    Laupland, Kevin B; Parkins, Michael D; Church, Deirdre L; Gregson, Daniel B; Louie, Thomas J; Conly, John M; Elsayed, Sameer; Pitout, Johann D D

    2005-11-01

    A study was conducted in the Calgary Health Region between May 2002 and April 2004 to define the population-based epidemiological characteristics of infections caused by imipenem-resistant Pseudomonas aeruginosa and to explore the clinical outcomes due to metallo- beta -lactamase (MBL)-producing and non-MBL-producing strains. Detailed clinical information was obtained by chart review, and phenotypic and molecular characterizations were performed using the MBL E-test, polymerase chain reaction with sequencing, and pulsed-field gel electrophoresis. A total of 228 patients with infections caused by imipenem-resistant P. aeruginosa were identified (annual incidence, 10.5 cases/100,000 population), with the highest incidence rate in those >or=75 years old. MBL-producing strains (98/228) were associated with higher rates of multidrug resistance and bacteremia. Ninety MBL-producing strains also produced VIM-2, 4 produced IMP-7, and 4 were unclassified. A cluster of VIM-2-producing strains was responsible for a nosocomial outbreak during 2003. The case-fatality rate was significantly higher for infections caused by MBL-producing strains than for those caused by non-MBL-producing strains (25% vs. 13%; relative risk, 1.98 [95% confidence interval, 1.00-3.90]; P=.05). MBL-producing P. aeruginosa strains were associated with a higher case-fatality rate and invasive disease. Our study highlights the potential importance of molecular laboratory techniques in infection control and patient care.

  8. Comparison of arbitrarily primed PCR and macrorestriction (pulsed-field gel electrophoresis) typing of Pseudomonas aeruginosa strains from cystic fibrosis patients.

    PubMed Central

    Kersulyte, D; Struelens, M J; Deplano, A; Berg, D E

    1995-01-01

    Arbitrarily primed PCR fingerprinting was carried out on 43 Pseudomonas aeruginosa isolates from cystic fibrosis (CF) patients. Seventeen major groups of strains that coincided with groups also distinguished by macrorestriction (pulsed-field gel electrophoresis) typing were identified. Our results illustrated that a CF patient can carry more than one strain and can carry a given strain for long periods of time and that strains can evolve by changes in drug resistance or other phenotypic traits during long-term colonization. The arbitrarily primed PCR method is recommended for first-pass screening of P. aeruginosa isolates from CF patients, especially when many strains are to be typed, because of its sensitivity and efficiency. PMID:7559985

  9. Anti-Pseudomonas aeruginosa compound, 1,2,3,4-tetrahydro-1,3,5-triazine derivative, exerts its action by primarily targeting MreB.

    PubMed

    Yamachika, Shinichiro; Sugihara, Chika; Tsuji, Hayato; Muramatsu, Yasunori; Kamai, Yasuki; Yamashita, Makoto

    2012-01-01

    In order to find new anti-Pseudomonas agents, we carried out whole-cell based P. aeruginosa growth assay, and identified 1,2,3,4-tetrahydro-1,3,5-triazine (Compound A). This compound showed anti-Pseudomonas activity against wild as well as pumpless strain equally at a same concentration. Also, this compound was structurally very similar to A22, which is known to inhibit the bacterial actin-like protein MreB. By the analysis of resistant strains, the primary target of this compound in P. aeruginosa was definitely confirmed to be MreB. In addition, these compounds showed a bacteriostatic effect, and induced the morphology changes in P. aeruginosa from rod shape to sphere shape, which leads to be clinically favorable in terms of susceptibility to phagocytosis and release of endotoxin. These results display that Compound A is a very attractive compound which shows anti-P. aeruginosa activity based on inhibition of MreB without being affected by efflux pumps, and could provide a new step toward development of new promising anti-Pseudomonas agents, MreB inhibitors.

  10. Unexpected diversity in the mobilome of a Pseudomonas aeruginosa strain isolated from a dental unit waterline revealed by SMRT Sequencing.

    PubMed

    Vincent, Antony T; Charette, Steve J; Barbeau, Jean

    2018-05-01

    The Gram-negative bacterium Pseudomonas aeruginosa is found in several habitats, both natural and human-made, and is particularly known for its recurrent presence as a pathogen in the lungs of patients suffering from cystic fibrosis, a genetic disease. Given its clinical importance, several major studies have investigated the genomic adaptation of P. aeruginosa in lungs and its transition as acute infections become chronic. However, our knowledge about the diversity and adaptation of the P. aeruginosa genome to non-clinical environments is still fragmentary, in part due to the lack of accurate reference genomes of strains from the numerous environments colonized by the bacterium. Here, we used PacBio long-read technology to sequence the genome of PPF-1, a strain of P. aeruginosa isolated from a dental unit waterline. Generating this closed genome was an opportunity to investigate genomic features that are difficult to accurately study in a draft genome (contigs state). It was possible to shed light on putative genomic islands, some shared with other reference genomes, new prophages, and the complete content of insertion sequences. In addition, four different group II introns were also found, including two characterized here and not listed in the specialized group II intron database.

  11. Role of TonB1 in Pyoverdine-Mediated Signaling in Pseudomonas aeruginosa▿

    PubMed Central

    Shirley, Matt; Lamont, Iain L.

    2009-01-01

    Pyoverdines are siderophores secreted by Pseudomonas aeruginosa. Uptake of ferripyoverdine in P. aeruginosa PAO1 occurs via the FpvA receptor protein and requires the energy-transducing protein TonB1. Interaction of (ferri)pyoverdine with FpvA activates pyoverdine gene expression in a signaling process involving the cytoplasmic-membrane-spanning anti-sigma factor FpvR and the sigma factor PvdS. Here, we show that mutation of a region of FpvA that interacts with TonB1 (the TonB box) prevents this signaling process, as well as inhibiting bacterial growth in the presence of the iron-chelating compound ethylenediamine-di(o-hydroxy-phenylacetic acid). Signaling via wild-type FpvA was also eliminated in strains lacking TonB1 but was unaffected in strains lacking either (or both) of two other TonB proteins in P. aeruginosa, TonB2 and TonB3. An absence of pyoverdine-mediated signaling corresponded with proteolysis of PvdS. These data show that interactions between FpvA and TonB1 are required for (ferri)pyoverdine signal transduction, as well as for ferripyoverdine transport, consistent with a mechanistic link between the signaling and transport functions of FpvA. PMID:19592589

  12. Pseudomonas aeruginosa Trent and zinc homeostasis.

    PubMed

    Davies, Corey B; Harrison, Mark D; Huygens, Flavia

    2017-09-01

    Pseudomonas aeruginosa is a Gram-negative pathogen and the major cause of mortality in patients with cystic fibrosis. The mechanisms that P. aeruginosa strains use to regulate intracellular zinc have an effect on infection, antibiotic resistance and the propensity to form biofilms. However, zinc homeostasis in P. aeruginosa strains of variable infectivity has not been compared. In this study, zinc homeostasis in P. aeruginosa Trent, a highly infectious clinical strain, was compared to that of a laboratory P. aeruginosa strain, ATCC27853. Trent was able to tolerate higher concentrations of additional zinc in rich media than ATCC27853. Further, pre-adaptation to additional zinc enhanced the growth of Trent at non-inhibitory concentrations but the impact of pre-adaption on the growth of ATCC27853 under the same conditions was minimal. The results establish clear differences in zinc-induced responses in Trent and ATCC27853, and how zinc homeostasis can be a promising target for the development of novel antimicrobial strategies for P. aeruginosa infection in cystic fibrosis patients. © FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  13. Phenotypic and Genetic Evaluation of the Influence of Pseudomonas aeruginosa Culture Fractions on the Human Mesenchymal Stem Cells Viability, Apoptotic Pathways and Cytokine Profile.

    PubMed

    Holban, Alina Maria; Bleotu, Coralia; Chifiriuc, Mariana Carmen; Lazar, Veronica

    2017-01-01

    The objective of this study was to investigate the effects of P. aeruginosa PAO1 cellular and soluble culture fractions on human mesenchymal stem cells (MSCs) death signaling pathways and cytokine profile. The bone marrow isolated MSCs, incubated for different periods of time with one of the three P. aeruginosa PAO1 culture fractions, i.e. low density whole cultures, heat inactivated bacterial cultures sediments and sterile supernatants, were submitted to the following assays: i) fluorescence microscopy evaluation of cellular morphology and viability; ii) bax, caspase 9, relA and bcl-2 genes expression analysis by qRT-PCR; and iii) quantification of the level of IL-1β, IL-6, IL-8 and IL-10 cytokines released in the MSCs supernatants determined by ELISA. Results were statistically analyzed using the GraphPad In Stat software. The PAO1 whole cultures exhibited the most relevant influences, impacting on MSCs morphology and viability, interfering with apoptotic pathways and significantly stimulating the production of IL-1β and IL-10, while decreasing the production of IL-6 and IL-8. The culture supernatants increased the production of IL-1β and reduced the secretion of all other tested cytokines, while heat-inactivated bacterial cells significantly stimulated both IL-1β and IL-10 production. These data could suggest that in vivo, the fate of P. aeruginosa infection depends on the proportion between different bacterial culture fractions (i.e. the number of viable bacterial cells, the number of dead cells and the amount of bacterial soluble products accumulated locally) that could be influenced by the initial infective dose, by the host defense mechanisms, and also by the administered antimicrobial treatment that may thus interfere with the evolution and magnitude of the induced lesions. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  14. Purification and characterization of an eggshell membrane decomposing protease from Pseudomonas aeruginosa strain ME-4.

    PubMed

    Cheng, Minyi; Takenaka, Shinji; Aoki, Shunsuke; Murakami, Shuichiro; Aoki, Kenji

    2009-04-01

    A bacterial strain, ME-4, isolated from farm soil and identified as Pseudomonas aeruginosa, grew well on a medium containing eggshell membrane (ESM). P. aeruginosa strain ME-4 decomposed the ESM by producing an extracellular protease able to solubilize it. The protease was purified to homogeneity from culture supernatant by fractionation with (NH(4))(2)SO(4), as well as CM52 cellulose and DE52 cellulose column chromatography, with a final yield of 47%. The molecular mass of the enzyme was 33 kDa. The isolated enzyme was a metalloprotease and was strongly inhibited by EDTA, o-phenanthroline, and phosphoramidon. The enzyme inhibited by these reagents was reactivated in the presence of several metal ions. The enzyme acted on various proteins and showed higher activity with collagen than collagenase from Clostridium histolyticum. Results of assays with the FRETS combinatorial libraries revealed that the enzyme preferred Ser at the P1 position and Lys at the P2 position. It also preferred hydrophobic amino acid residues at the P1' and P2' positions. The enzyme showed a much higher solubilization activity with the ESM substrate than commercially obtained enzymes. The enzyme decomposed ESM to produce water-soluble peptides, Val-Leu-Pro-Pro and (X)-Val-Pro-Pro, and a free amino acid, tryptophan.

  15. Biodegradation of isoproturon using a novel Pseudomonas aeruginosa strain JS-11 as a multi-functional bioinoculant of environmental significance.

    PubMed

    Dwivedi, Sourabh; Singh, Braj Raj; Al-Khedhairy, Abdulaziz A; Musarrat, Javed

    2011-01-30

    Biodegradation of phenylurea herbicide isoproturon was studied in soil microcosm bioaugmented with a novel bacterial strain JS-11 isolated from wheat rhizosphere. The molecular characterization based on 16SrDNA sequence homology confirmed its identity as Pseudomonas aeruginosa strain JS-11. The herbicide was completely degraded within 20 days at ambient temperature with the rate constant of 0.08 day(-1), following the first-order rate kinetics. In stationary phase, at a cell density of 6.5 × 10(9) CFU mL(-1), the bacteria produced substantially increased amounts of indole acetic acid (IAA) in the presence of tryptophan as compared with the control. Also, the bacteria exhibited a time-dependent increase in the amount of tri-calcium phosphate solubilization in Pikovskaya's medium. Further screening of the strain JS-11 for auxiliary activities revealed its remarkable capability of producing the siderophores and hydrogen cyanide (HCN), besides antifungal activity against a common phytopathogen Fusarium oxysporum. Thus, the versatile P. aeruginosa strain JS-11 with innate potential for multifarious biological activities is envisaged as a super-bioinoculant for exploitation in the integrated bioremediation, plant growth and disease management (IBPDM) in contaminated agricultural soils. Copyright © 2010 Elsevier B.V. All rights reserved.

  16. Enhancement of bioelectricity generation via heterologous expression of IrrE in Pseudomonas aeruginosa-inoculated MFCs.

    PubMed

    Luo, Jianmei; Wang, Tingting; Li, Xiao; Yang, Yanan; Zhou, Minghua; Li, Ming; Yan, Zhongli

    2018-05-30

    Low electricity power output (EPT) is still the main bottleneck limited the industrial application of microbial fuel cells (MFCs). Herein, EPT enhancement by introducing an exogenous global regulator IrrE derived from Deinococcus radiodurans into electrochemically active bacteria (EAB) was explored using Pseudomonas aeruginosa PAO1 as a model strain, achieving a power density 71% higher than that of the control strain. Moreover, IrrE-expressing strain exhibited a remarkable increase in the total amount of electron shuttles (majorly phenazines compounds) and a little decrease in internal resistance, which should underlie the enhancement in extracellular electron transfer (EET) efficiency and EPT. Strikingly, IrrE significantly affected substrate utilization profiling, improved cell growth characterization and cell tolerance to various stresses. Further quantitative RT-PCR analysis revealed that IrrE led to many differentially expressed genes, which were responsible for phenazines core biosynthesis, biofilm formation, QS systems, transcriptional regulation, glucose metabolism and general stress response. The results substantiated that targeting cellular regulatory network by the introduction of exogenous global regulators could be a facile and promising approach for the enhancement of bioelectricity generation and cell multiple phenotypes, and thus would be of great potential application in the practical MFCs. Copyright © 2018 Elsevier B.V. All rights reserved.

  17. Agaricus blazei hot water extract shows anti quorum sensing activity in the nosocomial human pathogen Pseudomonas aeruginosa.

    PubMed

    Soković, Marina; Ćirić, Ana; Glamočlija, Jasmina; Nikolić, Miloš; van Griensven, Leo J L D

    2014-04-03

    The edible mushroom Agaricus blazei Murill is known to induce protective immunomodulatory action against a variety of infectious diseases. In the present study we report potential anti-quorum sensing properties of A. blazei hot water extract. Quorum sensing (QS) plays an important role in virulence, biofilm formation and survival of many pathogenic bacteria, including the Gram negative Pseudomonas aeruginosa, and is considered as a novel and promising target for anti-infectious agents. In this study, the effect of the sub-MICs of Agaricus blazei water extract on QS regulated virulence factors and biofilm formation was evaluated against P. aeruginosa PAO1. Sub-MIC concentrations of the extract which did not kill P. aeruginosa nor inhibited its growth, demonstrated a statistically significant reduction of virulence factors of P. aeruginosa, such as pyocyanin production, twitching and swimming motility. The biofilm forming capability of P. aeruginosa was also reduced in a concentration-dependent manner at sub-MIC values. Water extract of A. blazei is a promising source of antiquorum sensing and antibacterial compounds.

  18. Histopathological comparisons of Staphylococcus aureus and Pseudomonas aeruginosa experimental infected porcine burn wounds.

    PubMed

    Chaney, Sarah B; Ganesh, Kasturi; Mathew-Steiner, Shomita; Stromberg, Paul; Roy, Sashwati; Sen, Chandan K; Wozniak, Daniel J

    2017-05-01

    Chronic skin wounds are a significant human health concern and are often complicated by infection with Pseudomonas aeruginosa and Staphylococcus aureus, particularly methicillin resistant S. aureus (MRSA). Translating the knowledge gained from extensive study of virulence mechanisms and pathogenesis of these bacterial species to new treatment modalities has been lacking in part due to a paucity of animal models able to recapitulate human disease. Our groups recently described a novel porcine chronic burn wound model for the study of bacterial infection; however, the histopathology of infection has yet to be described. The objective of this study is to define the histopathology of this model using important human chronic wound bacterial isolates. Porcine full-thickness burn wounds topically inoculated with P. aeruginosa strain PAO1, MRSA S. aureus strain USA300 or both bacteria were used to define and quantify histopathologic lesions. The development of a systemic, well-defined rubric for analysis allowed for evaluation of differences between infection groups. These differences, which included epithelial migration and proliferation, stromal necrosis, fluid accumulation and intensity and character of the innate and adaptive inflammatory cell responses, were identified temporally between infection groups. Mono-species infected wounds developed a hyper-proliferative wound edge. Coinfected wounds at day 35 had the largest wound sizes, increased amounts of neutrophilic inflammation, immaturity of the wound bed, and retention of necrotic tissue. Infection, regardless of species, inhibited wound contracture at all time points evaluated. Most importantly, this model recapitulated key features of chronic human wounds. Thus, this model will allow researchers to study novel treatment modalities in a biologically relevant animal model while monitoring both host and bacterial responses. © 2017 by the Wound Healing Society.

  19. Hospital costs of nosocomial multi-drug resistant Pseudomonas aeruginosa acquisition.

    PubMed

    Morales, Eva; Cots, Francesc; Sala, Maria; Comas, Mercè; Belvis, Francesc; Riu, Marta; Salvadó, Margarita; Grau, Santiago; Horcajada, Juan P; Montero, Maria Milagro; Castells, Xavier

    2012-05-23

    We aimed to assess the hospital economic costs of nosocomial multi-drug resistant Pseudomonas aeruginosa acquisition. A retrospective study of all hospital admissions between January 1, 2005, and December 31, 2006 was carried out in a 420-bed, urban, tertiary-care teaching hospital in Barcelona (Spain). All patients with a first positive clinical culture for P. aeruginosa more than 48 h after admission were included. Patient and hospitalization characteristics were collected from hospital and microbiology laboratory computerized records. According to antibiotic susceptibility, isolates were classified as non-resistant, resistant and multi-drug resistant. Cost estimation was based on a full-costing cost accounting system and on the criteria of clinical Activity-Based Costing methods. Multivariate analyses were performed using generalized linear models of log-transformed costs. Cost estimations were available for 402 nosocomial incident P. aeruginosa positive cultures. Their distribution by antibiotic susceptibility pattern was 37.1% non-resistant, 29.6% resistant and 33.3% multi-drug resistant. The total mean economic cost per admission of patients with multi-drug resistant P. aeruginosa strains was higher than that for non-resistant strains (15,265 vs. 4,933 Euros). In multivariate analysis, resistant and multi-drug resistant strains were independently predictive of an increased hospital total cost in compared with non-resistant strains (the incremental increase in total hospital cost was more than 1.37-fold and 1.77-fold that for non-resistant strains, respectively). P. aeruginosa multi-drug resistance independently predicted higher hospital costs with a more than 70% increase per admission compared with non-resistant strains. Prevention of the nosocomial emergence and spread of antimicrobial resistant microorganisms is essential to limit the strong economic impact.

  20. Fosfomycin and Tobramycin in Combination Downregulate Nitrate Reductase Genes narG and narH, Resulting in Increased Activity against Pseudomonas aeruginosa under Anaerobic Conditions

    PubMed Central

    McCaughey, Gerard; Gilpin, Deirdre F.; Schneiders, Thamarai; Hoffman, Lucas R.; McKevitt, Matt; Elborn, J. Stuart

    2013-01-01

    The activity of aminoglycosides, which are used to treat Pseudomonas aeruginosa respiratory infection in cystic fibrosis (CF) patients, is reduced under the anaerobic conditions that reflect the CF lung in vivo. In contrast, a 4:1 (wt/wt) combination of fosfomycin and tobramycin (F:T), which is under investigation for use in the treatment of CF lung infection, has increased activity against P. aeruginosa under anaerobic conditions. The aim of this study was to elucidate the mechanisms underlying the increased activity of F:T under anaerobic conditions. Microarray analysis was used to identify the transcriptional basis of increased F:T activity under anaerobic conditions, and key findings were confirmed by microbiological tests, including nitrate utilization assays, growth curves, and susceptibility testing. Notably, growth in subinhibitory concentrations of F:T, but not tobramycin or fosfomycin alone, significantly downregulated (P < 0.05) nitrate reductase genes narG and narH, which are essential for normal anaerobic growth of P. aeruginosa. Under anaerobic conditions, F:T significantly decreased (P < 0.001) nitrate utilization in P. aeruginosa strains PAO1, PA14, and PA14 lasR::Gm, a mutant known to exhibit increased nitrate utilization. A similar effect was observed with two clinical P. aeruginosa isolates. Growth curves indicate that nitrate reductase transposon mutants had reduced growth under anaerobic conditions, with these mutants also having increased susceptibility to F:T compared to the wild type under similar conditions. The results of this study suggest that downregulation of nitrate reductase genes resulting in reduced nitrate utilization is the mechanism underlying the increased activity of F:T under anaerobic conditions. PMID:23959314

  1. Contact lens disinfecting solutions antibacterial efficacy: comparison between clinical isolates and the standard ISO ATCC strains of Pseudomonas aeruginosa and Staphylococcus aureus.

    PubMed

    Mohammadinia, M; Rahmani, S; Eslami, G; Ghassemi-Broumand, M; Aghazadh Amiri, M; Aghaie, Gh; Tabatabaee, S M; Taheri, S; Behgozin, A

    2012-02-01

    To evaluate the disinfectant properties of the three multipurpose contact lens disinfecting solutions available in Iran, against clinical isolates and the standard ISO ATCC strains of Pseudomonas aeruginosa and Staphylococcus aureus, based on the international organization for standardization (ISO) 14729 guidelines. Three multipurpose solutions that were tested were ReNu Multiplus, Solo Care Aqua and All-Clean Soft. The test solutions were challenged with clinical isolates and the standard strains of P. aeruginosa(ATCC 9027) and S. aureus(ATCC 6538), based on the ISO Stand-alone procedure for disinfecting products. Solutions were sampled for surviving microorganisms at manufacturer's minimum recommended disinfection time. The number of viable organisms was determined and log reductions calculated. All of the three test solutions in this study provided a reduction greater than the required mean 3.0 logarithmic reduction against the recommended standard ATCC strains of P. aeruginosa and S. aureus. Antibacterial effectiveness of Solo Care Aqua and All-Clean Soft against clinical isolates of P. aeruginosa and S. aureus were acceptable based on ISO 14729 Stand-alone test. ReNu MultiPlus showed a minimum acceptable efficacy against the clinical isolate of S. aureus, but did not reduce the clinical isolate by the same amount. Although the contact lens disinfecting solutions meet/exceed the ISO 14729 Stand-alone primary acceptance criteria for standard strains of P. aeruginosa and S. aureus, their efficacy may be insufficient against clinical isolates of these organisms.

  2. [Survival elongation of Pseudomonas aeruginosa improves power output of microbial fuel cells].

    PubMed

    You, Ting; Liu, Jihua; Liang, Rubing; Liu, Jianhua

    2017-04-25

    The secondary metabolites, phenazine products, produced by Pseudomonas aeruginosa can mediate the electrons transfer in microbial fuel cells (MFCs). How increase the total electricity production in MFCs by improving the characteristics of Pseudomonas aeruginosa is one of research hot spots and problems. In this study, P. aeruginosa strain SJTD-1 and its knockout mutant strain SJTD-1 (ΔmvaT) were used to construct MFCs, and the discharge processes of the two MFCs were analyzed to determine the key factors to electricity yields. Results indicated that not only phenazine but also the viable cells in the fermentation broth were essential for the discharge of MFCs. The mutant strain SJTD-1 (ΔmvaT) could produce more phenazine products and continue discharging over 160 hours in MFCs, more than that of the wild-type SJTD-1 strain (90 hours discharging time). The total electricity generated by SJTD-1 (ΔmvaT) strain could achieve 2.32 J in the fermentation process, much higher than the total 1.30 J electricity of the wild-type SJTD-1 strain. Further cell growth analysis showed that the mutant strain SJTD-1 (ΔmvaT) could keep a longer stationary period, survive much longer in MFCs and therefore, discharge more electron than those of the wild-type SJTD-1 strain. Therefore, the cell survival elongation of P. aeruginosa in MFCs could enhance its discharging time and improve the overall energy yield. This work could give a clue to improve the characteristics of MFCs using genetic engineering strain, and could promote related application studies on MFCs.

  3. Analysis of the swimming activity of Pseudomonas aeruginosa by using photonic force microscope

    NASA Astrophysics Data System (ADS)

    Chan, Chia-Han; Chang, Bo-Jui; Huang, Ying-Jung; Fan, Chia-Chieh; Peng, Hwei-Ling; Chi, Sien; Hsu, Long

    2005-08-01

    Swimming activity of flagella is a main factor of the motility of bacteria. Flagella expressed on the surface of bacterial species serve as a primary means of motility including swimming. We propose to use optical tweezers to analyze the swimming activity of bacteria. The sample bacteria in the work is Pseudomonas aeruginosa, and it is a gram-negative bacterium and often causes leading to burn wound infections, urinary-tract infections, and pneumonia. The single polar flagellum of P. aeruginosa has been demonstrated to be important virulence and colonization factor of this opportunistic pathogen. We demonstrate a gene to regulate the bacterial swimming activity in P. aeruginosa PAO1 by biological method. However, the change of flagellar morphology was not observed by electron microscopy analysis, suggesting that the gene regulates the flagellar rotation that could not be detected by biological method. PFM exhibits a spatial resolution of a few nanometers to detect the relative position of the probe at an acquisition rate over 1 MHz. By binding a probe such as a bead or a quantum dot on the flagella, we expect the rotation of the probe due to the flagella could be detected. It is expected that the study of the swimming activity of P. aeruginosa provide potent method for the pathogenic role of the flagella in P. aeruginosa.

  4. Pharmacodynamics of levofloxacin in a murine pneumonia model of Pseudomonas aeruginosa infection: determination of epithelial lining fluid targets.

    PubMed

    Louie, Arnold; Fregeau, Christine; Liu, Weiguo; Kulawy, Robert; Drusano, G L

    2009-08-01

    The dose choice for Pseudomonas aeruginosa remains a matter of debate. The actual exposure targets required for multilog killing of organisms at the primary infection site have not been delineated. We studied Pseudomonas aeruginosa PAO1 using a murine model of pneumonia. We employed a large mathematical model to fit all the concentration-time data in plasma and epithelial lining fluid (ELF) as well as colony counts in lung simultaneously for all drug doses. Penetration into ELF was calculated to be approximately 77.7%, as indexed to the ratio of the area under the concentration-time curve for ELF (AUC(ELF)) to the AUC(plasma). We determined the ELF concentration-time profile required to drive a stasis response as well as 1-, 2-, or 3-log(10)(CFU/g) kill. AUC/MIC ratios of 12.4, 31.2, 62.8, and 127.6 were required to drive these bacterial responses. Emergence of resistance was seen only at the two lowest doses (three of five animals at 50 mg/kg [body weight] and one of five animals at 100 mg/kg). The low exposure targets were likely driven by a low mutational frequency to resistance. Bridging to humans was performed using Monte Carlo simulation. With a 750-mg levofloxacin dose, target attainment rates fell below 90% at 4 mg/liter, 1 mg/liter, and 0.5 mg/liter for 1-, 2-, and 3-log kills, respectively. Given the low exposure targets seen with this strain, we conclude that levofloxacin at a 750-mg dose is not adequate for serious Pseudomonas aeruginosa pneumonia as a single agent. More isolates need to be studied to make these observations more robust.

  5. Pharmacodynamics of Levofloxacin in a Murine Pneumonia Model of Pseudomonas aeruginosa Infection: Determination of Epithelial Lining Fluid Targets▿

    PubMed Central

    Louie, Arnold; Fregeau, Christine; Liu, Weiguo; Kulawy, Robert; Drusano, G. L.

    2009-01-01

    The dose choice for Pseudomonas aeruginosa remains a matter of debate. The actual exposure targets required for multilog killing of organisms at the primary infection site have not been delineated. We studied Pseudomonas aeruginosa PAO1 using a murine model of pneumonia. We employed a large mathematical model to fit all the concentration-time data in plasma and epithelial lining fluid (ELF) as well as colony counts in lung simultaneously for all drug doses. Penetration into ELF was calculated to be approximately 77.7%, as indexed to the ratio of the area under the concentration-time curve for ELF (AUCELF) to the AUCplasma. We determined the ELF concentration-time profile required to drive a stasis response as well as 1-, 2-, or 3-log10(CFU/g) kill. AUC/MIC ratios of 12.4, 31.2, 62.8, and 127.6 were required to drive these bacterial responses. Emergence of resistance was seen only at the two lowest doses (three of five animals at 50 mg/kg [body weight] and one of five animals at 100 mg/kg). The low exposure targets were likely driven by a low mutational frequency to resistance. Bridging to humans was performed using Monte Carlo simulation. With a 750-mg levofloxacin dose, target attainment rates fell below 90% at 4 mg/liter, 1 mg/liter, and 0.5 mg/liter for 1-, 2-, and 3-log kills, respectively. Given the low exposure targets seen with this strain, we conclude that levofloxacin at a 750-mg dose is not adequate for serious Pseudomonas aeruginosa pneumonia as a single agent. More isolates need to be studied to make these observations more robust. PMID:19364849

  6. Hospital costs of nosocomial multi-drug resistant Pseudomonas aeruginosa acquisition

    PubMed Central

    2012-01-01

    Background We aimed to assess the hospital economic costs of nosocomial multi-drug resistant Pseudomonas aeruginosa acquisition. Methods A retrospective study of all hospital admissions between January 1, 2005, and December 31, 2006 was carried out in a 420-bed, urban, tertiary-care teaching hospital in Barcelona (Spain). All patients with a first positive clinical culture for P. aeruginosa more than 48 h after admission were included. Patient and hospitalization characteristics were collected from hospital and microbiology laboratory computerized records. According to antibiotic susceptibility, isolates were classified as non-resistant, resistant and multi-drug resistant. Cost estimation was based on a full-costing cost accounting system and on the criteria of clinical Activity-Based Costing methods. Multivariate analyses were performed using generalized linear models of log-transformed costs. Results Cost estimations were available for 402 nosocomial incident P. aeruginosa positive cultures. Their distribution by antibiotic susceptibility pattern was 37.1% non-resistant, 29.6% resistant and 33.3% multi-drug resistant. The total mean economic cost per admission of patients with multi-drug resistant P. aeruginosa strains was higher than that for non-resistant strains (15,265 vs. 4,933 Euros). In multivariate analysis, resistant and multi-drug resistant strains were independently predictive of an increased hospital total cost in compared with non-resistant strains (the incremental increase in total hospital cost was more than 1.37-fold and 1.77-fold that for non-resistant strains, respectively). Conclusions P. aeruginosa multi-drug resistance independently predicted higher hospital costs with a more than 70% increase per admission compared with non-resistant strains. Prevention of the nosocomial emergence and spread of antimicrobial resistant microorganisms is essential to limit the strong economic impact. PMID:22621745

  7. Role of Iron Uptake Systems in Pseudomonas aeruginosa Virulence and Airway Infection

    PubMed Central

    Minandri, Fabrizia; Imperi, Francesco; Frangipani, Emanuela; Bonchi, Carlo; Visaggio, Daniela; Facchini, Marcella; Pasquali, Paolo; Bragonzi, Alessandra

    2016-01-01

    Pseudomonas aeruginosa is a leading cause of hospital-acquired pneumonia and chronic lung infections in cystic fibrosis patients. Iron is essential for bacterial growth, and P. aeruginosa expresses multiple iron uptake systems, whose role in lung infection deserves further investigation. P. aeruginosa Fe3+ uptake systems include the pyoverdine and pyochelin siderophores and two systems for heme uptake, all of which are dependent on the TonB energy transducer. P. aeruginosa also has the FeoB transporter for Fe2+ acquisition. To assess the roles of individual iron uptake systems in P. aeruginosa lung infection, single and double deletion mutants were generated in P. aeruginosa PAO1 and characterized in vitro, using iron-poor media and human serum, and in vivo, using a mouse model of lung infection. The iron uptake-null mutant (tonB1 feoB) and the Fe3+ transport mutant (tonB1) did not grow aerobically under low-iron conditions and were avirulent in the mouse model. Conversely, the wild type and the feoB, hasR phuR (heme uptake), and pchD (pyochelin) mutants grew in vitro and caused 60 to 90% mortality in mice. The pyoverdine mutant (pvdA) and the siderophore-null mutant (pvdA pchD) grew aerobically in iron-poor media but not in human serum, and they caused low mortality in mice (10 to 20%). To differentiate the roles of pyoverdine in iron uptake and virulence regulation, a pvdA fpvR double mutant defective in pyoverdine production but expressing wild-type levels of pyoverdine-regulated virulence factors was generated. Deletion of fpvR in the pvdA background partially restored the lethal phenotype, indicating that pyoverdine contributes to the pathogenesis of P. aeruginosa lung infection by combining iron transport and virulence-inducing capabilities. PMID:27271740

  8. Pseudomonas aeruginosa Regulated Intramembrane Proteolysis (RIP): Protease MucP can Overcome Mutations in the AlgO Periplasmic Protease to Restore Alginate Production in Nonmucoid Revertants.

    PubMed

    Delgado, Camila; Florez, Laura; Lollett, Ivonne; Lopez, Christine; Kangeyan, Shiva; Kumari, Hansi; Stylianou, Marios; Smiddy, Robert J; Schneper, Lisa; Sautter, Robert T; Szatmari, George; Mathee, Kalai

    2018-05-21

    The progression of cystic fibrosis (CF) from an acute to a chronic disease is often associated with the conversion of the opportunistic pathogen Pseudomonas aeruginosa from a nonmucoid form to a mucoid form in the lung. This conversion involves the overproduction of the exopolysaccharide alginate, whose production is under control of the AlgT/U sigma factor. This factor is regulated posttranslationally by an extremely unstable process and has been commonly attributed to mutations in the algT/U gene. By exploiting this unstable phenotype, we isolated 34 spontaneous nonmucoid variants arising from the mucoid strain PDO300, a PAO1 derivative containing the mucA22 allele commonly found in mucoid CF isolates. Complementation analysis using a minimal tiling path cosmid library revealed that most of these mutants mapped to two protease-encoding genes, algO also known as prc or PA3257 , and mucP. Interestingly, our algO mutations were complemented by both mucP and algO , leading us to delete, clone and overexpress mucP , algO , mucE and mucD in both wild-type PAO1 and in PDO300 backgrounds to better understand the regulation of this complex regulatory mechanism. Our findings suggest the regulatory proteases follow two pathways for regulated intramembrane proteolysis (RIP), where both the AlgO/MucP pathway and MucE/AlgW pathway are required in the wild type strain, but where the AlgO/MucP pathway can bypass the MucE/AlgW pathway in mucoid strains with membrane-associated forms of MucA with shortened C-termini, such as the MucA22 variant. This work gives us a better understanding of how alginate production is regulated in the clinically important mucoid variants of Pseudomonas aeruginosa. IMPORTANCE: Infection by the opportunistic pathogen Pseudomonas aeruginosa is the leading cause of morbidity and mortality seen in cystic fibrosis (CF) patients. Poor patient prognosis correlates with the genotypic and phenotypic change of the bacteria from a typical nonmucoid to a mucoid

  9. The Stringent Response Is Essential for Pseudomonas aeruginosa Virulence in the Rat Lung Agar Bead and Drosophila melanogaster Feeding Models of Infection▿†

    PubMed Central

    Vogt, Stefanie L.; Green, Christopher; Stevens, Katarzyna M.; Day, Brad; Erickson, David L.; Woods, Donald E.; Storey, Douglas G.

    2011-01-01

    The stringent response is a regulatory system that allows bacteria to sense and adapt to nutrient-poor environments. The central mediator of the stringent response is the molecule guanosine 3′,5′-bispyrophosphate (ppGpp), which is synthesized by the enzymes RelA and SpoT and which is also degraded by SpoT. Our laboratory previously demonstrated that a relA mutant of Pseudomonas aeruginosa, the principal cause of lung infections in cystic fibrosis patients, was attenuated in virulence in a Drosophila melanogaster feeding model of infection. In this study, we examined the role of spoT in P. aeruginosa virulence. We generated an insertion mutation in spoT within the previously constructed relA mutant, thereby producing a ppGpp-devoid strain. The relA spoT double mutant was unable to establish a chronic infection in D. melanogaster and was also avirulent in the rat lung agar bead model of infection, a model in which the relA mutant is fully virulent. Synthesis of the virulence determinants pyocyanin, elastase, protease, and siderophores was impaired in the relA spoT double mutant. This mutant was also defective in swarming and twitching, but not in swimming motility. The relA spoT mutant and, to a lesser extent, the relA mutant were less able to withstand stresses such as heat shock and oxidative stress than the wild-type strain PAO1, which may partially account for the inability of the relA spoT mutant to successfully colonize the rat lung. Our results indicate that the stringent response, and SpoT in particular, is a crucial regulator of virulence processes in P. aeruginosa. PMID:21788391

  10. Virulence attributes in Brazilian clinical isolates of Pseudomonas aeruginosa.

    PubMed

    Silva, Lívia V; Galdino, Anna Clara M; Nunes, Ana Paula F; dos Santos, Kátia R N; Moreira, Beatriz M; Cacci, Luciana C; Sodré, Cátia L; Ziccardi, Mariangela; Branquinha, Marta H; Santos, André L S

    2014-11-01

    Pseudomonas aeruginosa is an opportunistic human pathogen responsible for causing a huge variety of acute and chronic infections with significant levels of morbidity and mortality. Its success as a pathogen comes from its genetic/metabolic plasticity, intrinsic/acquired antimicrobial resistance, capacity to form biofilm and expression of numerous virulence factors. Herein, we have analyzed the genetic variability, antimicrobial susceptibility as well as the production of metallo-β-lactamases (MBLs) and virulence attributes (elastase, pyocyanin and biofilm) in 96 strains of P. aeruginosa isolated from different anatomical sites of patients attended at Brazilian hospitals. Our results revealed a great genetic variability, in which 86 distinct RAPD types (89.6% of polymorphisms) were detected. Regarding the susceptibility profile, 48 strains (50%) were resistant to the antimicrobials, as follows: 22.92% to the three tested antibiotics, 12.5% to both imipenem and meropenem, 11.46% to ceftazidime only, 2.08% to imipenem only and 1.04% to both ceftazidime and meropenem. Out of the 34 clinical strains of P. aeruginosa resistant to both imipenem and meropenem, 25 (73.53%) were MBL producers by phenotypic method while 12 (35.29%) were PCR positive for the MBL gene SPM-1. All P. aeruginosa strains produced pyocyanin, elastase and biofilm, although in different levels. Some associations were demonstrated among the susceptibility and/or production of these virulence traits with the anatomical site of strain isolation. For instance, almost all strains isolated from urine (85.71%) were resistant to the three antibiotics, while the vast majority of strains isolated from rectum (95%) and mouth (66.67%) were susceptible to all tested antibiotics. Urine isolates produced the highest pyocyanin concentration (20.15±5.65 μg/ml), while strains isolated from pleural secretion and mouth produced elevated elastase activity (1441.43±303.08 FAU) and biofilm formation (OD590 0.676±0

  11. Detecting bacterial lung infections: in vivo evaluation of in vitro volatile fingerprints.

    PubMed

    Zhu, Jiangjiang; Bean, Heather D; Wargo, Matthew J; Leclair, Laurie W; Hill, Jane E

    2013-03-01

    The identification of bacteria by their volatilomes is of interest to many scientists and clinicians as it holds the promise of diagnosing infections in situ, particularly lung infections via breath analysis. While there are many studies reporting various bacterial volatile biomarkers or fingerprints using in vitro experiments, it has proven difficult to translate these data to in vivo breath analyses. Therefore, we aimed to create secondary electrospray ionization-mass spectrometry (SESI-MS) pathogen fingerprints directly from the breath of mice with lung infections. In this study we demonstrated that SESI-MS is capable of differentiating infected versus uninfected mice, P. aeruginosa-infected versus S. aureus-infected mice, as well as distinguish between infections caused by P. aeruginosa strains PAO1 versus FRD1, with statistical significance (p < 0.05). In addition, we compared in vitro and in vivo volatiles and observed that only 25-34% of peaks are shared between the in vitro and in vivo SESI-MS fingerprints. To the best of our knowledge, these are the first breath volatiles measured for P. aeruginosa PAO1, FRD1, and S. aureus RN450, and the first comparison of in vivo and in vitro volatile profiles from the same strains using the murine infection model.

  12. Photodynamic antimicrobial therapy to inhibit pseudomonas aeruginosa of corneal isolates (Conference Presentation)

    NASA Astrophysics Data System (ADS)

    Durkee, Heather A.; Relhan, Nidhi; Arboleda, Alejandro; Halili, Francisco; De Freitas, Carolina; Alawa, Karam; Aguilar, Mariela C.; Amescua, Guillermo; Miller, Darlene; Parel, Jean-Marie

    2016-03-01

    Keratitis associated with Pseudomonas aeruginosa is difficult to manage. Treatment includes antibiotic eye drops, however, some strains of Pseudomonas aeruginosa are resistant. Current research efforts are focused on finding alternative and adjunct therapies to treat multi-drug resistant bacteria. One promising alternate technique is photodynamic therapy (PDT). The purpose of this study was to evaluate the effect of riboflavin- and rose bengal-mediated PDT on Pseudomonas aeruginosa keratitis isolates in vitro. Two isolates (S+U- and S-U+) of Pseudomonas aeruginosa were derived from keratitis patients and exposed to five experimental groups: (1) Control (dark, UV-A irradiation, 525nm irradiation); (2) 0.1% riboflavin (dark, UV-A irradiation); and (3) 0.1% rose bengal, (4) 0.05% rose bengal and (5) 0.01% rose bengal (dark, 525nm irradiation). Three days after treatment, in dark conditions of all concentration of riboflavin and rose bengal showed no inhibition in both S+U- and S-U+ strains of Pseudomonas aeruginosa. In 0.1% and 0.05% rose bengal irradiated groups, for both S+U- and S-U+ strains, there was complete inhibition of bacterial growth in the central 50mm zone corresponding to the diameter of the green light source. These in vitro results suggest that rose bengal photodynamic therapy may be an effective adjunct treatment for Pseudomonas aeruginosa keratitis.

  13. FIM-1, a new acquired metallo-β-lactamase from a Pseudomonas aeruginosa clinical isolate from Italy.

    PubMed

    Pollini, Simona; Maradei, Simona; Pecile, Patrizia; Olivo, Giuseppe; Luzzaro, Francesco; Docquier, Jean-Denis; Rossolini, Gian Maria

    2013-01-01

    Acquired metallo-β-lactamases (MBLs) are resistance determinants of increasing clinical importance in Gram-negative bacterial pathogens, which confer a broad-spectrum β-lactam resistance, including carbapenems. Several such enzymes have been described since the 1990s. In the present study, a novel acquired MBL, named FIM-1, was identified and characterized. The bla(FIM-1) gene was cloned from a multidrug-resistant Pseudomonas aeruginosa clinical isolate (FI-14/157) cultured from a patient with a vascular graft infection in Florence, Italy. The isolate belonged in the sequence type 235 epidemic clonal lineage. The FIM-1 enzyme is a member of subclass B1 and, among acquired MBLs, exhibited the highest similarity (ca. 40% amino acid identity) with NDM-type enzymes. In P. aeruginosa FI-14/157, the bla(FIM-1) gene was apparently inserted into the chromosome and associated with ISCR19-like elements that were likely involved in the capture and mobilization of this MBL gene. Transfer experiments of the bla(FIM-1) gene to an Escherichia coli strain or another P. aeruginosa strain by conjugation or electrotransformation were not successful. The FIM-1 protein was produced in E. coli and purified by two chromatography steps. Analysis of the kinetic parameters, carried out with the purified enzyme, revealed that FIM-1 has a broad substrate specificity, with a preference for penicillins (except the 6α-methoxy derivative temocillin) and carbapenems. Aztreonam was not hydrolyzed. Detection of this novel type of acquired MBL in a P. aeruginosa clinical isolate underscores the increasing diversity of such enzymes that can be encountered in the clinical setting.

  14. Pseudomonas aeruginosa KUCD1, a possible candidate for cadmium bioremediation

    PubMed Central

    Sinha, Sangram; Mukherjee, Samir Kumar

    2009-01-01

    A cadmium (8 mM) resistant Pseudomonas aeruginosa strain KUCd1 exhibiting high Cd accumulation under in vitro aerobic condition has been reported. The isolate showed a significant ability to remove more than 75% and 89% of the soluble cadmium during the active growth phase from the growth medium and from Cd-amended industrial wastewater under growth supportive condition. Transmission electron microscopy (TEM) and energy dispersive X-ray spectroscopy (EDXS) suggest the presence of Cd in the cells from mid stationary phase. The cell fractionation study revealed membrane and periplasm to be the major accumulating site in this strain. The chemical nature of the accumulated Cd was studied by X-ray powder diffraction analysis. PMID:24031411

  15. Evaluation of Synergistic Interactions Between Cell-Free Supernatant of Lactobacillus Strains and Amikacin and Genetamicin Against Pseudomonas aeruginosa

    PubMed Central

    Aminnezhad, Sargol; Kermanshahi, Rouha Kasra; Ranjbar, Reza

    2015-01-01

    Background: The indiscriminate use of antibiotics in the treatment of infectious diseases can increase the development of antibiotic resistance. Therefore, there is a big demand for new sources of antimicrobial agents and alternative treatments for reduction of antibiotic dosage required to decrease the associated side effects. Objectives: In this study, the synergistic action of aminoglycoside antibiotics and cell-free supernatant (CFS) of probiotic (Lactobacillus rahmnosus and L. casei) against Pseudomonas aeruginosa PTCC 1430 was evaluated. Materials and Methods: A growth medium for culturing of probiotic bacteria was separated by centrifugation. The antimicrobial effects of CFS of probiotic bacteria were evaluated using the agar well diffusion assay. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) were evaluated using the micro dilution method. Finally, an interaction between CFS and amikacin or gentamicin against P. aeruginosa PTCC 1430 was examined through the checkerboard method and fractional inhibitory concentration (FIC). Furthermore, CFSs from Lactobacillus strains were analyzed by reversed phase HPLC (RP-HPLC) for antimicrobial compounds. Results: The results showed a significant effect of CFS on the growth of P. aeruginosa. The MIC and MBC of CFS from L. casei were 62.5 µL⁄mL while the MIC and MBC of CFS from L. rhamnosus were 62.5 μL⁄mL and 125 μL⁄mL, respectively. Using the FIC indices, synergistic interactions were observed in combination of CFS and antibiotics. Fractional Inhibitory Concentration indices of CFS from L. casei and aminoglycoside antibiotics were 0.124 and 0.312 while FIC indices of CFS from L. rhamnosus and aminoglycoside antibiotics were 0.124 and 0.56, respectively showing a synergism effect. The results of RP-HPLC showed that CFS of Lactobacillus strains contained acetic acid, lactic acid and hydrogen peroxide (H2O2). Conclusions: Our findings indicate that probiotic bacterial

  16. Evaluation of Synergistic Interactions Between Cell-Free Supernatant of Lactobacillus Strains and Amikacin and Genetamicin Against Pseudomonas aeruginosa.

    PubMed

    Aminnezhad, Sargol; Kermanshahi, Rouha Kasra; Ranjbar, Reza

    2015-04-01

    The indiscriminate use of antibiotics in the treatment of infectious diseases can increase the development of antibiotic resistance. Therefore, there is a big demand for new sources of antimicrobial agents and alternative treatments for reduction of antibiotic dosage required to decrease the associated side effects. In this study, the synergistic action of aminoglycoside antibiotics and cell-free supernatant (CFS) of probiotic (Lactobacillus rahmnosus and L. casei) against Pseudomonas aeruginosa PTCC 1430 was evaluated. A growth medium for culturing of probiotic bacteria was separated by centrifugation. The antimicrobial effects of CFS of probiotic bacteria were evaluated using the agar well diffusion assay. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) were evaluated using the micro dilution method. Finally, an interaction between CFS and amikacin or gentamicin against P. aeruginosa PTCC 1430 was examined through the checkerboard method and fractional inhibitory concentration (FIC). Furthermore, CFSs from Lactobacillus strains were analyzed by reversed phase HPLC (RP-HPLC) for antimicrobial compounds. The results showed a significant effect of CFS on the growth of P. aeruginosa. The MIC and MBC of CFS from L. casei were 62.5 µL⁄mL while the MIC and MBC of CFS from L. rhamnosus were 62.5 μL⁄mL and 125 μL⁄mL, respectively. Using the FIC indices, synergistic interactions were observed in combination of CFS and antibiotics. Fractional Inhibitory Concentration indices of CFS from L. casei and aminoglycoside antibiotics were 0.124 and 0.312 while FIC indices of CFS from L. rhamnosus and aminoglycoside antibiotics were 0.124 and 0.56, respectively showing a synergism effect. The results of RP-HPLC showed that CFS of Lactobacillus strains contained acetic acid, lactic acid and hydrogen peroxide (H2O2). Our findings indicate that probiotic bacterial strains of Lactobacillus have a significant inhibitory effect on the

  17. Biofilm Formation by Otopathogenic Strains of P. aeruginosa is not Consistently Inhibited by EDTA

    PubMed Central

    Zenga, Joseph; Gagnon, Patricia M.; Vogel, Joseph; Chole, Richard A.

    2012-01-01

    Hypothesis Biofilm formation in otopathogenic of P. aeruginosa (OPPA) strains is inhibited by ethylenediaminetetraacetic acid (EDTA). Background EDTA, a widely used chelating agent, has been shown to inhibit biofilm formation in a number of bacteria. Since EDTA may be a well-tolerated reagent to inhibit biofilm formation in cases of suppurative otitis media, we asked if it might be effective in all OPPA strains isolated from chronically infected cholesteatomas. Methods OPPA strains were isolated from patients with infected cholesteatomas. These strains were grown into log phase then were placed in minimal media with varying concentrations of EDTA, and incubated for varying periods. Biofilm production was measured colorimetrically by staining with crystal violet. Results Without added EDTA, most otopathogenic PA exhibited a distinct, but varying, time-course of biofilm formation and dissolution with peak production at 12–18 hours. Addition of 1 mM EDTA resulted in a delay in the time to peak biofilm formation for most strains, although the amount of biofilm was not decreased. In contrast, some strains showed greater biofilm production with 1 mM EDTA compared to the untreated bacteria. Addition of 10 mM EDTA resulted in a similar effect. Some strains increased biofilm production over controls. Moreover, EDTA inhibited planktonic growth of all OPPA strains at the concentrations studied. Conclusion Our hypothesis was disproven: EDTA tends to delay biofilm development while it consistently inhibits planktonic growth. Since EDTA does not cause suppression of biofilm production in all isolates of OPPA, usefulness as an antimicrobial is questioned. PMID:22772018

  18. Molecular epidemiology of SPM-1-producing Pseudomonas aeruginosa by rep-PCR in hospitals in Parana, Brazil.

    PubMed

    Kalluf, K O; Arend, L N; Wuicik, T E; Pilonetto, M; Tuon, F F

    2017-04-01

    Infections caused by multidrug resistant microorganisms are a global health problem, and Pseudomonas aeruginosa is an important nosocomial pathogen, easily disseminated in the hospital environment. The aim of this study was to determine SPM-1 in P. aeruginosa strains in 30 Brazilian hospitals and the genetic similarity of isolates. We analyzed 161 isolates of carbapenem-resistant P. aeruginosa. Imipenem/EDTA and imipenem strip were used for phenotypic detection of MBL production; and real-time polymerase chain reaction (PCR) for genetic detection. Genetic similarity was determined by rep-PCR. We obtained 136/161 (84.5%) isolates with positive phenotypic result for metallo-β-lactamase (MBL) and the bla SPM-1 gene was identified in 41 isolates. There was a predominant profile (>95% of genetic similarity) in 92.7% of isolates. This predominant profile was widely disseminated in Paraná state. SPM-1 is the main MBL identified in carbapenem-resistant P. aeruginosa in Southern Brazil. The genetic similarity among some isolates suggests a clonal expansion. Copyright © 2016 Elsevier B.V. All rights reserved.

  19. Biosynthesis of Pyocyanine by a Paraffin Hydrocarbon-oxidizing Strain of Pseudomonas aeruginosa

    PubMed Central

    Lee, E. G.-H.; Walden, C. C.

    1969-01-01

    A paraffin-oxidizing bacterium, designated as Pseudomonas aeruginosa ATS-14, was isolated from soil samples obtained from the Athabasca “tar sands.” This strain utilized kerosene as the only carbon source of energy and produced a high concentration of pyocyanine in the culture medium. Aromatic carbons were not attacked, but C10 to C17n-alkanes were readily oxidized by the pseudomonad and formed pyocyanine. The highest yield of the pigment was obtained from hexadecane and heptadecane. PMID:4977219

  20. Inhibition of Biofilm Formation by Esomeprazole in Pseudomonas aeruginosa and Staphylococcus aureus

    PubMed Central

    Singh, Vandana; Arora, Vaneet; Alam, M. Jahangir

    2012-01-01

    Staphylococcus aureus and Pseudomonas aeruginosa are common nosocomial pathogens responsible for biofilm-associated infections. Proton pump inhibitors (PPI), such as esomeprazole, may have novel antimicrobial properties. The objective of this study was to assess whether esomeprazole prevents sessile bacterial growth and biofilm formation and whether it may have synergistic killing effects with standard antibiotics. The antibiofilm activity of esomeprazole at 0.25 mM was tested against two strains each of S. aureus and P. aeruginosa. Bacterial biofilms were prepared using a commercially available 96-peg-plate Calgary biofilm device. Sessile bacterial CFU counts and biomass were assessed during 72 hours of esomeprazole exposure. The killing activities after an additional 24 hours of vancomycin (against S. aureus) and meropenem (against P. aeruginosa) treatment with or without preexposure to esomeprazole were also assessed by CFU and biomass analyses. P. aeruginosa and S. aureus strains exposed to esomeprazole displayed decreased sessile bacterial growth and biomass (P < 0.001, each parameter). After 72 h of exposure, there was a 1-log10 decrease in the CFU/ml of esomeprazole-exposed P. aeruginosa and S. aureus strains compared to controls (P < 0.001). After 72 h of exposure, measured absorbance was 100% greater in P. aeruginosa control strains than in esomeprazole-exposed strains (P < 0.001). Increased killing and decreased biomass were observed for esomeprazole-treated bacteria compared to untreated controls exposed to conventional antibiotics (P < 0.001, each parameter). Reduced biofilm growth after 24 h was visibly apparent by light micrographs for P. aeruginosa and S. aureus isolates exposed to esomeprazole compared to untreated controls. In conclusion, esomeprazole demonstrated an antibiofilm effect against biofilm-producing S. aureus and P. aeruginosa. PMID:22664967

  1. Reduction of virulence factor pyocyanin production in multidrug-resistant Pseudomonas aeruginosa.

    PubMed

    Fuse, Katsuhiro; Fujimura, Shigeru; Kikuchi, Toshiaki; Gomi, Kazunori; Iida, Yasuhiro; Nukiwa, Toshihiro; Watanabe, Akira

    2013-02-01

    Nosocomial infections caused by metallo-β-lactamase (MBL)-producing multidrug-resistant (MDR) Pseudomonas aeruginosa have become a worldwide problem. Pyocyanin, a representative pigment produced by P. aeruginosa, is the major virulence factor of this organismThe aim of this study was to investigate the pyocyanin-producing ability of MBL-producing MDR P. aeruginosa. A total of 50 clinical isolates of P. aeruginosa, including 20 MDR strains, were collected at 18 general hospitals in Japan. The chromaticity and luminosity produced by pyocyanin in each isolate were measured. The quantity of pyocyanin and the expression of the phzM and phzS genes coding a pyocyanin synthesis enzyme were measured. MDR strains showed a bright yellow-green, while non-MDR strains tended to show a dark blue-green. The quantities of pyocyanin in MBL-producing strains and non-producing strains were 0.015 ± 0.002 and 0.41 ± 0.10 μg, respectively. The expression of the phzM and phzS genes in the MDR strains was 11 and 14 %, respectively, of the expression in the non-MDR strains. When the MBL gene was transduced into P. aeruginosa and it acquired multidrug resistance, it was shown that the pyocyanin-producing ability decreased. The pathogenicity of MBL-producing MDR P. aeruginosa may be lower than that of non-MDR strains. These MBL-producing MDR strains may be less pathogenic than non-MDR strains. This may explain why MDR-P. aeruginosa is unlikely to cause infection but, rather, causes subclinical colonization only.

  2. [Antibiotics sensitivity and characteristics of the esculin-positive Pseudomonas aeruginosa biovar].

    PubMed

    Sivolodskiĭ, E P

    2000-01-01

    Strains of Pseudomonas aeruginosa hydrolyzing esculin were isolated for the first time. They amount to 17.1 +/- 2.0% (60 from 325) of the investigated P. aeruginosa strains isolated from the clinical material in St. Petersburg. Esculin hydrolysis was measured by micromethod in plates, results were analysed after 3-hours incubation at 37 degrees C. Esculin-positive strains possesed biovar properties: they are widely spread, demonstrated other characteristic features (absence of triethylamine odour, specific colonies lysis), are stable on ability to hydrolyse esculin while culture storage and after repeated culturing. Typical strain of esculinolytica biovar was deposited into the culture collection of the National Research Institute of Agricultural Microbiology as P. aeruginosa ARRIAM 64-A. Susceptibility testing of the esculin-positive strains by disk-diffusion method revealed that most strains were inhibited by imipenem (86.6%), amikacin (75.0%), ceftazidime (65.0%), meropenem (60.0%), aztreonam (51.6%). The percent of strains susceptible to other antibiotics was lower: azlocillin--33.3%, netilmycin--33.3%, piperacillin--26.6%, ceftriaxon--18.3%. Only small number of strains were inhibited by ciprofloxacin (8.3%), gentamycin (3.4%), cefoperazone (1.7%) and carbenicillin (1.7%). The results may be used for empiric therapy before the isolated strain susceptibility is tested but only according to positive esculin-hydrolysis express-test evaluated in 3-hours period.

  3. Quorum quenching properties of Actinobacteria isolated from Malaysian tropical soils.

    PubMed

    Devaraj, Kavimalar; Tan, Geok Yuan Annie; Chan, Kok-Gan

    2017-08-01

    In this study, a total of 147 soil actinobacterial strains were screened for their ability to inhibit response of Chromobacterium violaceum CV026 to short chain N-acyl homoserine lactone (AHL) which is a quorum sensing molecule. Of these, three actinobacterial strains showed positive for violacein inhibition. We further tested these strains for the inhibition of Pseudomonas aeruginosa PAO1 quorum sensing-regulated phenotypes, namely, swarming and pyocyanin production. The three strains were found to inhibit at least one of the quorum sensing-regulated phenotypes of PAO1. Phylogenetic analysis of the 16S rRNA gene sequences indicated that these strains belong to the genera Micromonospora, Rhodococcus and Streptomyces. This is the first report presenting quorum quenching activity by a species of the genus Micromonospora. Our data suggest that Actinobacteria may be a rich source of active compounds that can act against bacterial quorum sensing system.

  4. Identification of extensive drug resistant Pseudomonas aeruginosa strains: New clone ST1725 and high-risk clone ST233

    PubMed Central

    Aguilar-Rodea, Pamela; Zúñiga, Gerardo; Rodríguez-Espino, Benjamín Antonio; Olivares Cervantes, Alma Lidia; Gamiño Arroyo, Ana Estela; Moreno-Espinosa, Sarbelio; de la Rosa Zamboni, Daniela; López Martínez, Briceida; Castellanos-Cruz, María del Carmen; Parra-Ortega, Israel; Jiménez Rojas, Verónica Leticia; Vigueras Galindo, Juan Carlos; Velázquez-Guadarrama, Norma

    2017-01-01

    Several microorganisms produce nosocomial infections (NIs), among which Pseudomonas aeruginosa stands out as an opportunist pathogen with the capacity to develop multiresistance to first-choice antibiotics. From 2007 to 2013, forty-six NIs produced by P. aeruginosa were detected at a pediatric tertiary care hospital in Mexico with a significant mortality rate (17.39%). All isolates (n = 58/46 patients) were characterized by evaluating their response to several antibiotics as panresistant (PDR), extensively resistant (XDR), multiresistant (MDR) or sensitive (S). In addition, all isolates were typified through multilocus sequencing of seven genes: acsA, aroE, guaA, mutL, nuoD, ppsA and trpE. Furthermore, to establish the genetic relationships among these isolates, we carried out a phylogenetic inference analysis using maximum likelihood to construct a phylogenetic network. To assess evolutionary parameters, recombination was evaluated using the PHI test, and the ratio of nonsynonymous to synonymous substitutions was determined. Two of the strains were PDR (ST1725); 42 were XDR; four were MDR; and ten were S. Twenty-one new sequence types were detected. Thirty-three strains exhibited novel sequence type ST1725. The ratio of nonsynonym to synonym substitutions was 1:1 considering all genes. Phylogenetic analysis showed that the genetic relationship of the PDR, XDR and MDR strains was mainly clonal; however, the PHI test and the phylogenetic network suggest that recombination events occurred to produce a non-clonal population. This study aimed not only to determine the genetic diversity of clinical P. aeruginosa but also to provide a warning regarding the identification and spreading of clone ST1725, its ability to cause outbreaks with high mortality rates, and to remain in the hospital environment for over seven years. These characteristics highlight the need to identify clonal outbreaks, especially where high resistance to most antibiotics is observed, and control

  5. Physiological responses of Microcystis aeruginosa against the algicidal bacterium Pseudomonas aeruginosa.

    PubMed

    Zhou, Su; Yin, Hua; Tang, Shaoyu; Peng, Hui; Yin, Donggao; Yang, Yixuan; Liu, Zehua; Dang, Zhi

    2016-05-01

    Proliferation of cyanobacteria in aquatic ecosystems has caused water security problems throughout the world. Our preliminary study has showed that Pseudomonas aeruginosa can inhibit the growth of cyanobacterium, Microcystis aeruginosa. In order to explore the inhibitory mechanism of P. aeruginosa on the cell growth and synthesis of intracellular substances of M. aeruginosa, concentrations of Chlorophyll-a, intracellular protein, carbohydrate, enzyme activities and ion metabolism of M. aeruginosa, were investigated. The results indicated that 83.84% algicidal efficiency of P. aeruginosa was achieved after treatment for 7 days. The strain inhibited the reproduction of M. aeruginosa by impeding the synthesis of intracellular protein and carbohydrate of cyanobacterium, and only a very small part of intracellular protein and carbohydrate was detected after exposure to P. aeruginosa for 5 days. P. aeruginosa caused the alteration of intracellular antioxidant enzyme activity of M. aeruginosa, such as catalase, peroxidase. The accumulation of malondialdehyde aggravated membrane injury after treatment for 3 days. P. aeruginosa also affected the ion metabolism of cyanobacteria. The release of Na(+) and Cl(-) was significantly enhanced while the uptake of K(+), Ca(2+), Mg(2+), NO3(-) and SO4(2)(-) decreased. Surface morphology and intracellular structure of cyanobacteria and bacterial cells changed dramatically over time as evidenced by electron microscope (SEM) and transmission electron microscope (TEM) analysis. These results revealed that the algicidal activity of P. aeruginosa was primarily due to the fermentation liquid of P. aeruginosa that impeded the synthesis of intracellular protein and carbohydrate, and damaged the cell membrane through membrane lipid peroxidation. Copyright © 2016 Elsevier Inc. All rights reserved.

  6. [Antimicrobial susceptibility of Pseudomonas aeruginosa isolated in Fukushima Prefecture].

    PubMed

    Niitsuma, K; Saitoh, M; Kojimabara, M; Kashiwabara, N; Aoki, T; Tomizawa, M; Maeda, J; Kosenda, T

    2001-02-01

    We investigated the susceptibility of Pseudomonas aeruginosa (isolated from the sputum of patients with respiratory infection in 4 medical institutions in Fukushima Prefecture) to 8 beta-lactam antibiotics including three carbapenems and relationships among MICs of antibiotics tested. The MIC90 values for a total of 216 strains were 6.25 micrograms/ml for meropenem, 12.5 micrograms/ml for imipenem and ceftazidime, 25 micrograms/ml for panipenem and cefsulodin, 50 micrograms/ml for cefpirome and over than 200 micrograms/ml for cefoperazone and piperacillin. The frequency of resistance of these strains to each antibiotic was as follows: The resistant strains were 19 (8.8%) for meropenem, 34 (15.7%) for imipenem and ceftazidime, 50 (23.1%) for cefsulodin, 72 (33.3%) for panipenem, 76 (35.2%) for piperacillin and 90 (41.7%) for cefpirome. Eighteen strains (18.3%) of 19 meropenem resitant straisn were resistant to imipenem and panipenem, but 16 strains of the 34 imipenem-resistant strains and 54 strains of the 72 panipenem-resistant strains were susceptible to meropenem. In investigation of isolation of multi-resistant Pseudomonas aeruginosa, the susceptibility of strains tested to 7 antibiotics except cefoperazone was as follows: The strains susceptible to all the 7 antibiotics were 92 strains (42.6%), and 33 strains (15.2%) were resistant to 2 antibiotics, 31 strains (14.4%) were resistant to 1 antibiotic, 21 strains (9.7%) were resistant to 3 antibiotics, 13 strains (6.0%) were resistant to 5 antibiotics, 9 (4.2%) were resistant to 4 and 7 antibiotics, and 8 strains (3.7%) were reistant to 6 antibiotics. Since the emergence of these multi-resistant strains is closely related to frequent use of antibiotics for nosocomial infections, special attention should be paid to the antimicrobial susceptibility of Pseudomonas aeruginosa and the situation of antibiotic resistant strains.

  7. Glycan involvement in the adhesion of Pseudomonas aeruginosa to tears.

    PubMed

    Kautto, Liisa; Nguyen-Khuong, Terry; Everest-Dass, Arun; Leong, Andrea; Zhao, Zhenjun; Willcox, Mark D P; Packer, Nicolle H; Peterson, Robyn

    2016-04-01

    The human eye is constantly bathed by tears, which protect the ocular surface via a variety of mechanisms. The O-linked glycans of tear mucins have long been considered to play a role in binding to pathogens and facilitating their removal in the tear flow. Other conjugated glycans in tears could similarly contribute to pathogen binding and removal but have received less attention. In the work presented here we assessed the contribution of glycan moieties, in particular the protein attached N-glycans, presented by the broad complement of tear proteins to the adhesion of the opportunistic pathogen Pseudomonas aeruginosa, a leading cause of microbial keratitis and ulceration of the cornea. Our adhesion assay involved immobilising the macromolecular components of tears into the wells of a polyvinyl difluoride (PVDF) microtitre filter plate and probing the binding of fluorescently labelled bacteria. Three P. aeruginosa strains were studied: a cytotoxic strain (6206) and an invasive strain (6294) from eye infections, and an invasive strain (320) from a urinary tract infection (UTI). The ocular isolates adhered two to three times more to human tears than to human saliva or porcine gastric mucin, suggesting ocular niche-specific adaptation. Support for the role of the N-glycans carried by human tear proteins in the binding and removal of P. aeruginosa from the eye was shown by: 1) pre-incubation of the bacteria with free component sugars, galactose, mannose, fucose and sialyl lactose (or combination thereof) inhibiting adhesion of all the P. aeruginosa strains to the immobilised tear proteins, with the greatest inhibition of binding of the ocular cytotoxic 6206 and least for the invasive 6294 strain; 2) pre-incubation of the bacteria with N-glycans released from the commercially available human milk lactoferrin, an abundant protein that carries N-linked glycans in tears, inhibiting the adhesion to tears of the ocular bacteria by up to 70%, which was significantly more

  8. A comparative intracellular proteomic profiling of Pseudomonas aeruginosa strain ASP-53 grown on pyrene or glucose as sole source of carbon and identification of some key enzymes of pyrene biodegradation pathway.

    PubMed

    Mukherjee, Ashis K; Bhagowati, Pabitra; Biswa, Bhim Bahadur; Chanda, Abhishek; Kalita, Bhargab

    2017-09-07

    Pseudomonas aeruginosa strain ASP-53, isolated from a petroleum oil-contaminated soil sample, was found to be an efficient degrader of pyrene. PCR amplification of selected hydrocarbon catabolic genes (alkB gene, which encodes for monooxygenase, and the C12O, C23O, and PAH-RHDα genes encoding for the dioxygenase enzyme) from the genomic DNA of P. aeruginosa strain ASP-53 suggested its hydrocarbon degradation potential. The GC-MS analysis demonstrated 30.1% pyrene degradation by P. aeruginosa strain ASP-53 after 144h of incubation at pH6.5, 37°C. Expressions of 115 and 196 intracellular proteins were unambiguously identified and quantitated by shotgun proteomics analysis when the isolate was grown in medium containing pyrene and glucose, respectively. The pyrene-induced uniquely expressed and up-regulated proteins in P. aeruginosa strain ASP-53 in addition to substrate (pyrene) metabolism are also likely to be associated with different cellular functions for example-related to protein folding (molecular chaperone), stress response, metabolism of carbohydrate, proteins and amino acids, and fatty acids; transport of metabolites, energy generation such as ATP synthesis, electron transport and nitrate assimilation, and other oxidation-reduction reactions. Proteomic analyses identified some important enzymes involved in pyrene degradation by P. aeruginosa ASP-53 which shows that this bacterium follows the salicylate pathway of pyrene degradation. This study is the first report on proteomic analysis of pyrene biodegradation pathway by Pseudomonas aeruginosa, isolated from a petroleum-oil contaminated soil sample. The pathway displays partial similarity with deduced pyrene degradation mechanisms of Mycobacterium vanbaalenii PYR-1. The GC-MS analysis as well as PCR amplification of hydrocarbon catabolic genes substantiated the potency of the bacterium under study to effectively degrade high molecular weight, toxic PAH such as pyrene for its filed scale bioremediation

  9. Pathogenic Phenotype and Genotype of Pseudomonas aeruginosa Isolates from Spontaneous Canine Ocular Infections

    PubMed Central

    Ledbetter, Eric C.; Mun, James J.; Kowbel, David; Fleiszig, Suzanne M. J.

    2009-01-01

    Purpose This study was designed to determine whether the ability to adversely affect corneal epithelial cell health is a factor common to Pseudomonas aeruginosa keratitis strains and to assess the prevalence of each pathogenic phenotype and genotype in a canine model of naturally-acquired P. aeruginosa ocular infection. Methods P. aeruginosa ocular isolates were collected by sampling 100 dogs without disease (six isolates collected) and by sampling dogs with conjunctivitis (two isolates), endophthalmitis (one isolate), active keratitis (12 isolates), and resolved P. aeruginosa keratitis (four isolates). Phenotype was determined in vitro by quantifying corneal epithelial cell invasion by gentamicin survival assays, and cytotoxic activity by Trypan blue exclusion assays. Genotyping was performed for genes encoding the type III secreted effectors. Results The ratio of invasive to cytotoxic strains with 95% confidence intervals (CI) was 0.83 (CI, 0.42– 0.99) for conjunctival microflora isolates, 0.80 (CI, 0.54 – 0.94) for ocular infection isolates, and 1.0 (CI, 0.45–1.0) for strains isolated post-resolution of keratitis. Among ocular infection isolates, invasive and cytotoxic strains were significantly (P ≤ 0.02) associated with older and younger dogs, respectively. Visible adverse effects on epithelial cells were significantly (P ≤ 0.03) more frequent for keratitis strains (6/12) than other strains (1/13), but only three of these keratitis strains and the single non-keratitis strain possessed ExoU. Conclusions Invasive strains predominated in the dogs of this study. Only keratitis strains had visible adverse effects on epithelial cells without overt cytotoxicity, suggesting virulence strategies affecting live corneal epithelial cell health are selected for among keratitis strains. PMID:18836164

  10. [Effect of Pseudomonas aeruginosa exometabolites on planktonic and biofilm cultures of Escherichia coli].

    PubMed

    Kuznetsova, M V; Karpunina, T I; Maslennikova, I L; Nesterova, L Iu; Demakov, V A

    2012-01-01

    Study the effect of P. aeruginosa exometabolites on planktonic and biofilm cultures of bioluminescent E. coli strain. E. coli K12 TG1 (pF1 lux+ Ap(r)) recombinant bioluminescent strain, P. aeruginosa ATCC 27853 reference strain and 2 nosocomial isolates were used. Pyocyanin and pyoverdin content in supernatant of P. aeruginosa over-night cultures was evaluated according to E. Deziel et al. (2001). Planktonic and biofilm cultures of E. coli were obtained in 96-well plates (LB, statically, 37 degrees C), optical density of plankton, film biomass (OD600, OD580) and bioluminescence in plankton and biofilm were evaluated in microplate reader Infiniti M200 (Tecan, Austria). P. aeruginosa exometabolites increased the duration of lag-phase in E. coli, and short term exposition inhibited luminescence of planktonic cells. These effects are determined by bactericidal action ofpyocyanin and pyoverdin. Supernatants ofover-night cultures of P. aeruginosa inhibit formation of biofilm and disrupt the formed biofilm of E. coli. Effect of pyocyanin and pyoverdin on these processes is not established, other factors may have higher significance. Bioluminescence of E. coli K12 TGI that reflects the energetic status of the cell allows to evaluate and prognose the character of coexistence of P. aeruginosa in combined with E. coli planktonic and biofilm culture.

  11. Effect of Light Intensity on the Relative Dominance of Toxigenic and Nontoxigenic Strains of Microcystis aeruginosa

    PubMed Central

    LeBlanc Renaud, Susan; Pick, Frances R.; Fortin, Nathalie

    2011-01-01

    In aquatic ecosystems, the factors that regulate the dominance of toxin-producing cyanobacteria over non-toxin-producing strains of the same species are largely unknown. One possible hypothesis is that limiting resources lead to the dominance of the latter because of the metabolic costs associated with toxin production. In this study, we tested the effect of light intensity on the performance of a microcystin-producing strain of Microcystis aeruginosa (UTCC 300) when grown in mixed cultures with non-microcystin-producing strains with similar intrinsic growth rates (UTCC 632 and UTCC 633). The endpoints measured included culture growth rates, microcystin concentrations and composition, and mcyD gene copy numbers determined using quantitative PCR (Q-PCR). In contrast to the predicted results, under conditions of low light intensity (20 μmol·m−2·s−1), the toxigenic strain became dominant in both of the mixed cultures based on gene copy numbers and microcystin concentrations. When grown under conditions of high light intensity (80 μmol·m−2·s−1), the toxigenic strain still appeared to dominate over nontoxigenic strain UTCC 632 but less so over strain UTCC 633. Microcystins may not be so costly to produce that toxigenic cyanobacteria are at a disadvantage in competition for limiting resources. PMID:21841026

  12. Enhanced thermophysical properties via PAO superstructure

    NASA Astrophysics Data System (ADS)

    Pournorouz, Zahra; Mostafavi, Amirhossein; Pinto, Aditya; Bokka, Apparao; Jeon, Junha; Shin, Donghyun

    2017-01-01

    For the last few years, molten salt nanomaterials have attracted many scientists for their enhanced specific heat by doping a minute concentration of nanoparticles (up to 1% by weight). Likewise, enhancing the specific heat of liquid media is important in many aspects of engineering such as engine oil, coolant, and lubricant. However, such enhancement in specific heat was only observed for molten salts, yet other engineering fluids such as water, ethylene glycol, and oil have shown a decrease of specific heat with doped nanoparticles. Recent studies have shown that the observed specific heat enhancement resulted from unique nanostructures that were formed by molten salt molecules when interacting with nanoparticles. Thus, such enhancement in specific heat is only possible for molten salts because other fluids may not naturally form such nanostructures. In this study, we hypothesized such nanostructures can be mimicked through in situ formation of fabricated nano-additives, which are putative nanoparticles coated with useful organic materials (e.g., polar-group-ended organic molecules) leading to superstructures, and thus can be directly used for other engineering fluids. We first applied this approach to polyalphaolefin (PAO). A differential scanning calorimeter (DSC), a rheometer, and a customized setup were employed to characterize the heat capacity, viscosity, and thermal conductivity of PAO and PAO with fabricated nano-additives. Results showed 44.5% enhanced heat capacity and 19.8 and 22.98% enhancement for thermal conductivity and viscosity, respectively, by an addition of only 2% of fabricated nanostructures in comparison with pure PAO. Moreover, a partial melting of the polar-group-ended organic molecules was observed in the first thermal cycle and the peak disappeared in the following cycles. This indicates that the in situ formation of fabricated nano-additives spontaneously occurs in the thermal cycle to form nanostructures. Figure of merit analyses have

  13. Enhanced thermophysical properties via PAO superstructure.

    PubMed

    Pournorouz, Zahra; Mostafavi, Amirhossein; Pinto, Aditya; Bokka, Apparao; Jeon, Junha; Shin, Donghyun

    2017-12-01

    For the last few years, molten salt nanomaterials have attracted many scientists for their enhanced specific heat by doping a minute concentration of nanoparticles (up to 1% by weight). Likewise, enhancing the specific heat of liquid media is important in many aspects of engineering such as engine oil, coolant, and lubricant. However, such enhancement in specific heat was only observed for molten salts, yet other engineering fluids such as water, ethylene glycol, and oil have shown a decrease of specific heat with doped nanoparticles. Recent studies have shown that the observed specific heat enhancement resulted from unique nanostructures that were formed by molten salt molecules when interacting with nanoparticles. Thus, such enhancement in specific heat is only possible for molten salts because other fluids may not naturally form such nanostructures. In this study, we hypothesized such nanostructures can be mimicked through in situ formation of fabricated nano-additives, which are putative nanoparticles coated with useful organic materials (e.g., polar-group-ended organic molecules) leading to superstructures, and thus can be directly used for other engineering fluids. We first applied this approach to polyalphaolefin (PAO). A differential scanning calorimeter (DSC), a rheometer, and a customized setup were employed to characterize the heat capacity, viscosity, and thermal conductivity of PAO and PAO with fabricated nano-additives. Results showed 44.5% enhanced heat capacity and 19.8 and 22.98% enhancement for thermal conductivity and viscosity, respectively, by an addition of only 2% of fabricated nanostructures in comparison with pure PAO. Moreover, a partial melting of the polar-group-ended organic molecules was observed in the first thermal cycle and the peak disappeared in the following cycles. This indicates that the in situ formation of fabricated nano-additives spontaneously occurs in the thermal cycle to form nanostructures. Figure of merit analyses have

  14. Pulmonary Bacteriophage Therapy on Pseudomonas aeruginosa Cystic Fibrosis Strains: First Steps Towards Treatment and Prevention

    PubMed Central

    Morello, Eric; Saussereau, Emilie; Maura, Damien; Huerre, Michel; Touqui, Lhousseine; Debarbieux, Laurent

    2011-01-01

    Multidrug-resistant bacteria are the cause of an increasing number of deadly pulmonary infections. Because there is currently a paucity of novel antibiotics, phage therapy—the use of specific viruses that infect bacteria—is now more frequently being considered as a potential treatment for bacterial infections. Using a mouse lung-infection model caused by a multidrug resistant Pseudomonas aeruginosa mucoid strain isolated from a cystic fibrosis patient, we evaluated bacteriophage treatments. New bacteriophages were isolated from environmental samples and characterized. Bacteria and bacteriophages were applied intranasally to the immunocompetent mice. Survival was monitored and bronchoalveolar fluids were analysed. Quantification of bacteria, bacteriophages, pro-inflammatory and cytotoxicity markers, as well as histology and immunohistochemistry analyses were performed. A curative treatment (one single dose) administrated 2 h after the onset of the infection allowed over 95% survival. A four-day preventive treatment (one single dose) resulted in a 100% survival. All of the parameters measured correlated with the efficacy of both curative and preventive bacteriophage treatments. We also showed that in vitro optimization of a bacteriophage towards a clinical strain improved both its efficacy on in vivo treatments and its host range on a panel of 20 P. aeruginosa cystic fibrosis strains. This work provides an incentive to develop clinical studies on pulmonary bacteriophage therapy to combat multidrug-resistant lung infections. PMID:21347240

  15. Evaluation of five selective media for the detection of Pseudomonas aeruginosa using a strain panel from clinical, environmental and industrial sources.

    PubMed

    Weiser, Rebecca; Donoghue, Denise; Weightman, Andrew; Mahenthiralingam, Eshwar

    2014-04-01

    Isolation and correct identification of the opportunistic pathogen and industrial contaminant Pseudomonas aeruginosa are very important and numerous selective media are available for this purpose. A novel comparison of five selective media having positive (acetamide-based agars), negative (Pseudomonas CN selective agar [Oxoid Ltd.] and Pseudomonas Isolation agar [Sigma-Aldrich Company Ltd.]) and chromogenic (chromID® P. aeruginosa [bioMérieux]) selection strategies was performed using a systematically designed bacterial test panel (58 P. aeruginosa and 90 non-P. aeruginosa strains including those commonly misidentified as P. aeruginosa by culture-dependent techniques). Standardised inocula were added to the selective media and the results were recorded after 24 and 72h. After 72h of incubation at 37°C chromID® P. aeruginosa displayed the highest specificity (70%) and had good sensitivity (95%), although the sensitivity was negatively impacted by the large variation in colour of P. aeruginosa colonies, which hampered interpretation. Both media containing inhibitory selective agents performed very similarly, both having 100% sensitivity and a specificity of approximately 30%. Raising the incubation temperature to 42°C increased the specificity of Pseudomonas CN selective agar and Pseudomonas isolation agar (61% and 47% respectively after 72h), but increased the number of false positives encountered with the chromogenic medium, decreasing its specificity to 68% after 72h. Growth on the acetamide agars was weak for all strains and it was often difficult to determine whether true growth had occurred. This, compounded by the low specificity of the acetamide agars (<26%), suggested they were less suitable for application to clinical or industrial settings without further modification. Overall, the chromogenic agar was the most selective but further consideration is required to optimise interpretation of results. Copyright © 2014 Elsevier B.V. All rights reserved.

  16. [Effect of Pseudomonas aeruginosa melanin on antibiotic activity].

    PubMed

    Rozhavin, M A

    1978-08-01

    The properties of microbial melanines are very diverse. Melanine of P. aeruginosa is little studied. The pigment was isolated from a strain of P. aeruginosa possessing all characteristic properties of the species. Interaction of P. aeruginosa melanine with various antibiotics was determined by the method of serial dilutions in beaf-peptone broth, using Staph. aureus 209 as a test-microbe, which was added to the medium in an amount of 10(6) cells to each tube. It was found that P. aeruginosa melanine differed from DOPA-melanine in a concentration of 1 mg/ml and did not change the activity of penicillin, tetracycline, oleandomycin, kanamycin and gentamicin with respect to Staph. aureus.

  17. In Vivo Activities of Ceftolozane, a New Cephalosporin, with and without Tazobactam against Pseudomonas aeruginosa and Enterobacteriaceae, Including Strains with Extended-Spectrum β-Lactamases, in the Thighs of Neutropenic Mice

    PubMed Central

    Andes, D. R.

    2013-01-01

    Ceftolozane is a new cephalosporin with potent activity against Pseudomonas aeruginosa and Enterobacteriaceae. A neutropenic murine thigh infection model was used to determine which pharmacokinetic/pharmacodynamic index and magnitude drives the efficacy of ceftolozane with Gram-negative bacilli, to compare the rates of in vivo killing of P. aeruginosa by ceftolozane and ceftazidime, and to determine the impact of different ratios of ceftolozane plus tazobactam on Enterobacteriaceae containing extended-spectrum β-lactamases (ESBLs). Neutropenic mice had 106.2-7.1 CFU/thigh when treated with ceftolozane for 24 h with (i) various doses (3.12 to 1,600 mg/kg) and dosage intervals (3, 6, 12, and 24 h) against two Enterobacteriaceae strains, (ii) 0.39 to 800 mg/kg every 6 h for four Enterobacteriaceae and four P. aeruginosa strains, and (iii) 400 or 800 mg/kg with 2:1. 4:1, and 8:1 ratios of tazobactam against five Enterobacteriaceae strains with ESBLs. The pharmacokinetics of ceftolozane at 25, 100, and 400 mg/kg were linear with peak/dose values of 1.0 to 1.4 and half-lives of 12 to 14 min. T>MIC was the primary index driving efficacy. For stasis (1 log kill), T>MIC was 26.3% ± 2.1% (31.6% ± 1.6%) for wild-type Enterobacteriaceae, 31.1% ± 4.9% (34.8% ± 4.4%) for Enterobacteriaceae with ESBLs, and 24.0% ± 3.3% (31.5% ± 3.9%) for P. aeruginosa. At 200 mg/kg every 3 h, the rate of in vivo killing of P. aeruginosa was faster with ceftolozane than with ceftazidime (−0.34 to −0.41 log10 CFU/thigh/h versus −0.21 to −0.24 log10 CFU/thigh/h). The 2:1 ratio of ceftolozane with tazobactam was the most potent combination studied. The T>MIC required for ceftolozane is less than with other cephalosporins and may be due to more rapid killing. PMID:23274659

  18. Real-Time Monitoring of nfxB Mutant Occurrence and Dynamics in Pseudomonas aeruginosa Biofilm Exposed to Subinhibitory Concentrations of Ciprofloxacin

    PubMed Central

    Zaborskyte, Greta; Andersen, Jens Bo; Kragh, Kasper Nørskov

    2016-01-01

    ABSTRACT Biofilm infections caused by Pseudomonas aeruginosa are frequently treated with ciprofloxacin (CIP); however, resistance rapidly develops. One of the primary resistance mechanisms is the overexpression of the MexCD-OprJ pump due to a mutation in nfxB, encoding the transcriptional repressor of this pump. The aim of this study was to investigate the effect of subinhibitory concentrations of CIP on the occurrence of nfxB mutants in the wild-type PAO1 flow cell biofilm model. For this purpose, we constructed fluorescent reporter strains (PAO1 background) with an mCherry tag for constitutive red fluorescence and chromosomal transcriptional fusion between the PmexCD promoter and gfp leading to green fluorescence upon mutation of nfxB. We observed a rapid development of nfxB mutants by live confocal laser scanning microscopy (CLSM) imaging of the flow cell biofilm (reaching 80 to 90% of the whole population) when treated with 1/10 minimal biofilm inhibitory concentration of CIP for 24 h and 96 h. Based on the observed developmental stages, we propose that nfxB mutants emerged de novo in the biofilm during CIP treatment from filamentous cells, which might have arisen due to the stress responses induced by CIP. Identical nfxB mutations were found in fluorescent colonies from the same flow cell biofilm, especially in 24-h biofilms, suggesting selection and clonal expansion of the mutants during biofilm growth. Our findings point at the significant role of high-enough antibiotic dosages or appropriate combination therapy to avoid the emergence of resistant mutants in biofilms. PMID:27993856

  19. From the Environment to the Host: Re-Wiring of the Transcriptome of Pseudomonas aeruginosa from 22°C to 37°C

    PubMed Central

    Bielecki, Piotr; Suárez-Diez, María; Puchałka, Jacek; Albertí, Sebastian; dos Santos, Vitor Martins; Goldberg, Joanna B.

    2014-01-01

    Pseudomonas aeruginosa is a highly versatile opportunistic pathogen capable of colonizing multiple ecological niches. This bacterium is responsible for a wide range of both acute and chronic infections in a variety of hosts. The success of this microorganism relies on its ability to adapt to environmental changes and re-program its regulatory and metabolic networks. The study of P. aeruginosa adaptation to temperature is crucial to understanding the pathogenesis upon infection of its mammalian host. We examined the effects of growth temperature on the transcriptome of the P. aeruginosa PAO1. Microarray analysis of PAO1 grown in Lysogeny broth at mid-exponential phase at 22°C and 37°C revealed that temperature changes are responsible for the differential transcriptional regulation of 6.4% of the genome. Major alterations were observed in bacterial metabolism, replication, and nutrient acquisition. Quorum-sensing and exoproteins secreted by type I, II, and III secretion systems, involved in the adaptation of P. aeruginosa to the mammalian host during infection, were up-regulated at 37°C compared to 22°C. Genes encoding arginine degradation enzymes were highly up-regulated at 22°C, together with the genes involved in the synthesis of pyoverdine. However, genes involved in pyochelin biosynthesis were up-regulated at 37°C. We observed that the changes in expression of P. aeruginosa siderophores correlated to an overall increase in Fe2+ extracellular concentration at 37°C and a peak in Fe3+ extracellular concentration at 22°C. This suggests a distinct change in iron acquisition strategies when the bacterium switches from the external environment to the host. Our work identifies global changes in bacterial metabolism and nutrient acquisition induced by growth at different temperatures. Overall, this study identifies factors that are regulated in genome-wide adaptation processes and discusses how this life-threatening pathogen responds to temperature. PMID:24587139

  20. Pyocine typing as an epidemiological marker in Pseudomonas aeruginosa mastitis in cattle

    PubMed Central

    Ziv, G.; Mushin, Rose; Tagg, J. R.

    1971-01-01

    Pyocine typing was used for the characterization of 134 Pseudomonas aeruginosa strains isolated from bovine mastitis. The scheme of Gillies & Govan (1966) was adopted with some modifications, and the procedure gave 89·6% typability. Pyocine type 1 strains were most commonly encountered and were followed in frequency by types 10 and 3. The introduction of two additional indicator strains allowed for division of these types into subtypes. In spite of some limitations, discussed in the paper, the pyocine typing scheme proved to be useful in `marking' P. aeruginosa strains and in following their association with bovine mastitis in various herds. PMID:4996924

  1. Type VI Secretion System in Pseudomonas aeruginosa

    PubMed Central

    Hachani, Abderrahman; Lossi, Nadine S.; Hamilton, Alexander; Jones, Cerith; Bleves, Sophie; Albesa-Jové, David; Filloux, Alain

    2011-01-01

    Pseudomonas aeruginosa is a Gram-negative bacterium causing chronic infections in cystic fibrosis patients. Such infections are associated with an active type VI secretion system (T6SS), which consists of about 15 conserved components, including the AAA+ ATPase, ClpV. The T6SS secretes two categories of proteins, VgrG and Hcp. Hcp is structurally similar to a phage tail tube component, whereas VgrG proteins show similarity to the puncturing device at the tip of the phage tube. In P. aeruginosa, three T6SSs are known. The expression of H1-T6SS genes is controlled by the RetS sensor. Here, 10 vgrG genes were identified in the PAO1 genome, among which three are co-regulated with H1-T6SS, namely vgrG1a/b/c. Whereas VgrG1a and VgrG1c were secreted in a ClpV1-dependent manner, secretion of VgrG1b was ClpV1-independent. We show that VgrG1a and VgrG1c form multimers, which confirmed the VgrG model predicting trimers similar to the tail spike. We demonstrate that Hcp1 secretion requires either VgrG1a or VgrG1c, which may act independently to puncture the bacterial envelope and give Hcp1 access to the surface. VgrG1b is not required for Hcp1 secretion. Thus, VgrG1b does not require H1-T6SS for secretion nor does H1-T6SS require VgrG1b for its function. Finally, we show that VgrG proteins are required for secretion of a genuine H1-T6SS substrate, Tse3. Our results demonstrate that VgrG proteins are not only secreted components but are essential for secretion of other T6SS substrates. Overall, we emphasize variability in behavior of three P. aeruginosa VgrGs, suggesting that, although very similar, distinct VgrGs achieve specific functions. PMID:21325275

  2. Current therapies for pseudomonas aeruginosa.

    PubMed

    Giamarellou, Helen; Kanellakopoulou, Kyriaki

    2008-04-01

    Based on the worldwide prevalence of multidrug-resistant strains of Pseudomas aeruginosa and the fact that no newer antipseudomonal agents are available, this article aims to investigate therapeutic solutions for combating infections caused by P aeruginosa, including multidrug-resistant strains. The article focuses mainly on colistin, the re-emerging old antibiotic that possesses prominent antipseudomonal activity in vitro and on doripenem, a newer carbapenem that seems to be close to its global marketing. Regarding older antipseudomonal antibiotics that have been reviewed extensively, only newer aspects on their use are considered in this article.

  3. The PAPI-1 pathogenicity island-encoded small RNA PesA influences Pseudomonas aeruginosa virulence and modulates pyocin S3 production

    PubMed Central

    Ferrara, Silvia; Falcone, Marilena; Macchi, Raffaella; Bragonzi, Alessandra; Girelli, Daniela; Cariani, Lisa; Cigana, Cristina

    2017-01-01

    Small non-coding RNAs (sRNAs) are post-transcriptional regulators of gene expression that have been recognized as key contributors to bacterial virulence and pathogenic mechanisms. In this study, we characterized the sRNA PesA of the opportunistic human pathogen Pseudomonas aeruginosa. We show that PesA, which is transcribed within the pathogenicity island PAPI-1 of P. aeruginosa strain PA14, contributes to P. aeruginosa PA14 virulence. In fact, pesA gene deletion resulted in a less pathogenic strain, showing higher survival of cystic fibrosis human bronchial epithelial cells after infection. Moreover, we show that PesA influences positively the expression of pyocin S3 whose genetic locus comprises two structural genes, pyoS3A and pyoS3I, encoding the killing S3A and the immunity S3I proteins, respectively. Interestingly, the deletion of pesA gene results in increased sensitivity to UV irradiation and to the fluoroquinolone antibiotic ciprofloxacin. The degree of UV sensitivity displayed by the PA14 strain lacking PesA is comparable to that of a strain deleted for pyoS3A-I. These results suggest an involvement of pyocin S3 in DNA damage repair and a regulatory role of PesA on this function. PMID:28665976

  4. General and inducible hypermutation facilitate parallel adaptation in Pseudomonas aeruginosa despite divergent mutation spectra.

    PubMed

    Weigand, Michael R; Sundin, George W

    2012-08-21

    The successful growth of hypermutator strains of bacteria contradicts a clear preference for lower mutation rates observed in the microbial world. Whether by general DNA repair deficiency or the inducible action of low-fidelity DNA polymerases, the evolutionary strategies of bacteria include methods of hypermutation. Although both raise mutation rate, general and inducible hypermutation operate through distinct molecular mechanisms and therefore likely impart unique adaptive consequences. Here we compare the influence of general and inducible hypermutation on adaptation in the model organism Pseudomonas aeruginosa PAO1 through experimental evolution. We observed divergent spectra of single base substitutions derived from general and inducible hypermutation by sequencing rpoB in spontaneous rifampicin-resistant (Rif(R)) mutants. Likewise, the pattern of mutation in a draft genome sequence of a derived inducible hypermutator isolate differed from those of general hypermutators reported in the literature. However, following experimental evolution, populations of both mutator types exhibited comparable improvements in fitness across varied conditions that differed from the highly specific adaptation of nonmutators. Our results suggest that despite their unique mutation spectra, general and inducible hypermutation can analogously influence the ecology and adaptation of bacteria, significantly shaping pathogenic populations where hypermutation has been most widely observed.

  5. Duckweed (Lemna minor) as a model plant system for the study of human microbial pathogenesis.

    PubMed

    Zhang, Yong; Hu, Yangbo; Yang, Baoyu; Ma, Fang; Lu, Pei; Li, Lamei; Wan, Chengsong; Rayner, Simon; Chen, Shiyun

    2010-10-25

    Plant infection models provide certain advantages over animal models in the study of pathogenesis. However, current plant models face some limitations, e.g., plant and pathogen cannot co-culture in a contained environment. Development of such a plant model is needed to better illustrate host-pathogen interactions. We describe a novel model plant system for the study of human pathogenic bacterial infection on a large scale. This system was initiated by co-cultivation of axenic duckweed (Lemna minor) plants with pathogenic bacteria in 24-well polystyrene cell culture plate. Pathogenesis of bacteria to duckweed was demonstrated with Pseudomonas aeruginosa and Staphylococcus aureus as two model pathogens. P. aeruginosa PAO1 caused severe detriment to duckweed as judged from inhibition to frond multiplication and chlorophyll formation. Using a GFP-marked PAO1 strain, we demonstrated that bacteria colonized on both fronds and roots and formed biofilms. Virulence of PAO1 to duckweed was attenuated in its quorum sensing (QS) mutants and in recombinant strains overexpressing the QS quenching enzymes. RN4220, a virulent strain of S. aureus, caused severe toxicity to duckweed while an avirulent strain showed little effect. Using this system for antimicrobial chemical selection, green tea polyphenols exhibited inhibitory activity against S. aureus virulence. This system was further confirmed to be effective as a pathogenesis model using a number of pathogenic bacterial species. Our results demonstrate that duckweed can be used as a fast, inexpensive and reproducible model plant system for the study of host-pathogen interactions, could serve as an alternative choice for the study of some virulence factors, and could also potentially be used in large-scale screening for the discovery of antimicrobial chemicals.

  6. Pseudomonas aeruginosa Reduces VX-809 Stimulated F508del-CFTR Chloride Secretion by Airway Epithelial Cells

    PubMed Central

    Stanton, Bruce A.; Coutermarsh, Bonita; Barnaby, Roxanna; Hogan, Deborah

    2015-01-01

    Background P. aeruginosa is an opportunistic pathogen that chronically infects the lungs of 85% of adult patients with Cystic Fibrosis (CF). Previously, we demonstrated that P. aeruginosa reduced wt-CFTR Cl secretion by airway epithelial cells. Recently, a new investigational drug VX-809 has been shown to increase F508del-CFTR Cl secretion in human bronchial epithelial (HBE) cells, and, in combination with VX-770, to increase FEV1 (forced expiratory volume in 1 second) by an average of 3-5% in CF patients homozygous for the F508del-CFTR mutation. We propose that P. aeruginosa infection of CF lungs reduces VX-809 + VX-770- stimulated F508del-CFTR Cl secretion, and thereby reduces the clinical efficacy of VX-809 + VX-770. Methods and Results F508del-CFBE cells and primary cultures of CF-HBE cells (F508del/F508del) were exposed to VX-809 alone or a combination of VX-809 + VX-770 for 48 hours and the effect of P. aeruginosa on F508del-CFTR Cl secretion was measured in Ussing chambers. The effect of VX-809 on F508del-CFTR abundance was measured by cell surface biotinylation and western blot analysis. PAO1, PA14, PAK and 6 clinical isolates of P. aeruginosa (3 mucoid and 3 non-mucoid) significantly reduced drug stimulated F508del-CFTR Cl secretion, and plasma membrane F508del-CFTR. Conclusion The observation that P. aeruginosa reduces VX-809 and VX-809 + VX-770 stimulated F508del CFTR Cl secretion may explain, in part, why VX-809 + VX-770 has modest efficacy in clinical trials. PMID:26018799

  7. Pseudomonas aeruginosa Reduces VX-809 Stimulated F508del-CFTR Chloride Secretion by Airway Epithelial Cells.

    PubMed

    Stanton, Bruce A; Coutermarsh, Bonita; Barnaby, Roxanna; Hogan, Deborah

    2015-01-01

    P. aeruginosa is an opportunistic pathogen that chronically infects the lungs of 85% of adult patients with Cystic Fibrosis (CF). Previously, we demonstrated that P. aeruginosa reduced wt-CFTR Cl secretion by airway epithelial cells. Recently, a new investigational drug VX-809 has been shown to increase F508del-CFTR Cl secretion in human bronchial epithelial (HBE) cells, and, in combination with VX-770, to increase FEV1 (forced expiratory volume in 1 second) by an average of 3-5% in CF patients homozygous for the F508del-CFTR mutation. We propose that P. aeruginosa infection of CF lungs reduces VX-809 + VX-770- stimulated F508del-CFTR Cl secretion, and thereby reduces the clinical efficacy of VX-809 + VX-770. F508del-CFBE cells and primary cultures of CF-HBE cells (F508del/F508del) were exposed to VX-809 alone or a combination of VX-809 + VX-770 for 48 hours and the effect of P. aeruginosa on F508del-CFTR Cl secretion was measured in Ussing chambers. The effect of VX-809 on F508del-CFTR abundance was measured by cell surface biotinylation and western blot analysis. PAO1, PA14, PAK and 6 clinical isolates of P. aeruginosa (3 mucoid and 3 non-mucoid) significantly reduced drug stimulated F508del-CFTR Cl secretion, and plasma membrane F508del-CFTR. The observation that P. aeruginosa reduces VX-809 and VX-809 + VX-770 stimulated F508del CFTR Cl secretion may explain, in part, why VX-809 + VX-770 has modest efficacy in clinical trials.

  8. UCST-Type Thermoresponsive Polymers in Synthetic Lubricating Oil Polyalphaolefin (PAO)

    DOE PAGES

    Fu, Wenxin; Bai, Wei; Jiang, Sisi; ...

    2018-02-20

    Here, this article reports a family of UCST-type thermoresponsive polymers, poly(alkyl methacrylate)s with an appropriate alkyl pendant length, in an industrially important non-volatile organic liquid polyalphaolefin (PAO). The cloud point (CP) can be readily tuned over a wide temperature range by changing the alkyl pendant length; at a concentration of 1 wt% and similar polymer molecular weights, the CP varies linearly with the (average) number of carbon atoms in the alkyl pendant. PAO solutions of ABA triblock copolymers, composed of a PAO-philic middle block and thermoresponsive outer blocks with appropriate block lengths, undergo thermoreversible sol-gel transitions at sufficiently high concentrations.more » The discovery of thermoresponsive polymers in PAO makes it possible to explore new applications by utilizing PAO’s unique characteristics such as thermal stability, non-volatility, superior lubrication properties, etc. Lastly, two examples are presented: thermoresponsive physical gels for control of optical transmittance and injectable gel lubricants.« less

  9. UCST-Type Thermoresponsive Polymers in Synthetic Lubricating Oil Polyalphaolefin (PAO)

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Fu, Wenxin; Bai, Wei; Jiang, Sisi

    Here, this article reports a family of UCST-type thermoresponsive polymers, poly(alkyl methacrylate)s with an appropriate alkyl pendant length, in an industrially important non-volatile organic liquid polyalphaolefin (PAO). The cloud point (CP) can be readily tuned over a wide temperature range by changing the alkyl pendant length; at a concentration of 1 wt% and similar polymer molecular weights, the CP varies linearly with the (average) number of carbon atoms in the alkyl pendant. PAO solutions of ABA triblock copolymers, composed of a PAO-philic middle block and thermoresponsive outer blocks with appropriate block lengths, undergo thermoreversible sol-gel transitions at sufficiently high concentrations.more » The discovery of thermoresponsive polymers in PAO makes it possible to explore new applications by utilizing PAO’s unique characteristics such as thermal stability, non-volatility, superior lubrication properties, etc. Lastly, two examples are presented: thermoresponsive physical gels for control of optical transmittance and injectable gel lubricants.« less

  10. Chemical Inhibition of Kynureninase Reduces Pseudomonas aeruginosa Quorum Sensing and Virulence Factor Expression.

    PubMed

    Kasper, Stephen H; Bonocora, Richard P; Wade, Joseph T; Musah, Rabi Ann; Cady, Nathaniel C

    2016-04-15

    The opportunistic pathogen Pseudomonas aeruginosa utilizes multiple quorum sensing (QS) pathways to coordinate an arsenal of virulence factors. We previously identified several cysteine-based compounds inspired by natural products from the plant Petiveria alliacea which are capable of antagonizing multiple QS circuits as well as reducing P. aeruginosa biofilm formation. To understand the global effects of such compounds on virulence factor production and elucidate their mechanism of action, RNA-seq transcriptomic analysis was performed on P. aeruginosa PAO1 exposed to S-phenyl-l-cysteine sulfoxide, the most potent inhibitor from the prior study. Exposure to this inhibitor down-regulated expression of several QS-regulated virulence operons (e.g., phenazine biosynthesis, type VI secretion systems). Interestingly, many genes that were differentially regulated pertain to the related metabolic pathways that yield precursors of pyochelin, tricarboxylic acid cycle intermediates, phenazines, and Pseudomonas quinolone signal (PQS). Activation of the MexT-regulon was also indicated, including the multidrug efflux pump encoded by mexEF-oprN, which has previously been shown to inhibit QS and pathogenicity. Deeper investigation of the metabolites involved in these systems revealed that S-phenyl-l-cysteine sulfoxide has structural similarity to kynurenine, a precursor of anthranilate, which is critical for P. aeruginosa virulence. By supplementing exogenous anthranilate, the QS-inhibitory effect was reversed. Finally, it was shown that S-phenyl-l-cysteine sulfoxide competitively inhibits P. aeruginosa kynureninase (KynU) activity in vitro and reduces PQS production in vivo. The kynurenine pathway has been implicated in P. aeruginosa QS and virulence factor expression; however, this is the first study to show that targeted inhibition of KynU affects P. aeruginosa gene expression and QS, suggesting a potential antivirulence strategy.

  11. Use of bacteriophage to prevent Pseudomonas aeruginosa contamination and fouling in Jet A aviation fuel.

    PubMed

    Bojanowski, Caitlin L; Crookes-Goodson, Wendy J; Robinson, Jayne B

    2016-11-01

    In the present study, the use of bacteriophages to prevent growth and/or biofouling by Pseudomonas aeruginosa PAO1 was investigated in microcosms containing Jet A aviation fuel as the carbon source. Bacteriophages were found to be effective at preventing biofilm formation but did not always prevent planktonic growth in the microcosms. This result was at odds with experiments conducted in nutrient-rich medium, demonstrating the necessity to test antimicrobial and antifouling strategies under conditions as near as possible to the 'real world'. The success of the bacteriophages at preventing biofilm formation makes them potential candidates as antifouling agents for fuel systems.

  12. Detection of restriction fragment length polymorphisms in clinical isolates and serially passaged Pseudomonas aeruginosa strains.

    PubMed Central

    Hjelm, L N; Branstrom, A A; Warren, R L

    1990-01-01

    An 800-base-pair HindIII-PstI fragment that flanks a hot spot for Tn7 insertion was isolated from the chromosome of Pseudomonas aeruginosa and cloned into pUC12. The fragment was used to probe XhoI digests of genomic DNA from 18 P. aeruginosa isolates collected from sputum samples of seven cystic fibrosis patients. Only two XhoI restriction fragment length polymorphisms (RFLPs), of 3.7 and 7.7 kilobases (kb), were detected. Isolate WSU3531-1 (3.7-kb XhoI fragment) and WSU3860 (7.7-kb XhoI fragment), while isolated from the same patient, showed different RFLPs. Serial passages of isolate WSU3531-1 demonstrated that this strain was phenotypically stable. In contrast, colony and pigment variants were readily isolated at a frequency of 1% from serial passages of isolate WSU3860. When XhoI-digested genomic DNA from phenotypic variants of serially passaged WSU3860 were probed with the 800-base-pair HindIII-PstI fragment, the probe hybridized to a 10.4-kb XhoI fragment from three isolates. Restriction analysis of the genomic DNA digested with a variety of restriction enzymes showed that a 2.7-kb insertion occurred in the same region for all three isolates. There appeared to be no correlation between changes in the RFLP and changes in colony morphology. Images PMID:1977762

  13. Pseudomonas Genome Database: facilitating user-friendly, comprehensive comparisons of microbial genomes.

    PubMed

    Winsor, Geoffrey L; Van Rossum, Thea; Lo, Raymond; Khaira, Bhavjinder; Whiteside, Matthew D; Hancock, Robert E W; Brinkman, Fiona S L

    2009-01-01

    Pseudomonas aeruginosa is a well-studied opportunistic pathogen that is particularly known for its intrinsic antimicrobial resistance, diverse metabolic capacity, and its ability to cause life threatening infections in cystic fibrosis patients. The Pseudomonas Genome Database (http://www.pseudomonas.com) was originally developed as a resource for peer-reviewed, continually updated annotation for the Pseudomonas aeruginosa PAO1 reference strain genome. In order to facilitate cross-strain and cross-species genome comparisons with other Pseudomonas species of importance, we have now expanded the database capabilities to include all Pseudomonas species, and have developed or incorporated methods to facilitate high quality comparative genomics. The database contains robust assessment of orthologs, a novel ortholog clustering method, and incorporates five views of the data at the sequence and annotation levels (Gbrowse, Mauve and custom views) to facilitate genome comparisons. A choice of simple and more flexible user-friendly Boolean search features allows researchers to search and compare annotations or sequences within or between genomes. Other features include more accurate protein subcellular localization predictions and a user-friendly, Boolean searchable log file of updates for the reference strain PAO1. This database aims to continue to provide a high quality, annotated genome resource for the research community and is available under an open source license.

  14. Biofilm inhibition formation of clinical strains of Pseudomonas aeruginosa mutans, photocatalytic activity of azo dye and GC-MS analysis of leaves of Lagerstroemia speciosa.

    PubMed

    Sai Saraswathi, V; Kamarudheen, Neethu; Bhaskara Rao, K V; Santhakumar, K

    2017-04-01

    The investigation was conducted to analyse the bioactive compounds from the leaf extracts of L. speciosa by GC-MS. The extracts were screened for antibacterial and antibiofilm activities against potential clinical strains. The bioactive compounds from the leaves of L. speciosa were extracted by soxhlet continuous extraction method and their chemical composition was analysed by Gas Chromatography-Mass Spectroscopy (GC-MS). The antibacterial activity was evaluated against clinical strain like Staphylococcus aureus, Escherichia coli, P. aeruginosa and Salmonella typhi by well diffusion technique. We also screened for antibacterial property against common food borne pathogens namely Listeria monocytogenes and Bacillus cereus at varied concentration 250μml -1 to 1000μml -1 . Thereafter antibiofilm assay was carried out at from 250 to 1000μg/ml against P. aeruginosa (high biofilm forming pathogen) clinical strain by cover slip technique and the morphology of the pathogen was observed using Scanning Electron Microscopy-(SEM). It was observed that diverse class of secondary metabolites were found by GC-MS analysis for all the extracts upon the continuous extraction. It was found that only minimum inhibition was seen in alcoholic extract for antibacterial activity, whereas all other extracts showed negligible activity. P. aeruginosa biofilm inhibited to 93.0±2% and 91±2% at higher concentration (1000μg/ml) for methanolic and ethanolic extract respectively. Absence of extracellular matrix structure and the surface cracking of biofilm were viewed by SEM, which confirmed the antibiofilm activity. Hence this study reveals that L. speciosa showed significant antibiofilm activity against P. aeruginosa due to the phytoconstituents present in the leaf extracts which was well documented in the alcoholic extracts by GC-MS analysis. The methanolic and ethanolic extract showed good photocatalytic activity of 77.44% and 96.66% against azo dye degradation respectively. Further

  15. Purification and antibiofilm activity of AHL-lactonase from endophytic Enterobacter aerogenes VT66.

    PubMed

    Rajesh, P S; Rai, V Ravishankar

    2015-11-01

    The opportunistic pathogen Pseudomonas aeruginosa uses biofilm lifestyle to resist antibiotic treatment. In our study, endophytic bacterium Enterobacter aerogenes VT66 quenched the N-acyl homoserine lactone (AHL) molecules produced by P. aeruginosa PAO1. The quorum quenching activity was attributed to the presence of AHL-lactonase. The AHL-lactonase was purified using column chromatography and purified AHL-lactonase was applied for the control of biofilm formation in P. aeruginosa PAO1. The results showed that purified AHL-lactonase obtained with a molecular weight about 30kDa was able to inhibit more than 70% of biofilm in P. aeruginosa PAO1 (P<0.001). Antibiofilm activity of AHL-lactonase was correlated well with results from staining technique used to determine inhibition of biomass and viable cell activity. Therefore, results unambiguously confirm that the AHL-lactonase from E. aerogenes VT66 could be used as antibiofilm therapeutics in P. aeruginosa associated biomedical applications. Copyright © 2015 Elsevier B.V. All rights reserved.

  16. Rearrangement of a large novel Pseudomonas aeruginosa gene island in strains isolated from a patient developing ventilator-associated pneumonia.

    PubMed

    Singh, G; Srinivasan, R; Cheng, J; Peng, Z; Fujimura, K; Baek, M S; Panzer, A R; Tringe, S G; Chen, F; Sorek, R; Weng, L; Bristow, J; Wiener-Kronish, J P; Lynch, S V

    2014-07-01

    Bacterial gene islands add to the genetic repertoire of opportunistic pathogens. Here, we perform comparative analyses of three Pseudomonas aeruginosa strains isolated sequentially over a 3-week period from a patient with ventilator-associated pneumonia (VAP) who received clindamycin and piperacillin-tazobactam as part of their treatment regime. While all three strains appeared to be clonal by standard pulsed-field gel electrophoresis, whole-genome sequencing revealed subtle alterations in the chromosomal organization of the last two strains; specifically, an inversion event within a novel 124-kb gene island (PAGI 12) composed of 137 open reading frames [ORFs]. Predicted ORFs in the island included metabolism and virulence genes. Overexpression of a gene island-borne putative β-lactamase gene was observed following piperacillin-tazobactam exposure and only in those strains that had undergone the inversion event, indicating altered gene regulation following genomic remodeling. Examination of a separate cohort of 76 patients with VAP for integration at this tRNA(lys) recombination site demonstrated that patients exhibiting evidence of integration at this site had significantly higher 28-day mortality. These findings provide evidence that P. aeruginosa can integrate, rapidly remodel, and express exogenous genes, which likely contributes to its fitness in a clinical setting. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

  17. Rearrangement of a Large Novel Pseudomonas aeruginosa Gene Island in Strains Isolated from a Patient Developing Ventilator-Associated Pneumonia

    PubMed Central

    Singh, G.; Srinivasan, R.; Cheng, J.; Peng, Z.; Fujimura, K.; Baek, M. S.; Panzer, A. R.; Tringe, S. G.; Chen, F.; Sorek, R.; Weng, L.; Bristow, J.; Wiener-Kronish, J. P.

    2014-01-01

    Bacterial gene islands add to the genetic repertoire of opportunistic pathogens. Here, we perform comparative analyses of three Pseudomonas aeruginosa strains isolated sequentially over a 3-week period from a patient with ventilator-associated pneumonia (VAP) who received clindamycin and piperacillin-tazobactam as part of their treatment regime. While all three strains appeared to be clonal by standard pulsed-field gel electrophoresis, whole-genome sequencing revealed subtle alterations in the chromosomal organization of the last two strains; specifically, an inversion event within a novel 124-kb gene island (PAGI 12) composed of 137 open reading frames [ORFs]. Predicted ORFs in the island included metabolism and virulence genes. Overexpression of a gene island-borne putative β-lactamase gene was observed following piperacillin-tazobactam exposure and only in those strains that had undergone the inversion event, indicating altered gene regulation following genomic remodeling. Examination of a separate cohort of 76 patients with VAP for integration at this tRNAlys recombination site demonstrated that patients exhibiting evidence of integration at this site had significantly higher 28-day mortality. These findings provide evidence that P. aeruginosa can integrate, rapidly remodel, and express exogenous genes, which likely contributes to its fitness in a clinical setting. PMID:24789195

  18. Chronic infection sustained by a Pseudomonas aeruginosa High-Risk clone producing the VIM-1 metallo-β-lactamase in a cystic fibrosis patient after lung transplantation.

    PubMed

    Pollini, Simona; Mugnaioli, Claudia; Dolce, Daniela; Campana, Silvia; Neri, Anna Silvia; Taccetti, Giovanni; Rossolini, Gian Maria

    2018-02-12

    The significance of chronic lung infection by multidrug-resistant (MDR) pathogens in Cystic Fibrosis (CF) transplanted patients remains controversial, and the available information is overall limited. Here we describe the case of a chronic infection, sustained by a metallo-β-lactamase (MBL)-producing P. aeruginosa strain, in a CF patient following lung transplantation. Twelve P. aeruginosa isolates collected from a CF patient over a 15-years follow-up period after lung transplantation were analysed for their antibiotic susceptibility profile, MBL production and clonal relatedness. Available clinical and microbiological records were reviewed. The transplanted CF patient was chronically infected by an MBL-producing P. aeruginosa strain which harboured a bla VIM-1 determinant inserted into a novel class 1 integron. The strain exhibited an MDR phenotype and belonged to the globally widespread ST235 epidemic clonal lineage, which however is not a typical CF-associated epidemic clone. Despite the chronic infection, the long-term outcome of this patient during the post-transplant period was characterized by the absence of acute exacerbations and by a mostly stable pulmonary function. This report provides one of the few descriptions of MBL-producing P. aeruginosa infections in CF patients, and the first description of such an infection after lung transplantation in these patients. Infection with the MBL-producing strain apparently did not significantly affect the patient pulmonary function. Copyright © 2018 European Cystic Fibrosis Society. Published by Elsevier B.V. All rights reserved.

  19. Proteome Profiles of Outer Membrane Vesicles and Extracellular Matrix of Pseudomonas aeruginosa Biofilms.

    PubMed

    Couto, Narciso; Schooling, Sarah R; Dutcher, John R; Barber, Jill

    2015-10-02

    In the present work, two different proteomic platforms, gel-based and gel-free, were used to map the matrix and outer membrane vesicle exoproteomes of Pseudomonas aeruginosa PAO1 biofilms. These two proteomic strategies allowed us a confident identification of 207 and 327 proteins from enriched outer membrane vesicles and whole matrix isolated from biofilms. Because of the physicochemical characteristics of these subproteomes, the two strategies showed complementarity, and thus, the most comprehensive analysis of P. aeruginosa exoproteome to date was achieved. Under our conditions, outer membrane vesicles contribute approximately 20% of the whole matrix proteome, demonstrating that membrane vesicles are an important component of the matrix. The proteomic profiles were analyzed in terms of their biological context, namely, a biofilm. Accordingly relevant metabolic processes involved in cellular adaptation to the biofilm lifestyle as well as those related to P. aeruginosa virulence capabilities were a key feature of the analyses. The diversity of the matrix proteome corroborates the idea of high heterogeneity within the biofilm; cells can display different levels of metabolism and can adapt to local microenvironments making this proteomic analysis challenging. In addition to analyzing our own primary data, we extend the analysis to published data by other groups in order to deepen our understanding of the complexity inherent within biofilm populations.

  20. Biofilm and metallo beta-lactamase production among the strains of Pseudomonas aeruginosa and Acinetobacter spp. at a Tertiary Care Hospital in Kathmandu, Nepal.

    PubMed

    Baniya, Bandana; Pant, Narayan Dutt; Neupane, Sanjeev; Khatiwada, Saroj; Yadav, Uday Narayan; Bhandari, Nisha; Khadka, Rama; Bhatta, Sabita; Chaudhary, Raina

    2017-11-02

    Pseudomonas aeruginosa and Acinetobacter spp. are found to be associated with biofilm and metallo-β-lactamase production and are the common causes of serious infections mainly in hospitalized patients. So, the main aims of this study were to determine the rates of biofilm production and metallo beta-lactamase production (MBL) among the strains of Pseudomonas aeruginosa and Acinetobacter spp. isolated from hospitalized patients. A total of 85 P. aeruginosa isolates and 50 Acinetobacter spp. isolates isolated from different clinical specimens from patients admitted to Shree Birendra Hospital, Kathmandu, Nepal from July 2013 to May 2014 were included in this study. The bacterial isolates were identified with the help of biochemical tests. Modified Kirby-Bauer disc diffusion technique was used for antimicrobial susceptibility testing. Combined disc diffusion technique was used for the detection of MBL production, while Congo red agar method and tube adherence method were used for detection of biofilm production. Around 16.4% of P. aeruginosa isolates and 22% of the strains of Acinetobacter spp. were metallo β-lactamase producers. Out of 85 P. aeruginosa isolates, 23 (27.05%) were biofilm producers according to tube adherence test while, only 13 (15.29%) were biofilm producers as per Congo red agar method. Similarly, out of 50 Acinetobacter spp. 7 (14%) isolates were biofilm producers on the basis of tube adherence test, while only 5 (10%) were positive for biofilm production by Congo red agar method. Highest rates of susceptibility of P. aeruginosa as well as Acinetobacter spp. were seen toward colistin. In our study, biofilm production and metallo beta-lactamase production were observed among Pseudomonas aeruginosa and Acinetobacter spp. However, no statistically significant association could be established between biofilm production and metallo beta-lactamase production.

  1. Detection of P. aeruginosa harboring bla CTX-M-2, bla GES-1 and bla GES-5, bla IMP-1 and bla SPM-1 causing infections in Brazilian tertiary-care hospital

    PubMed Central

    2012-01-01

    Background Nosocomial infections caused by Pseudomonas aeruginosa presenting resistance to beta-lactam drugs are one of the most challenging targets for antimicrobial therapy, leading to substantial increase in mortality rates in hospitals worldwide. In this context, P. aeruginosa harboring acquired mechanisms of resistance, such as production of metallo-beta-lactamase (MBLs) and extended-spectrum beta-lactamases (ESBLs) have the highest clinical impact. Hence, this study was designed to investigate the presence of genes codifying for MBLs and ESBLs among carbapenem resistant P. aeruginosa isolated in a Brazilian 720-bed teaching tertiary care hospital. Methods Fifty-six carbapenem-resistant P. aeruginosa strains were evaluated for the presence of MBL and ESBL genes. Strains presenting MBL and/or ESBL genes were submitted to pulsed-field gel electrophoresis for genetic similarity evaluation. Results Despite the carbapenem resistance, genes for MBLs (blaSPM-1 or blaIMP-1) were detected in only 26.7% of isolates. Genes encoding ESBLs were detected in 23.2% of isolates. The blaCTX-M-2 was the most prevalent ESBL gene (19.6%), followed by blaGES-1 and blaGES-5 detected in one isolate each. In all isolates presenting MBL phenotype by double-disc synergy test (DDST), the blaSPM-1 or blaIMP-1 genes were detected. In addition, blaIMP-1 was also detected in three isolates which did not display any MBL phenotype. These isolates also presented the blaCTX-M-2 gene. The co-existence of blaCTX-M-2 with blaIMP-1 is presently reported for the first time, as like as co-existence of blaGES-1 with blaIMP-1. Conclusions In this study MBLs production was not the major mechanism of resistance to carbapenems, suggesting the occurrence of multidrug efflux pumps, reduction in porin channels and production of other beta-lactamases. The detection of blaCTX-M-2,blaGES-1 and blaGES-5 reflects the recent emergence of ESBLs among antimicrobial resistant P. aeruginosa and the extraordinary

  2. Degradation of paracetamol by Pseudomonas aeruginosa strain HJ1012.

    PubMed

    Hu, Jun; Zhang, Li L; Chen, Jian M; Liu, Yu

    2013-01-01

    Pseudomonas aeruginosa strain HJ1012 was isolated on paracetamol as a sole carbon and energy source. This organism could completely degrade paracetamol as high as 2200 mg/L. Following paracetamol consumption, a CO₂ yield rate up to 71.4% proved that the loss of paracetamol was mainly via mineralization. Haldane's equation adequately described the relationship between the specific growth rate and substrate concentration. The maximum specific growth rate and yield coefficient were 0.201 g-Paracetamol/g-VSS·h and 0.101 mg of biomass yield/mg of paracetamol consumed, respectively. A total of 8 metabolic intermediates was identified and classified into aromatic compounds, carboxylic acids, and inorganic species (nitrite and nitrate ions). P-aminophenol and hydroquinone are the two key metabolites of the initial steps in the paracetamol catabolic pathway. Paracetamol is degraded predominantly via p-aminophenol to hydroquinone with subsequent ring fission, suggesting partially new pathways for paracetamol-degrading bacteria.

  3. Genetic and Functional Diversity of Pseudomonas aeruginosa Lipopolysaccharide

    PubMed Central

    Lam, Joseph S.; Taylor, Véronique L.; Islam, Salim T.; Hao, Youai; Kocíncová, Dana

    2011-01-01

    Lipopolysccharide (LPS) is an integral component of the Pseudomonas aeruginosa cell envelope, occupying the outer leaflet of the outer membrane in this Gram-negative opportunistic pathogen. It is important for bacterium–host interactions and has been shown to be a major virulence factor for this organism. Structurally, P. aeruginosa LPS is composed of three domains, namely, lipid A, core oligosaccharide, and the distal O antigen (O-Ag). Most P. aeruginosa strains produce two distinct forms of O-Ag, one a homopolymer of D-rhamnose that is a common polysaccharide antigen (CPA, formerly termed A band), and the other a heteropolymer of three to five distinct (and often unique dideoxy) sugars in its repeat units, known as O-specific antigen (OSA, formerly termed B band). Compositional differences in the O units among the OSA from different strains form the basis of the International Antigenic Typing Scheme for classification via serotyping of different strains of P. aeruginosa. The focus of this review is to provide state-of-the-art knowledge on the genetic and resultant functional diversity of LPS produced by P. aeruginosa. The underlying factors contributing to this diversity will be thoroughly discussed and presented in the context of its contributions to host–pathogen interactions and the control/prevention of infection. PMID:21687428

  4. The genome and structural proteome of YuA, a new Pseudomonas aeruginosa phage resembling M6.

    PubMed

    Ceyssens, Pieter-Jan; Mesyanzhinov, Vadim; Sykilinda, Nina; Briers, Yves; Roucourt, Bart; Lavigne, Rob; Robben, Johan; Domashin, Artem; Miroshnikov, Konstantin; Volckaert, Guido; Hertveldt, Kirsten

    2008-02-01

    Pseudomonas aeruginosa phage YuA (Siphoviridae) was isolated from a pond near Moscow, Russia. It has an elongated head, encapsulating a circularly permuted genome of 58,663 bp, and a flexible, noncontractile tail, which is terminally and subterminally decorated with short fibers. The YuA genome is neither Mu- nor lambda-like and encodes 78 gene products that cluster in three major regions involved in (i) DNA metabolism and replication, (ii) host interaction, and (iii) phage particle formation and host lysis. At the protein level, YuA displays significant homology with phages M6, phiJL001, 73, B3, DMS3, and D3112. Eighteen YuA proteins were identified as part of the phage particle by mass spectrometry analysis. Five different bacterial promoters were experimentally identified using a promoter trap assay, three of which have a sigma54-specific binding site and regulate transcription in the genome region involved in phage particle formation and host lysis. The dependency of these promoters on the host sigma54 factor was confirmed by analysis of an rpoN mutant strain of P. aeruginosa PAO1. At the DNA level, YuA is 91% identical to the recently (July 2007) annotated phage M6 of the Lindberg typing set. Despite this level of DNA homology throughout the genome, both phages combined have 15 unique genes that do not occur in the other phage. The genome organization of both phages differs substantially from those of the other known Pseudomonas-infecting Siphoviridae, delineating them as a distinct genus within this family.

  5. The Pseudomonas aeruginosa Periplasmic Protease CtpA Can Affect Systems That Impact Its Ability To Mount Both Acute and Chronic Infections

    PubMed Central

    Seo, Jin

    2013-01-01

    Proteases play important roles in the virulence of Pseudomonas aeruginosa. Some are exported to act on host targets and facilitate tissue destruction and bacterial dissemination. Others work within the bacterial cell to process virulence factors and regulate virulence gene expression. Relatively little is known about the role of one class of bacterial serine proteases known as the carboxyl-terminal processing proteases (CTPs). The P. aeruginosa genome encodes two CTPs annotated as PA3257/Prc and PA5134/CtpA in strain PAO1. Prc degrades mutant forms of the anti-sigma factor MucA to promote mucoidy in some cystic fibrosis lung isolates. However, nothing is known about the role or importance of CtpA. We have now found that endogenous CtpA is a soluble periplasmic protein and that a ctpA null mutant has specific phenotypes consistent with an altered cell envelope. Although a ctpA null mutation has no major effect on bacterial growth in the laboratory, CtpA is essential for the normal function of the type 3 secretion system (T3SS), for cytotoxicity toward host cells, and for virulence in a mouse model of acute pneumonia. Conversely, increasing the amount of CtpA above its endogenous level induces an uncharacterized extracytoplasmic function sigma factor regulon, an event that has been reported to attenuate P. aeruginosa in a rat model of chronic lung infection. Therefore, a normal level of CtpA activity is critical for T3SS function and acute virulence, whereas too much activity can trigger an apparent stress response that is detrimental to chronic virulence. PMID:24082078

  6. Pseudomonas aeruginosa biofilm-associated homoserine lactone C12 rapidly activates apoptosis in airway epithelia

    PubMed Central

    Schwarzer, Christian; Fu, Zhu; Patanwala, Maria; Hum, Lauren; Lopez-Guzman, Mirielle; Illek, Beate; Kong, Weidong; Lynch, Susan V.; Machen, Terry E.

    2014-01-01

    Pseudomonas aeruginosa (PA) forms biofilms in lungs of cystic fibrosis CF) patients, a process regulated by quorum sensing molecules including N-(3-oxododecanoyl)-L-homoserine lactone, C12. C12 (10–100 μM) rapidly triggered events commonly associated with the intrinsic apoptotic pathway in JME (CFΔF508CFTR, nasal surface) epithelial cells: depolarization of mitochondrial (mito) membrane potential (Δψmito) and release of cytochrome C (cytoC) from mitos into cytosol and activation of caspases 3/7, 8 and 9. C12 also had novel effects on the endoplasmic reticulum (release of both Ca2+ and ER-targeted GFP and oxidized contents into the cytosol). Effects began within 5 minutes and were complete in 1–2 hrs. C12 caused similar activation of caspases and release of cytoC from mitos in Calu-3 (wtCFTR, bronchial gland) cells, showing that C12-triggered responses occurred similarly in different airway epithelial types. C12 had nearly identical effects on three key aspects of the apoptosis response (caspase 3/7, depolarization of Δψmito and reduction of redox potential in the ER) in JME and CFTR-corrected JME cells (adenoviral expression), showing that CFTR was likely not an important regulator of C12-triggered apoptosis in airway epithelia. Exposure of airway cultures to biofilms from PAO1wt caused depolarization of Δψmito and increases in Cacyto like 10–50 μM C12. In contrast, biofilms from PAO1ΔlasI (C12 deficient) had no effect, suggesting that C12 from P. aeruginosa biofilms may contribute to accumulation of apoptotic cells that cannot be cleared from CF lungs. A model to explain the effects of C12 is proposed. PMID:22233488

  7. Geographical differences in first acquisition of Pseudomonas aeruginosa in cystic fibrosis.

    PubMed

    Ranganathan, Sarath C; Skoric, Billy; Ramsay, Kay A; Carzino, Rosemary; Gibson, Anne-Marie; Hart, Emily; Harrison, Jo; Bell, Scott C; Kidd, Timothy J

    2013-04-01

    Risk of infection with Pseudomonas aeruginosa in cystic fibrosis (CF) may be associated with environmental factors. To determine whether residential location is associated with risk of first acquisition of P. aeruginosa. We performed bronchoalveolar lavage and upper airway cultures in children newly diagnosed with CF to identify infection with P. aeruginosa during infancy and early childhood. Children were assessed according to their residence in a regional or metropolitan area. Multilocus sequence typing was used to determine P. aeruginosa genotype. An environmental questionnaire was also administered. A total of 105 of 120 (87.5%) infants diagnosed with CF were included in this study. Diagnosis in 65 infants (61.9%) followed newborn screening at mean age of 4.6 weeks. Sixty subjects (57.1%) were homozygous ΔF508, and 47 (44.8%) were female. Fifty-five (52.3%) infants were regional, of whom 26 (47.3%), compared with 9 of 50 (18.0%) metropolitan children, acquired infection with P. aeruginosa (odds ratio, 4.084; 95% confidence interval, 1.55-11.30). Age at acquisition was similar (regional: median, 2.31 yr; range, 0.27-5.96 yr; metropolitan: median, 3.10 yr, range, 0.89-3.70 yr). Strain typing identified P. aeruginosa genotypes often encountered in different ecological settings and little evidence of cross-infection. Ninety questionnaires (85.7%) were completed. Those who acquired P. aeruginosa were more likely to be living in a household that used water sprinkler systems (P = 0.032), but no differences were identified to explain increased risk of acquisition of P. aeruginosa in regional children. Geographical difference in residence of children with CF was associated with increased risk of first acquisition of P. aeruginosa, usually with strains associated with the environment rather than with cross-infection.

  8. Pao Pereira Extract Suppresses Castration-Resistant Prostate Cancer Cell Growth, Survival, and Invasion Through Inhibition of NFκB Signaling.

    PubMed

    Chang, Cunjie; Zhao, Wei; Xie, Bingxian; Deng, Yongming; Han, Tao; Cui, Yangyan; Dai, Yundong; Zhang, Zhen; Gao, Jimin; Guo, Hongqian; Yan, Jun

    2014-05-01

    Pao extract, derived from bark of Amazonian tree Pao Pereira, is commonly used in South American medicine. A recent study showed that Pao extract repressed androgen-dependent LNCaP prostate cancer cell growth. We hypothesize that Pao extract asserts its anticancer effects on metastatic castration-resistant prostate cancer (CRPC) cells. Pao extract suppressed CRPC PC3 cell growth in a dose- and time-dependent manner, through induction of apoptosis and cell cycle arrest. Pao extract treatment induced cell cycle inhibitors, p21 and p27, and repressed PCNA, Cyclin A and Cyclin D1. Furthermore, Pao extract also induced the upregulation of pro-apoptotic Bax, reduction of anti-apoptotic Bcl-2, Bcl-xL, and XIAP expression, which were associated with the cleavage of PARP protein. Moreover, Pao extract treatment blocked PC3 cell migration and invasion. Mechanistically, Pao extract suppressed phosphorylation levels of AKT and NFκB/p65, NFκB DNA binding activity, and luciferase reporter activity. Pao inhibited TNFα-induced relocation of NFκB/p65 to the nucleus, NFκB/p65 transcription activity, and MMP9 activity as shown by zymography. Consistently, NFκB/p65 downstream targets involved in proliferation (Cyclin D1), survival (Bcl-2, Bcl-xL, and XIAP), and metastasis (VEGFa, MMP9, and GROα/CXCL1) were also downregulated by Pao extract. Finally, forced expression of NFκB/p65 reversed the growth inhibitory effect of Pao extract. Overall, Pao extract induced cell growth arrest, apoptosis, partially through inhibiting NFκB activation in prostate cancer cells. These data suggest that Pao extract may be beneficial for protection against CRPC. © The Author(s) 2013.

  9. Inhibition of Biofilm Formation, Quorum Sensing and Infection in Pseudomonas aeruginosa by Natural Products-Inspired Organosulfur Compounds

    PubMed Central

    Cady, Nathaniel C.; McKean, Kurt A.; Behnke, Jason; Kubec, Roman; Mosier, Aaron P.; Kasper, Stephen H.; Burz, David S.; Musah, Rabi A.

    2012-01-01

    Using a microplate-based screening assay, the effects on Pseudomonas aeruginosa PAO1 biofilm formation of several S-substituted cysteine sulfoxides and their corresponding disulfide derivatives were evaluated. From our library of compounds, S-phenyl-L-cysteine sulfoxide and its breakdown product, diphenyl disulfide, significantly reduced the amount of biofilm formation by P. aeruginosa at levels equivalent to the active concentration of 4-nitropyridine-N-oxide (NPO) (1 mM). Unlike NPO, which is an established inhibitor of bacterial biofilms, our active compounds did not reduce planktonic cell growth and only affected biofilm formation. When used in a Drosophila-based infection model, both S-phenyl-L-cysteine sulfoxide and diphenyl disulfide significantly reduced the P. aeruginosa recovered 18 h post infection (relative to the control), and were non-lethal to the fly hosts. The possibility that the observed biofilm inhibitory effects were related to quorum sensing inhibition (QSI) was investigated using Escherichia coli-based reporters expressing P. aeruginosa lasR or rhIR response proteins, as well as an endogenous P. aeruginosa reporter from the lasI/lasR QS system. Inhibition of quorum sensing by S-phenyl-L-cysteine sulfoxide was observed in all of the reporter systems tested, whereas diphenyl disulfide did not exhibit QSI in either of the E. coli reporters, and showed very limited inhibition in the P. aeruginosa reporter. Since both compounds inhibit biofilm formation but do not show similar QSI activity, it is concluded that they may be functioning by different pathways. The hypothesis that biofilm inhibition by the two active compounds discovered in this work occurs through QSI is discussed. PMID:22715388

  10. Duckweed (Lemna minor) as a Model Plant System for the Study of Human Microbial Pathogenesis

    PubMed Central

    Zhang, Yong; Hu, Yangbo; Yang, Baoyu; Ma, Fang; Lu, Pei; Li, Lamei; Wan, Chengsong; Rayner, Simon; Chen, Shiyun

    2010-01-01

    Background Plant infection models provide certain advantages over animal models in the study of pathogenesis. However, current plant models face some limitations, e.g., plant and pathogen cannot co-culture in a contained environment. Development of such a plant model is needed to better illustrate host-pathogen interactions. Methodology/Principal Findings We describe a novel model plant system for the study of human pathogenic bacterial infection on a large scale. This system was initiated by co-cultivation of axenic duckweed (Lemna minor) plants with pathogenic bacteria in 24-well polystyrene cell culture plate. Pathogenesis of bacteria to duckweed was demonstrated with Pseudomonas aeruginosa and Staphylococcus aureus as two model pathogens. P. aeruginosa PAO1 caused severe detriment to duckweed as judged from inhibition to frond multiplication and chlorophyll formation. Using a GFP-marked PAO1 strain, we demonstrated that bacteria colonized on both fronds and roots and formed biofilms. Virulence of PAO1 to duckweed was attenuated in its quorum sensing (QS) mutants and in recombinant strains overexpressing the QS quenching enzymes. RN4220, a virulent strain of S. aureus, caused severe toxicity to duckweed while an avirulent strain showed little effect. Using this system for antimicrobial chemical selection, green tea polyphenols exhibited inhibitory activity against S. aureus virulence. This system was further confirmed to be effective as a pathogenesis model using a number of pathogenic bacterial species. Conclusions/Significance Our results demonstrate that duckweed can be used as a fast, inexpensive and reproducible model plant system for the study of host-pathogen interactions, could serve as an alternative choice for the study of some virulence factors, and could also potentially be used in large-scale screening for the discovery of antimicrobial chemicals. PMID:21049039

  11. Evaluation of anti-quorum-sensing activity of edible plants and fruits through inhibition of the N-acyl-homoserine lactone system in Chromobacterium violaceum and Pseudomonas aeruginosa.

    PubMed

    Musthafa, K Syed; Ravi, A Veera; Annapoorani, A; Packiavathy, I Sybiya Vasantha; Pandian, S Karutha

    2010-01-01

    To find out an alternative strategy to antibiotic usage against bacterial infection. The purpose of this study is to describe the quorum-sensing (QS) inhibitory activity of edible plants and fruits against N-acyl-homoserine lactone (AHL)-mediated violacein production in Chromobacterium violaceum and virulence factor expression in Pseudomonas aeruginosa PAO1. Aqueous extracts of Ananas comosus (Bromeliaceae), Musa paradiciaca (Musaceae), Manilkara zapota (Sapotaceae) and Ocimum sanctum (Lamiaceae) were prepared and anti-QS activity of each extract was tested against AHL-mediated phenotypic expressions of C. violaceum and PAO1. Most of these extracts showed significant reduction in AHL-mediated violacein production in C. violaceum as well as pyocyanin pigment, staphylolytic protease, elastase production and biofilm formation in PAO1. However, these extracts were not inhibitory to bacterial growth, revealing that the QS inhibition by the extracts is not related to static or killing effects on the bacteria. The present study identified the anti-QS activity of A. comosus, M. paradiciaca, M. zapota and O. sanctum. An AHL-inactivating compound from these plant sources can be used as an alternative to antibiotic compounds to prevent AHL-mediated bacterial infection in higher organisms. Copyright © 2010 S. Karger AG, Basel.

  12. An outbreak of hospital-acquired Pseudomonas aeruginosa infection caused by contaminated bottled water in intensive care units.

    PubMed

    Eckmanns, T; Oppert, M; Martin, M; Amorosa, R; Zuschneid, I; Frei, U; Rüden, H; Weist, K

    2008-05-01

    This study describes an outbreak of Pseudomonas aeruginosa infections caused by contaminated bottled still water (BSW) in six intensive care units (ICUs) of a German university hospital. Clinical and environmental samples from these units were cultured and genotyped by amplified fragment-length polymorphism and pulsed-field gel electrophoresis analysis. Microbiological results were reviewed on a weekly basis to determine the number of P. aeruginosa infections and colonisations of ICU patients. Clinical specimens from 19 ICU patients--15 infections and four colonisations--yielded the same strain of P. aeruginosa. Furthermore, four of 103 environmental samples also yielded P. aeruginosa. However, only a P. aeruginosa strain isolated from unopened BSW was genetically identical to the P. aeruginosa strain isolated from the patients. In the 42-week period before the outbreak, the mean weekly number of new ICU patients infected or colonised with P. aeruginosa was 46.9 (95% CI 40.7-53.1)/1000 bed-days. During the 6-week period of the outbreak, the weekly number of new patients with P. aeruginosa was 88.9 (95% CI 54.3-122.2)/1000 bed-days. This number returned to the previous level after removal of the BSW. Thus, the microbiological and epidemiological findings revealed that the outbreak was related to BSW contaminated with P. aeruginosa. It was concluded that all untested BSW should be removed from ICUs.

  13. Antagonistic interactions peak at intermediate genetic distance in clinical and laboratory strains of Pseudomonas aeruginosa.

    PubMed

    Schoustra, Sijmen E; Dench, Jonathan; Dali, Rola; Aaron, Shawn D; Kassen, Rees

    2012-03-22

    Bacteria excrete costly toxins to defend their ecological niche. The evolution of such antagonistic interactions between individuals is expected to depend on both the social environment and the strength of resource competition. Antagonism is expected to be weak among highly similar genotypes because most individuals are immune to antagonistic agents and among dissimilar genotypes because these are unlikely to be competing for the same resources and antagonism should not yield much benefit. The strength of antagonism is therefore expected to peak at intermediate genetic distance. We studied the ability of laboratory strains of Pseudomonas aeruginosa to prevent growth of 55 different clinical P. aeruginosa isolates derived from cystic fibrosis patients. Genetic distance was determined using genetic fingerprints. We found that the strength of antagonism was maximal among genotypes of intermediate genetic distance and we show that genetic distance and resource use are linked. Our results suggest that the importance of social interactions like antagonism may be modulated by the strength of resource competition.

  14. Sensitivity of Scenedesmus obliquus and Microcystis aeruginosa to atrazine: effects of acclimation and mixed cultures, and their removal ability.

    PubMed

    Chalifour, Annie; LeBlanc, André; Sleno, Lekha; Juneau, Philippe

    2016-12-01

    Atrazine is an herbicide frequently detected in watercourses that can affect the phytoplankton community, thus impacting the whole food chain. This study aims, firstly, to measure the sensitivity of monocultures of the green alga Scenedemus obliquus and toxic and non-toxic strains of the cyanobacteria Microcystis aeruginosa before, during and after a 30-day acclimation period to 0.1 µM of atrazine. Secondly, the sensitivity of S. obliquus and M. aeruginosa to atrazine in mixed cultures was evaluated. Finally, the ability of these strains to remove atrazine from the media was measured. We demonstrated that both strains of M. aeruginosa had higher growth rate-based EC 50 values than S. obliquus when exposed to atrazine, even though their photosynthesis-based EC 50 values were lower. After being exposed to 0.1 µM of atrazine for 1 month, only the photosynthesis-based EC 50 of S. obliquus increased significantly. In mixed cultures, the growth rate of the non-toxic strain of M. aeruginosa was higher than S. obliquus at high concentrations of atrazine, resulting in a ratio of M. aeruginosa to total cell count of 0.6. This lower sensitivity might be related to the higher growth rate of cyanobacteria at low light intensity. Finally, a negligible fraction of atrazine was removed from the culture media by S. obliquus or M. aeruginosa over 6 days. These results bring new insights on the acclimation of some phytoplankton species to atrazine and its effect on the competition between S. obliquus and M. aeruginosa in mixed cultures.

  15. Pseudomonas aeruginosa inactivation mechanism is affected by capsular extracellular polymeric substances reactivity with chlorine and monochloramine.

    PubMed

    Xue, Zheng; Hessler, Christopher M; Panmanee, Warunya; Hassett, Daniel J; Seo, Youngwoo

    2013-01-01

    The reactivity of capsular extracellular polymeric substances (EPS) to chlorine and monochloramine was assessed and compared in this study. The impact of capsular EPS on Gram-negative bacteria Pseudomonas aeruginosa inactivation mechanisms was investigated both qualitatively and quantitatively using a combination of batch experiments, viability tests with LIVE/DEAD staining, and Fourier transform infrared spectroscopy (FTIR). Both wild-type and isogenic mutant strains with different alginate EPS production capabilities were used to evaluate their susceptibility to chlorine and monochloramine. The mucA22 mutant strain, which overproduces the EPS composed largely of acidic polysaccharide alginate, exhibited high resistance and prolonged inactivation time to both chlorine and monochloramine relative to PAO1 (wild-type) and algT(U) mutant strains (alginate EPS deficient). Multiple analyses were combined to better understand the mechanistic role of EPS against chlorine-based disinfectants. The extracted EPS exhibited high reactivity with chlorine and very low reactivity with monochloramine, suggesting different mechanism of protection against disinfectants. Moreover, capsular EPS on cell membrane appeared to reduce membrane permeabilization by disinfectants as suggested by deformation of key functional groups in EPS and cell membrane (the C-O-C stretching of carbohydrate and the C=O stretching of ester group). The combined results supported that capsular EPS, acting either as a disinfectant consumer (for chlorine inactivation) or limiting access to reactive sites on cell membrane (for monochloramine inactivation), provide a protective role for bacterial cells against regulatory residual disinfectants by reducing membrane permeabilization. © 2012 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.

  16. Identification of Genes and Proteins Necessary for Catabolism of Acyclic Terpenes and Leucine/Isovalerate in Pseudomonas aeruginosa

    PubMed Central

    Förster-Fromme, Karin; Höschle, Birgit; Mack, Christina; Bott, Michael; Armbruster, Wolfgang; Jendrossek, Dieter

    2006-01-01

    Geranyl-coenzyme A (CoA)-carboxylase (GCase; AtuC/AtuF) and methylcrotonyl-CoA-carboxylase (MCase; LiuB/LiuD) are characteristic enzymes of the catabolic pathway of acyclic terpenes (citronellol and geraniol) and of saturated methyl-branched compounds, such as leucine or isovalerate, respectively. Proteins encoded by two gene clusters (atuABCDEFGH and liuRABCDE) of Pseudomonas aeruginosa PAO1 were essential for acyclic terpene utilization (Atu) and for leucine and isovalerate utilization (Liu), respectively, as revealed by phenotype analysis of 10 insertion mutants, two-dimensional gel electrophoresis, determination of GCase and MCase activities, and Western blot analysis of wild-type and mutant strains. Analysis of the genome sequences of other pseudomonads (P. putida KT2440 and P. fluorescens Pf-5) revealed candidate genes for Liu proteins for both species and candidate genes for Atu proteins in P. fluorescens. This result concurred with the finding that P. fluorescens, but not P. putida, could grow on acyclic terpenes (citronellol and citronellate), while both species were able to utilize leucine and isovalerate. A regulatory gene, atuR, was identified upstream of atuABCDEFGH and negatively regulated expression of the atu gene cluster. PMID:16820476

  17. Anti-infective properties of Lactobacillus fermentum against Staphylococcus aureus and Pseudomonas aeruginosa.

    PubMed

    Varma, Parvathi; Nisha, N; Dinesh, Kavitha R; Kumar, Anil V; Biswas, Raja

    2011-01-01

    Surgical wounds and implant-associated Staphylococcus aureus and Pseudomonas aeruginosa infections are often difficult to treat because of limited susceptibility of several of these strains to conventional antibiotics. As a result, there is a constant need for new alternative drugs. The aim of this study was to investigate the antimicrobial properties of Lactobacillus fermentum, a probiotic bacterium, which we have isolated from colonic biopsies. The inhibition of S. aureus and P. aeruginosa growth was evaluated by coincubating with L. fermentum strains. Growth inhibition was tested for several of their clinical isolates using agar well diffusion assays. For biofilm assay S. aureus and P. aeruginosa were grown on the glass slides and in 96-well plates in presence of 2.5 μg/ml culture filtrate of L. fermentum. Biofilms were photographed using confocal microscope or stained with 0.1% crystal violet. Reduction in the cytotoxicity of S. aureus and P. aeruginosa was observed in presence of 2.5 μg/ml L. fermentum-spent media. Using in vitroexperiments, we showed that L. fermentum-secreted compound(s) inhibits the growth, cytotoxicity and biofilm formation of several S. aureus and P. aeruginosa strains. Compound(s) present in the culture supernatant of L. fermentum may have promising applications in treating hospital-acquired infections. Copyright © 2011 S. Karger AG, Basel.

  18. Wheat Bran Enhances the Cytotoxicity of Immobilized Alcaligenes aquatilis F8 against Microcystis aeruginosa

    PubMed Central

    Sun, Pengfei; Lin, Hui; Wang, Guan; Zhang, Ximing; Zhang, Qichun; Zhao, Yuhua

    2015-01-01

    Algicidal bacteria offer a promising option for killing cyanobacteria. Therefore, a new Alcaligenes aquatilis strain F8 was isolated to control Microcystis aeruginosa in this study. The algicidal activity of strain F8 was dependent on the cell density of M. aeruginosa, and the maximal algicidal rate of the free bacterium reached 88.45% within 72 h. With a view to its application to the control of M. aeruginosa in the natural environment, strain F8 was immobilized in sodium alginate beads, but immobilization of the strain decreased its algicidal rate compared to that of the free bacterium. However, addition of wheat bran to the sodium alginate matrix used to immobilize strain F8 not only eliminated the adverse effects of immobilization on the bacteria but also resulted in an 8.83% higher algicidal rate of the immobilized than free bacteria. Exclusion and recovery methods were used to identify key ingredients of wheat bran and gain insight into the mechanism underlying the observed enhancement of algicidal activity. This analysis indicated that certain factors in wheat bran, including vitamins B1, B2, B9, and E were responsible for promoting bacterial growth and thereby improving the algicidal rate of immobilized strain F8. Our findings indicate that wheat bran is able to improve the algicidal efficiency of A. aquatilis strain F8 for killing M. aeruginosa and is a good source of not only carbon and nitrogen but also vitamins for bacteria. PMID:26295573

  19. Activity of Ceftazidime-Avibactam against Fluoroquinolone-Resistant Enterobacteriaceae and Pseudomonas aeruginosa

    PubMed Central

    Pitart, C.; Marco, F.; Keating, T. A.; Nichols, W. W.

    2015-01-01

    Ceftazidime-avibactam and comparator antibiotics were tested by the broth microdilution method against 200 Enterobacteriaceae and 25 Pseudomonas aeruginosa strains resistant to fluoroquinolones (including strains with the extended-spectrum β-lactamase [ESBL] phenotype and ceftazidime-resistant strains) collected from our institution. The MICs and mechanisms of resistance to fluoroquinolone were also studied. Ninety-nine percent of fluoroquinolone-resistant Enterobacteriaceae strains were inhibited at a ceftazidime-avibactam MIC of ≤4 mg/liter (using the susceptible CLSI breakpoint for ceftazidime alone as a reference). Ceftazidime-avibactam was very active against ESBL Escherichia coli (MIC90 of 0.25 mg/liter), ESBL Klebsiella pneumoniae (MIC90 of 0.5 mg/liter), ceftazidime-resistant AmpC-producing species (MIC90 of 1 mg/liter), non-ESBL E. coli (MIC90 of ≤0.125 mg/liter), non-ESBL K. pneumoniae (MIC90 of 0.25 mg/liter), and ceftazidime-nonresistant AmpC-producing species (MIC90 of ≤0.5 mg/liter). Ninety-six percent of fluoroquinolone-resistant P. aeruginosa strains were inhibited at a ceftazidime-avibactam MIC of ≤8 mg/liter (using the susceptible CLSI breakpoint for ceftazidime alone as a reference), with a MIC90 of 8 mg/liter. Additionally, fluoroquinolone-resistant mutants from each species tested were obtained in vitro from two strains, one susceptible to ceftazidime and the other a β-lactamase producer with a high MIC against ceftazidime but susceptible to ceftazidime-avibactam. Thereby, the impact of fluoroquinolone resistance on the activity of ceftazidime-avibactam could be assessed. The MIC90 values of ceftazidime-avibactam for the fluoroquinolone-resistant mutant strains of Enterobacteriaceae and P. aeruginosa were ≤4 mg/liter and ≤8 mg/liter, respectively. We conclude that the presence of fluoroquinolone resistance does not affect Enterobacteriaceae and P. aeruginosa susceptibility to ceftazidime-avibactam; that is, there is no cross

  20. Inhibitory effect of biofilm-forming Lactobacillus kunkeei strains against virulent Pseudomonas aeruginosa in vitro and in honeycomb moth (Galleria mellonella) infection model.

    PubMed

    Berríos, P; Fuentes, J A; Salas, D; Carreño, A; Aldea, P; Fernández, F; Trombert, A N

    2018-02-27

    Biofilms correspond to complex communities of microorganisms embedded in an extracellular polymeric matrix. Biofilm lifestyle predominates in Pseudomonas aeruginosa, an opportunistic Gram negative pathogen responsible for a wide spectrum of infections in humans, plants and animals. In this context, anti-biofilm can be considered a key strategy to control P. aeruginosa infections, thereby more research in the field is required. On the other hand, Lactobacillus species have been described as beneficial due to their anti-biofilm properties and their consequent effect against a wide spectrum of pathogens. In fact, biofilm-forming Lactobacilli seem to be more efficient than their planktonic counterpart to antagonise pathogenic bacteria. In this work, we demonstrated that Lactobacillus kunkeei, a novel Lactobacillus species isolated from honeybee guts, can form biofilms in vitro. In addition, the L. kunkeei biofilm can, in turn, inhibit the formation of P. aeruginosa biofilms. Finally, we found that L. kunkeei strains attenuate infection of P. aeruginosa in the Galleria mellonella model, presumably by affecting P. aeruginosa biofilm formation and/or their stability. Since L. kunkeei presents characteristics of a probiotic, this work provides evidence arguing that the use of this Lactobacillus species in both animals (including insects) and humans could contribute to impair P. aeruginosa biofilm formation.

  1. Pseudomonas aeruginosa genotypes acquired by children with cystic fibrosis by age 5-years.

    PubMed

    Kidd, Timothy J; Ramsay, Kay A; Vidmar, Suzanna; Carlin, John B; Bell, Scott C; Wainwright, Claire E; Grimwood, Keith

    2015-05-01

    We describe Pseudomonas aeruginosa acquisitions in children with cystic fibrosis (CF) aged ≤5-years, eradication treatment efficacy, and genotypic relationships between upper and lower airway isolates and strains from non-CF sources. Of 168 CF children aged ≤5-years in a bronchoalveolar lavage (BAL)-directed therapy trial, 155 had detailed microbiological results. Overall, 201/271 (74%) P. aeruginosa isolates from BAL and oropharyngeal cultures were available for genotyping, including those collected before and after eradication therapy. Eighty-two (53%) subjects acquired P. aeruginosa, of which most were unique strains. Initial eradication success rate was 90%, but 36 (44%) reacquired P. aeruginosa, with genotypic substitutions more common in BAL (12/14) than oropharyngeal (3/11) cultures. Moreover, oropharyngeal cultures did not predict BAL genotypes reliably. CF children acquire environmental P. aeruginosa strains frequently. However, discordance between BAL and oropharyngeal strains raises questions over upper airway reservoirs and how to best determine eradication in non-expectorating children. Copyright © 2014 European Cystic Fibrosis Society. Published by Elsevier B.V. All rights reserved.

  2. Antimicrobial susceptibilities and bacteriological characteristics of bovine Pseudomonas aeruginosa and Serratia marcescens isolates from mastitis.

    PubMed

    Ohnishi, Mamoru; Sawada, Takuo; Hirose, Kazuhiko; Sato, Reiichiro; Hayashimoto, Mizuki; Hata, Eiji; Yonezawa, Chizuko; Kato, Hajime

    2011-12-29

    The presence of metallo-β-lactamase (MBL)-producing and multidrug-resistant Pseudomonas aeruginosa (MDRP) strains among bovine isolates of Gram-negative bacilli, and O-serotypes of bovine Serratia marcescens and P. aeruginosa isolates have been reported rarely. The aims of this study were to (1) elucidate antimicrobial susceptibilities and O-serotypes of P. aeruginosa and S. marcescens isolates from bovine mastitis and the presence of MBL-producers and MDRP strains among them and (2) evaluate their relationships to human isolates. We investigated the MICs of 24 antimicrobials and O-serotypes for 116 P. aeruginosa and 55 S. marcescens isolates in Japan, primarily in 2006. A total of 171 isolates exhibited high antimicrobial susceptibilities with the exception of a partial drug. P. aeruginosa isolates exhibited high susceptibilities of ≥ 95.7% to ciprofloxacin, imipenem, meropenem, piperacillin, ceftazidime, cefepime, cefoperazone/sulbactam, amikacin, tobramycin, and gentamicin; however, they exhibited a susceptibility of only 69.8% to aztreonam. They exhibited substantial resistances to ceftriaxone, enrofloxacin, cefotaxime, and moxalactam. S. marcescens isolates exhibited high susceptibilities of ≥ 90.9% to kanamycin, ceftiofur, sulfamethoxazole-trimethoprim, and the 15 aforementioned drugs, but exhibited resistance to minocycline. Neither MBL-producers nor MDRP strains were detected among the 171 strains. The dominant serotypes of P. aeruginosa isolates were OG, OA, OB, OI, OF, OE, and OK; those of S. marcescens isolates were O6 and O5. Every S. marcescens isolate was pigmented. These findings suggest that bovine P. aeruginosa and S. marcescens isolates differ from human isolates from both antibiogram and phenotypic perspectives, and could help to evaluate differences in bacteriological characteristics between bovine and human isolates. Copyright © 2011 Elsevier B.V. All rights reserved.

  3. Pseudomonas aeruginosa Lifestyle: A Paradigm for Adaptation, Survival, and Persistence

    PubMed Central

    Moradali, M. Fata; Ghods, Shirin; Rehm, Bernd H. A.

    2017-01-01

    Pseudomonas aeruginosa is an opportunistic pathogen affecting immunocompromised patients. It is known as the leading cause of morbidity and mortality in cystic fibrosis (CF) patients and as one of the leading causes of nosocomial infections. Due to a range of mechanisms for adaptation, survival and resistance to multiple classes of antibiotics, infections by P. aeruginosa strains can be life-threatening and it is emerging worldwide as public health threat. This review highlights the diversity of mechanisms by which P. aeruginosa promotes its survival and persistence in various environments and particularly at different stages of pathogenesis. We will review the importance and complexity of regulatory networks and genotypic-phenotypic variations known as adaptive radiation by which P. aeruginosa adjusts physiological processes for adaptation and survival in response to environmental cues and stresses. Accordingly, we will review the central regulatory role of quorum sensing and signaling systems by nucleotide-based second messengers resulting in different lifestyles of P. aeruginosa. Furthermore, various regulatory proteins will be discussed which form a plethora of controlling systems acting at transcriptional level for timely expression of genes enabling rapid responses to external stimuli and unfavorable conditions. Antibiotic resistance is a natural trait for P. aeruginosa and multiple mechanisms underlying different forms of antibiotic resistance will be discussed here. The importance of each mechanism in conferring resistance to various antipseudomonal antibiotics and their prevalence in clinical strains will be described. The underlying principles for acquiring resistance leading pan-drug resistant strains will be summarized. A future outlook emphasizes the need for collaborative international multidisciplinary efforts to translate current knowledge into strategies to prevent and treat P. aeruginosa infections while reducing the rate of antibiotic resistance

  4. Mitigation of a nitrate reducing Pseudomonas aeruginosa biofilm and anaerobic biocorrosion using ciprofloxacin enhanced by D-tyrosine.

    PubMed

    Jia, Ru; Yang, Dongqing; Xu, Dake; Gu, Tingyue

    2017-07-31

    Pseudomonas aeruginosa (PA) is a ubiquitous microbe. It can form recalcitrant biofilms in clinical and industrial settings. PA biofilms cause infections in patients. They also cause biocorrosion of medical implants. In this work, D-tyrosine (D-tyr) was investigated as an antimicrobial enhancer for ciprofloxacin (CIP) against a wild-type PA biofilm (strain PAO1) on C1018 carbon steel in a strictly anaerobic condition. Seven-day biofilm prevention test results demonstrated that 2 ppm (w/w) D-tyr enhanced 30 ppm CIP by achieving extra 2-log sessile cell reduction compared with the 30 ppm CIP alone treatment. The cocktail of 30 ppm CIP + 2 ppm D-tyr achieved similar efficacy as the 80 ppm CIP alone treatment in the biofilm prevention test. Results also indicated that the enhanced antimicrobial treatment reduced weight loss and pitting corrosion. In the 3-hour biofilm removal test, the cocktail of 80 ppm CIP + 5 ppm D-tyr achieved extra 1.5-log reduction in sessile cell count compared with the 80 ppm CIP alone treatment. The cocktail of 80 ppm CIP + 5 ppm D-tyr achieved better efficacy than the 150 ppm CIP alone treatment in the biofilm removal test.

  5. Description of IMP-31, a novel metallo-β-lactamase found in an ST235 Pseudomonas aeruginosa strain in Western Germany.

    PubMed

    Pfennigwerth, Niels; Geis, Gabriele; Gatermann, Sören G; Kaase, Martin

    2015-07-01

    The objective of this study was to characterize a novel IMP-type metallo-β-lactamase (MBL) found in an MDR clinical isolate of Pseudomonas aeruginosa. The P. aeruginosa isolate NRZ-00156 was recovered from an inguinal swab from a patient hospitalized in Western Germany and showed high MICs of carbapenems. MBL production was analysed by Etest for MBLs, an EDTA combined disc test and an EDTA bioassay. Typing of the isolate was performed by MLST. Genetic characterization of the new blaIMP gene was performed by sequencing the PCR products. A phylogenetic tree was constructed. The novel blaIMP gene was expressed in Escherichia coli TOP10 and the enzyme was subjected to biochemical characterization. The P. aeruginosa isolate NRZ-00156 expressed the ST235 allelic profile and was resistant to all the β-lactams tested except aztreonam. The isolate was positive for MBL production and harboured a new IMP allele, blaIMP-31, located on a disrupted class I integron [also carrying the blaOXA-35, aac(6')-Ib, aac(3)-Ic and aphA15 genes]. Its closest relative was IMP-35, with 96.7% amino acid identity. Expression of blaIMP-31 demonstrated that E. coli TOP10 producing IMP-31 had elevated resistance to all the β-lactams tested except aztreonam. Kinetic data were obtained for both IMP-31 and IMP-1. In comparison with IMP-1, IMP-31 showed weaker hydrolytic activity against all the β-lactams tested, which resulted from lower kcat values. The characterization of the new IMP-type gene blaIMP-31 from an ST235 P. aeruginosa isolate indicates an ongoing spread of highly divergent IMP-type carbapenemases in clinical P. aeruginosa strains and highlights the continuous need for the prevention of nosocomial infections caused by MDR Gram-negative bacteria. © The Author 2015. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  6. The Effect of Phenazine-1-Carboxylic Acid on Mycelial Growth of Botrytis cinerea Produced by Pseudomonas aeruginosa LV Strain

    PubMed Central

    Simionato, Ane S.; Navarro, Miguel O. P.; de Jesus, Maria L. A.; Barazetti, André R.; da Silva, Caroline S.; Simões, Glenda C.; Balbi-Peña, Maria I.; de Mello, João C. P.; Panagio, Luciano A.; de Almeida, Ricardo S. C.; Andrade, Galdino; de Oliveira, Admilton G.

    2017-01-01

    One of the most important postharvest plant pathogens that affect strawberries, grapes and tomatoes is Botrytis cinerea, known as gray mold. The fungus remains in latent form until spore germination conditions are good, making infection control difficult, causing great losses in the whole production chain. This study aimed to purify and identify phenazine-1-carboxylic acid (PCA) produced by the Pseudomonas aeruginosa LV strain and to determine its antifungal activity against B. cinerea. The compounds produced were extracted with dichloromethane and passed through a chromatographic process. The purity level of PCA was determined by reversed-phase high-performance liquid chromatography semi-preparative. The structure of PCA was confirmed by nuclear magnetic resonance and electrospray ionization mass spectrometry. Antifungal activity was determined by the dry paper disk and minimum inhibitory concentration (MIC) methods and identified by scanning electron microscopy and confocal microscopy. The results showed that PCA inhibited mycelial growth, where MIC was 25 μg mL-1. Microscopic analysis revealed a reduction in exopolysaccharide (EPS) formation, showing distorted and damaged hyphae of B. cinerea. The results suggested that PCA has a high potential in the control of B. cinerea and inhibition of EPS (important virulence factor). This natural compound is a potential alternative to postharvest control of gray mold disease. PMID:28659907

  7. The Effect of Phenazine-1-Carboxylic Acid on Mycelial Growth of Botrytis cinerea Produced by Pseudomonas aeruginosa LV Strain.

    PubMed

    Simionato, Ane S; Navarro, Miguel O P; de Jesus, Maria L A; Barazetti, André R; da Silva, Caroline S; Simões, Glenda C; Balbi-Peña, Maria I; de Mello, João C P; Panagio, Luciano A; de Almeida, Ricardo S C; Andrade, Galdino; de Oliveira, Admilton G

    2017-01-01

    One of the most important postharvest plant pathogens that affect strawberries, grapes and tomatoes is Botrytis cinerea , known as gray mold. The fungus remains in latent form until spore germination conditions are good, making infection control difficult, causing great losses in the whole production chain. This study aimed to purify and identify phenazine-1-carboxylic acid (PCA) produced by the Pseudomonas aeruginosa LV strain and to determine its antifungal activity against B. cinerea . The compounds produced were extracted with dichloromethane and passed through a chromatographic process. The purity level of PCA was determined by reversed-phase high-performance liquid chromatography semi-preparative. The structure of PCA was confirmed by nuclear magnetic resonance and electrospray ionization mass spectrometry. Antifungal activity was determined by the dry paper disk and minimum inhibitory concentration (MIC) methods and identified by scanning electron microscopy and confocal microscopy. The results showed that PCA inhibited mycelial growth, where MIC was 25 μg mL -1 . Microscopic analysis revealed a reduction in exopolysaccharide (EPS) formation, showing distorted and damaged hyphae of B. cinerea . The results suggested that PCA has a high potential in the control of B. cinerea and inhibition of EPS (important virulence factor). This natural compound is a potential alternative to postharvest control of gray mold disease.

  8. The distribution of a phage-related insertion sequence element in the cyanobacterium, Microcystis aeruginosa.

    PubMed

    Kuno, Sotaro; Yoshida, Takashi; Kamikawa, Ryoma; Hosoda, Naohiko; Sako, Yoshihiko

    2010-01-01

    The cyanophage Ma-LMM01, specifically-infecting Microcystis aeruginosa, has an insertion sequence (IS) element that we named IS607-cp showing high nucleotide similarity to a counterpart in the genome of the cyanobacterium Cyanothece sp. We tested 21 strains of M. aeruginosa for the presence of IS607-cp using PCR and detected the element in strains NIES90, NIES112, NIES604, and RM6. Thermal asymmetric interlaced PCR (TAIL-PCR) revealed each of these strains has multiple copies of IS607-cp. Some of the ISs were classified into three types based on their inserted positions; IS607-cp-1 is common in strains NIES90, NIES112 and NIES604, whereas IS607-cp-2 and IS607-cp-3 are specific to strains NIES90 and RM6, respectively. This multiplicity may reflect the replicative transposition of IS607-cp. The sequence of IS607-cp in Ma-LMM01 showed robust affinity to those found in M. aeruginosa and Cyanothece spp. in a phylogenetic tree inferred from counterparts of various bacteria. This suggests the transfer of IS607-cp between the cyanobacterium and its cyanophage. We discuss the potential role of Ma-LMM01-related phages as donors of IS elements that may mediate the transfer of IS607-cp; and thereby partially contribute to the genome plasticity of M. aeruginosa.

  9. Volatile fingerprinting of pseudomonas aeruginosa and respiratory syncytial virus infection in an in vitro cystic fibrosis co-infection model.

    PubMed

    Purcaro, Giorgia; Rees, Christiaan; Melvin, Jeffrey A; Bomberg, Jennifer M; Hill, Jane E

    2018-05-08

    Volatile molecules in exhaled breath represent potential biomarkers in the setting of infectious diseases, particularly those affecting the respiratory tract. In particular, Pseudomonas aeruginosa is a critically-important respiratory pathogen in specific subsets of the population, such as those with cystic fibrosis. Infections caused by P. aeruginosa can be particularly problematic when co-infection with respiratory syncytial virus (RSV) occurs, as this is correlated with the establishment of chronic P. aeruginosa infection. In the present study, we evaluate the volatile metabolites produced by P. Aeruginosa (PAO1)-infected, RSV-infected, co-infected, or uninfected cystic fibrosis bronchial epithelial (CFBE) cells, in vitro. We identified a volatile metabolic signature that could discriminate between P. aeruginosa-infected and non-P. aeruginosa-infected CFBE with an area under the receiver operating characteristic curve (AUROC) of 0.850, using the machine learning algorithm Random Forest (RF). Although we could not discriminate between RSV-infected and non-RSV-infected CFBE (AUROC = 0.431), we note that sample classification probabilities for RSV-infected cell, generated using RF, were between those of uninfected CFBE and P. aeruginosa-infected CFBE, suggesting that RSV infection may result in a volatile metabolic profile that shares attributes with both of these groups. To more precisely elucidate the biological origins of the volatile metabolites that were discriminatory between P. aeruginosa-infected and non-P. aeruginosa-infected CFBE, we measured the volatile metabolites produced by P. aeruginosa grown in the absence of CFBE. Our findings suggest that the discriminatory metabolites produced likely result from the interaction of P. aeruginosa with the CFBE cells, rather than the metabolism of media components by the bacterium. Taken together, our findings support the notion that P. aeruginosa interacting with CFBE yields a particular volatile metabolic

  10. Purification and molecular and biological characterisation of the 1-hydroxyphenazine, produced by an environmental strain of Pseudomonas aeruginosa.

    PubMed

    Prabhu, Meghanath S; Walawalkar, Yogesh D; Furtado, Irene

    2014-12-01

    Pseudomonas aeruginosa--an opportunistic pathogen, perhaps best known for chronic lung infections, produces wide range of pigments that possess specific activities which either assist the organism's survival or bring about changes within host. A similar blue-green diffusible pigment producing P. aeruginosa was isolated from dug-well water, so as to extract 1-hydroxyphenazine from its crude pigment. The compound was purified from the crude pigment using column chromatography followed by a preparative thin layer chromatography that showed a single yellow spot. Further molecular characterisation of the purified component was carried out using UV-Vis spectrophotometer, Fourier transform infrared spectroscopy, nuclear magnetic resonance spectroscopy and mass spectroscopy which showed respective peaks corresponding to 1-hydroxyphenazine. Biological characterisation using in vitro assays revealed that 1-hydroxyphenazine showed anti-bacterial activity only against Bacillus sp. and a concentration of 30 µg/ml induced noticeable morphological alteration in A549 human lung adenocarcinoma cells followed by cell death after 48 h. Thus, such active components within bacterial pigments can be characterized and used as possible anti-bacterial or anti-cancer agents.

  11. An investigation of Pseudomonas aeruginosa biofilm growth on novel nanocellulose fibre dressings.

    PubMed

    Powell, Lydia C; Khan, Saira; Chinga-Carrasco, Gary; Wright, Chris J; Hill, Katja E; Thomas, David W

    2016-02-10

    Nanocellulose from wood is a novel biomaterial, which is highly fibrillated at the nanoscale. This affords the material a number of advantages, including self-assembly, biodegradability and the ability to absorb and retain moisture, which highlights its potential usefulness in clinical wound-dressing applications. In these in vitro studies, the wound pathogen Pseudomonas aeruginosa PAO1 was used to assess the ability of two nanocellulose materials to impair bacterial growth (<48 h). The two nanocelluloses had a relatively small fraction of residual fibres (<4%) and thus a large fraction of nanofibrils (widths <20 nm). Scanning electron microscopy and confocal laser scanning microscopy imaging demonstrated impaired biofilm growth on the nanocellulose films and increased cell death when compared to a commercial control wound dressing, Aquacel(®). Nanocellulose suspensions inhibited bacterial growth, whilst UV-vis spectrophotometry and laser profilometry also revealed the ability of nanocellulose to form smooth, translucent films. Atomic force microscopy studies of the surface properties of nanocellulose demonstrated that PAO1 exhibited markedly contrasting morphology when grown on the nanocellulose film surfaces compared to an Aquacel(®) control dressing (p<0.05). This study highlights the potential utility of these biodegradable materials, from a renewable source, for wound dressing applications in the prevention and treatment of biofilm development. Copyright © 2015 Elsevier Ltd. All rights reserved.

  12. Interspecific Small Molecule Interactions between Clinical Isolates of Pseudomonas aeruginosa and Staphylococcus aureus from Adult Cystic Fibrosis Patients

    PubMed Central

    Mitchell, Gabriel; Déziel, Eric; Dekimpe, Valérie; Cantin, André M.; Frost, Eric; Malouin, François

    2014-01-01

    Pseudomonas aeruginosa and Staphylococcus aureus are the most prevalent pathogens in airway infections of cystic fibrosis (CF) patients. We studied how these pathogens coexist and interact with each other. Clinical isolates of both species were retrieved from adult CF patients. Culture supernatants from 63 P. aeruginosa isolates triggered a wide range of biofilm-stimulatory activities when added to the culture of a control S. aureus strain. The extent of biofilm formation by S. aureus was positively correlated to the levels of the 2-alkyl-4-(1H)-quinolones (AQs) Pseudomonas Quinolone Signal (PQS) and 2-heptyl-4-hydroxy quinoline N-oxide (HQNO) produced by the P. aeruginosa isolates. Supernatants from P. aeruginosa isogenic mutants deficient in PQS and HQNO production stimulated significantly less biofilm formation by S. aureus than that seen with the parental strain PA14. When studying co-isolated pairs of P. aeruginosa and S. aureus retrieved from patients showing both pathogens, P. aeruginosa supernatants stimulated less biofilm production by the S. aureus counterparts compared to that observed using the control S. aureus strain. Accordingly, some P. aeruginosa isolates produced low levels of exoproducts and also some of the clinical S. aureus isolates were not stimulated by their co-isolates or by PA14 despite adequate production of HQNO. This suggests that colonization of the CF lungs promotes some type of strain selection, or that co-existence requires specific adaptations by either or both pathogens. Results provide insights on bacterial interactions in CF. PMID:24466207

  13. Biodegradation of petroleum hydrocarbons by oleophilic strain of Pseudomonas aeruginosa NCIM 5514.

    PubMed

    Varjani, Sunita J; Upasani, Vivek N

    2016-12-01

    The aim of this work was to study the potential of an indigenous strain of Pseudomonas aeruginosa NCIM 5514, isolated from petroleum-polluted soil, for the biodegradation of crude petroleum oil. The isolate completely decolorized 2,6-dichlorophenol indophenol in 120h when grown at (37±1°C), indicating its hydrocarbon utilizing nature. Ex situ biodegradation study was performed to find out quantitative utilization and biodegradation of paraffin(s) present in crude oil. When the culture was grown in Bushnell-Hass medium containing crude oil (3%,v/v) at 37°C, 180rpm for 60days, the viscosity of the oil was reduced from 1883cp to 1002cp. Gravimetric and gas chromatographic analysis showed 61.03% and 60.63% of biodegradation of C8-C36+ hydrocarbons, respectively. These results indicated that the isolate has potential to be used for ex-situ and in-situ bioremediation of hydrocarbon pollutants and could have promising applications in petrochemical industry. Copyright © 2016 Elsevier Ltd. All rights reserved.

  14. [Susceptibility and resistence of Pseudomonas aeruginosa to antimicrobial agents].

    PubMed

    Gamero Delgado, M C; García-Mayorgas, A D; Rodríguez, F; Ibarra, A; Casal, M

    2007-06-01

    Pseudomonas aeruginosa is an opportunistic microorganism that is frequently the cause of nosocomial infections. Multiple mechanisms are involved in its natural and acquired resistance to many of the antimicrobial agents commonly used in clinical practice. The objective of this study was to assess the susceptibility and resistance patterns of P. aeruginosa strains isolated in Hospital Reina Sofia between 2000 and 2005, as well as to analyze the differences between intrahospital and extrahospital isolates in 2005 and to compare the results with those obtained in other studies. A total of 3,019 strains of P. aeruginosa from different hospitals and nonhospital settings were evaluated, taking into consideration their degree of sensitivity to different antibiotics. The MICs were determined by means of the Wider I automated system (Soria Melguizo), taking into consideration the criteria of susceptibility and resistance recommended by MENSURA. Results of the analysis showed that P. aeruginosa maintained similar levels of antimicrobial susceptibility during the period 2000-2005, with increased susceptibility to amikacin, gentamicin and tobramycin. There were also important differences in the degree of susceptibility between intrahospital and extrahospital strains, except for imipenem and fosfomycin. The intrahospital difference in susceptibility was also evaluated, emphasizing the importance of periodically studying susceptibility and resistance patterns of P. aeruginosa in each setting in order to evaluate different therapeutic guidelines, as it is not always advisable to extrapolate data from different regions. These differences can be explained by the different use of antibiotics in each center and the geographic variations of the resistance mechanisms of P. aeruginosa.

  15. Mechanisms decreasing in vitro susceptibility to the LpxC inhibitor CHIR-090 in the gram-negative pathogen Pseudomonas aeruginosa.

    PubMed

    Caughlan, Ruth E; Jones, Adriana K; Delucia, Angela M; Woods, Angela L; Xie, Lili; Ma, Bing; Barnes, S Whitney; Walker, John R; Sprague, Elizabeth R; Yang, Xia; Dean, Charles R

    2012-01-01

    Testing P. aeruginosa efflux pump mutants showed that the LpxC inhibitor CHIR-090 is a substrate for MexAB-OprM, MexCD-OprJ, and MexEF-OprN. Utilizing P. aeruginosa PAO1 with a chromosomal mexC::luxCDABE fusion, luminescent mutants arose on medium containing 4 μg/ml CHIR-090, indicating upregulation of MexCD-OprJ. These mutants were less susceptible to CHIR-090 (MIC, 4 μg/ml) and had mutations in the mexCD-oprJ repressor gene nfxB. Nonluminescent mutants (MIC, 4 μg/ml) that had mutations in the mexAB-oprM regulator gene mexR were also observed. Plating the clinical isolate K2153 on 4 μg/ml CHIR-090 selected mutants with alterations in mexS (immediately upstream of mexT), which upregulates MexEF-OprN. A mutant altered in the putative1ribosomal binding site (RBS) upstream of lpxC and overexpressing LpxC was selected on a related LpxC inhibitor and exhibited reduced susceptibility to CHIR-090. Overexpression of LpxC from a plasmid reduced susceptibility to CHIR-090, and introduction of the altered RBS in this construct further increased expression of LpxC and decreased susceptibility to CHIR-090. Using a mutS (hypermutator) strain, a mutant with an altered lpxC target gene (LpxC L18V) was also selected. Purified LpxC L18V had activity similar to that of wild-type LpxC in an in vitro assay but had reduced inhibition by CHIR-090. Finally, an additional class of mutant, typified by an extreme growth defect, was identified. These mutants had mutations in fabG, indicating that alteration in fatty acid synthesis conferred resistance to LpxC inhibitors. Passaging experiments showed progressive decreases in susceptibility to CHIR-090. Therefore, P. aeruginosa can employ several strategies to reduce susceptibility to CHIR-090 in vitro.

  16. Pseudomonas protegens Pf-5 favours self-produced siderophore over free-loading in interspecies competition for iron.

    PubMed

    Sexton, D Joseph; Glover, Rochelle C; Loper, Joyce E; Schuster, Martin

    2017-09-01

    Many microorganisms compete for extracellular iron using strain-specific chelators known as siderophores. The ferric-siderophore complex limits local access to iron because import requires a suitable cognate receptor. Interestingly, many species carry receptors that enable 'cross-feeding' on heterologous siderophores made by neighboring organisms, although little is known about how this ubiquitous behaviour is regulated. Here, we investigated the soil bacterium Pseudomonas protegens Pf-5, a strain remarkable for its ability to use dozens of heterologous siderophores. We characterized the expression of six pyoverdine-type (PVD) siderophore receptors in response to their cognate PVD. In general, we found expression is tightly regulated to reflect availability of their cognate PVD. In contrast, Pf-5 continues to secrete its own primary siderophore, PVD Pf-5 , despite the capability and opportunity to cross-feed. We demonstrate that this strategy is beneficial in co-culture with a competing PVD PAO1 -producer, P. aeruginosa PAO1. Although Pf-5 can cross-feed on PVD PAO1 , production of PVD Pf-5 is required to maintain a competitive advantage. We attribute this to an antagonistic effect of PVD Pf-5 on the growth of PAO1, presumably through limiting access to iron. Our results demonstrate the benefits of excluding competitors out-weigh the incentives associated with a free-loader lifestyle for Pf-5. © 2017 Society for Applied Microbiology and John Wiley & Sons Ltd.

  17. Identification of pilin pools in the membranes of Pseudomonas aeruginosa.

    PubMed Central

    Watts, T H; Worobec, E A; Paranchych, W

    1982-01-01

    The proteins of purified inner and outer membranes obtained from Pseudomonas aeruginosa strains PAK and PAK/2Pfs were subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis, transferred to nitrocellulose, and treated with antiserum raised against pure pili. Bound antipilus antibodies were visualized by reaction with 125I-labeled protein A from Staphylococcus aureus. The results showed that there are pools of pilin in both the inner and outer membranes of P. aeruginosa and that the pool size in the multipiliated strain is comparable with that of the wild-type strain. Images PMID:6813311

  18. Prevalence of ESBL-producing Pseudomonas aeruginosa isolates in Warsaw, Poland, detected by various phenotypic and genotypic methods

    PubMed Central

    Róg, Patrycja; Smolińska-Król, Katarzyna; Ćmiel, Milena; Słoczyńska, Alicja; Patzer, Jan; Dzierżanowska, Danuta; Wolinowska, Renata; Starościak, Bohdan; Tyski, Stefan

    2017-01-01

    Knowledge of the prevalence of ESBL enzymes among P. aeruginosa strains compared to the Enterobacteraiceae family is limited. The phenotypic tests recommended by EUCAST for the detection of ESBL-producing Enterobacteriaceae are not always suited for P. aeruginosa strains. This is mainly due to the presence of other families of ESBLs in P. aeruginosa isolates more often than in Enterobacteriaceae, production of natural AmpC cephalosporinase and its overexpression, and co-production of metallo-β-lactamases. The aim of this study was to determine the occurrence of ESBLs in P. aeruginosa isolated from patients from hospitals in Warsaw, to evaluate the ESBL production of these isolates using currently available phenotypic tests, their modifications, multiplex PCR and molecular typing of ESBL-positive isolates by PFGE. Clinical isolates of P. aeruginosa were collected in 2000-2014 from four Warsaw hospitals. Based on the data obtained in this study, we suggest using three DDST methods with inhibitors, such as clavulanic acid, sulbactam and imipenem, to detect ESBL-producing P. aeruginosa strains. Depending on the appearance of the plates, we suggest a reduction in the distance between discs with antibiotics to 15 mm and the addition of boronic acid at 0.4 mg per disc. The analysed isolates carried genes encoding ESBL from the families VEB (69 isolates with VEB-9), GES (6 with GES-1, 1 GES-5, 5 GES-13 and 2 with GES-15), OXA-2 (12 with OXA-15, 1 OXA-141, 1 OXA-210, 1 OXA-543 and 1 with OXA-544) and OXA-10 (5 isolates with OXA-74 and one with OXA-142). The most important result of this study was the discovery of three new genes, blaGES-15, blaOXA-141 and blaOXA-142; their nucleotide sequences have been submitted to the NCBI GenBank. It is also very important to note that this is the first report on the epidemiological problem of VEB-9-producing bacterial strains, not only in Poland but also worldwide. PMID:28658322

  19. Genomic islands 1 and 2 play key roles in the evolution of extensively drug-resistant ST235 isolates of Pseudomonas aeruginosa

    PubMed Central

    Scott, Martin; Worden, Paul; Huntington, Peter; Hudson, Bernard; Karagiannis, Thomas; Charles, Ian G.; Djordjevic, Steven P.

    2016-01-01

    Pseudomonas aeruginosa are noscomially acquired, opportunistic pathogens that pose a major threat to the health of burns patients and the immunocompromised. We sequenced the genomes of P. aeruginosa isolates RNS_PA1, RNS_PA46 and RNS_PAE05, which displayed resistance to almost all frontline antibiotics, including gentamicin, piperacillin, timentin, meropenem, ceftazidime and colistin. We provide evidence that the isolates are representatives of P. aeruginosa sequence type (ST) 235 and carry Tn6162 and Tn6163 in genomic islands 1 (GI1) and 2 (GI2), respectively. GI1 disrupts the endA gene at precisely the same chromosomal location as in P. aeruginosa strain VR-143/97, of unknown ST, creating an identical CA direct repeat. The class 1 integron associated with Tn6163 in GI2 carries a blaGES-5–aacA4–gcuE15–aphA15 cassette array conferring resistance to carbapenems and aminoglycosides. GI2 is flanked by a 12 nt direct repeat motif, abuts a tRNA-gly gene, and encodes proteins with putative roles in integration, conjugative transfer as well as integrative conjugative element-specific proteins. This suggests that GI2 may have evolved from a novel integrative conjugative element. Our data provide further support to the hypothesis that genomic islands play an important role in de novo evolution of multiple antibiotic resistance phenotypes in P. aeruginosa. PMID:26962050

  20. [In vitro indirect pathogenesis of Pseudomonas aeruginosa against anti MRSA chemotherapy].

    PubMed

    Satoh, Naotake; Kondo, Shigemi; Yamada, Toshihiko; Saionji, Katsu; Oguri, Toyoko; Igari, Jun

    2004-09-01

    In the patient with a chronic respiratory disease, both Pseudomonas aeruginosa and methicillin-resistant Staphylococcus aureus (MRSA) are frequently detected from expectoration. Vancomycin (VCM) and arbekacin (ABK) are both recommended for the chemotherapy of MRSA infection in Japan. Minocycline (MINO) is also selected for the treatment of MRSA infection. While rifampicin (RFP) and a trimetoprim-sulfamethoxazole combination (ST) are also recommended in Europe and USA but not recommended in Japan for the chemotherapy of MRSA infection. It is pointed out that coexistence bacteria affect chemotherapy as an indirect pathogen. Not only an antibacterial action but the immunological action or the metabolic effect against chronic P. aeruginosa infection such as DPB is known by the administration of 14-membered ring macrolides including erythromycin (EM). We considered the influence of P. aeruginosa isolated with MRSA on the activity against anti-MRSA agents by the disk diffusion method with bilayer flat agar in vitro. Moreover, we also examined the influence of EM against the activity of the anti-MRSA agents when P. aeruginosa was coexistence. One strain of MRSA as an indicator strain and 100 strains of P. aeruginosa as test strains, which were obtained from clinical materials, were used for the following experiment. P. aeruginosa was streaked on to the Mueller-Hinton agar culture medium (MHA), and they incubated at 35 degrees C for 24 hours. Then, the blood agar plate was piled up, MRSA was streaked on the blood agar surface, the susceptibility test disks (VCM, ABK, MINO, RFP, ST) were put on it, and incubated at 35 degrees C for a further 24 hours. The diameter of the zone of inhibition around the susceptibility disks against MRSA was measured and compared with P. aeruginosa free experiments. The anti-MRSA activity of MINO, ST and ABK was reduced by coexistence of P. aeruginosa. In RFP and VCM, the anti-MRSA activity was reinforced by coexistence of P. aeruginosa

  1. Relating the physical properties of Pseudomonas aeruginosa lipopolysaccharides to virulence by atomic force microscopy.

    PubMed

    Ivanov, Ivan E; Kintz, Erica N; Porter, Laura A; Goldberg, Joanna B; Burnham, Nancy A; Camesano, Terri A

    2011-03-01

    Lipopolysaccharides (LPS) are an important class of macromolecules that are components of the outer membrane of Gram-negative bacteria such as Pseudomonas aeruginosa. P. aeruginosa contains two different sugar chains, the homopolymer common antigen (A band) and the heteropolymer O antigen (B band), which impart serospecificity. The characteristics of LPS are generally assessed after isolation rather than in the context of whole bacteria. Here we used atomic force microscopy (AFM) to probe the physical properties of the LPS of P. aeruginosa strain PA103 (serogroup O11) in situ. This strain contains a mixture of long and very long polymers of O antigen, regulated by two different genes. For this analysis, we studied the wild-type strain and four mutants, ΔWzz1 (producing only very long LPS), ΔWzz2 (producing only long LPS), DΔM (with both the wzz1 and wzz2 genes deleted), and Wzy::GM (producing an LPS core oligosaccharide plus one unit of O antigen). Forces of adhesion between the LPS on these strains and the silicon nitride AFM tip were measured, and the Alexander and de Gennes model of steric repulsion between a flat surface and a polymer brush was used to calculate the LPS layer thickness (which we refer to as length), compressibility, and spacing between the individual molecules. LPS chains were longest for the wild-type strain and ΔWzz1, at 170.6 and 212.4 nm, respectively, and these values were not statistically significantly different from one another. Wzy::GM and DΔM have reduced LPS lengths, at 34.6 and 37.7 nm, respectively. Adhesion forces were not correlated with LPS length, but a relationship between adhesion force and bacterial pathogenicity was found in a mouse acute pneumonia model of infection. The adhesion forces with the AFM probe were lower for strains with LPS mutations, suggesting that the wild-type strain is optimized for maximal adhesion. Our research contributes to further understanding of the role of LPS in the adhesion and virulence of

  2. Pseudomonas aeruginosa gshA Mutant Is Defective in Biofilm Formation, Swarming, and Pyocyanin Production

    PubMed Central

    Van Laar, Tricia A.; Esani, Saika; Birges, Tyler J.; Hazen, Bethany; Thomas, Jason M.

    2018-01-01

    ABSTRACT Pseudomonas aeruginosa is a ubiquitous Gram-negative bacterium that can cause severe opportunistic infections. The principal redox buffer employed by this organism is glutathione (GSH). To assess the role of GSH in the virulence of P. aeruginosa, a number of analyses were performed using a mutant strain deficient in gshA, which does not produce GSH. The mutant strain exhibited a growth delay in minimal medium compared to the wild-type strain. Furthermore, the gshA mutant was defective in biofilm and persister cell formation and in swimming and swarming motility and produced reduced levels of pyocyanin, a key virulence factor. Finally, the gshA mutant strain demonstrated increased sensitivity to methyl viologen (a redox cycling agent) as well as the thiol-reactive antibiotics fosfomycin and rifampin. Taken together, these data suggest a key role for GSH in the virulence of P. aeruginosa. IMPORTANCE Pseudomonas aeruginosa is a ubiquitous bacterium that can cause severe opportunistic infections, including many hospital-acquired infections. It is also a major cause of infections in patients with cystic fibrosis. P. aeruginosa is intrinsically resistant to a number of drugs and is capable of forming biofilms that are difficult to eradicate with antibiotics. The number of drug-resistant strains is also increasing, making treatment of P. aeruginosa infections very difficult. Thus, there is an urgent need to understand how P. aeruginosa causes disease in order to find novel ways to treat infections. We show that the principal redox buffer, glutathione (GSH), is involved in intrinsic resistance to the fosfomycin and rifampin antibiotics. We further demonstrate that GSH plays a role in P. aeruginosa disease and infection, since a mutant lacking GSH has less biofilm formation, is less able to swarm, and produces less pyocyanin, a pigment associated with infection. PMID:29669887

  3. [Analysis of drug resistance and drug resistance genes of imipenem-resistant Pseudomonas aeruginosa strains isolated from burn wards].

    PubMed

    Liu, Shuhua; Liu, Pinghong; Xue, Xiaodong; Chen, Zhaojun; Pei, Decui

    2014-02-01

    To analyze the drug resistance and drug resistance genes of imipenem-resistant Pseudomonas aeruginosa (IRPA) strains isolated from burn wards. From June 2011 to June 2012, 30 strains of IRPA were isolated from wound excretion, sputum, and venous catheter attachment from burn patients hospitalized in Guangzhou Hospital of Integrated Traditional Chinese and Western Medicine. Drug resistance of the IRPA to 12 antibiotics commonly used in clinic, including ceftazidime, amikacin, ciprofloxacin, etc., was tested with K-B paper agar disk diffusion method. Metallo-β-lactamase (MBL)-producing IRPA was detected by synergism test with imipenem-2-mercaptoethanol. Plasmid of IRPA was extracted, and it was inserted into competent cells, producing transformation strains (TSs). Drug resistance of TSs to imipenem and the MBL-producing TSs were detected. The genes blaIMP, blaVIM, blaOXA-1, blaOXA-2 and blaOXA-10 of IRPA and the TSs were detected by polymerase chain reaction. The drug resistance of IRPA producing MBL or OXA enzyme was summed up. The sensitive rates of the 30 strains of IRPA to the 12 antibiotics were equal to or above 60.0%. Six strains of MBL-producing IRPA were screened. Twenty-four TSs were resistant to imipenem, and 6 strains among them were MBL-producing positive. Among the 30 strains of IRPA, 6 strains and their corresponding TSs carried blaVIM; 20 strains and their corresponding TSs carried blaOXA-10; no strain was detected to carry blaIMP, blaOXA-1 or blaOXA-2. Two strains and their corresponding TSs were detected carrying both blaVIM and blaOXA-10. No significant difference of drug resistance was observed between strains producing only MBL or OXA enzyme, with the same high resistance to β-lactam antibiotics and some degree of sensitivity to aminoglycoside antibiotics. Strains producing enzymes MBL and OXA were all resistant to the 12 antibiotics. IRPA strains isolated from burn wards of Guangzhou Hospital of Integrated Traditional Chinese and Western

  4. Expression of Fap amyloids in Pseudomonas aeruginosa, P. fluorescens, and P. putida results in aggregation and increased biofilm formation

    PubMed Central

    Dueholm, Morten S; Søndergaard, Mads T; Nilsson, Martin; Christiansen, Gunna; Stensballe, Allan; Overgaard, Michael T; Givskov, Michael; Tolker-Nielsen, Tim; Otzen, Daniel E; Nielsen, Per H

    2013-01-01

    The fap operon, encoding functional amyloids in Pseudomonas (Fap), is present in most pseudomonads, but so far the expression and importance for biofilm formation has only been investigated for P. fluorescens strain UK4. In this study, we demonstrate the capacity of P. aeruginosa PAO1, P. fluorescens Pf-5, and P. putida F1 to express Fap fibrils, and investigated the effect of Fap expression on aggregation and biofilm formation. The fap operon in all three Pseudomonas species conferred the ability to express Fap fibrils as shown using a recombinant approach. This Fap overexpression consistently resulted in highly aggregative phenotypes and in increased biofilm formation. Detailed biophysical investigations of purified fibrils confirmed FapC as the main fibril monomer and supported the role of FapB as a minor, nucleating constituent as also indicated by bioinformatic analysis. Bioinformatics analysis suggested FapF and FapD as a potential β-barrel membrane pore and protease, respectively. Manipulation of the fap operon showed that FapA affects monomer composition of the final amyloid fibril, and that FapB is an amyloid protein, probably a nucleator for FapC polymerization. Our study highlights the fap operon as a molecular machine for functional amyloid formation. PMID:23504942

  5. In vitro management of hospital Pseudomonas aeruginosa biofilm using indigenous T7-like lytic phage.

    PubMed

    Ahiwale, Sangeeta; Tamboli, Nilofer; Thorat, Kiran; Kulkarni, Rajendra; Ackermann, Hans; Kapadnis, Balasaheb

    2011-02-01

    Pseudomonas aeruginosa, a human pathogen capable of forming biofilm and contaminating medical settings, is responsible for 65% mortality in the hospitals all over the world. This study was undertaken to isolate lytic phages against biofilm forming Ps. aeruginosa hospital isolates and to use them for in vitro management of biofilms in the microtiter plate. Multidrug resistant strains of Ps. aeruginosa were isolated from the hospital environment in and around Pimpri-Chinchwad, Maharashtra by standard microbiological methods. Lytic phages against these strains were isolated from the Pavana river water by double agar layer plaque assay method. A wide host range phage bacterial virus Ps. aeruginosa phage (BVPaP-3) was selected. Electron microscopy revealed that BVPaP-3 phage is a T7-like phage and is a relative of phage species gh-1. A phage at MOI-0.001 could prevent biofilm formation by Ps. aeruginosa hospital strain-6(HS6) on the pegs within 24 h. It could also disperse pre-formed biofilms of all hospital isolates (HS1-HS6) on the pegs within 24 h. Dispersion of biofilm was studied by monitoring log percent reduction in cfu and log percent increase in pfu of respective bacterium and phage on the peg as well as in the well. Scanning electron microscopy confirmed that phage BVPaP-3 indeed causes biofilm reduction and bacterial cell killing. Laboratory studies prove that BVPaP-3 is a highly efficient phage in preventing and dispersing biofilms of Ps. aeruginosa. Phage BVPaP-3 can be used as biological disinfectant to control biofilm problem in medical devices.

  6. Recruitment of a chromosomally encoded maleylacetate reductase for degradation of 2,4-dichlorophenoxyacetic acid by plasmid pJP4.

    PubMed Central

    Kukor, J J; Olsen, R H; Siak, J S

    1989-01-01

    When Pseudomonas aeruginosa PAO1c or P. putida PPO200 or PPO300 carry plasmid pJP4, which encodes enzymes for the degradation of 2,4-dichlorophenoxyacetic acid (TFD) to 2-chloromaleylacetate, cells do not grow on TFD and UV-absorbing material with spectral characteristics of chloromaleylacetate accumulates in the culture medium. Using plasmid pRO1727, we cloned from the chromosome of a nonfluorescent pseudomonad, Pseudomonas sp. strain PKO1, 6- and 0.5-kilobase BamHI DNA fragments which contain the gene for maleylacetate reductase. When carrying either of the recombinant plasmids, pRO1944 or pRO1945, together with pJP4, cells of P. aeruginosa or P. putida were able to utilize TFD as a sole carbon source for growth. A novel polypeptide with an estimated molecular weight of 18,000 was detected in cell extracts of P. aeruginosa carrying either plasmid pRO1944 or plasmid pRO1945. Maleylacetate reductase activity was induced in cells of P. aeruginosa or P. putida carrying plasmid pRO1945, as well as in cells of Pseudomonas strain PKO1, when grown on L-tyrosine, suggesting that the tyrosine catabolic pathway might be the source from which maleylacetate reductase is recruited for the degradation of TFD in pJP4-bearing cells of Pseudomonas sp. strain PKO1. Images PMID:2722753

  7. Antibacterial properties of Chinese herbal medicines against nosocomial antibiotic resistant strains of Pseudomonas aeruginosa in Taiwan.

    PubMed

    Liu, Ching-Shen; Cham, Thau-Ming; Yang, Cheng-Hong; Chang, Hsueh-Wei; Chen, Chia-Hong; Chuang, Li-Yeh

    2007-01-01

    Pseudomonas aeruginosa is well-recognized as a nosocomial pathogen, which exhibits inherent drug resistance. In this study, the antibacterial activity of ethanol extracts of 58 Chinese herbal medicines used in Taiwan were tested against 89 nosocomial antibiotic resistant strains of Pseudomonas aeruginosa. The results gathered by the disc diffusion method showed that 26 out of the 58 herbal extracts exhibited antibacterial activity. Among the 26 herbal extracts, 10 extracts showed broad-spectrum antibacterial activities and were selected for further antibacterial property assay. The minimum inhibitory concentrations (MIC) of the active partition fractions ranged from 0.25 to 11.0 mg/L. The presence of flavonoid compounds in the active fractions of test herbal extracts was observed by the TLC-bioautography. The results from the time-kill assay revealed that most of the herbal extracts completely killed the test organisms within 4 hours. Exposure of the test strains to a sub-MIC level of the herbal extracts for 10 consecutive subcultures did not induce resistance to the active components. A combination of the active herbal fractions with antibiotics showed that one of the herbal medicines, the hexane fraction of Ramulus Cinnamomi, possessed a synergistic effect with tetracycline, gentamycin, and streptomycin. In conclusion, the tested Chinese medical herbs have the potential to be developed into natural antibiotics. This is the first evaluation for screening large amounts of medical plants against nosocomial antibiotic resistant bacteria in Taiwan.

  8. Pseudomonas aeruginosa clinical and environmental isolates constitute a single population with high phenotypic diversity

    PubMed Central

    2014-01-01

    Background Pseudomonas aeruginosa is an opportunistic pathogen with a high incidence of hospital infections that represents a threat to immune compromised patients. Genomic studies have shown that, in contrast to other pathogenic bacteria, clinical and environmental isolates do not show particular genomic differences. In addition, genetic variability of all the P. aeruginosa strains whose genomes have been sequenced is extremely low. This low genomic variability might be explained if clinical strains constitute a subpopulation of this bacterial species present in environments that are close to human populations, which preferentially produce virulence associated traits. Results In this work, we sequenced the genomes and performed phenotypic descriptions for four non-human P. aeruginosa isolates collected from a plant, the ocean, a water-spring, and from dolphin stomach. We show that the four strains are phenotypically diverse and that this is not reflected in genomic variability, since their genomes are almost identical. Furthermore, we performed a detailed comparative genomic analysis of the four strains studied in this work with the thirteen previously reported P. aeruginosa genomes by means of describing their core and pan-genomes. Conclusions Contrary to what has been described for other bacteria we have found that the P. aeruginosa core genome is constituted by a high proportion of genes and that its pan-genome is thus relatively small. Considering the high degree of genomic conservation between isolates of P. aeruginosa from diverse environments, including human tissues, some implications for the treatment of infections are discussed. This work also represents a methodological contribution for the genomic study of P. aeruginosa, since we provide a database of the comparison of all the proteins encoded by the seventeen strains analyzed. PMID:24773920

  9. Control of Attachment of Pseudomonas aeruginosa and Burkholderia cepacia to Surfaces by Shear Force.

    PubMed

    Hui, Yew Woh; Narayanan, Kumaran; Dykes, Gary A

    2016-11-01

      The effect of physical shearing on the attachment of six Pseudomonas aeruginosa strains and six Burkholderia cepacia strains to glass, stainless steel, polystyrene and Teflon® was determined. A significant (p < 0.05) decrease in hydrophobicity was apparent for all P. aeruginosa strains (17-36%) and B. cepacia, MS 5 (20%) after shearing. A significant (p < 0.05) decrease in attachment of some P. aeruginosa (0.2-0.5 log CFU/cm2) and B. cepacia (0.2-0.4 log CFU/cm2) strains to some surface types was apparent after shearing. Significant (p < 0.05) correlation was observed for both numbers of flagellated cells and hydrophobicity against attachment to glass, stainless steel and polystyrene for P. aeruginosa while only hydrophobicity showed significant correlation against the same surfaces for B. cepacia. Scanning electron microscopy and protein analysis showed that shearing removed surface proteins from the cells and may have led to the observed changes in hydrophobicity and attachment to abiotic surfaces.

  10. SERS detection of the biomarker hydrogen cyanide from Pseudomonas aeruginosa cultures isolated from cystic fibrosis patients

    NASA Astrophysics Data System (ADS)

    Lauridsen, Rikke Kragh; Sommer, Lea M.; Johansen, Helle Krogh; Rindzevicius, Tomas; Molin, Søren; Jelsbak, Lars; Engelsen, Søren Balling; Boisen, Anja

    2017-03-01

    Pseudomonas aeruginosa is the primary cause of chronic airway infections in cystic fibrosis (CF) patients. Persistent infections are seen from the first P. aeruginosa culture in about 75% of young CF patients, and it is important to discover new ways to detect P. aeruginosa at an earlier stage. The P. aeruginosa biomarker hydrogen cyanide (HCN) contains a triple bond, which is utilized in this study because of the resulting characteristic C≡N peak at 2135 cm-1 in a Raman spectrum. The Raman signal was enhanced by surface-enhanced Raman spectroscopy (SERS) on a Au-coated SERS substrate. After long-term infection, a mutation in the patho-adaptive lasR gene can alter the expression of HCN, which is why it is sometimes not possible to detect HCN in the breath of chronically infected patients. Four P. aeruginosa reference strains and 12 clinical P. aeruginosa strains isolated from CF children were evaluated, and HCN was clearly detected from overnight cultures of all wild type-like isolates and half of the later isolates from the same patients. The clinical impact could be that P. aeruginosa infections could be detected at an earlier stage, because daily breath sampling with an immediate output could be possible with a point-of-care SERS device.

  11. Antibiotic resistance pattern and evaluation of metallo-beta lactamase genes (VIM and IMP) in Pseudomonas aeruginosa strains producing MBL enzyme, isolated from patients with secondary immunodeficiency.

    PubMed

    Shirani, Kiana; Ataei, Behrouz; Roshandel, Fardad

    2016-01-01

    One of the most common causes of hospital-acquired secondary infections in hospitalized patients is Pseudomonas aeruginosa. The aim of this study is to evaluate the expression of IMP and VIM in Pseudomonas aeruginosa strains (carbapenem resistant and producer MBL enzyme) in patients with secondary immunodeficiency. In a cross sectional study, 96 patients with secondary immunodeficiency hospitalized in the Al-Zahra hospital were selected. Carbapenem resistant strains isolated and modified Hodge test was performed in order to confirm the presence of the metallo carbapenemase enzyme. Under the standard conditions they were sent to the central laboratory for investigating nosocomial infection Multiplex PCR. Of 96 samples 28.1% were IMP positive, 5.2% VIM positive and 3.1% both VIM and IMP positive. The prevalence of multidrug resistance in the IMP and/or VIM negative samples was 29%, while all 5 VIM positive samples have had multidrug resistance. Also the prevalence of multi-drug resistance in IMP positive samples were 96.3% and in IMP and VIM positive samples were 100%. According to Fisher's test, the prevalence of multi-drug resistance based on gene expression has significant difference (P < 0.001). Based on the results of this study it can be concluded that, a significant percentage of patients with secondary immunodeficiency that suffer nosocomial infections with multidrug resistance, especially Pseudomonas aeruginosa, are probably MBL-producing gene positive. Therefore the cause of infection should be considered in the hospital care system to identify their features, the presence of genes involved in the development of multi-drug resistance and antibiotic therapy.

  12. Beta-Lactamases Produced by a Pseudomonas aeruginosa Strain Highly Resistant to Carbenicillin

    PubMed Central

    Labia, Roger; Guionie, Marlène; Masson, Jean-Michel; Philippon, Alain; Barthelemy, Michel

    1977-01-01

    A Pseudomonas aeruginosa strain isolated at Besançon Hospital, France, proved to be highly resistant to carbenicillin and showed a high hydrolytic activity toward this antibiotic. We clearly demonstrated that two β-lactamases were synthetized: one of them, constitutive, has its enzymatic activity directed mainly toward penicillins, and carbenicillin appears to be its best substrate (higher Vmax); thus, this β-lactamase is a “carbenicillinase” that differs from the well-known “TEM-like” enzymes. The isoelectric point of this carbenicillinase is 5.30 ± 0.03. The other one is an inducible cephalosporinase, very similar to the cephalosporinases usually found in these organisms. Its isoelectric point is 8.66 ± 0.04. These two enzymes have been separated by affinity chromatography and isoelectric focusing. The kinetic constants were measured by computerized microacidimetry. Images PMID:406828

  13. Long-term clinical outcomes of 'Prairie Epidemic Strain' Pseudomonas aeruginosa infection in adults with cystic fibrosis.

    PubMed

    Somayaji, Ranjani; Lam, John C; Surette, Michael G; Waddell, Barbara; Rabin, Harvey R; Sibley, Christopher D; Purighalla, Swathi; Parkins, Michael D

    2017-04-01

    Epidemic Pseudomonas aeruginosa (PA) plays an important role in cystic fibrosis (CF) lung disease. A novel strain, the 'Prairie Epidemic Strain' (PES), has been identified in up to 30% of patients in Prairie-based Canadian CF centres. To determine the incidence, prevalence and long-term clinical impact of PES infection. A cohort of adults with CF was followed from 1980 to 2014 where bacteria isolated from clinical encounters were prospectively collected. Strain typing was performed using pulse-field gel electrophoresis and multilocus sequence typing. Patients were divided into one of four cohorts: no PA, transient PA, chronic PA with unique strains and chronic PES. Proportional Cox hazard and linear mixed models were used to assess for CF-associated respiratory death or transplantation, and rates of %FEV 1 and body mass index (BMI) decline. 274 patients (51.7% male) were analysed: 44--no PA, 29--transient PA, 137--unique PA, 64--PES. A total of 92 patients (33.6%) died or underwent lung transplantation (2423.0 patient-years). PES infection was associated with greater risk of respiratory death or lung transplant compared with the no PA group (aHR, 3.94 (95% CI 1.18 to 13.1); p=0.03) and unique PA group (aHR, 1.75 (95% CI 1.05 to 2.92) p=0.03). Rate of lung function decline (%FEV 1 predicted) was greatest in the PES group (1.73%/year (95% CI 1.63% to 1.82%); p<0.001). BMI improved over time but at an attenuated rate in the PES group (p=0.001). Infection with PES was associated with increased patient morbidity through three decades and manifested in an increased risk of respiratory death and/or lung transplantation. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/.

  14. Application of protein typing in molecular epidemiological investigation of nosocomial infection outbreak of aminoglycoside-resistant Pseudomonas aeruginosa.

    PubMed

    Song, Min; Tang, Min; Ding, Yinghuan; Wu, Zecai; Xiang, Chengyu; Yang, Kui; Zhang, Zhang; Li, Baolin; Deng, Zhenghua; Liu, Jinbo

    2017-12-16

    Pseudomonas aeruginosan has emerged as an important pathogen elated to serious infections and nosocomial outbreaks worldwide. This study was conducted to understand the prevalence of aminoglycoside (AMG)-resistant P. aeruginosa in our hospital and to provide a scientific basis for control measures against nosocomial infections. Eighty-two strains of P. aeruginosa were isolated from clinical departments and divided into AMG-resistant strains and AMG-sensitive strains based on susceptibility test results. AMG-resistant strains were typed by drug resistance gene typing (DRGT) and protein typing. Five kinds of aminoglycoside-modifying enzyme (AME) genes were detected in the AMG-resistant group. AMG-resistant P. aeruginosa strains were classified into three types and six subtypes by DRGT. Four protein peaks, namely, 9900.02, 7600.04, 9101.25 and 10,372.87 Da, were significantly and differentially expressed between the two groups. AMG-resistant P. aeruginosa strains were also categorised into three types and six subtypes at the distance level of 10 by protein typing. AMG-resistant P. aeruginosa was cloned spread in our hospital; the timely implementation of nosocomial infection prevention and control strategies were needed in preventing outbreaks and epidemic of AMG-resistant P. aeruginosa. SELDI-TOF MS technology can be used for bacterial typing, which provides a new method of clinical epidemiological survey and nosocomial infection control.

  15. Dissecting the machinery that introduces disulfide bonds in Pseudomonas aeruginosa.

    PubMed

    Arts, Isabelle S; Ball, Geneviève; Leverrier, Pauline; Garvis, Steven; Nicolaes, Valérie; Vertommen, Didier; Ize, Bérengère; Tamu Dufe, Veronica; Messens, Joris; Voulhoux, Romé; Collet, Jean-François

    2013-12-10

    Disulfide bond formation is required for the folding of many bacterial virulence factors. However, whereas the Escherichia coli disulfide bond-forming system is well characterized, not much is known on the pathways that oxidatively fold proteins in pathogenic bacteria. Here, we report the detailed unraveling of the pathway that introduces disulfide bonds in the periplasm of the human pathogen Pseudomonas aeruginosa. The genome of P. aeruginosa uniquely encodes two DsbA proteins (P. aeruginosa DsbA1 [PaDsbA1] and PaDsbA2) and two DsbB proteins (PaDsbB1 and PaDsbB2). We found that PaDsbA1, the primary donor of disulfide bonds to secreted proteins, is maintained oxidized in vivo by both PaDsbB1 and PaDsbB2. In vitro reconstitution of the pathway confirms that both PaDsbB1 and PaDsbB2 shuttle electrons from PaDsbA1 to membrane-bound quinones. Accordingly, deletion of both P. aeruginosa dsbB1 (PadsbB1) and PadsbB2 is required to prevent the folding of several P. aeruginosa virulence factors and to lead to a significant decrease in pathogenicity. Using a high-throughput proteomic approach, we also analyzed the impact of PadsbA1 deletion on the global periplasmic proteome of P. aeruginosa, which allowed us to identify more than 20 new potential substrates of this major oxidoreductase. Finally, we report the biochemical and structural characterization of PaDsbA2, a highly oxidizing oxidoreductase, which seems to be expressed under specific conditions. By fully dissecting the machinery that introduces disulfide bonds in P. aeruginosa, our work opens the way to the design of novel antibacterial molecules able to disarm this pathogen by preventing the proper assembly of its arsenal of virulence factors. The human pathogen Pseudomonas aeruginosa causes life-threatening infections in immunodepressed and cystic fibrosis patients. The emergence of P. aeruginosa strains resistant to all of the available antibacterial agents calls for the urgent development of new antibiotics

  16. Bacterial infection and acute lung injury in hamsters.

    PubMed

    Seidenfeld, J J; Mullins, R C; Fowler, S R; Johanson, W G

    1986-07-01

    Bacterial pneumonia is a common complication of lung injury that can be an important determinant of outcome. We studied experimental lung injury produced in hamsters by injecting 20 mg/kg paraquat (PQ) intraperitoneally; control animals received saline vehicle. Three days later, Pseudomonas aeruginosa (PAO1), 10(8) organisms in 0.25 ml, or saline, 0.25 ml, was inoculated intratracheally. Lung and systemic antibacterial defenses were studied at death 24 h later. Paraquat alone produced focal interstitial pneumonitis and neutrophilic alveolitis, and resulted in a 12% (3 of 26) mortality. PAO1 alone caused focal pneumonias and no deaths. Animals receiving both agents (PAO1/PQ) had extensive diffuse alveolar damage characterized by alveolar hemorrhage, edema, influx of neutrophils, and vasculitis; 50% (16 of 32) died within 96 h of PQ injection. Mean lung counts of PAO1 at death were 7.6 X 10(4) colony forming units/g in PAO1 and 2.8 X 10(7) in PAO1/PQ animals (p less than 0.05). PAO1 colony counts in liver were increased nearly 100-fold in PAO1/PQ animals (p less than 0.05). Half-time of clearance of P. aeruginosa from the blood was prolonged in PAO1 and in PAO1/PQ animals (p less than 0.05) but not in PQ animals. Phagocytosis of Staphylococcus aureus by leukocytes lavaged from the lung was not impaired in any group compared with that in control animals, but intracellular killing was impaired in PAO1 and PAO1/PQ but not in PQ animals. Paraquat injury impairs lung antibacterial defenses by uncertain mechanisms. Superinfection of PQ-injured lungs by PAO1 appears responsible for defects in intrapulmonary and systemic antibacterial defenses.

  17. Antimicrobial Activity of Ceftolozane-Tazobactam Tested against Enterobacteriaceae and Pseudomonas aeruginosa with Various Resistance Patterns Isolated in U.S. Hospitals (2011-2012)

    PubMed Central

    Flamm, Robert K.; Sader, Helio S.; Jones, Ronald N.

    2013-01-01

    Ceftolozane/tazobactam, a novel antimicrobial agent with activity against Pseudomonas aeruginosa (including drug-resistant strains) and other common Gram-negative pathogens (including most extended-spectrum-β-lactamase [ESBL]-producing Enterobacteriaceae strains), and comparator agents were susceptibility tested by a reference broth microdilution method against 7,071 Enterobacteriaceae and 1,971 P. aeruginosa isolates. Isolates were collected consecutively from patients in 32 medical centers across the United States during 2011 to 2012. Overall, 15.7% and 8.9% of P. aeruginosa isolates were classified as multidrug resistant (MDR) and extensively drug resistant (XDR), and 8.4% and 1.2% of Enterobacteriaceae were classified as MDR and XDR. No pandrug-resistant (PDR) Enterobacteriaceae isolates and only one PDR P. aeruginosa isolate were detected. Ceftolozane/tazobactam was the most potent (MIC50/90, 0.5/2 μg/ml) agent tested against P. aeruginosa and demonstrated good activity against 310 MDR strains (MIC50/90, 2/8 μg/ml) and 175 XDR strains (MIC50/90, 4/16 μg/ml). Ceftolozane/tazobactam exhibited high overall activity (MIC50/90, 0.25/1 μg/ml) against Enterobacteriaceae and retained activity (MIC50/90, 4/>32 μg/ml) against many 601 MDR strains but not against the 86 XDR strains (MIC50, >32 μg/ml). Ceftolozane/tazobactam was highly potent (MIC50/90, 0.25/0.5 μg/ml) against 2,691 Escherichia coli isolates and retained good activity against most ESBL-phenotype E. coli isolates (MIC50/90, 0.5/4 μg/ml), but activity was low against ESBL-phenotype Klebsiella pneumoniae isolates (MIC50/90, 32/>32 μg/ml), explained by the high rate (39.8%) of meropenem coresistance observed in this species phenotype. In summary, ceftolozane/tazobactam demonstrated high potency and broad-spectrum activity against many contemporary Enterobacteriaceae and P. aeruginosa isolates collected in U.S. medical centers. Importantly, ceftolozane/tazobactam retained potency against many MDR and

  18. Algicidal activity of Bacillus sp. Lzh-5 and its algicidal compounds against Microcystis aeruginosa.

    PubMed

    Li, Zhenghua; Geng, Mengxin; Yang, Hong

    2015-01-01

    A freshwater algicidal bacterial strain, Lzh-5, isolated from Lake Taihu, with strong algicidal activity against Microcystis aeruginosa, was identified as Bacillus sp. based on its phenotypic characteristics and 16S ribosomal RNA (rRNA) gene sequence. The algicidal mode of Bacillus sp. Lzh-5 was indirect, attacking M. aeruginosa cells by releasing algicidal compounds. Two algicidal compounds (S-5A and S-5B) produced by Bacillus sp. Lzh-5 were purified with ethyl acetate extraction, column chromatography, and high-performance liquid chromatography and identified as hexahydropyrrolo[1,2-a]pyrazine-1,4-dione and 3-isopropyl-hexahydropyrrolo[1,2-a]pyrazine-1,4-dione based on liquid chromatography-mass spectrometry, gas chromatography-mass spectrometry, and nuclear magnetic resonance analyses. The active algicidal compounds S-5A (hexahydropyrrolo[1,2-a]pyrazine-1,4-dione) and S-5B (3-isopropyl-hexahydropyrrolo[1,2-a]pyrazine-1,4-dione) displayed high levels of algicidal activity against M. aeruginosa 9110, with LD50 values of 5.7 and 19.4 μg/ml, respectively. This is the first report of 3-isopropyl-hexahydropyrrolo[1,2-a]pyrazine-1,4-dione as an algicidal compound. Compounds S-5A and S-5B also induced obvious morphological changes in M. aeruginosa 9110. In cocultures of M. aeruginosa 9110 and Bacillus sp. Lzh-5, the cell density of Bacillus sp. Lzh-5 and the concentrations of S-5A and S-5B correlated positively with the algicidal activity. Our results indicate that strain Lzh-5 and its two algicidal compounds are potentially useful for controlling cyanobacterial blooms in Lake Taihu.

  19. Evaluation of the PaO2/FiO2 ratio after cardiac surgery as a predictor of outcome during hospital stay.

    PubMed

    Esteve, Francisco; Lopez-Delgado, Juan C; Javierre, Casimiro; Skaltsa, Konstantina; Carrio, Maria Ll; Rodríguez-Castro, David; Torrado, Herminia; Farrero, Elisabet; Diaz-Prieto, Antonio; Ventura, Josep Ll; Mañez, Rafael

    2014-09-26

    The arterial partial pressure of O2 and the fraction of inspired oxygen (PaO2/FiO2) ratio is widely used in ICUs as an indicator of oxygenation status. Although cardiac surgery and ICU scores can predict mortality, during the first hours after cardiac surgery few instruments are available to assess outcome. The aim of this study was to evaluate the usefulness of PaO2/FIO2 ratio to predict mortality in patients immediately after cardiac surgery. We prospectively studied 2725 consecutive cardiac surgery patients between 2004 and 2009. PaO2/FiO2 ratio was measured on admission and at 3 h, 6 h, 12 h and 24 h after ICU admission, together with clinical data and outcomes. All PaO2/FIO2 ratio measurements differed between survivors and non-survivors (p < 0.001). The PaO2/FIO2 at 3 h after ICU admission was the best predictor of mortality based on area under the curve (p < 0.001) and the optimum threshold estimation gave an optimal cut-off of 222 (95% Confidence interval (CI): 202-242), yielding three groups of patients: Group 1, with PaO2/FIO2 > 242; Group 2, with PaO2/FIO2 from 202 to 242; and Group 3, with PaO2/FIO2 < 202. Group 3 showed higher in-ICU mortality and ICU length of stay and Groups 2 and 3 also showed higher respiratory complication rates. The presence of a PaO2/FIO2 ratio < 202 at 3 h after admission was shown to be a predictor of in-ICU mortality (OR:1.364; 95% CI:1.212-1.625, p < 0.001) and of worse long-term survival (88.8% vs. 95.8%; Log rank p = 0.002. Adjusted Hazard ratio: 1.48; 95% CI:1.293-1.786; p = 0.004). A simple determination of PaO2/FIO2 at 3 h after ICU admission may be useful to identify patients at risk immediately after cardiac surgery.

  20. Molecular epidemiology of Pseudomonas aeruginosa.

    PubMed

    Speert, David P

    2002-10-01

    Pseudomonas aeruginosa is a serious opportunistic pathogen in certain compromised hosts, such as those with cystic fibrosis, thermal burns and cancer. It also causes less severe noninvasive disease, such as otitis externa and hot tub folliculitis, in normal hosts. P. aeruginosa is phenotypically very unstable, particularly in patients with chronic infection. Phenotypic typing techniques are useful for understanding the epidemiology of acute infections, but they are limited by their discriminatory power and by their inability to group isolates that are phenotypically unrelated but genetically homologous. Molecular typing techniques, developed over the past decade, are highly discriminatory and are useful for typing strains from patients with chronic infection where the bacterial phenotype is unstable; this is particularly true in cystic fibrosis, where patients often are infected with the same strain for several decades, but the bacteria undergo phenotypic alteration. Molecular typing techniques, which have proven useful in typing P. aeruginosa for epidemiological purposes, include pulsed field gel electrophoresis, restriction fragment length polymorphic DNA analysis, random amplified polymorphic DNA analysis, repetitive extrapalindromic PCR analysis, and multilocus sequence typing. These methods are generally only available in specialized laboratories, but they should be used when data from phenotypic typing analysis are ambiguous or when phenotypic methods are unreliable, such as in cystic fibrosis.

  1. The Pseudomonas aeruginosa PA14 ABC Transporter NppA1A2BCD Is Required for Uptake of Peptidyl Nucleoside Antibiotics.

    PubMed

    Pletzer, Daniel; Braun, Yvonne; Dubiley, Svetlana; Lafon, Corinne; Köhler, Thilo; Page, Malcolm G P; Mourez, Michael; Severinov, Konstantin; Weingart, Helge

    2015-07-01

    Analysis of the genome sequence of Pseudomonas aeruginosa PA14 revealed the presence of an operon encoding an ABC-type transporter (NppA1A2BCD) showing homology to the Yej transporter of Escherichia coli. The Yej transporter is involved in the uptake of the peptide-nucleotide antibiotic microcin C, a translation inhibitor that targets the enzyme aspartyl-tRNA synthetase. Furthermore, it was recently shown that the Opp transporter from P. aeruginosa PAO1, which is identical to Npp, is required for uptake of the uridyl peptide antibiotic pacidamycin, which targets the enzyme translocase I (MraY), which is involved in peptidoglycan synthesis. We used several approaches to further explore the substrate specificity of the Npp transporter. Assays of growth in defined minimal medium containing peptides of various lengths and amino acid compositions as sole nitrogen sources, as well as Biolog Phenotype MicroArrays, showed that the Npp transporter is not required for di-, tri-, and oligopeptide uptake. Overexpression of the npp operon increased susceptibility not just to pacidamycin but also to nickel chloride and the peptidyl nucleoside antibiotic blasticidin S. Furthermore, heterologous expression of the npp operon in a yej-deficient mutant of E. coli resulted in increased susceptibility to albomycin, a naturally occurring sideromycin with a peptidyl nucleoside antibiotic. Additionally, heterologous expression showed that microcin C is recognized by the P. aeruginosa Npp system. Overall, these results suggest that the NppA1A2BCD transporter is involved in the uptake of peptidyl nucleoside antibiotics by P. aeruginosa PA14. One of the world's most serious health problems is the rise of antibiotic-resistant bacteria. There is a desperate need to find novel antibiotic therapeutics that either act on new biological targets or are able to bypass known resistance mechanisms. Bacterial ABC transporters play an important role in nutrient uptake from the environment. These uptake

  2. [Immunization with Bifidobacterium bifidum-vectored OprI vaccine of Pseudomonas aeruginosa enhances inhibitory effect on P. aeruginosa in mice].

    PubMed

    Liu, Xiao; Li, Wengui

    2017-08-01

    Objective To study the pulmonary bacterial loads, splenocyte proliferation, distributions of T cell subsets and cell apoptosis in mice immunized with Bifidobacterium bifidum-vectored OprI (Bb-OprI) vaccine of Pseudomonas aeruginosa and challenged with P. aeruginosa PA01 strain. Methods BALB/c mice were immunized with 5×10 9 CFUs of vaccine by intragastric administration, 3 times a week for 3 weeks, and challenged intranasally with 5×10 6 CFUs of PA01 strain at the fourth week after the first immunization. At the second week after the challenge, all mice were sacrificed to separate their lungs and spleens, and the pulmonary bacterial loads were counted. The proliferation of the splenocytes was determined by MTT assay. The splenic CD4 + , CD8 + T cell subsets and the apoptotic rate of splenocytes were detected by flow cytometry. Results The number of pulmonary bacterial colonies in the mice immunized with the vaccine and challenged with PA01 strain decreased, while the proliferation of splenocytes and the proportion of CD4 + T cells markedly increased, and the apoptosis of splenocytes was notably reduced. Conclusion The intragastric vaccination of recombinant Bb-OprI vaccine can increase the proportion of CD4 + T cells and enhance the inhibitory effect on P. aeruginosa.

  3. Heavy metals resistant plasmid-mediated utilization of solar by Pseudomonas aeruginosa AA301.

    PubMed

    Abo-Amer, Aly E; Mohamed, Rehab M

    2006-01-01

    Solar-degrading bacteria, Pseudomonas aeruginosa strains, were isolated from Egyptian soil by Mineral Salt Medium (MSM) supplemented with Solar (motor fuel) from different oil-contaminated sites in Sohag province. The strain AA301 of Pseudomonas aeruginosa showed appreciable growth in MSM medium containing high concentrations of Solar ranging from 0.5 to 3% (v/v), with optimum concentration at 1.5%. Solar was used as a sole carbon source and a source of energy by the bacterium. The ability to degrade Solar was found to be associated with a single 60-kb plasmid designated pSOL15. The plasmid-cured variant, which was obtained by culturing in LB broth with kanamycin, lost the plasmid indicative the ability to degrade Solar must depend on this plasmid. The wild type isolate, Pseudomonas aeruginosa AA301 and transformant strain, have maximum growth (OD600 = approximately 2) on Solar, however the plasmid-cured variant did not have any significant growth on Solar. Moreover, resistance to a wide range of heavy metals such as Mn2+, Hg2+, Mg2+, Cd2+, Zn2+, and Ni2+ was also 60-kb plasmid-mediated. Therefore, the strain AA301 could be good candidate for remediation of some heavy metals and oil hydrocarbons in heavily polluted sites.

  4. Efficacy of methanolic extract of green and black teas against extended-spectrum β-Lactamase-producing Pseudomonas aeruginosa.

    PubMed

    Taherpour, Arezou; Hashemi, Ali; Erfanimanesh, Soroor; Taki, Elahe

    2016-07-01

    Pseudomonas aeruginosa is one of the major bacteria causing acute infections. β-Lactamase production is the principal defense mechanism in gram-negative bacteria. The aim of our study was to evaluate the antibacterial activity of Methanolic Extracts of Green and Black Teas on P. aeruginosa Extended Spectrum-β-Lactamases (ESBLs) production. This research was carried out on burn wounds of 245 hospitalized patients in Kerman, Iran. P. aeruginosa ESBLs and MBL producing strains were detected by Combination Disk Diffusion Test (CDDT) and Epsilometer test (E-test) strips, respectively. Minimum inhibitory concentration (MIC) was measured for Ceftazidime, Meropenem, Imipenem, Aztreonam, Cefotaxime and methanollic extracts of Camellia Sinensis (Green Tea). From 245 patients in the burn ward, 120 cases were infected with P. aeruginosa. 41 isolates contained ESBL while MBL was not detected. P. aeruginosa were resistant to Cefotaxime, Aztreonam, Ceftazidime, Meropenem and Imipenem, 72 (60%), 50 (41.66%), 79 (65.83%), 33 (27.5%) and 24 (20%), respectively. Green tea extract had the highest anti-bacterial effect on standard and P. aeruginosa strains in 1.25mg/ml concentration. This study determined that the methanolic extract of green tea has a higher effect against ESBL producing P. aeruginosa than Cefotaxime, Aztreonam and Ceftazidime.

  5. [Molecular epidemiology of beta-lactamases in ceftazidime-resistant Pseudomonas aeruginosa isolates].

    PubMed

    Er, Halil; Altındiş, Mustafa; Aşık, Gülşah; Demir, Cengiz

    2015-04-01

    Pseudomonas aeruginosa is an important opportunistic pathogen that cause mainly nosocomial infections especially in the immunocompromised patients, the elderly and patients with severe burns. The bacterial feature of developing high degree of resistance against several antibiotics leads to increased morbidity and mortality of P.aeruginosa infections. The aims of this study were to investigate the antibiotic susceptibilities of P.aeruginosa strains isolated from hospitalized patients and to determine the presence of resistance enzymes namely PER, GES, KPC, VIM, IMP and OXA. A total of 195 P.aeruginosa strains isolated from different clinical samples (29 sputum, 67 wound, 53 tracheal aspirate, 23 blood, 18 urine, 3 cerebrospinal fluid, 2 pleural fluid) of inpatients (134 male, 61 female) in Afyon Kocatepe University School of Medicine Hospital between 2010-2012, were included in the study. The isolates were identified by conventional methods and automated systems (VITEK 2, BioMerieux, France), and their antibiotic susceptibilities were detected by disk diffusion and E-test methods. Inducible beta-lactamase (IBL), extended-spectrum beta-lactamase (ESBL) and metallo-beta-lactamase (MBL) productions of the isolates were phenotypically investigated by double disk induction, double disk synergy and E-test methods, respectively. The presence of resistance genes encoding PER, GES, KPC, VIM, IMP and OXA enzymes were determined by real-time polymerase chain reaction, and sequence analysis was applied to positive samples. In our study, the antibiotic resistance rates of 195 P.aeruginosa strains were found as follows: ceftazidime 100%, tazobactam/piperacillin 90.8%, aztreonam 60.5%, cefepime 50.2%, imipenem 48.2%, meropenem 47.2%, ofloxacin 47.2%, piperacillin 44.1%, levofloxacin 31.3%, ciprofloxacin 26.2%, gentamicin 11.8%, amikacin 8.7% and tobramycin 6.2%. With the use of phenotypical methods, IBL, ESBL and MBL production rates in the isolates were detected as 89.2% (174

  6. Anti-quorum sensing potential of Adenanthera pavonina

    PubMed Central

    Vasavi, Halkare Suryanarayana; Arun, Ananthapadmanabha Bhagwath; Rekha, Punchappady-Devasya

    2015-01-01

    Background: Quorum sensing (QS) in Pseudomonas aeruginosa plays a key role in virulence factor production, biofilm formation and antimicrobial resistance. Because of emerging antimicrobial resistance in P. aeruginosa, there is a need to find an alternate nonantibiotic agent for the control of infections caused by this organism. Objective: To evaluate anti-QS activity of Adenanthera pavonina L., a medicinal plant used in traditional medicine. Materials and Methods: Preliminary screening for anti-QS activity of ethanol extract of A. pavonina was carried out using Chromobacterium violaceum CV026 biosensor strain and inhibition of QS-regulated violacein production was quantified using C. violaceum ATCC12472. Bioassay guided fractionation of ethanol extract resulted in ethyl acetate fraction (AEF) with strong anti-QS activity and AEF was evaluated for inhibition of QS-regulated pyocyanin production, proteolytic, elastolytic activity, swarming motility and biofilm formation in P. aeruginosa PAO1. Results: AEF, at 0.5 mg/ml, inhibited pyocyanin production completely and at 1 mg/ml of AEF, complete inhibition of proteolytic and elastolytic activities were observed. However, viability of P. aeruginosa PAO1 was not affected at the tested concentrations of AEF as observed by cell count. Swarming motility was inhibited at the concentration of 0.1 mg/ml of AEF. Thin layer chromatography and biosensor overlay of AEF showed violacein inhibition zone at Rf value 0.63. Conclusion: From the results of this study, it can be concluded that A. pavonina extracts can be used as effective anti-QS agents. PMID:25598643

  7. Key role of an ADP - ribose - dependent transcriptional regulator of NAD metabolism for fitness and virulence of Pseudomonas aeruginosa.

    PubMed

    Okon, Elza; Dethlefsen, Sarah; Pelnikevich, Anna; Barneveld, Andrea van; Munder, Antje; Tümmler, Burkhard

    2017-01-01

    NAD is an essential co-factor of redox reactions and metabolic conversions of NAD-dependent enzymes. NAD biosynthesis in the opportunistic pathogen Pseudomonas aeruginosa has yet not been experimentally explored. The in silico search for orthologs in the P. aeruginosa PAO1 genome identified the operon pncA - pncB1-nadE (PA4918-PA4920) to encode the nicotinamidase, nicotinate phosporibosyltransferase and Nad synthase of salvage pathway I. The functional role of the preceding genes PA4917 and PA4916 was resolved by the characterization of recombinant protein. PA4917 turned out to encode the nicotinate mononucleotide adenylyltransferase NadD2 and PA4916 was determined to encode the transcriptional repressor NrtR that binds to an intergenic sequence between nadD2 and pncA. Complex formation between the catalytically inactive Nudix protein NrtR and its DNA binding site was suppressed by the antirepressor ADP-ribose. NrtR plasposon mutagenesis abrogated virulence of P. aeruginosa TBCF10839 in a murine acute airway infection model and constrained its metabolite profile. When grown together with other isogenic plasposon mutants, the nrtR knock-out was most compromised in competitive fitness to persist in nutrient-rich medium in vitro or murine airways in vivo. This example demonstrates how tightly metabolism and virulence can be intertwined by key elements of metabolic control. Copyright © 2016 Elsevier GmbH. All rights reserved.

  8. New options of antibiotic combination therapy for multidrug-resistant Pseudomonas aeruginosa.

    PubMed

    Nakamura, I; Yamaguchi, T; Tsukimori, A; Sato, A; Fukushima, S; Matsumoto, T

    2015-01-01

    Several antibiotic combinations have demonstrated increased activity against multidrug-resistant Pseudomonas aeruginosa (MDRP) in vitro compared with a single antibiotic. The aim of this study was to investigate the activity against MDRP of some aminoglycosides in combination with monobactam, piperacillin (PIPC), and carbapenem. Clinical isolates of MDRP were collected between November 2010 and October 2012 from patients in Tokyo Medical University Hospital, Tokyo (1,015 beds). Our new method was designed to evaluate three concentrations around the breakpoint of each drug using the Checkerboard method. The aminoglycosides tested were amikacin (AMK), tobramycin (TOB), and arbekacin (ABK). Ciprofloxacin, PIPC, and biapenem (BIPM), which have been reported to demonstrate combination effects, were also tested. Sixty-six MDRP strains were identified from the 2,417 P. aeruginosa strains. Of the 66, 27 tested positive for metallo-β-lactamase (MBL). Aztreonam (AZT) with AMK or ABK was the most effective against MDRP. PIPC with AMK or ABK were somewhat effective. AZT with AMK or ABK were more effective against MBL-positive strains than MBL-negative strains. However, PIPC with AMK or ABK were more effective against MBL-negative strains than MBL-positive strains. Combination activities showed differences between MBL-positive and MBL-negative strains.

  9. Draft Genome Sequences of Pseudomonas aeruginosa Isolates from Wounded Military Personnel.

    PubMed

    Arivett, Brock A; Ream, Dave C; Fiester, Steven E; Kidane, Destaalem; Actis, Luis A

    2016-08-11

    Pseudomonas aeruginosa, a Gram-negative bacterium that causes severe hospital-acquired infections, is grouped as an ESKAPE (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter species) pathogen because of its extensive drug resistance phenotypes and effects on human health worldwide. Five multidrug resistant P. aeruginosa strains isolated from wounded military personnel were sequenced and annotated in this work. Copyright © 2016 Arivett et al.

  10. Emergence of Pseudomonas aeruginosa with class 1 integron carrying blaVIM-2 and blaVIM-4 in the University Clinical Hospital of Bialystok (northeastern Poland).

    PubMed

    Michalska-Falkowska, Anna; Sacha, Paweł Tomasz; Grześ, Henryk; Hauschild, Tomasz; Wieczorek, Piotr; Ojdana, Dominika; Tryniszewska, Elżbieta Anna

    2017-07-11

    The effectiveness of carbapenems, considered as last-resort antimicrobials in severe infections, becomes compromised by bacterial resistance. The production of metallo-β-lactamases (MBLs) is the most significant threat to carbapenems activity among Pseudomonas aeruginosa. The aim of this study was to assess the presence and type of MBLs genes in carbapenem-resistant P. aeruginosa clinical strains, to identify the location of MBLs genes and to determine genetic relatedness between MBL-producers using pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST). The first identified MBL-positive (with blaVIM genes) P. aeruginosa strains were isolated from patients hospitalized in the University Clinical Hospital of Bialystok in the period from September 2012 to December 2013. Variants of MBLs genes and variable integron regions were characterized by PCR and sequencing. PFGE was performed after digesting of bacterial genomes by XbaI enzyme. By MLST seven housekeeping genes were analyzed for the determination of sequence type (ST). Three strains carried the blaVIM-2 gene and one harbored the blaVIM-4 gene. The blaVIM genes resided within class 1 integrons. PCR mapping of integrons revealed the presence of four different cassette arrays. Genetic relatedness analysis by PFGE classified VIM-positive strains into four unrelated pulsotypes (A-D). MLST demonstrated the presence of four (ST 111, ST27, and ST17) different sequence type including one previously undescribed new type of ST 2342. Antimicrobial susceptibility testing showed that VIM-positive strains were resistant to carbapenems, cephalosporins, aminoglycosides, and quinolones, intermediate to aztreonam, and susceptible only to colistin. Integrons mapping, PFGE, and MLST results may point to different origin of these strains and independent introduction into hospitalized patients.

  11. RAPD- and ERIC-Based Typing of Clinical and Environmental Pseudomonas aeruginosa Isolates.

    PubMed

    Auda, Ibtesam Ghadban; Al-Kadmy, Israa M S; Kareem, Sawsan Mohammed; Lafta, Aliaa Khyuon; A'Affus, Mustafa Hussein Obeid; Khit, Ibrahim Abd Aloahd; Al Kheraif, Abdulaziz Abdullah; Divakar, Darshan Devang; Ramakrishnaiah, Ravikumar

    2017-03-01

    Pseudomonas aeruginosa is a major cause of nosocomial infection in children and adults, resulting in significant morbidity and mortality due to its ability to acquire drug resistance. The ability of P. aeruginosa in the environment to cause infection in individuals has been reported previously; henceforth, surveillance of the emergence and transmission of P. aeruginosa strains among patients is important for infection control in a clinical setup. Various gene-typing methods have been used for epidemiological typing of P. aeruginosa isolates for the purpose of surveillance. In this work, the suitability and comparability of two typing methods, enterobacterial repetitive intergenic consensus (ERIC)-PCR and random amplification of polymorphic DNA (RAPD)-PCR fingerprinting, were studied to characterize P. aeruginosa strains isolated from clinical and environmental sources. Forty-four clinical and environmental bacterial isolates of P. aeruginosa were collected between October 2015 and January 2016. DNA extraction, ERIC-PCR and RAPD-PCR, agarose gel electrophoresis, and phylogenetic analyses were carried using the unweighted pair-group method with mean. RAPD typing revealed less clonality among clinical isolates, whereas the ERIC method showed greater similarity in comparison with RAPD. Environmental isolates, however, showed greater similarity using RAPD compared with ERIC typing. With only a few exceptions, most clinical isolates were distinct from environmental isolates, irrespective of the typing method. In conclusion, both the RAPD and ERIC typing methods proved to be good tools in understanding clonal diversity. The results also suggest that there is no relationship between clinical and environmental isolates. The absence of clonality among the clinical isolates may indicate that most P. aeruginosa infection cases could be endemic and not epidemic and that endemic infections may be due to nonclonal strains of P. aeruginosa.

  12. Resistance of Pseudomonas aeruginosa Isolates to Hydrogel Contact Lens Disinfection Correlates with Cytotoxic Activity

    PubMed Central

    Lakkis, Carol; Fleiszig, Suzanne M. J.

    2001-01-01

    One of the most common pathogens in infection of hydrogel contact lens wearers is Pseudomonas aeruginosa, which can gain access to the eye via contamination of the lens, lens case, and lens care solutions. Only one strain per species is used in current regulatory testing for the marketing of chemical contact lens disinfectants. The aim of this study was to determine whether P. aeruginosa strains vary in their susceptibility to hydrogel contact lens disinfectants. A method for rapidly screening bacterial susceptibility to contact lens disinfectants was developed, based on measurement of the MIC. The susceptibility of 35 P. aeruginosa isolates to two chemical disinfectants was found to vary among strains. MICs ranged from 6.25 to 100% for both disinfectants at 37°C, and a number of strains were not inhibited by a 100% disinfectant concentration in the lens case environment at room temperature (22°C). Resistance to disinfection appeared to be an inherent rather than acquired trait, since some resistant strains had been isolated prior to the introduction of the disinfectants and some susceptible P. aeruginosa strains could not be made more resistant by repeated disinfectant exposure. A number of P. aeruginosa strains which were comparatively more resistant to short-term disinfectant exposure also demonstrated the ability to grow to levels above the initial inoculum in one chemical disinfectant after long-term (24 to 48 h) disinfectant exposure. Resistance was correlated with acute cytotoxic activity toward corneal epithelial cells and with exsA, which encodes a protein that regulates cytotoxicity via a complex type III secretion system. These results suggest that chemical disinfection solutions may select for contamination with cytotoxic strains. Further investigation of the mechanisms and factors responsible for resistance may also lead to strategies for reducing adverse responses to contact lens wear. PMID:11283074

  13. Resistance of Pseudomonas aeruginosa isolates to hydrogel contact lens disinfection correlates with cytotoxic activity.

    PubMed

    Lakkis, C; Fleiszig, S M

    2001-04-01

    One of the most common pathogens in infection of hydrogel contact lens wearers is Pseudomonas aeruginosa, which can gain access to the eye via contamination of the lens, lens case, and lens care solutions. Only one strain per species is used in current regulatory testing for the marketing of chemical contact lens disinfectants. The aim of this study was to determine whether P. aeruginosa strains vary in their susceptibility to hydrogel contact lens disinfectants. A method for rapidly screening bacterial susceptibility to contact lens disinfectants was developed, based on measurement of the MIC. The susceptibility of 35 P. aeruginosa isolates to two chemical disinfectants was found to vary among strains. MICs ranged from 6.25 to 100% for both disinfectants at 37 degrees C, and a number of strains were not inhibited by a 100% disinfectant concentration in the lens case environment at room temperature (22 degrees C). Resistance to disinfection appeared to be an inherent rather than acquired trait, since some resistant strains had been isolated prior to the introduction of the disinfectants and some susceptible P. aeruginosa strains could not be made more resistant by repeated disinfectant exposure. A number of P. aeruginosa strains which were comparatively more resistant to short-term disinfectant exposure also demonstrated the ability to grow to levels above the initial inoculum in one chemical disinfectant after long-term (24 to 48 h) disinfectant exposure. Resistance was correlated with acute cytotoxic activity toward corneal epithelial cells and with exsA, which encodes a protein that regulates cytotoxicity via a complex type III secretion system. These results suggest that chemical disinfection solutions may select for contamination with cytotoxic strains. Further investigation of the mechanisms and factors responsible for resistance may also lead to strategies for reducing adverse responses to contact lens wear.

  14. Expansion of Antibacterial Spectrum of Muraymycins toward Pseudomonas aeruginosa.

    PubMed

    Takeoka, Yusuke; Tanino, Tetsuya; Sekiguchi, Mitsuaki; Yonezawa, Shuji; Sakagami, Masahiro; Takahashi, Fumiyo; Togame, Hiroko; Tanaka, Yoshikazu; Takemoto, Hiroshi; Ichikawa, Satoshi; Matsuda, Akira

    2014-05-08

    It is urgent to develop novel anti-Pseudomonas agents that should also be active against multidrug resistant P. aeruginosa. Expanding the antibacterial spectrum of muraymycins toward P. aeruginosa was investigated by the systematic structure-activity relationship study. It was revealed that two functional groups, a lipophilic side chain and a guanidino group, at the accessory moiety of muraymycins were important for the anti-Pseudomonas activity, and analogue 29 exhibited antibacterial activity against a range of P. aeruginosa strains with the minimum inhibitory concentration values of 4-8 μg/mL.

  15. Expansion of Antibacterial Spectrum of Muraymycins toward Pseudomonas aeruginosa

    PubMed Central

    2014-01-01

    It is urgent to develop novel anti-Pseudomonas agents that should also be active against multidrug resistant P. aeruginosa. Expanding the antibacterial spectrum of muraymycins toward P. aeruginosa was investigated by the systematic structure–activity relationship study. It was revealed that two functional groups, a lipophilic side chain and a guanidino group, at the accessory moiety of muraymycins were important for the anti-Pseudomonas activity, and analogue 29 exhibited antibacterial activity against a range of P. aeruginosa strains with the minimum inhibitory concentration values of 4–8 μg/mL. PMID:24900879

  16. Antibiotic resistance pattern and evaluation of metallo-beta lactamase genes (VIM and IMP) in Pseudomonas aeruginosa strains producing MBL enzyme, isolated from patients with secondary immunodeficiency

    PubMed Central

    Shirani, Kiana; Ataei, Behrouz; Roshandel, Fardad

    2016-01-01

    Background: One of the most common causes of hospital-acquired secondary infections in hospitalized patients is Pseudomonas aeruginosa. The aim of this study is to evaluate the expression of IMP and VIM in Pseudomonas aeruginosa strains (carbapenem resistant and producer MBL enzyme) in patients with secondary immunodeficiency. Materials and Methods: In a cross sectional study, 96 patients with secondary immunodeficiency hospitalized in the Al-Zahra hospital were selected. Carbapenem resistant strains isolated and modified Hodge test was performed in order to confirm the presence of the metallo carbapenemase enzyme. Under the standard conditions they were sent to the central laboratory for investigating nosocomial infection Multiplex PCR. Results: Of 96 samples 28.1% were IMP positive, 5.2% VIM positive and 3.1% both VIM and IMP positive. The prevalence of multidrug resistance in the IMP and/or VIM negative samples was 29%, while all 5 VIM positive samples have had multidrug resistance. Also the prevalence of multi-drug resistance in IMP positive samples were 96.3% and in IMP and VIM positive samples were 100%. According to Fisher’s test, the prevalence of multi-drug resistance based on gene expression has significant difference (P < 0.001). Conclusion: Based on the results of this study it can be concluded that, a significant percentage of patients with secondary immunodeficiency that suffer nosocomial infections with multidrug resistance, especially Pseudomonas aeruginosa, are probably MBL-producing gene positive. Therefore the cause of infection should be considered in the hospital care system to identify their features, the presence of genes involved in the development of multi-drug resistance and antibiotic therapy. PMID:27563634

  17. A Novel RNase 3/ECP Peptide for Pseudomonas aeruginosa Biofilm Eradication That Combines Antimicrobial, Lipopolysaccharide Binding, and Cell-Agglutinating Activities

    PubMed Central

    Prats-Ejarque, Guillem; Villalba, Clara; Albacar, Marcel; González-López, Juan J.; Torrent, Marc; Moussaoui, Mohammed

    2016-01-01

    Eradication of established biofilm communities of pathogenic Gram-negative species is one of the pending challenges for the development of new antimicrobial agents. In particular, Pseudomonas aeruginosa is one of the main dreaded nosocomial species, with a tendency to form organized microbial communities that offer an enhanced resistance to conventional antibiotics. We describe here an engineered antimicrobial peptide (AMP) which combines bactericidal activity with a high bacterial cell agglutination and lipopolysaccharide (LPS) affinity. The RN3(5-17P22-36) peptide is a 30-mer derived from the eosinophil cationic protein (ECP), a host defense RNase secreted by eosinophils upon infection, with a wide spectrum of antipathogen activity. The protein displays high biofilm eradication activity that is not dependent on its RNase catalytic activity, as evaluated by using an active site-defective mutant. On the other hand, the peptide encompasses both the LPS-binding and aggregation-prone regions from the parental protein, which provide the appropriate structural features for the peptide's attachment to the bacterial exopolysaccharide layer and further improved removal of established biofilms. Moreover, the peptide's high cationicity and amphipathicity promote the cell membrane destabilization action. The results are also compared side by side with other reported AMPs effective against either planktonic and/or biofilm forms of Pseudomonas aeruginosa strain PAO1. The ECP and its derived peptide are unique in combining high bactericidal potency and cell agglutination activity, achieving effective biofilm eradication at a low micromolar range. We conclude that the designed RN3(5-17P22-36) peptide is a promising lead candidate against Gram-negative biofilms. PMID:27527084

  18. Glycolipid-Dependent, Protease Sensitive Internalization of Pseudomonas aeruginosa Into Cultured Human Respiratory Epithelial Cells

    PubMed Central

    Emam, Aufaugh; Carter, William G; Lingwood, Clifford

    2010-01-01

    Internalization of PAK strain Pseudomonas aeruginosa into human respiratory epithelial cell lines and HeLa cervical cancer cells in vitro was readily demonstrable via a gentamycin protection assay. Depletion of target cell glycosphingolipids (GSLs) using a glucosyl ceramide synthase inhibitor, P4, completely prevented P. aeruginosa internalization. In contrast, P4 treatment had no effect on the internalization of Salmonella typhimurium into HeLa cells. Internalized P. aeruginosa were within membrane vacuoles, often containing microvesicles, between the bacterium and the limiting membrane. P. aeruginosa internalization was markedly enhanced by target cell pretreatment with the exogenous GSL, deacetyl gangliotetraosyl ceramide (Gg4). Gg4 binds the lipid raft marker, GM1 ganglioside. Target cell pretreatment with TLCK, but not other (serine) protease inhibitors, prevented both P. aeruginosa host cell binding and internalization. NFkB inhibition also prevented internalization. A GSL-containing lipid-raft model of P. aeruginosa host cell binding/internalization is proposed PMID:21270937

  19. Enhancement of Rhamnolipid Production in Residual Soybean Oil by an Isolated Strain of Pseudomonas aeruginosa

    NASA Astrophysics Data System (ADS)

    de Lima, C. J. B.; França, F. P.; Sérvulo, E. F. C.; Resende, M. M.; Cardoso, V. L.

    In the present work, the production of rhamnolipid from residual soybean oil (RSO) from food frying facilities was studied using a strain of Pseudomonas aeruginosa of contaminated lagoon, isolated from a hydrocarbon contaminated soil. The optimization of RSO, amonium nitrate, and brewery residual yeast concentrations was accomplished by a central composite experimental design and surface response analysis. The experiments were performed in 500-mL Erlenmeyer flasks containing 50mL of mineral medium, at 170 rpm and 30±1°C, for a 48-h fermentation period. Rhamnolipid production has been monitored by measurements of surface tension, rhamnose concentration, and emulsifying activity. The best-planned results, located on the central point, have corresponded to 22g/L of RSO, 5.625 g/ L of NH4NO3' and 11.5 g/L of brewery yeast. At the maximum point the values for rhamnose and emulsifying index were 2.2g/L and 100%, respectively.

  20. The consistent differential expression of genetic pathways following exposure of an industrial Pseudomonas aeruginosa strain to preservatives and a laundry detergent formulation.

    PubMed

    Green, Angharad E; Amézquita, Alejandro; Le Marc, Yvan; Bull, Matthew J; Connor, Thomas R; Mahenthiralingam, Eshwar

    2018-05-01

    Pseudomonas aeruginosa is a common contaminant associated with product recalls in the home and personal care industry. Preservation systems are used to prevent spoilage and protect consumers, but greater knowledge is needed of preservative resistance mechanisms used by P. aeruginosa contaminants. We aimed to identify genetic pathways associated with preservative exposure by using an industrial P. aeruginosa strain and implementing RNA-Seq to understand gene expression changes in response to industry relevant conditions. The consistent differential expression of five genetic pathways during exposure to multiple industrial growth conditions associated with benzisothiazolone (BIT) and phenoxyethanol (POE) preservatives, and a laundry detergent (LD) formulation, was observed. A MexPQ-OpmE Resistance Nodulation Division efflux pump system was commonly upregulated in response to POE, a combination of BIT and POE, and LD together with BIT. In response to all industry conditions, a putative sialic acid transporter and isoprenoid biosynthesis gnyRDBHAL operon demonstrated consistent upregulation. Two operons phnBA and pqsEDCBA involved in Pseudomonas quinolone signaling production and quorum-sensing were also consistently downregulated during exposure to all the industry conditions. The ability to identify consistently differentially expressed genetic pathways in P. aeruginosa can inform the development of future targeted preservation systems that maintain product safety and minimise resistance development.

  1. The consistent differential expression of genetic pathways following exposure of an industrial Pseudomonas aeruginosa strain to preservatives and a laundry detergent formulation

    PubMed Central

    Amézquita, Alejandro; Le Marc, Yvan; Bull, Matthew J; Connor, Thomas R; Mahenthiralingam, Eshwar

    2018-01-01

    Abstract Pseudomonas aeruginosa is a common contaminant associated with product recalls in the home and personal care industry. Preservation systems are used to prevent spoilage and protect consumers, but greater knowledge is needed of preservative resistance mechanisms used by P. aeruginosa contaminants. We aimed to identify genetic pathways associated with preservative exposure by using an industrial P. aeruginosa strain and implementing RNA-Seq to understand gene expression changes in response to industry relevant conditions. The consistent differential expression of five genetic pathways during exposure to multiple industrial growth conditions associated with benzisothiazolone (BIT) and phenoxyethanol (POE) preservatives, and a laundry detergent (LD) formulation, was observed. A MexPQ-OpmE Resistance Nodulation Division efflux pump system was commonly upregulated in response to POE, a combination of BIT and POE, and LD together with BIT. In response to all industry conditions, a putative sialic acid transporter and isoprenoid biosynthesis gnyRDBHAL operon demonstrated consistent upregulation. Two operons phnBA and pqsEDCBA involved in Pseudomonas quinolone signaling production and quorum-sensing were also consistently downregulated during exposure to all the industry conditions. The ability to identify consistently differentially expressed genetic pathways in P. aeruginosa can inform the development of future targeted preservation systems that maintain product safety and minimise resistance development. PMID:29548026

  2. Investigation of a pseudo-outbreak of orthopedic infections caused by Pseudomonas aeruginosa.

    PubMed

    Forman, W; Axelrod, P; St John, K; Kostman, J; Khater, C; Woodwell, J; Vitagliano, R; Truant, A; Satishchandran, V; Fekete, T

    1994-10-01

    To report a pseudoepidemic of Pseudomonas aeruginosa infections discovered during an investigation of postoperative joint infections. A retrospective review of case patients' hospital charts, operative reports, and laboratory data, as well as environmental culturing, polymerase chain reaction (PCR) ribotyping of outbreak isolates, and in vitro analysis of P aeruginosa growth characteristics. A 510-bed, university-affiliated adult tertiary care hospital. Between October 1 and December 1, 1992, seven postsurgical joint infections were diagnosed, including four caused by P aeruginosa. A bottle of "sterile" saline used to process tissue specimens was found to be contaminated with P aeruginosa. Further investigation revealed that P aeruginosa had grown from seven additional tissue cultures, all of which had been processed with the contaminated saline. PCR ribotypes of the contaminant matched those of the clinical isolates. In vitro, P aeruginosa strains were viable in commercial nonbacteriostatic saline, but never caused visible turbidity. Six patients received antibiotics for their presumed infections; four patients had peripherally inserted central catheters placed, and one experienced severe anaphylactic reactions to several antibiotics. Pseudoepidemics due to common organisms are often difficult to detect, and delayed recognition can result in substantial morbidity. This outbreak investigation illustrates the potential for contamination of diluents in the microbiology laboratory and emphasizes the need for meticulous quality control.

  3. Persistence and Epidemic Propagation of a Pseudomonas aeruginosa Sequence Type 235 Clone Harboring an IS26 Composite Transposon Carrying the blaIMP-1 Integron in Hiroshima, Japan, 2005 to 2012

    PubMed Central

    Shimizu, Wataru; Kayama, Shizuo; Kouda, Shuntaro; Ogura, Yoshitoshi; Kobayashi, Kanao; Shigemoto, Norifumi; Shimada, Norimitsu; Yano, Raita; Hisatsune, Junzo; Kato, Fuminori; Hayashi, Tetsuya; Sueda, Taijiro; Ohge, Hiroki

    2015-01-01

    A 9-year surveillance for multidrug-resistant (MDR) Pseudomonas aeruginosa in the Hiroshima region showed that the number of isolates harboring the metallo-β-lactamase gene blaIMP-1 abruptly increased after 2004, recorded the highest peak in 2006, and showed a tendency to decline afterwards, indicating a history of an epidemic. PCR mapping of the variable regions of the integrons showed that this epidemic was caused by the clonal persistence and propagation of an MDR P. aeruginosa strain harboring the blaIMP-1 gene and an aminoglycoside 6′-N-acetyltransferase gene, aac(6′)-Iae in a class I integron (In113), whose integrase gene intl1 was disrupted by an IS26 insertion. Sequence analysis of the representative strain PA058447 resistance element containing the In113-derived gene cassette array showed that the element forms an IS26 transposon embedded in the chromosome. It has a Tn21 backbone and is composed of two segments sandwiched by three IS26s. In Japan, clonal nationwide expansion of an MDR P. aeruginosa NCGM2.S1 harboring chromosomally encoded In113 with intact intl1 is reported. Multilocus sequence typing and genomic comparison strongly suggest that PA058447 and NCGM2.S1 belong to the same clonal lineage. Moreover, the structures of the resistance element in the two strains are very similar, but the sites of insertion into the chromosome are different. Based on tagging information of the IS26 present in both resistance elements, we suggest that the MDR P. aeruginosa clone causing the epidemic in Hiroshima for the past 9 years originated from a common ancestor genome of PA058447 and NCGM2.S1 through an IS26 insertion into intl1 of In113 and through IS26-mediated genomic rearrangements. PMID:25712351

  4. Evaluating the use of Spectral Induced Conductivity to Detect Biofilm Development within Porous Media

    NASA Astrophysics Data System (ADS)

    Rosier, C. L.; Atekwana, E. A.; Price, A.; Sharma, S.; Patrauchan, M.

    2015-12-01

    Microbial biomass accumulation in subsurface sediments dynamically alters porosity/permeability; factors critical to contaminant transport and management of bioremediation efforts. Current methodologies (i.e. plate counts, tracer/slug tests) offer some understanding of biofilm effect on subsurface hydrology, yet do not provide real-time information regarding biofilm development. Due to these limitations there is interest in assessing the near surface geophysical technique Spectral Induced Polarization (SIP), to measure biofilm formation. Our study assesses the influence of cell density and biofilm production on SIP response. Laboratory experiments monitored changes in SIP, measured colony forming units (CFU), and cellular protein levels on sand packed columns inoculated with either Pseudomonas aeruginosa PAO1 (non-mucoid strain) or Pseudomonas aeruginosa FRD1 (biofilm-overproducing mucoid strain) cells over one month. Confocal laser scanning microscopy (CLSM) and scanning electron microscopy (SEM) were used to confirm the presence of biofilm. Our results indicate that phase and imaginary conductivity remained stable in PAO1 treatments as cell densities and cellular protein levels remained low (1.7x105 CFUml-1; 111 μg ml-1). However, we observed a significant decrease in both phase (0.5 to -0.20 mrad) and imaginary conductivity (0.0 to -3.0x10-5 S m-1) when both cell densities and cellular protein levels increased. In FRD1 treatments we observed an immediate decrease in phase (0.1 mrad) and imaginary conductivity (-2.0x10-6 S m-1) as cell densities were an order of magnitude greater then PAO1 treatments and cellular protein levels surpassed 500 μg ml-1. CLSM and SEM analysis confirmed the presence of biofilm and cells within both PAO1 and FRD1 treatments. Our findings suggest that the ratio of cells to cellular protein production is an important factor influencing both phase and imaginary conductivity response. However, our results are not in agreement with

  5. [Epidemiological profile and antibiotic resistance of Pseudomonas aeruginosa isolates in burn and traumatology center in Tunisia over a three-year period].

    PubMed

    Zoghlami, Ayoub; Kanzari, Lamia; Boukadida, Jalel; Messadi, Amen Allah; Ghanem, Abdelraouef

    2012-11-01

    Pseudomonas aeruginosa is a known opportunistic pathogen frequently causing serious infections in burned patients. To analyze the epidemiological profile of Pseudomonas aeruginosa isolated in a Tunisian burn unit. During a 3-year period (from 01 July 2008 to 30 June 2011), 544 non repetitive strains of P. aeruginosa were isolated from burn patients. Susceptibility to antibiotics was assessed according to CA-SFM guidelines. Serotypes were identified by slide agglutination test using P.aeruginosa O antisera (Biorad). Producing carbapenemase was analyzed for 202 imipenem resistant isolates by EDTA test. Susceptibility testing data were stored in a laboratory data base using whonet 5.3 software. The most frequent sites of isolation were cutaneous infections and blood cultures (83.4%). The percentages of resistant isolates were as follows: ceftazidime: 34%; imipenem: 37.1%, ciprofloxacin: 27.1% and amikacin: 29.6%. The most prevalent serotypes were: 011(51%), 06(17%), 03 (8%), 04(12%), 012(5%). Among the 202 imipenem resistant strains, 58% expressed a metallocarbapenemase. All theses strains were resistant to all tested antibiotics except colistin and belonged to the serotype O11. The dissemination of carbapenemases strains must be contained by implementation of timely identification, strict isolation methods and better hygienic procedures.

  6. Pseudomonas aeruginosa Population Structure Revisited

    PubMed Central

    Pirnay, Jean-Paul; Bilocq, Florence; Pot, Bruno; Cornelis, Pierre; Zizi, Martin; Van Eldere, Johan; Deschaght, Pieter; Vaneechoutte, Mario; Jennes, Serge; Pitt, Tyrone; De Vos, Daniel

    2009-01-01

    At present there are strong indications that Pseudomonas aeruginosa exhibits an epidemic population structure; clinical isolates are indistinguishable from environmental isolates, and they do not exhibit a specific (disease) habitat selection. However, some important issues, such as the worldwide emergence of highly transmissible P. aeruginosa clones among cystic fibrosis (CF) patients and the spread and persistence of multidrug resistant (MDR) strains in hospital wards with high antibiotic pressure, remain contentious. To further investigate the population structure of P. aeruginosa, eight parameters were analyzed and combined for 328 unrelated isolates, collected over the last 125 years from 69 localities in 30 countries on five continents, from diverse clinical (human and animal) and environmental habitats. The analysed parameters were: i) O serotype, ii) Fluorescent Amplified-Fragment Length Polymorphism (FALFP) pattern, nucleotide sequences of outer membrane protein genes, iii) oprI, iv) oprL, v) oprD, vi) pyoverdine receptor gene profile (fpvA type and fpvB prevalence), and prevalence of vii) exoenzyme genes exoS and exoU and viii) group I pilin glycosyltransferase gene tfpO. These traits were combined and analysed using biological data analysis software and visualized in the form of a minimum spanning tree (MST). We revealed a network of relationships between all analyzed parameters and non-congruence between experiments. At the same time we observed several conserved clones, characterized by an almost identical data set. These observations confirm the nonclonal epidemic population structure of P. aeruginosa, a superficially clonal structure with frequent recombinations, in which occasionally highly successful epidemic clones arise. One of these clones is the renown and widespread MDR serotype O12 clone. On the other hand, we found no evidence for a widespread CF transmissible clone. All but one of the 43 analysed CF strains belonged to a ubiquitous P

  7. Electron transfer of Pseudomonas aeruginosa CP1 in electrochemical reduction of nitric oxide.

    PubMed

    Zhou, Shaofeng; Huang, Shaobin; He, Jiaxin; Li, Han; Zhang, Yongqing

    2016-10-01

    This study reports catalytic electro-chemical reduction of nitric oxide (NO) enhanced by Pseudomonas aeruginosa strain CP1. The current generated in the presence of bacteria was 4.36times that in the absence of the bacteria. The strain was able to catalyze electro-chemical reduction of NO via indirect electron transfer with an electrode, revealed by a series of cyclic voltammetry experiments. Soluble electron shuttles secreted into solution by live bacteria were responsible for the catalytic effects. The enhancement of NO reduction was also confirmed by detection of nitrous oxide; the level of this intermediate was 46.4% higher in the presence of bacteria than in controls, illustrated that the electron transfer pathway did not directly reduce nitric oxide to N2. The findings of this study may offer a new model for bioelectrochemical research in the field of NO removal by biocatalysts. Copyright © 2016 Elsevier Ltd. All rights reserved.

  8. Electrochemical sensors for identifying pyocyanin production in clinical Pseudomonas aeruginosa isolates.

    PubMed

    Sismaet, Hunter J; Pinto, Ameet J; Goluch, Edgar D

    2017-11-15

    In clinical practice, delays in obtaining culture results impact patient care and the ability to tailor antibiotic therapy. Despite the advancement of rapid molecular diagnostics, the use of plate cultures inoculated from swab samples continues to be the standard practice in clinical care. Because the inoculation culture process can take between 24 and 48h before a positive identification test can be run, there is an unmet need to develop rapid throughput methods for bacterial identification. Previous work has shown that pyocyanin can be used as a rapid, redox-active biomarker for identifying Pseudomonas aeruginosa in clinical infections. However, further validation is needed to confirm pyocyanin production occurs in all clinical strains of P. aeruginosa. Here, we validate this electrochemical detection strategy using clinical isolates obtained from patients with hospital-acquired infections or with cystic fibrosis. Square-wave voltammetric scans of 94 different clinical P. aeruginosa isolates were taken to measure the concentration of pyocyanin. The results showed that all isolates produced measureable concentrations of pyocyanin with production rates correlated with patient symptoms and comorbidity. Further bioinformatics analysis confirmed that 1649 genetically sequenced strains (99.9%) of P. aeruginosa possess the two genes (PhzM and PhzS) necessary to produce pyocyanin, supporting the specificity of this biomarker. Confirming the production of pyocyanin by all clinically-relevant strains of P. aeruginosa is a significant step towards validating this strategy for rapid, point-of-care diagnostics. Copyright © 2017 Elsevier B.V. All rights reserved.

  9. Highly plastic genome of Microcystis aeruginosa PCC 7806, a ubiquitous toxic freshwater cyanobacterium.

    PubMed

    Frangeul, Lionel; Quillardet, Philippe; Castets, Anne-Marie; Humbert, Jean-François; Matthijs, Hans C P; Cortez, Diego; Tolonen, Andrew; Zhang, Cheng-Cai; Gribaldo, Simonetta; Kehr, Jan-Christoph; Zilliges, Yvonne; Ziemert, Nadine; Becker, Sven; Talla, Emmanuel; Latifi, Amel; Billault, Alain; Lepelletier, Anthony; Dittmann, Elke; Bouchier, Christiane; de Marsac, Nicole Tandeau

    2008-06-05

    The colonial cyanobacterium Microcystis proliferates in a wide range of freshwater ecosystems and is exposed to changing environmental factors during its life cycle. Microcystis blooms are often toxic, potentially fatal to animals and humans, and may cause environmental problems. There has been little investigation of the genomics of these cyanobacteria. Deciphering the 5,172,804 bp sequence of Microcystis aeruginosa PCC 7806 has revealed the high plasticity of its genome: 11.7% DNA repeats containing more than 1,000 bases, 6.8% putative transposases and 21 putative restriction enzymes. Compared to the genomes of other cyanobacterial lineages, strain PCC 7806 contains a large number of atypical genes that may have been acquired by lateral transfers. Metabolic pathways, such as fermentation and a methionine salvage pathway, have been identified, as have genes for programmed cell death that may be related to the rapid disappearance of Microcystis blooms in nature. Analysis of the PCC 7806 genome also reveals striking novel biosynthetic features that might help to elucidate the ecological impact of secondary metabolites and lead to the discovery of novel metabolites for new biotechnological applications. M. aeruginosa and other large cyanobacterial genomes exhibit a rapid loss of synteny in contrast to other microbial genomes. Microcystis aeruginosa PCC 7806 appears to have adopted an evolutionary strategy relying on unusual genome plasticity to adapt to eutrophic freshwater ecosystems, a property shared by another strain of M. aeruginosa (NIES-843). Comparisons of the genomes of PCC 7806 and other cyanobacterial strains indicate that a similar strategy may have also been used by the marine strain Crocosphaera watsonii WH8501 to adapt to other ecological niches, such as oligotrophic open oceans.

  10. Cross-infections with Pseudomonas aeruginosa in patients with cystic fibrosis attending the Warsaw Centre.

    PubMed

    Milczewska, Justyna; Wołkowicz, Tomasz; Zacharczuk, Katarzyna; Kwiatkowska, Monika

    2015-01-01

    1. To assess the prevalence of cross-infections with P. aeruginosa in order to evaluate the epidemiological situation of this infection in patients with cystic fibrosis attending our centre; 2. To correlate the clinical features of the patients carrying a potentially transmissible strain with the entire study group in order to determine the risk factors and possible effects of its acquisition. 170 Pseudomonas aeruginosa strains obtained from the respiratory tract of 75 cystic fibrosis patients attending the Warsaw Centre in 2011 and 2012 were typed using restriction enzyme analysis-pulsed field gel electrophoresis (Spe I restriction enzyme was used). Simultaneously, the information concerning contacts between patients, as well as several clinical data regarding the course of the disease were collected. Twenty four clusters of strains were detected. The main cluster included 49 isolates derived from 21 patients. The other detected clusters included 2 to 12 isolates derived from 1 to 7 patients. Three clusters comprised the isolates derived from three pairs of siblings. There were 15 clusters containing 2 to 7 strains belonging to the same patient. The remaining 24 patients were infected with their own strains, not fitting any clonal group. Several clinical parameters showed that the 21 patients whose strains constituted the main cluster, were in worse clinical condition than the other patients in the study group. Moreover, the total duration of their hospitalizations in order to perform intravenous antibiotic treatment was longer. 1. Frequent hospitalizations of CF patients with a more severe course of the disease seem to be a risk factor of cross-infections with P. aeruginosa. 2. Intensification of measures to prevent cross-infection, such as hygienic precautions, patient segregation, introduction of home intravenous antibiotic therapy programme, as well as further education of patients and their parents should lead to the improvement of the epidemiological

  11. Microcystin production by Microcystis aeruginosa exposed to different stages of herbivorous zooplankton.

    PubMed

    Jang, Min-Ho; Ha, Kyong; Takamura, Noriko

    2008-04-01

    Microcystin (MC) production by four monoclonal Microcystis aeruginosa strains was evaluated in response to infochemicals (indirect exposure) released from different stages of herbivorous zooplankton (neonate/juvenile and adult Daphnia magna and Moina macrocopa). The intracellular MC and extracellular MC concentrations were significantly different among the control and treatments with zooplankton culture media filtrates (p<0.05), and in most cases MC production was significantly higher (p<0.05) in strains exposed to infochemicals released from adult zooplankton rather than those of neonate/juvenile zooplankton in four strains of M. aeruginosa. Compared to intracellular MC (385.0-5598.6microg g(-1)DW), very low concentrations of extracellular MC (9.9-737.6microg ml(-1)) were released, but both showed similar temporal patterns over the course of the experiment. This result might be attributed to the fact that adult zooplankton produced more infochemical signals than equal numbers of smaller juveniles and neonates. It is the first study to provide evidence that MC production might be impacted by infochemicals released from different stages of zooplankton, mediated with physiological characteristics, body size, and feeding habits.

  12. Pseudomonas aeruginosa adapts to octenidine in the laboratory and a simulated clinical setting, leading to increased tolerance to chlorhexidine and other biocides.

    PubMed

    Shepherd, M J; Moore, G; Wand, M E; Sutton, J M; Bock, L J

    2018-03-31

    Octenidine is frequently used for infection prevention in neonatal and burn intensive care units, where Pseudomonas aeruginosa has caused nosocomial outbreaks. To investigate the efficacy and impact of using octenidine against P. aeruginosa. Seven clinical isolates of P. aeruginosa were exposed to increasing concentrations of octenidine over several days. Fitness, minimum bactericidal concentrations after 1 min, 5 min and 24 h, and minimum inhibitory concentrations (MICs) of a variety of antimicrobials were measured for the parental and octenidine-adapted P. aeruginosa strains. Octenidine and chlorhexidine MICs of a population of P. aeruginosa isolated from a hospital drain trap, exposed to a diluted octenidine formulation four times daily for three months, were also tested. Some planktonic cultures of P. aeruginosa survived >50% of the working concentration of an in-use octenidine formulation at the recommended exposure time. Seven strains of P. aeruginosa stably adapted following continuous exposure to increasing concentrations of octenidine. Adaptation increased tolerance to octenidine formulations and chlorhexidine up to 32-fold. In one strain, it also led to increased MICs of antipseudomonal drugs. Subsequent to continuous octenidine exposure of a multi-species community in a simulated clinical setting, up to eight-fold increased tolerance to octenidine and chlorhexidine of P. aeruginosa was also found, which was lost upon removal of octenidine. Incorrect use of octenidine formulations may lead to inadequate decontamination, and even increased tolerance of P. aeruginosa to octenidine, with resulting cross-resistance to other biocides. Crown Copyright © 2018. Published by Elsevier Ltd. All rights reserved.

  13. Mannitol Enhances Antibiotic Sensitivity of Persister Bacteria in Pseudomonas aeruginosa Biofilms

    PubMed Central

    Barraud, Nicolas; Buson, Alberto; Jarolimek, Wolfgang; Rice, Scott A.

    2013-01-01

    The failure of antibiotic therapies to clear Pseudomonas aeruginosa lung infection, the key mortality factor for cystic fibrosis (CF) patients, is partly attributed to the high tolerance of P. aeruginosa biofilms. Mannitol has previously been found to restore aminoglycoside sensitivity in Escherichia coli by generating a proton-motive force (PMF), suggesting a potential new strategy to improve antibiotic therapy and reduce disease progression in CF. Here, we used the commonly prescribed aminoglycoside tobramycin to select for P. aeruginosa persister cells during biofilm growth. Incubation with mannitol (10–40 mM) increased tobramycin sensitivity of persister cells up to 1,000-fold. Addition of mannitol to pre-grown biofilms was able to revert the persister phenotype and improve the efficacy of tobramycin. This effect was blocked by the addition of a PMF inhibitor or in a P. aeruginosa mutant strain unable to metabolise mannitol. Addition of glucose and NaCl at high osmolarity also improved the efficacy of tobramycin although to a lesser extent compared to mannitol. Therefore, the primary effect of mannitol in reverting biofilm associated persister cells appears to be an active, physiological response, associated with a minor contribution of osmotic stress. Mannitol was tested against clinically relevant strains, showing that biofilms containing a subpopulation of persister cells are better killed in the presence of mannitol, but a clinical strain with a high resistance to tobramycin was not affected by mannitol. Overall, these results suggest that in addition to improvements in lung function by facilitating mucus clearance in CF, mannitol also affects antibiotic sensitivity in biofilms and does so through an active, physiological response. PMID:24349568

  14. Pseudomonas aeruginosa outbreak in a pediatric oncology care unit caused by an errant water jet into contaminated siphons.

    PubMed

    Schneider, Henriette; Geginat, Gernot; Hogardt, Michael; Kramer, Alexandra; Dürken, Matthias; Schroten, Horst; Tenenbaum, Tobias

    2012-06-01

    We analyzed an outbreak of invasive infections with an exotoxin U positive Pseudomonas aeruginosa strain within a pediatric oncology care unit. Environmental sampling and molecular characterization of the Pseudomonas aeruginosa strains led to identification of the outbreak source. An errant water jet into the sink within patient rooms was observed. Optimized outbreak management resulted in an abundance of further Pseudomonas aeruginosa infections within the pediatric oncology care unit.

  15. Prevalence of Virulence Genes Among Bulgarian Nosocomial and Cystic Fibrosis Isolates of Pseudomonas Aeruginosa

    PubMed Central

    Mitov, Ivan; Strateva, Tanya; Markova, Boyka

    2010-01-01

    The aim of this study was to evaluate the prevalence of some virulence genes among 202 Pseudomonas aeruginosa isolates from cystic fibrosis (CF) patients (n=42) and non-CF in-patients (n=160) and to analyze the values according to the patient groups, infection localization and antimicrobial resistance. The following frequencies in all studied strains were established: algD (encoding GDP-mannose 6-dehydrogenase AlgD) – 91.1%, pilB (type IV fimbrial biogenesis protein PilB) – 23.8%, nan1 (neuraminidase) – 21.3%, lasB (elastase LasB) – 100%, plcH (haemolytic phospholipase C precursor) – 91.6%, exoS (exoenzyme S) – 62.4%, and exoU (exoenzyme U) – 30.2%. The prevalence of nan1 was significantly higher (P<0.01) in CF isolates (38.1%) than that in non-CF isolates (16.9%). The nan1–positive CF strains were cultured from 16 patients with recurrent lung exacerbations. This study revealed a statistically significant difference (P<0.01) between the portion of multidrug-resistant (MDR) nosocomial P. aeruginosa strains containing a large number (≥5) of virulence genes (38.1%) and the respective part of non-MDR isolates (17.6%). Moreover, pilB, exoU and nan1 manifested a higher spread (P<0.001) among MDR than in non-MDR strains (respectively, 39.1% vs. 13.2%; 40.2% vs. 17.7% and 26.1% vs. 4.4%). In conclusion, the dissemination of nan1 in CF isolates was moderate and correlated with the lower proportion of patients with lung exacerbations. The molecular-genetic detection of this gene may be used as an indirect measure of CF pulmonary disease evolution. Simultaneous determination of virulence factors and antimicrobial resistance is the contemporary approach for examination of the microbiological aspects of infections caused by P. aeruginosa. PMID:24031533

  16. The Effect of Strict Segregation on Pseudomonas aeruginosa in Cystic Fibrosis Patients

    PubMed Central

    van Mansfeld, Rosa; de Vrankrijker, Angelica; Brimicombe, Roland; Heijerman, Harry; Teding van Berkhout, Ferdinand; Spitoni, Cristian; Grave, Sanne; van der Ent, Cornelis; Wolfs, Tom; Willems, Rob; Bonten, Marc

    2016-01-01

    Introduction Segregation of patients with cystic fibrosis (CF) was implemented to prevent chronic infection with epidemic Pseudomonas aeruginosa strains with presumed detrimental clinical effects, but its effectiveness has not been carefully evaluated. Methods The effect of strict segregation on the incidence of P. aeruginosa infection in CF patients was investigated through longitudinal protocolized follow-up of respiratory tract infection before and after segregation. In two nested cross-sectional studies in 2007 and 2011 the P. aeruginosa population structure was investigated and clinical parameters were determined in patients with and without infection with the Dutch epidemic P. aeruginosa clone (ST406). Results Of 784 included patients 315 and 382 were at risk for acquiring chronic P. aeruginosa infection before and after segregation. Acquisition rates were, respectively, 0.14 and 0.05 per 1,000 days at risk (HR: 0.66, 95% CI [0.2548–1.541]; p = 0.28). An exploratory subgroup analysis indicated lower acquisition after segregation in children < 15 years of age (HR: 0.43, 95% CI[0.21–0.95]; p = 0.04). P. aeruginosa population structure did not change after segregation and ST406 was not associated with lung function decline, death or lung transplantation. Conclusions Strict segregation was not associated with a statistically significant lower acquisition of chronic P. aeruginosa infection and ST406 was not associated with adverse clinical outcome. After segregation there were no new acquisitions of ST406. In an unplanned exploratory analysis chronic acquisition of P. aeruginosa was lower after implementation of segregation in patients under 15 years of age. PMID:27280467

  17. Candida albicans and Pseudomonas aeruginosa adhesion on soft contact lenses.

    PubMed

    Onurdağ, Fatma Kaynak; Ozkan, Semiha; Ozgen, Selda; Olmuş, Hülya; Abbasoğlu, Ufuk

    2011-04-01

    In this study it was aimed to determine the adherence of Pseudomonas and Candida to contact lens surfaces, and to determine the difference in adherence between five contact lens types. Biofilm-negative control strains were also used to emphasize the difference between biofilm-positive and biofilm-negative strains in adherence. Five different soft contact lenses were used to investigate the adherence of Pseudomonas aeruginosa and Candida albicans strains. P. aeruginosa ATCC 27853, P. aeruginosa ATCC 10145, C.albicans ATCC 10231 standard strains and C. albicans clinical isolate were included in the study. Slime formation was investigated by two methods; modified Christensen macrotube method, and a modified microtiter plate test. P. aeruginosa and C. albicans slime formation on soft contact lenses was studied in adherence and separation phases. Pseudomonas and Candida suspensions were serially diluted and inoculated to blood agar and sabouraud dextrose agar surfaces respectively. After overnight incubation, the colonies were counted. Sterile unworn contact lenses were used as negative controls, and bacterial and fungal culture suspensions were used as positive controls. The experiments were conducted in three parallel series. The number of adherent Pseudomonas was as follows from high to low in polymacon, etafilcon A, hilafilcon, ocufilcon and lotrafilcon contact lenses respectively. However, the number of adherent yeast were determined higher in lotrafilcon and ocufilcon contact lenses, followed by hilafilcon, etafilcon A and polymacon contact lenses. Biofilm-negative Pseudomonas ATCC standard strain and Candida clinical isolate were used to confirm that the number of adherent cells were lower than the biofilm-positive ones. This study demonstrates that in addition to the contact lens properties, the microorganisms themselves and their interactions with the lens material also play an important role in adherence.

  18. Pseudomonas aeruginosa Genotype Prevalence in Dutch Cystic Fibrosis Patients and Age Dependency of Colonization by Various P. aeruginosa Sequence Types ▿

    PubMed Central

    van Mansfeld, Rosa; Willems, Rob; Brimicombe, Roland; Heijerman, Harry; van Berkhout, Ferdinand Teding; Wolfs, Tom; van der Ent, Cornelis; Bonten, Marc

    2009-01-01

    The patient-to-patient transmission of highly prevalent Pseudomonas aeruginosa clones which are associated with enhanced disease progression has led to strict segregation policies for cystic fibrosis (CF) patients in many countries. However, little is known about the population structure of P. aeruginosa among CF patients. The aim of the present cross-sectional study was to determine the prevalence and genetic relatedness of P. aeruginosa isolates from CF patients who visited two major CF centers in The Netherlands in 2007 and 2008. These patients represented 45% of the Dutch CF population. P. aeruginosa carriage in the respiratory tract was determined by standard microbiological culture techniques, and all phenotypically different isolates in the first specimens recovered in 2007 and 2008 were genotyped by multilocus sequence typing. A total of 313 (57%) of 551 patients whose samples were cultured carried P. aeruginosa. Two sequence types (STs), ST406 and ST497, were found in 15% and 5% of the patients, respectively, and 60% of the patients harbored a strain that was also found in at least two other patients. The risk ratios for carrying ST406 and ST497 were 17.8 (95% confidence interval [CI], 7.2 to 43.6) for those aged between 15 and 24 years and 6 (95% CI, 1.4 to 26.1) for those aged >25 years. ST406 and ST497 were not genetically linked to previously described epidemic clones, which were also not found in this CF population. The population structure of P. aeruginosa in Dutch CF patients is characterized by the presence of two prevalent STs that are associated with certain age groups and that are not genetically linked to previously described epidemic clones. PMID:19828746

  19. Pyoverdine and Proteases Affect the Response of Pseudomonas aeruginosa to Gallium in Human Serum

    PubMed Central

    Bonchi, Carlo; Frangipani, Emanuela; Imperi, Francesco

    2015-01-01

    Gallium is an iron mimetic which has recently been repurposed as an antibacterial agent due to its capability to disrupt bacterial iron metabolism. In this study, the antibacterial activity of gallium nitrate [Ga(NO3)3] was investigated in complement-free human serum (HS) on 55 Pseudomonas aeruginosa clinical isolates from cystic fibrosis and non-cystic fibrosis patients. The susceptibility of P. aeruginosa to Ga(NO3)3 in HS was dependent on the bacterial ability to acquire iron from serum binding proteins (i.e., transferrin). The extent of serum protein degradation correlated well with P. aeruginosa growth in HS, while pyoverdine production did not. However, pyoverdine-deficient P. aeruginosa strains were unable to grow in HS and overcome iron restriction, albeit capable of releasing proteases. Predigestion of HS with proteinase K promoted the growth of all strains, irrespective of their ability to produce proteases and/or pyoverdine. The MICs of Ga(NO3)3 were higher in HS than in an iron-poor Casamino Acids medium, where proteolysis does not affect iron availability. Coherently, strains displaying high proteolytic activity were less susceptible to Ga(NO3)3 in HS. Our data support a model in which both pyoverdine and proteases affect the response of P. aeruginosa to Ga(NO3)3 in HS. The relatively high Ga(NO3)3 concentration required to inhibit the growth of highly proteolytic P. aeruginosa isolates in HS poses a limitation to the potential of Ga(NO3)3 in the treatment of P. aeruginosa bloodstream infections. PMID:26149986

  20. Two Genetic Loci Produce Distinct Carbohydrate-Rich Structural Components of the Pseudomonas aeruginosa Biofilm Matrix

    PubMed Central

    Friedman, Lisa; Kolter, Roberto

    2004-01-01

    Pseudomonas aeruginosa forms biofilms, which are cellular aggregates encased in an extracellular matrix. Molecular genetics studies of three common autoaggregative phenotypes, namely wrinkled colonies, pellicles, and solid-surface-associated biofilms, led to the identification of two loci, pel and psl, that are involved in the production of carbohydrate-rich components of the biofilm matrix. The pel gene cluster is involved in the production of a glucose-rich matrix material in P. aeruginosa strain PA14 (L. Friedman and R. Kolter, Mol. Microbiol. 51:675-690, 2004). Here we investigate the role of the pel gene cluster in P. aeruginosa strain ZK2870 and identify a second genetic locus, termed psl, involved in the production of a mannose-rich matrix material. The 11 predicted protein products of the psl genes are homologous to proteins involved in carbohydrate processing. P. aeruginosa is thus able to produce two distinct carbohydrate-rich matrix materials. Either carbohydrate-rich matrix component appears to be sufficient for mature biofilm formation, and at least one of them is required for mature biofilm formation in P. aeruginosa strains PA14 and ZK2870. PMID:15231777

  1. Two genetic loci produce distinct carbohydrate-rich structural components of the Pseudomonas aeruginosa biofilm matrix.

    PubMed

    Friedman, Lisa; Kolter, Roberto

    2004-07-01

    Pseudomonas aeruginosa forms biofilms, which are cellular aggregates encased in an extracellular matrix. Molecular genetics studies of three common autoaggregative phenotypes, namely wrinkled colonies, pellicles, and solid-surface-associated biofilms, led to the identification of two loci, pel and psl, that are involved in the production of carbohydrate-rich components of the biofilm matrix. The pel gene cluster is involved in the production of a glucose-rich matrix material in P. aeruginosa strain PA14 (L. Friedman and R. Kolter, Mol. Microbiol. 51:675-690, 2004). Here we investigate the role of the pel gene cluster in P. aeruginosa strain ZK2870 and identify a second genetic locus, termed psl, involved in the production of a mannose-rich matrix material. The 11 predicted protein products of the psl genes are homologous to proteins involved in carbohydrate processing. P. aeruginosa is thus able to produce two distinct carbohydrate-rich matrix materials. Either carbohydrate-rich matrix component appears to be sufficient for mature biofilm formation, and at least one of them is required for mature biofilm formation in P. aeruginosa strains PA14 and ZK2870. Copyright 2004 American Society for Microbiology

  2. Characterization of 2-(2-nitro-4-trifluoromethylbenzoyl)-1,3-cyclohexanedione resistance in pyomelanogenic Pseudomonas aeruginosa DKN343

    PubMed Central

    Ketelboeter, Laura M.

    2017-01-01

    Pyomelanin is a reddish-brown pigment that provides bacteria and fungi protection from oxidative stress, and is reported to contribute to infection persistence. Production of this pigment can be inhibited by the anti-virulence agent 2-(2-nitro-4-trifluoromethylbenzoyl)-1,3-cyclohexanedione (NTBC). The Pseudomonas aeruginosa clinical isolate DKN343 exhibited high levels of resistance to NTBC, and the mechanism of pyomelanin production in this strain was uncharacterized. We determined that pyomelanin production in the clinical Pseudomonas aeruginosa isolate DKN343 was due to a loss of function in homogentisate 1,2-dioxygenase (HmgA). Several potential resistance mechanisms were investigated, and the MexAB-OprM efflux pump is required for resistance to NTBC. DKN343 has a frameshift mutation in NalC, which is a known indirect repressor of the mexAB-oprM operon. This frameshift mutation may contribute to the increased resistance of DKN343 to NTBC. Additional studies investigating the prevalence of resistance in pyomelanogenic microbes are necessary to determine the future applications of NTBC as an anti-virulence therapy. PMID:28570601

  3. Clinical utilization of genomics data produced by the international Pseudomonas aeruginosa consortium

    PubMed Central

    Freschi, Luca; Jeukens, Julie; Kukavica-Ibrulj, Irena; Boyle, Brian; Dupont, Marie-Josée; Laroche, Jérôme; Larose, Stéphane; Maaroufi, Halim; Fothergill, Joanne L.; Moore, Matthew; Winsor, Geoffrey L.; Aaron, Shawn D.; Barbeau, Jean; Bell, Scott C.; Burns, Jane L.; Camara, Miguel; Cantin, André; Charette, Steve J.; Dewar, Ken; Déziel, Éric; Grimwood, Keith; Hancock, Robert E. W.; Harrison, Joe J.; Heeb, Stephan; Jelsbak, Lars; Jia, Baofeng; Kenna, Dervla T.; Kidd, Timothy J.; Klockgether, Jens; Lam, Joseph S.; Lamont, Iain L.; Lewenza, Shawn; Loman, Nick; Malouin, François; Manos, Jim; McArthur, Andrew G.; McKeown, Josie; Milot, Julie; Naghra, Hardeep; Nguyen, Dao; Pereira, Sheldon K.; Perron, Gabriel G.; Pirnay, Jean-Paul; Rainey, Paul B.; Rousseau, Simon; Santos, Pedro M.; Stephenson, Anne; Taylor, Véronique; Turton, Jane F.; Waglechner, Nicholas; Williams, Paul; Thrane, Sandra W.; Wright, Gerard D.; Brinkman, Fiona S. L.; Tucker, Nicholas P.; Tümmler, Burkhard; Winstanley, Craig; Levesque, Roger C.

    2015-01-01

    The International Pseudomonas aeruginosa Consortium is sequencing over 1000 genomes and building an analysis pipeline for the study of Pseudomonas genome evolution, antibiotic resistance and virulence genes. Metadata, including genomic and phenotypic data for each isolate of the collection, are available through the International Pseudomonas Consortium Database (http://ipcd.ibis.ulaval.ca/). Here, we present our strategy and the results that emerged from the analysis of the first 389 genomes. With as yet unmatched resolution, our results confirm that P. aeruginosa strains can be divided into three major groups that are further divided into subgroups, some not previously reported in the literature. We also provide the first snapshot of P. aeruginosa strain diversity with respect to antibiotic resistance. Our approach will allow us to draw potential links between environmental strains and those implicated in human and animal infections, understand how patients become infected and how the infection evolves over time as well as identify prognostic markers for better evidence-based decisions on patient care. PMID:26483767

  4. RX-P873, a Novel Protein Synthesis Inhibitor, Accumulates in Human THP-1 Monocytes and Is Active against Intracellular Infections by Gram-Positive (Staphylococcus aureus) and Gram-Negative (Pseudomonas aeruginosa) Bacteria

    PubMed Central

    Buyck, Julien M.; Peyrusson, Frédéric

    2015-01-01

    The pyrrolocytosine RX-P873, a new broad-spectrum antibiotic in preclinical development, inhibits protein synthesis at the translation step. The aims of this work were to study RX-P873's ability to accumulate in eukaryotic cells, together with its activity against extracellular and intracellular forms of infection by Staphylococcus aureus and Pseudomonas aeruginosa, using a pharmacodynamic approach allowing the determination of maximal relative efficacies (Emax values) and bacteriostatic concentrations (Cs values) on the basis of Hill equations of the concentration-response curves. RX-P873's apparent concentration in human THP-1 monocytes was about 6-fold higher than the extracellular one. In broth, MICs ranged from 0.125 to 0.5 mg/liter (S. aureus) and 2 to 8 mg/liter (P. aeruginosa), with no significant shift in these values against strains resistant to currently used antibiotics being noted. In concentration-dependent experiments, the pharmacodynamic profile of RX-P873 was not influenced by the resistance phenotype of the strains. Emax values (expressed as the decrease in the number of CFU from that in the initial inoculum) against S. aureus and P. aeruginosa reached more than 4 log units and 5 log units in broth, respectively, and 0.7 log unit and 2.7 log units in infected THP-1 cells, respectively, after 24 h. Cs values remained close to the MIC in all cases, making RX-P873 more potent than antibiotics to which the strains were resistant (moxifloxacin, vancomycin, and daptomycin for S. aureus; ciprofloxacin and ceftazidime for P. aeruginosa). Kill curves in broth showed that RX-P873 was more rapidly bactericidal against P. aeruginosa than against S. aureus. Taken together, these data suggest that RX-P873 may constitute a useful alternative for infections involving intracellular bacteria, especially Gram-negative species. PMID:26014952

  5. A freshwater bacterial strain, Shewanella sp. Lzh-2, isolated from Lake Taihu and its two algicidal active substances, hexahydropyrrolo[1,2-a]pyrazine-1,4-dione and 2, 3-indolinedione.

    PubMed

    Li, Zhenghua; Lin, Shengqin; Liu, Xianglong; Tan, Jing; Pan, Jianliang; Yang, Hong

    2014-05-01

    Cyanobacterial blooms have become a serious problem in Lake Taihu during the last 20 years, and Microcystis aeruginosa and Synechococcus sp. are the two dominant species in cyanobacterial blooms of Lake Taihu. A freshwater bacterial strain, Shewanella sp. Lzh-2, with strong algicidal properties against harmful cyanobacteria was isolated from Lake Taihu. Two substances with algicidal activity secreted extracellularly by Shewanella sp. Lzh-2, S-2A and S-2B, were purified from the bacterial culture of strain Lzh-2 using ethyl acetate extraction, column chromatography, and high performance liquid chromatography (HPLC) in turn. The substances S-2A and S-2B were identified as hexahydropyrrolo[1,2-a]pyrazine-1,4-dione and 2, 3-indolinedione (isatin), respectively, based on liquid chromatography-mass spectrometry (LC-MS), gas chromatography-mass spectrometry (GC-MS), and hydrogen-nuclear magnetic resonance (H-NMR) analyses, making this the first report of their algicidal activity toward cyanobacteria. S-2A (hexahydropyrrolo[1,2-a]pyrazine-1,4-dione) had no algicidal effects against Synechococcus sp. BN60, but had a high level of algicidal activity against M. aeruginosa 9110. The LD50 value of S-2A against M. aeruginosa 9110 was 5.7 μg/ml. S-2B (2, 3-indolinedione) showed a potent algicidal effect against both M. aeruginosa 9110 and Synechococcus sp. BN60, and the LD50 value of S-2B against M. aeruginosa 9110 and Synechococcus sp. BN60 was 12.5 and 34.2 μg/ml, respectively. Obvious morphological changes in M. aeruginosa 9110 and Synechococcus sp. BN60 were observed after they were exposed to S-2A (or S-2B) for 24 h. Approximately, the algicidal activity, the concentration of S-2A and S-2B, and the cell density of Lzh-2 were positively related to each other during the cocultivation process. Overall, these findings increase our knowledge about algicidal substances secreted by algicidal bacteria and indicate that strain Lzh-2 and its two algicidal substances have the

  6. Oxidation of Naphthenoaromatic and Methyl-Substituted Aromatic Compounds by Naphthalene 1,2-Dioxygenase

    PubMed Central

    Selifonov, S. A.; Grifoll, M.; Eaton, R. W.; Chapman, P. J.

    1996-01-01

    Oxidation of acenaphthene, acenaphthylene, and fluorene was examined with recombinant strain Pseudomonas aeruginosa PAO1(pRE695) expressing naphthalene dioxygenase genes cloned from plasmid NAH7. Acenaphthene underwent monooxygenation to 1-acenaphthenol with subsequent conversion to 1-acenaphthenone and cis- and trans-acenaphthene-1,2-diols, while acenaphthylene was dioxygenated to give cis-acenaphthene-1,2-diol. Nonspecific dehydrogenase activities present in the host strain led to the conversion of both of the acenaphthene-1,2-diols to 1,2-acenaphthoquinone. The latter was oxidized spontaneously to naphthalene-1,8-dicarboxylic acid. No aromatic ring dioxygenation products were detected from acenaphthene and acenaphthylene. Mixed monooxygenase and dioxygenase actions of naphthalene dioxygenase on fluorene yielded products of benzylic 9-monooxygenation, aromatic ring dioxygenation, or both. The action of naphthalene dioxygenase on a variety of methyl-substituted aromatic compounds, including 1,2,4-trimethylbenzene and isomers of dimethylnaphthalene, resulted in the formation of benzylic alcohols, i.e., methyl group monooxygenation products, which were subsequently converted to the corresponding carboxylic acids by dehydrogenase(s) in the host strain. Benzylic monooxygenation of methyl groups was strongly predominant over aromatic ring dioxygenation and essentially nonspecific with respect to the substitution pattern of the aromatic substrates. In addition to monooxygenating benzylic methyl and methylene groups, naphthalene dioxygenase behaved as a sulfoxygenase, catalyzing monooxygenation of the sulfur heteroatom of 3-methylbenzothiophene. PMID:16535238

  7. Adaptation of Pseudomonas aeruginosa in Cystic Fibrosis Airways Influences Virulence of Staphylococcus aureus In Vitro and Murine Models of Co-Infection

    PubMed Central

    Baldan, Rossella; Cigana, Cristina; Testa, Francesca; Bianconi, Irene; De Simone, Maura; Pellin, Danilo; Di Serio, Clelia

    2014-01-01

    Cystic fibrosis (CF) airways disease represents an example of polymicrobial infection whereby different bacterial species can interact and influence each other. In CF patients Staphylococcus aureus is often the initial pathogen colonizing the lungs during childhood, while Pseudomonas aeruginosa is the predominant pathogen isolated in adolescents and adults. During chronic infection, P. aeruginosa undergoes adaptation to cope with antimicrobial therapy, host response and co-infecting pathogens. However, S. aureus and P. aeruginosa often co-exist in the same niche influencing the CF pathogenesis. The goal of this study was to investigate the reciprocal interaction of P. aeruginosa and S. aureus and understand the influence of P. aeruginosa adaptation to the CF lung in order to gain important insight on the interplay occurring between the two main pathogens of CF airways, which is still largely unknown. P. aeruginosa reference strains and eight lineages of clinical strains, including early and late clonal isolates from different patients with CF, were tested for growth inhibition of S. aureus. Next, P. aeruginosa/S. aureus competition was investigated in planktonic co-culture, biofilm, and mouse pneumonia model. P. aeruginosa reference and early strains, isolated at the onset of chronic infection, outcompeted S. aureus in vitro and in vivo models of co-infection. On the contrary, our results indicated a reduced capacity to outcompete S. aureus of P. aeruginosa patho-adaptive strains, isolated after several years of chronic infection and carrying several phenotypic changes temporally associated with CF lung adaptation. Our findings provide relevant information with respect to interspecies interaction and disease progression in CF. PMID:24603807

  8. Post-antibiotic effect of orbifloxacin against Escherichia coli and Pseudomonas aeruginosa isolates from dogs

    PubMed Central

    2012-01-01

    Orbifloxacin is a fluoroquinolone drug used widely in companion animal medicine. In this study, we firstly determined post-antibiotic effects (PAEs) and post-antibiotic sub-minimum inhibitory concentrations (MIC) effects (PA-SMEs) of orbifloxacin for two strains each of Escherichia coli and Pseudomonas aeruginosa from dogs, and these parameters were compared with those of enrofloxacin. At twice the MIC, the PAEs of orbifloxacin ranged from -0.28-0.93 h (mean, 0.29 h) for E. coli and -0.18-1.18 h (mean, 0.37 h) for P. aeruginosa. These parameters were not significantly different for E. coli and shorter for P. aeruginosa, compared to enrofloxacin (P < 0.05). Continued exposure to 0.1, 0.2, and 0.3 the MIC of orbifloxacin resulted in average PA-SMEs of 0.55, 1.11, and 2.03 h, respectively, for E. coli, and 1.04, 1.40, and 2.47 h, respectively, for P. aeruginosa. These PA-SMEs, which had no significant differences with those of enrofloxacin, were significantly longer than the corresponding PAEs (P < 0.05). These results suggest that the PA-SME of orbifloxacin for E. coli and P. aeruginosa can be meaningfully prolonged by increase of sub-MICs. PMID:22433170

  9. Post-antibiotic effect of orbifloxacin against Escherichia coli and Pseudomonas aeruginosa isolates from dogs.

    PubMed

    Harada, Kazuki; Shimizu, Takae; Kataoka, Yasushi; Takahashi, Toshio

    2012-03-20

    Orbifloxacin is a fluoroquinolone drug used widely in companion animal medicine. In this study, we firstly determined post-antibiotic effects (PAEs) and post-antibiotic sub-minimum inhibitory concentrations (MIC) effects (PA-SMEs) of orbifloxacin for two strains each of Escherichia coli and Pseudomonas aeruginosa from dogs, and these parameters were compared with those of enrofloxacin. At twice the MIC, the PAEs of orbifloxacin ranged from -0.28-0.93 h (mean, 0.29 h) for E. coli and -0.18-1.18 h (mean, 0.37 h) for P. aeruginosa. These parameters were not significantly different for E. coli and shorter for P. aeruginosa, compared to enrofloxacin (P < 0.05). Continued exposure to 0.1, 0.2, and 0.3 the MIC of orbifloxacin resulted in average PA-SMEs of 0.55, 1.11, and 2.03 h, respectively, for E. coli, and 1.04, 1.40, and 2.47 h, respectively, for P. aeruginosa. These PA-SMEs, which had no significant differences with those of enrofloxacin, were significantly longer than the corresponding PAEs (P < 0.05). These results suggest that the PA-SME of orbifloxacin for E. coli and P. aeruginosa can be meaningfully prolonged by increase of sub-MICs.

  10. Covalent Dimer Species of β-Defensin Defr1 Display Potent Antimicrobial Activity against Multidrug-Resistant Bacterial Pathogens▿

    PubMed Central

    Taylor, Karen; McCullough, Bryan; Clarke, David J.; Langley, Ross J.; Pechenick, Tali; Hill, Adrian; Campopiano, Dominic J.; Barran, Perdita E.; Dorin, Julia R.; Govan, John R. W.

    2007-01-01

    Beta defensins comprise a family of cationic, cysteine-rich antimicrobial peptides, predominantly expressed at epithelial surfaces. Previously we identified a unique five-cysteine defensin-related peptide (Defr1) that, when synthesized, is a mixture of dimeric isoforms and exhibits potent antimicrobial activity against Escherichia coli and Pseudomonas aeruginosa. Here we report that Defr1 displays antimicrobial activity against an extended panel of multidrug-resistant nosocomial pathogens for which antimicrobial treatment is limited or nonexistent. Defr1 fractions were collected by high-pressure liquid chromatography and analyzed by gel electrophoresis and mass spectrometry. Antimicrobial activity was initially investigated with the type strain Pseudomonas aeruginosa PAO1. All fractions tested displayed equivalent, potent antimicrobial activity levels comparable with that of the unfractionated Defr1. However, use of an oxidized, monomeric six-cysteine analogue (Defr1 Y5C), or of reduced Defr1, gave diminished antimicrobial activity. These results suggest that the covalent dimer structure of Defr1 is crucial to antimicrobial activity; this hypothesis was confirmed by investigation of a synthetic one-cysteine variant (Defr1-1cys). This gave an activity profile similar to that of synthetic Defr1 but only in an oxidized, dimeric form. Thus, we have shown that covalent, dimeric molecules based on the Defr1 β-defensin sequence demonstrate antimicrobial activity even in the absence of the canonical cysteine motif. PMID:17353239

  11. Expression of Fap amyloids in Pseudomonas aeruginosa, P. fluorescens, and P. putida results in aggregation and increased biofilm formation.

    PubMed

    Dueholm, Morten S; Søndergaard, Mads T; Nilsson, Martin; Christiansen, Gunna; Stensballe, Allan; Overgaard, Michael T; Givskov, Michael; Tolker-Nielsen, Tim; Otzen, Daniel E; Nielsen, Per H

    2013-06-01

    The fap operon, encoding functional amyloids in Pseudomonas (Fap), is present in most pseudomonads, but so far the expression and importance for biofilm formation has only been investigated for P. fluorescens strain UK4. In this study, we demonstrate the capacity of P. aeruginosa PAO1, P. fluorescens Pf-5, and P. putida F1 to express Fap fibrils, and investigated the effect of Fap expression on aggregation and biofilm formation. The fap operon in all three Pseudomonas species conferred the ability to express Fap fibrils as shown using a recombinant approach. This Fap overexpression consistently resulted in highly aggregative phenotypes and in increased biofilm formation. Detailed biophysical investigations of purified fibrils confirmed FapC as the main fibril monomer and supported the role of FapB as a minor, nucleating constituent as also indicated by bioinformatic analysis. Bioinformatics analysis suggested FapF and FapD as a potential β-barrel membrane pore and protease, respectively. Manipulation of the fap operon showed that FapA affects monomer composition of the final amyloid fibril, and that FapB is an amyloid protein, probably a nucleator for FapC polymerization. Our study highlights the fap operon as a molecular machine for functional amyloid formation. © 2013 The Authors. Microbiology Open published by John Wiley & Sons Ltd.

  12. Glutathione-Disrupted Biofilms of Clinical Pseudomonas aeruginosa Strains Exhibit an Enhanced Antibiotic Effect and a Novel Biofilm Transcriptome.

    PubMed

    Klare, William; Das, Theerthankar; Ibugo, Amaye; Buckle, Edwina; Manefield, Mike; Manos, Jim

    2016-08-01

    Pseudomonas aeruginosa infections result in high morbidity and mortality rates for individuals with cystic fibrosis (CF), with premature death often occurring. These infections are complicated by the formation of biofilms in the sputum. Antibiotic therapy is stymied by antibiotic resistance of the biofilm matrix, making novel antibiofilm strategies highly desirable. Within P. aeruginosa biofilms, the redox factor pyocyanin enhances biofilm integrity by intercalating with extracellular DNA. The antioxidant glutathione (GSH) reacts with pyocyanin, disrupting intercalation. This study investigated GSH disruption by assaying the physiological effects of GSH and DNase I on biofilms of clinical CF isolates grown in CF artificial sputum medium (ASMDM+). Confocal scanning laser microscopy showed that 2 mM GSH, alone or combined with DNase I, significantly disrupted immature (24-h) biofilms of Australian epidemic strain (AES) isogens AES-1R and AES-1M. GSH alone greatly disrupted mature (72-h) AES-1R biofilms, resulting in significant differential expression of 587 genes, as indicated by RNA-sequencing (RNA-seq) analysis. Upregulated systems included cyclic diguanylate and pyoverdine biosynthesis, the type VI secretion system, nitrate metabolism, and translational machinery. Biofilm disruption with GSH revealed a cellular physiology distinct from those of mature and dispersed biofilms. RNA-seq results were validated by biochemical and quantitative PCR assays. Biofilms of a range of CF isolates disrupted with GSH and DNase I were significantly more susceptible to ciprofloxacin, and increased antibiotic effectiveness was achieved by increasing the GSH concentration. This study demonstrated that GSH, alone or with DNase I, represents an effective antibiofilm treatment when combined with appropriate antibiotics, pending in vivo studies. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  13. Glutathione-Disrupted Biofilms of Clinical Pseudomonas aeruginosa Strains Exhibit an Enhanced Antibiotic Effect and a Novel Biofilm Transcriptome

    PubMed Central

    Das, Theerthankar; Ibugo, Amaye; Buckle, Edwina; Manefield, Mike; Manos, Jim

    2016-01-01

    Pseudomonas aeruginosa infections result in high morbidity and mortality rates for individuals with cystic fibrosis (CF), with premature death often occurring. These infections are complicated by the formation of biofilms in the sputum. Antibiotic therapy is stymied by antibiotic resistance of the biofilm matrix, making novel antibiofilm strategies highly desirable. Within P. aeruginosa biofilms, the redox factor pyocyanin enhances biofilm integrity by intercalating with extracellular DNA. The antioxidant glutathione (GSH) reacts with pyocyanin, disrupting intercalation. This study investigated GSH disruption by assaying the physiological effects of GSH and DNase I on biofilms of clinical CF isolates grown in CF artificial sputum medium (ASMDM+). Confocal scanning laser microscopy showed that 2 mM GSH, alone or combined with DNase I, significantly disrupted immature (24-h) biofilms of Australian epidemic strain (AES) isogens AES-1R and AES-1M. GSH alone greatly disrupted mature (72-h) AES-1R biofilms, resulting in significant differential expression of 587 genes, as indicated by RNA-sequencing (RNA-seq) analysis. Upregulated systems included cyclic diguanylate and pyoverdine biosynthesis, the type VI secretion system, nitrate metabolism, and translational machinery. Biofilm disruption with GSH revealed a cellular physiology distinct from those of mature and dispersed biofilms. RNA-seq results were validated by biochemical and quantitative PCR assays. Biofilms of a range of CF isolates disrupted with GSH and DNase I were significantly more susceptible to ciprofloxacin, and increased antibiotic effectiveness was achieved by increasing the GSH concentration. This study demonstrated that GSH, alone or with DNase I, represents an effective antibiofilm treatment when combined with appropriate antibiotics, pending in vivo studies. PMID:27161630

  14. Pyoverdine and proteases affect the response of Pseudomonas aeruginosa to gallium in human serum.

    PubMed

    Bonchi, Carlo; Frangipani, Emanuela; Imperi, Francesco; Visca, Paolo

    2015-09-01

    Gallium is an iron mimetic which has recently been repurposed as an antibacterial agent due to its capability to disrupt bacterial iron metabolism. In this study, the antibacterial activity of gallium nitrate [Ga(NO3)3] was investigated in complement-free human serum (HS) on 55 Pseudomonas aeruginosa clinical isolates from cystic fibrosis and non-cystic fibrosis patients. The susceptibility of P. aeruginosa to Ga(NO3)3 in HS was dependent on the bacterial ability to acquire iron from serum binding proteins (i.e., transferrin). The extent of serum protein degradation correlated well with P. aeruginosa growth in HS, while pyoverdine production did not. However, pyoverdine-deficient P. aeruginosa strains were unable to grow in HS and overcome iron restriction, albeit capable of releasing proteases. Predigestion of HS with proteinase K promoted the growth of all strains, irrespective of their ability to produce proteases and/or pyoverdine. The MICs of Ga(NO3)3 were higher in HS than in an iron-poor Casamino Acids medium, where proteolysis does not affect iron availability. Coherently, strains displaying high proteolytic activity were less susceptible to Ga(NO3)3 in HS. Our data support a model in which both pyoverdine and proteases affect the response of P. aeruginosa to Ga(NO3)3 in HS. The relatively high Ga(NO3)3 concentration required to inhibit the growth of highly proteolytic P. aeruginosa isolates in HS poses a limitation to the potential of Ga(NO3)3 in the treatment of P. aeruginosa bloodstream infections. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  15. Divergence of a strain of Pseudomonas aeruginosa during an outbreak of ovine mastitis.

    PubMed

    Wright, Elli A; Di Lorenzo, Valeria; Trappetti, Claudia; Liciardi, Manuele; Orru, Germano; Viti, Carlo; Bronowski, Christina; Hall, Amanda J; Darby, Alistair C; Oggioni, Marco R; Winstanley, Craig

    2015-01-30

    Bacterial infections causing mastitis in sheep can result in severe economic losses for farmers. A large survey of milk samples from ewes with mastitis in Sardinia, Italy, indicated an increasing prevalence of Pseudomonas aeruginosa infections. It has been shown previously that during chronic, biofilm-associated infections P. aeruginosa populations diversify. We report the phenotypic and genomic characterisation of two clonal P. aeruginosa isolates (PSE305 and PSE306) from a mastitis infection outbreak, representing distinct colony morphology variants. In addition to pigment production, PSE305 and PSE306 differed in phenotypic characteristics including biofilm formation, utilisation of various carbon and nitrogen sources, twitching motility. We found higher levels of expression of genes associated with biofilm formation (pelB) and twitching motility (flgD) in PSE305, compared to the biofilm and twitching-defective PSE306. Comparative genomics analysis revealed single nucleotide polymorphisms (SNPs) and minor insertion/deletion variations between PSE305 and PSE306, including a SNP mutation in the pilP gene of PSE306. By introducing a wild-type pilP gene we were able to partially complement the defective twitching motility of PSE306. There were also three larger regions of difference between the two genomes, indicating genomic instability. Hence, we have demonstrated that P. aeruginosa population divergence can occur during an outbreak of mastitis, leading to significant variations in phenotype and genotype, and resembling the behaviour of P. aeruginosa during chronic biofilm-associated infections. Copyright © 2014 Elsevier B.V. All rights reserved.

  16. Outbreak of Pseudomonas aeruginosa folliculitis associated with a swimming pool inflatable.

    PubMed Central

    Tate, D.; Mawer, S.; Newton, A.

    2003-01-01

    On 18 February 2002, the Communicable Disease Unit was notified by the local Public Health Service Laboratory of a child with a positive skin swab for Pseudomonas aeruginosa. This child had attended the local swimming pool and played on an inflatable, subsequently presenting to a Primary Care Nurse Practitioner with folliculitis. A total of 35 cases was identified during the outbreak. This paper describes a case-control study and microbiological sampling of the cases, the suspected inflatable and a survey of 10 swimming pool inflatables in the local area. The odds ratio for developing folliculitis following use of the inflatable was 12 (95% CI 1.05-136.80). The strain of P. aeruginosa found on the inflatable was identical to that obtained from skin swabs of cases. Nine of 10 (90%) of the inflatables sampled were colonized by P. aeruginosa. Attention should be given to the problem of routine decontamination of swimming pool inflatables. P. aeruginosa folliculitis needs to be considered in the differential diagnosis of skin rashes in children, especially in Primary Care. PMID:12729186

  17. Imipenem Resistant Pseudomonas aeruginosa: The fall of the final quarterback.

    PubMed

    Ameen, Nadya; Memon, Zahida; Shaheen, Shehla; Fatima, Ghulam; Ahmed, Farah

    2015-01-01

    To isolate, determine the frequency, and study the demographic trends of MBL positive Pseudomonas aeruginosa from imipenem resistant isolates collected from clinical samples in a tertiary care hospital of Pakistan. In this cross sectional study a total of 230 strains of Pseudomonas were isolated from various clinical specimens on the basis of culture and biochemical tests. Imipenem resistant isolates were selected by Kirby Bauer Diffusion technique, followed by screening for MBL production by Imipenem EDTA Combined Disk Test. Demographic details of each patient were recorded on a separate questionnaire. Chi-Square goodness-of-fit test was computed to review the isolation of MBL positive isolates (P-value ≤ 0.05) in different specimen. Out of 230 strains of P. aeruginosa 49.5% were imipenem resistant; MBL production was confirmed in 64.9% of the resistant isolates. Resistance to polymyxin B (12.5%) was notable. Majority of the MBL positive strains were isolated from patients aged between 20-39 years (45.9%) and the predominant source was pus (43.24%) which was found to be statistically significant (P-value=0.04). Outpatient departments (24.3%) and burn unit (21.6%) were the major places for resistant isolates. MBL production is one of the major causes of IRPA. Increasing resistance to polymyxin B is grave. Due to acquisition of MBL strains MDR P. aeruginosa has become endemic in tertiary setups.

  18. Emergence and Spread of Epidemic Multidrug-Resistant Pseudomonas aeruginosa.

    PubMed

    Miyoshi-Akiyama, Tohru; Tada, Tatsuya; Ohmagari, Norio; Viet Hung, Nguyen; Tharavichitkul, Prasit; Pokhrel, Bharat Mani; Gniadkowski, Marek; Shimojima, Masahiro; Kirikae, Teruo

    2017-12-01

    Pseudomonas aeruginosa (P. aeruginosa) is one of the most common nosocomial pathogens worldwide. Although the emergence of multidrug-resistant (MDR) P. aeruginosa is a critical problem in medical practice, the key features involved in the emergence and spread of MDR P. aeruginosa remain unknown. This study utilized whole genome sequence (WGS) analyses to define the population structure of 185 P. aeruginosa clinical isolates from several countries. Of these 185 isolates, 136 were categorized into sequence type (ST) 235, one of the most common types worldwide. Phylogenetic analysis showed that these isolates fell within seven subclades. Each subclade harbors characteristic drug resistance genes and a characteristic genetic background confined to a geographic location, suggesting that clonal expansion following antibiotic exposure is the driving force in generating the population structure of MDR P. aeruginosa. WGS analyses also showed that the substitution rate was markedly higher in ST235 MDR P. aeruginosa than in other strains. Notably, almost all ST235 isolates harbor the specific type IV secretion system and very few or none harbor the CRISPR/CAS system. These findings may help explain the mechanism underlying the emergence and spread of ST235 P. aeruginosa as the predominant MDR lineage. © The Author(s) 2017. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

  19. Enterobactin-mediated iron transport in Pseudomonas aeruginosa.

    PubMed Central

    Poole, K; Young, L; Neshat, S

    1990-01-01

    A pyoverdine-deficient strain of Pseudomonas aeruginosa was unable to grow in an iron-deficient minimal medium in the presence of the nonmetabolizable iron chelator ethylene diamine-di(omega-hydroxyphenol acetic acid) (EDDHA), although addition of enterobactin to EDDHA-containing minimal media did restore growth of the pyoverdine-deficient P. aeruginosa. Consistent with the apparent ability of enterobactin to provide iron to P. aeruginosa, enterobactin-dependent 55Fe3+ uptake was observed in cells of P. aeruginosa previously grown in an iron-deficient medium containing enterobactin (or enterobactin-containing Escherichia coli culture supernatant). This uptake was energy dependent, was observable at low concentrations (60 nM) of FeCl3, and was absent in cells cultured without enterobactin. A novel protein with a molecular weight of approximately 80,000 was identified in the outer membranes of cells grown in iron-deficient minimal medium containing enterobactin, concomitant with the induction of enterobactin-dependent iron uptake. A Tn501 insertion mutant lacking this protein was isolated and shown to be deficient in enterobactin-mediated iron transport at 60 nM FeCl3, although it still exhibited enterobactin-dependent growth in iron-deficient medium containing EDDHA. It was subsequently observed that the mutant was, however, capable of enterobactin-mediated iron transport at much higher concentrations (600 nM) of FeCl3. Indeed, enterobactin-dependent iron uptake at this concentration of iron was observed in both the mutant and parent strains irrespective of whether they had been cultured in the presence of enterobactin.(ABSTRACT TRUNCATED AT 250 WORDS) Images PMID:2174865

  20. Inhibition of quorum sensing regulated bacterial functions by plant essential oils with special reference to clove oil.

    PubMed

    Khan, M S A; Zahin, M; Hasan, S; Husain, F M; Ahmad, I

    2009-09-01

    To evaluate quorum sensing (QS) inhibitory activity of plant essential oils using strains of Chromobacterium violaceum (CV12472 and CVO26) and Pseudomonas aeruginosa (PAO1). Inhibition of QS-controlled violacein production in C. violaceum was assayed using disc diffusion and agar well diffusion method. Of the 21 essential oils, four oils showed varying levels of anti-QS activity. Syzygium aromaticum (Clove) oil showed promising anti-QS activity on both wild and mutant strains with zones of pigment inhibition 19 and 17 mm, respectively, followed by activity in cinnamon, lavender and peppermint oils. The effect of clove oil on the extent of violacein production was estimated photometrically and found to be concentration dependent. At sub-MICs of clove oil, 78.4% reduction in violacein production over control and up to 78% reduction in swarming motility in PAO1 over control were recorded. Gas chromatography-mass spectrometry analysis of clove oil indicated presence of many phytocompounds. Eugenol, the major constituent of clove oil could not exhibit anti-QS activity. Presence of anti-QS activity in clove oil and other essential oils has indicated new anti-infective activity. The identification of anti-QS phytoconstituents is needed to assess the mechanism of action against both C. violaceum and Ps. aeruginosa. Essential oils having new antipathogenic drugs principle because of its anti-QS activity might be important in reducing virulence and pathogenicity of drug-resistant bacteria in vivo.

  1. Pseudomonas aeruginosa rugose small-colony variants evade host clearance, are hyper-inflammatory, and persist in multiple host environments.

    PubMed

    Pestrak, Matthew J; Chaney, Sarah B; Eggleston, Heather C; Dellos-Nolan, Sheri; Dixit, Sriteja; Mathew-Steiner, Shomita S; Roy, Sashwati; Parsek, Matthew R; Sen, Chandan K; Wozniak, Daniel J

    2018-02-01

    Pseudomonas aeruginosa causes devastating infections in immunocompromised individuals. Once established, P. aeruginosa infections become incredibly difficult to treat due to the development of antibiotic tolerant, aggregated communities known as biofilms. A hyper-biofilm forming clinical variant of P. aeruginosa, known as a rugose small-colony variant (RSCV), is frequently isolated from chronic infections and is correlated with poor clinical outcome. The development of these mutants during infection suggests a selective advantage for this phenotype, but it remains unclear how this phenotype promotes persistence. While prior studies suggest RSCVs could survive by evading the host immune response, our study reveals infection with the RSCV, PAO1ΔwspF, stimulated an extensive inflammatory response that caused significant damage to the surrounding host tissue. In both a chronic wound model and acute pulmonary model of infection, we observed increased bacterial burden, host tissue damage, and a robust neutrophil response during RSCV infection. Given the essential role of neutrophils in P. aeruginosa-mediated disease, we investigated the impact of the RSCV phenotype on neutrophil function. The RSCV phenotype promoted phagocytic evasion and stimulated neutrophil reactive oxygen species (ROS) production. We also demonstrate that bacterial aggregation and TLR-mediated pro-inflammatory cytokine production contribute to the immune response to RSCVs. Additionally, RSCVs exhibited enhanced tolerance to neutrophil-produced antimicrobials including H2O2 and the antimicrobial peptide LL-37. Collectively, these data indicate RSCVs elicit a robust but ineffective neutrophil response that causes significant host tissue damage. This study provides new insight on RSCV persistence, and indicates this variant may have a critical role in the recurring tissue damage often associated with chronic infections.

  2. Acquisition of 16S rRNA methylase gene in Pseudomonas aeruginosa.

    PubMed

    Yokoyama, Keiko; Doi, Yohei; Yamane, Kunikazu; Kurokawa, Hiroshi; Shibata, Naohiro; Shibayama, Keigo; Yagi, Tetsuya; Kato, Haru; Arakawa, Yoshichika

    2003-12-06

    Bacteria develop resistance to aminoglycosides by producing aminoglycoside-modifying enzymes such as acetyltransferase, phosphorylase, and adenyltransferase. These enzymes, however, cannot confer consistent resistance to various aminoglycosides because of their substrate specificity. Notwithstanding, a Pseudomonas aeruginosa strain AR-2 showing high-level resistance (minimum inhibitory concentration >1024 mg/L) to various aminoglycosides was isolated clinically. We aimed to clone and characterise the genetic determinant of this resistance. We used conventional methods for DNA manipulation, susceptibility testing, and gene analyses to clone and characterise the genetic determinant of the resistance seen. PCR detection of the gene was also done on a stock of P aeruginosa strains that were isolated clinically since 1997. An aminoglycoside-resistance gene, designated rmtA, was identified in P aeruginosa AR-2. The Escherichia coli transformant and transconjugant harbouring the rmtA gene showed very high-level resistance to various aminoglycosides, including amikacin, tobramycin, isepamicin, arbekacin, kanamycin, and gentamicin. The 756-bp nucleotide rmtA gene encoded a protein, RmtA. This protein showed considerable similarity to the 16S rRNA methylases of aminoglycoside-producing actinomycetes, which protect bacterial 16S rRNA from intrinsic aminoglycosides by methylation. Incorporation of radiolabelled methyl groups into the 30S ribosome was detected in the presence of RmtA. Of 1113 clinically isolated P aeruginosa strains, nine carried the rmtA gene, as shown by PCR analyses. Our findings strongly suggest intergeneric lateral gene transfer of 16S rRNA methylase gene from some aminoglycoside-producing microorganisms to P aeruginosa. Further dissemination of the rmtA gene in nosocomial bacteria could be a matter of concern in the future.

  3. Comparison of phenotypic tests for the detection of metallo-beta-lactamases in clinical isolates of Pseudomonas aeruginosa.

    PubMed

    Lucena, Andréa; Dalla Costa, Libera M; Nogueira, Keite da Silva; Matos, Adriana P; Gales, Ana C; Raboni, Sonia M

    2014-12-01

    Metallo-β-lactamase (MBL)-producing gram-negative bacteria are an increasing public health concern worldwide. Screening tests for the rapid and specific identification of these pathogens are essential, and should be included among routine diagnostics in laboratories. This study aimed to determine the MBL frequency among carbapenem-resistant Pseudomonas aeruginosa isolates, and to evaluate the accuracy of different tests in screening for MBL production. From January 2001 to December 2008, a total of 142 imipenem-non-susceptible P. aeruginosa strains were isolated from distinct clinical samples from hospitalized patients. These isolates were examined by PCR, MBL E-test, double-disk synergy test (DDST), and combined disk (CD) test. The minimal inhibitory concentration (MIC; μg/mL) was determined by agar dilution, and pulsed field gel electrophoresis (PFGE) was performed on all samples. Sequencing was performed to confirm and define the MBL variant and subtype. Using PCR and DNA sequence analysis, 93 strains were confirmed positive for MBLs, 91 strains for the blaSPM-1 gene, 1 strain for the blaIMP-1 gene, and 1 strain for the blaIMP-16 gene. PFGE displayed a clonal pattern. The sensitivities, specificities, positive and negative predictive values were evaluated for all tests. The DDST assay (CAZ-MPA) was the optimal method for screening MBL production in P. aeruginosa strains. However, the results of the CD assay (IMP/EDTA) showed close agreement with those of the DDST. In addition, the CD assay allowed a more objective interpretation and did not require the use of a toxic substance. Copyright © 2013 Elsevier España, S.L.U. y Sociedad Española de Enfermedades Infecciosas y Microbiología Clínica. All rights reserved.

  4. Isolation and characterization of Pseudomonas aeruginosa strain SJTD-2 for degrading long-chain n-alkanes and crude oil.

    PubMed

    Xu, Jing; Liu, Huan; Liu, Jianhua; Liang, Rubing

    2015-06-04

    Oil pollution poses a severe threat to ecosystems, and bioremediation is considered as a safe and efficient alternative to physicochemical. for eliminating this contaminant. In this study, a gram-negative bacteria strain SJTD-2 isolated from oil-contaminated soil was found capable of utilizing n-alkanes and crude oil as sole energy sources. The efficiency of this strain in degrading these pollutants was analyzed. Strain SJTD-2 was identified on the basis of its phenotype, its physiological features, and a comparative genetic analysis using 16S rRNA sequence. Growth of strain SJTD-2 with different carbon sources (n-alkanes of different lengths and crude oil) was assessed, and the gas chromatography-mass spectrometry method was used to analyze the degradation efficiency of strain SJTD-2 for n-alkanes and petroleum by detecting the residual n-alkane concentrations. Strain SJTD-2 was identified as Pseudomonas aeruginosa based on the phenotype, physiological features, and 16S rRNA sequence analysis. This strain can efficiently decompose medium-chain and long-chain n-alkanes (C10-C26), and petroleum as its sole carbon sources. It preferred the long-chain n-alkanes (C18-C22), and n-docosane was considered as the best carbon source for its growth. In 48 h, 500 mg/L n-docosane could be degraded completely, and 2 g/L n-docosane was decomposed to undetectable levels within 72 h. Moreover, strain SJTD-2 could utilize about 88% of 2 g/L crude oil in 7days. Compared with other alkane-utilizing strains, strain SJTD-2 showed outstanding degradation efficiency for long-chain n-alkanes and high tolerance to petroleum at elevated concentrations. The isolation and characterization of strain SJTD-2 would help researchers study the mechanisms underlying the biodegradation of n-alkanes, and this strain could be used as a potential strain for environmental governance and soil bioremediation.

  5. Molecular Epidemiology of a Pseudomonas aeruginosa Hospital Outbreak Driven by a Contaminated Disinfectant-Soap Dispenser

    PubMed Central

    Lanini, Simone; D'Arezzo, Silvia; Puro, Vincenzo; Martini, Lorena; Imperi, Francesco; Piselli, Pierluca; Montanaro, Marco; Paoletti, Simonetta; Visca, Paolo; Ippolito, Giuseppe

    2011-01-01

    Background and Objective Pseudomonas aeruginosa infection represents a main cause of morbidity and mortality among immunocompromised patients. This study describes a fatal epidemic of P. aeruginosa that occurred in a hematology unit in Italy. Methods Retrospective cohort study, prospective surveillance, auditing, extensive testing on healthcare workers and environmental investigation were performed to define the dynamics and potential causes of transmission. RAPD, macrorestriction analyses and sequence typing were used to define relationships between P. aeruginosa isolates. Results Eighteen cases of infection were identified in the different phases of the investigation. Of these, five constitute a significant molecular cluster of infection. A P. aeruginosa strain with the same genetic fingerprint and sequence type (ST175) as clinical isolates strain was also isolated from a heavily contaminated triclosan soap dispenser. Discussion and Conclusions Our results are consistent with the hypothesis that patients became indirectly infected, e.g., during central venous catheter handling through contaminated items, and that the triclosan soap dispenser acted as a common continuous source of P. aeruginosa infection. Since P. aeruginosa is intrinsically unsusceptible to triclosan, the use of triclosan-based disinfectant formulations should be avoided in those healthcare settings hosting patients at high risk of P. aeruginosa infection. PMID:21359222

  6. Surface association and the MreB cytoskeleton regulate pilus production, localization and function in Pseudomonas aeruginosa.

    PubMed

    Cowles, Kimberly N; Gitai, Zemer

    2010-06-01

    Spatial organization of bacterial proteins influences many cellular processes, including division, chromosome segregation and motility. Virulence-associated proteins also localize to specific destinations within bacterial cells. However, the functions and mechanisms of virulence factor localization remain largely unknown. In this work, we demonstrate that polar assembly of the Pseudomonas aeruginosa PAO1 type IV pilus is regulated by surface association in a manner that affects gene transcription, protein levels and protein localization. We also uncover one mechanism for this regulation that acts through the actin homologue MreB. Inactivation of MreB leads to mislocalization of the pilus retraction ATPase PilT, mislocalization of the pili themselves and a reduction in motility. Furthermore, the role of MreB in polar localization of PilT is modulated by surface association, corroborating our results that environmental factors influence the regulation of pilus production. Specifically, MreB mediates both the initiation and maintenance of PilT localization when cells are grown in suspension but only affects the initiation of localization when cells are grown on a surface. Together, these results suggest that the bacterial cytoskeleton provides a mechanism for the polar localization of P. aeruginosa pili and demonstrate that protein localization may represent an important aspect of virulence factor regulation in bacterial pathogens.

  7. Activity of Bacteriophages in Removing Biofilms of Pseudomonas aeruginosa Isolates from Chronic Rhinosinusitis Patients

    PubMed Central

    Fong, Stephanie A.; Drilling, Amanda; Morales, Sandra; Cornet, Marjolein E.; Woodworth, Bradford A.; Fokkens, Wytske J.; Psaltis, Alkis J.; Vreugde, Sarah; Wormald, Peter-John

    2017-01-01

    Introduction: Pseudomonas aeruginosa infections are prevalent amongst chronic rhinosinusitis (CRS) sufferers. Many P. aeruginosa strains form biofilms, leading to treatment failure. Lytic bacteriophages (phages) are viruses that infect, replicate within, and lyse bacteria, causing bacterial death. Aim: To assess the activity of a phage cocktail in eradicating biofilms of ex vivo P.aeruginosa isolates from CRS patients. Methods: P. aeruginosa isolates from CRS patients with and without cystic fibrosis (CF) across three continents were multi-locus sequence typed and tested for antibiotic resistance. Biofilms grown in vitro were treated with a cocktail of four phages (CT-PA). Biofilm biomass was measured after 24 and 48 h, using a crystal violet assay. Phage titrations were performed to confirm replication of the phages. A linear mixed effects model was applied to assess the effects of treatment, time, CF status, and multidrug resistance on the biomass of the biofilm. Results: The isolates included 44 strain types. CT-PA treatment significantly reduced biofilm biomass at both 24 and 48 h post-treatment (p < 0.0001), regardless of CF status or antibiotic resistance. Biomass was decreased by a median of 76% at 48 h. Decrease in biofilm was accompanied by a rise in phage titres for all except one strain. Conclusion: A single dose of phages is able to significantly reduce biofilms formed in vitro by a range of P.aeruginosa isolates from CRS patients. This represents an exciting potential and novel targeted treatment for P. aeruginosa biofilm infections and multidrug resistant bacteria. PMID:29018773

  8. Preparation, characterization and in vitro antimicrobial activity of liposomal ceftazidime and cefepime against Pseudomonas aeruginosa strains

    PubMed Central

    Torres, Ieda Maria Sapateiro; Bento, Etiene Barbosa; Almeida, Larissa da Cunha; de Sá, Luisa Zaiden Carvalho Martins; Lima, Eliana Martins

    2012-01-01

    Pseudomonas aeruginosa is an opportunistic microorganism with the ability to respond to a wide variety of environmental changes, exhibiting a high intrinsic resistance to a number of antimicrobial agents. This low susceptibility to antimicrobial substances is primarily due to the low permeability of its outer membrane, efflux mechanisms and the synthesis of enzymes that promote the degradation of these drugs. Cephalosporins, particularty ceftazidime and cefepime are effective against P. aeruginosa, however, its increasing resistance has limited the usage of these antibiotics. Encapsulating antimicrobial drugs into unilamellar liposomes is an approach that has been investigated in order to overcome microorganism resistance. In this study, antimicrobial activity of liposomal ceftazidime and cefepime against P. aeruginosa ATCC 27853 and P. aeruginosa SPM-1 was compared to that of the free drugs. Liposomal characterization included diameter, encapsulation efficiency and stability. Minimum Inhibitory Concentration (MIC) was determined for free and liposomal forms of both drugs. Minimum Bactericidal Concentration (MBC) was determined at concentrations 1, 2 and 4 times MIC. Average diameter of liposomes was 131.88 nm and encapsulation efficiency for cefepime and ceftazidime were 2.29% end 5.77%, respectively. Improved stability was obtained when liposome formulations were prepared with a 50% molar ratio for cholesterol in relation to the phospholipid. MIC for liposomal antibiotics for both drugs were 50% lower than that of the free drug, demonstrating that liposomal drug delivery systems may contribute to increase the antibacterial activity of these drugs. PMID:24031917

  9. IDENTIFICATION OF MICROCYSTIN TOXINS FROM A STRAIN OF MICROCYSTIS AERUGINOSA BY LIQUID CHROMATOGRAPHY INTRODUCTION INTO A HYBRID LINEAR ION TRAP-FOURIER TRANSFORM ION CYCLOTRON RESONANCE MASS SPECTROMETER

    EPA Science Inventory

    The cyclic heptapeptide microcystin toxins produced by a strain of Microcystis aeruginosa that has not been investigated previously were separated by liquid chromatography and identified by high-accuracy m/z measurements of their [M + H]+ ions and the fragment i...

  10. SPECT measurements with /sup 99m/Tc-HM-PAO in focal epilepsy

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Ryding, E.; Rosen, I.; Elmqvist, D.

    1988-12-01

    The ability of SPECT measurements with (/sup 99m/Tc)-HM-PAO (Ceretec) to find the location of the epileptic focus was studied in patients under consideration for neurosurgical treatment for therapy-resistant focal epilepsy. The location of low (/sup 99m/Tc)-HM-PAO uptake regions found at interictal measurements, and of high (/sup 99m/Tc)-HM-PAO uptake regions found at ictal measurements, was compared to the findings of extensive ictal and interictal EEG examinations, and to the results of CT and MRT. While EEG revealed focal epileptic activity in all of the 14 patients, SPECT showed regional abnormalities in 13 (93%). CT and MRT showed abnormal findings in 30%.

  11. VIM-1, VIM-2, and GES-5 Carbapenemases Among Pseudomonas aeruginosa Isolates at a Tertiary Hospital in Istanbul, Turkey.

    PubMed

    Malkoçoğlu, Gülşah; Aktaş, Elif; Bayraktar, Banu; Otlu, Bariş; Bulut, Mehmet Emin

    2017-04-01

    Worldwide increase in carbapenem resistance and transferable carbapenemases are significant challenges in treatment of Pseudomonas aeruginosa infections. In this study, investigation of carbapenemase production in carbapenem-resistant P. aeruginosa isolates recovered from clinical specimens in a tertiary hospital was aimed. A total of 84 carbapenem-resistant P. aeruginosa isolates were examined. "Carbapenem inactivation method" (CIM) was used for phenotypic detection of carbapenemase production. The existence of bla KPC , bla NDM , bla IMP , bla VIM , bla OXA-48 , and bla GES genes was investigated by polymerase chain reaction (PCR). Subtypes of the detected genes were identified by sequence analysis. Arbitrarily primed PCR (AP-PCR) was performed to evaluate the clonal relationship among the isolates. The presence of high-risk clones in carbapenemase producers was investigated by Matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS). Three isolates (3.5%) were identified as carbapenemase producers by CIM tests, while PCR tests demonstrated three isolates carrying carbapenemase genes as well. bla VIM gene was found in two isolates and bla GES gene was found in one isolate. Sequence analysis demonstrated that the carbapenemases were VIM-1, VIM-2, and GES-5. AP-PCR yielded high clonal diversity among the isolates. According to MALDI-TOF MS analysis, none of the carbapenemase-producing strains belonged to the high-risk clones. In conclusion, the presence of VIM-1, VIM-2, and GES-5 type carbapenemases in P. aeruginosa isolates was demonstrated for the first time in our hospital, GES-5 being reported for the second time in Turkey. Our results will lead strategies for controlling the spread of carbapenemases and contribute to epidemiological data from Turkey.

  12. Isolation of lactic acid bacteria from pao cai, a Chinese traditional fermented vegetable, with inhibitory activity against Salmonella associated with fresh-cut apple, using a modelling study.

    PubMed

    Luo, W; Chen, M; Chen, A; Dong, W; Hou, X; Pu, B

    2015-04-01

    To isolate lactic acid bacteria (LAB) from pao cai, a Chinese traditional fermented vegetable, with outstanding inhibitory activity against Salmonella inoculated on fresh-cut apple, using a modelling method. Four kinds of pao cai were selected. A total of 122 isolates exhibited typical LAB characteristics: Gram-positive and catalase negative, among which 104 (85·24%) colonies showed antibacterial activity against Salmonella by the well diffusion assay. Four colonies showing maximum antibacterial radius against Salmonella were selected to co-inoculate with Salmonella on fresh-cut apple and stored at 10°C, further identified as three strains of Lactobacillus plantarum and one strain of Lactobacillus brevis by 16s rRNA gene sequence analysis. The modified Gompertz model was employed to analyse the growth of the micro-organisms on apple wedges. Two of the four selected strains showed antagonistic activity against Salmonella on fresh-cut apple, one of which, RD1, exhibited best inhibitory activity (Salmonella were greatly inhibited when co-inoculated with RD1 at 10°C at 168 h). No deterioration in odour or appearance of the apple piece was observed by the triangle test when fresh-cut apple was inoculated with RD1. The mathematical modelling method is essential to select LAB with outstanding inhibitory activity against Salmonella associated with fresh-cut apple. LAB RD1 holds promise for the preservation of fresh-cut apple. This study provided a new method on fresh-cut product preservation. Besides, to make the LAB isolating procedure a more correct one, this study first added the mathematical modelling method to the isolating procedure. © 2014 The Society for Applied Microbiology.

  13. Methylene blue internalization and photodynamic action against clinical and ATCC Pseudomonas aeruginosa and Staphyloccocus aureus strains.

    PubMed

    Pereira, André Henrique Correia; Pinto, Juliana Guerra; Freitas, Mirian Aparecida Alves; Fontana, Letícia Corrêa; Pacheco Soares, Cristina; Ferreira-Strixino, Juliana

    2018-06-01

    Bacterial infections have been a major challenge to health. Increasing resistance to antimicrobial agents, according to World Health Organization, could be the major cause of death until 2050. Photodynamic therapy emerges as an alternative in microbial inactivation, due to its selectivity and to decreasing or dismissing antibiotic use. This study aimed at evaluating, in vitro, the internalization of the Methylene Blue and its photodynamic activity against a clinical and ATCC strain of Pseudomonas aeruginosa and Staphyloccocus aureus. Thus, the strains were incubated with MB in concentrations of 100, 300 e 500 μg/ml and then irradiated with a LED (±660 nm) at fluence of 10 and 25 J/cm 2 . The MB internalization was evaluated using a confocal microscope (Zeiss LSM 700), to capture the MB and the DAPI (for DNA staining). It was possible to observe that the MB was internalized by the bacterial cells, in all concentrations tested. The CFU/ml count demonstrated significant reduction (p ≤ 0,01) at the average 5.0 logs comparing with control group for the two species in all the tested concentrations. In conclusion, the strains tested were capable of internalizing the MB. PDT with MB was able to decrease the growth of the tested strains in vitro, being a promising alternative to the future treatment of infections caused by these species. Copyright © 2018 Elsevier B.V. All rights reserved.

  14. Activation of the lectin pathway of complement in experimental human keratitis with Pseudomonas aeruginosa.

    PubMed

    Osthoff, Michael; Brown, Karl D; Kong, David C M; Daniell, Mark; Eisen, Damon P

    2014-01-01

    Pseudomonas aeruginosa (P. aeruginosa) microbial keratitis (MK) is a sight-threatening disease. Previous animal studies have identified an important contribution of the complement system to the clearance of P. aeruginosa infection of the cornea. Mannose-binding lectin (MBL), a pattern recognition receptor of the lectin pathway of complement, has been implicated in the host defense against P. aeruginosa. However, studies addressing the role of the lectin pathway in P. aeruginosa MK are lacking. Hence, we sought to determine the activity of the lectin pathway in human MK caused by P. aeruginosa. Primary human corneal epithelial cells (HCECs) from cadaveric donors were exposed to two different P. aeruginosa strains. Gene expression of interleukin (IL)-6, IL-8, MBL, and other complement proteins was determined by reverse transcription-polymerase chain reaction (RT-PCR) and MBL synthesis by enzyme-linked immunosorbent assay and intracellular flow cytometry. MBL gene expression was not detected in unchallenged HCECs. Exposure of HCECs to P. aeruginosa resulted in rapid induction of the transcriptional expression of MBL, IL-6, and IL-8. In addition, expression of several complement proteins of the classical and lectin pathways, but not the alternative pathway, were upregulated after 5 h of challenge, including MBL-associated serine protease 1. However, MBL protein secretion was not detectable 18 h after challenge with P. aeruginosa. MK due to P. aeruginosa triggers activation of MBL and the lectin pathway of complement. However, the physiologic relevance of this finding is unclear, as corresponding MBL oligomer production was not observed.

  15. Activation of the lectin pathway of complement in experimental human keratitis with Pseudomonas aeruginosa

    PubMed Central

    Osthoff, Michael; Brown, Karl D.; Kong, David C.M.; Daniell, Mark

    2014-01-01

    Purpose Pseudomonas aeruginosa (P. aeruginosa) microbial keratitis (MK) is a sight-threatening disease. Previous animal studies have identified an important contribution of the complement system to the clearance of P. aeruginosa infection of the cornea. Mannose-binding lectin (MBL), a pattern recognition receptor of the lectin pathway of complement, has been implicated in the host defense against P. aeruginosa. However, studies addressing the role of the lectin pathway in P. aeruginosa MK are lacking. Hence, we sought to determine the activity of the lectin pathway in human MK caused by P. aeruginosa. Methods Primary human corneal epithelial cells (HCECs) from cadaveric donors were exposed to two different P. aeruginosa strains. Gene expression of interleukin (IL)-6, IL-8, MBL, and other complement proteins was determined by reverse transcription-polymerase chain reaction (RT–PCR) and MBL synthesis by enzyme-linked immunosorbent assay and intracellular flow cytometry. Results MBL gene expression was not detected in unchallenged HCECs. Exposure of HCECs to P. aeruginosa resulted in rapid induction of the transcriptional expression of MBL, IL-6, and IL-8. In addition, expression of several complement proteins of the classical and lectin pathways, but not the alternative pathway, were upregulated after 5 h of challenge, including MBL-associated serine protease 1. However, MBL protein secretion was not detectable 18 h after challenge with P. aeruginosa. Conclusions MK due to P. aeruginosa triggers activation of MBL and the lectin pathway of complement. However, the physiologic relevance of this finding is unclear, as corresponding MBL oligomer production was not observed. PMID:24426774

  16. Imipenem Resistant Pseudomonas aeruginosa: The fall of the final quarterback

    PubMed Central

    Ameen, Nadya; Memon, Zahida; Shaheen, Shehla; Fatima, Ghulam; Ahmed, Farah

    2015-01-01

    Objective: To isolate, determine the frequency, and study the demographic trends of MBL positive Pseudomonas aeruginosa from imipenem resistant isolates collected from clinical samples in a tertiary care hospital of Pakistan. Methods: In this cross sectional study a total of 230 strains of Pseudomonas were isolated from various clinical specimens on the basis of culture and biochemical tests. Imipenem resistant isolates were selected by Kirby Bauer Diffusion technique, followed by screening for MBL production by Imipenem EDTA Combined Disk Test. Demographic details of each patient were recorded on a separate questionnaire. Chi-Square goodness-of-fit test was computed to review the isolation of MBL positive isolates (P-value ≤ 0.05) in different specimen. Results: Out of 230 strains of P. aeruginosa 49.5% were imipenem resistant; MBL production was confirmed in 64.9% of the resistant isolates. Resistance to polymyxin B (12.5%) was notable. Majority of the MBL positive strains were isolated from patients aged between 20-39 years (45.9%) and the predominant source was pus (43.24%) which was found to be statistically significant (P-value=0.04). Outpatient departments (24.3%) and burn unit (21.6%) were the major places for resistant isolates. Conclusion: MBL production is one of the major causes of IRPA. Increasing resistance to polymyxin B is grave. Due to acquisition of MBL strains MDR P. aeruginosa has become endemic in tertiary setups. PMID:26150844

  17. Bacterial mutation affecting plasmid maintenance in Pseudomonas aeruginosa.

    PubMed Central

    Chang, B J; Holloway, B W

    1977-01-01

    A bacterial mutation, risA, in Pseudomonas aeruginosa caused growth inhibition at 43 degrees C of risA strains containing P2 plasmids. Incubation at 43 degrees C resulted in selection for clones that had lost P2 plasmids. PMID:122513

  18. Molecular Characterization of OXA-198 Carbapenemase-Producing Pseudomonas aeruginosa Clinical Isolates.

    PubMed

    Bonnin, Rémy A; Bogaerts, Pierre; Girlich, Delphine; Huang, Te-Din; Dortet, Laurent; Glupczynski, Youri; Naas, Thierry

    2018-06-01

    Carbapenemase-producing Pseudomonadaceae have increasingly been reported worldwide, with an ever-increasing heterogeneity of carbapenem resistance mechanisms, depending on the bacterial species and the geographical location. OXA-198 is a plasmid-encoded class D β-lactamase involved in carbapenem resistance in one Pseudomonas aeruginosa isolate from Belgium. In the setting of a multicenter survey of carbapenem resistance in P. aeruginosa strains in Belgian hospitals in 2013, three additional OXA-198-producing P. aeruginosa isolates originating from patients hospitalized in one hospital were detected. To reveal the molecular mechanism underlying the reduced susceptibility to carbapenems, MIC determinations, whole-genome sequencing, and PCR analyses to confirm the genetic organization were performed. The plasmid harboring the bla OXA-198 gene was characterized, along with the genetic relatedness of the four P. aeruginosa isolates. The bla OXA-198 gene was harbored on a class 1 integron carried by an ∼49-kb IncP-type plasmid proposed as IncP-11. The same plasmid was present in all four P. aeruginosa isolates. Multilocus sequence typing revealed that the isolates all belonged to sequence type 446, and single-nucleotide polymorphism analysis revealed only a few differences between the isolates. This report describes the structure of a 49-kb plasmid harboring the bla OXA-198 gene and presents the first description of OXA-198-producing P. aeruginosa isolates associated with a hospital-associated cluster episode. Copyright © 2018 American Society for Microbiology.

  19. Pseudomonas aeruginosa bacteraemia: independent risk factors for mortality and impact of resistance on outcome.

    PubMed

    Dantas, Raquel Cavalcanti; Ferreira, Melina Lorraine; Gontijo-Filho, Paulo Pinto; Ribas, Rosineide Marques

    2014-12-01

    The rates of multidrug-resistant, extensively drug-resistant and pandrug-resistant isolates amongst non-fermenting Gram-negative bacilli, particularly Pseudomonas aeruginosa, have risen worldwide. The clinical consequence of resistance and the impact of adverse treatment on the outcome of patients with P. aeruginosa bacteraemia remain unclear. To better understand the predictors of mortality, the clinical consequence of resistance and the impact of inappropriate therapy on patient outcomes, we analysed the first episode of P. aeruginosa bacteraemia in patients from a Brazilian tertiary-care hospital during the period from May 2009 to August 2011. Antimicrobial susceptibility testing was conducted; phenotypic detection of metallo-β-lactamase (MBL) and PCR of MBL genes were performed on carbapenem-resistant strains. Amongst the 120 P. aeruginosa isolates, 45.8 % were resistant to carbapenem and 36 strains were tested for MBL detection. A total of 30 % were phenotypically positive and, of these, 77.8 % expressed an MBL gene, bla(SPM-1) (57 %) and bla(VIM-type) (43 %). The resistance rates to ceftazidime, cefepime, piperacillin/tazobactam, carbapenem, fluoroquinolone and aminoglycoside were 55, 42.5, 35, 45.8, 44 and 44 %, respectively. Previous antibiotic use, length of a hospital stay ≥30 days prior to P. aeruginosa, haemodialysis, tracheostomy, pulmonary source of bacteraemia and Intensive Care Unit admission were common independent risk factors for antimicrobial resistance. Cefepime resistance, multidrug resistance and extensive drug resistance were independently associated with inappropriate therapy, which was an important predictor of mortality, being synergistic with the severity of the underlying disease. © 2014 The Authors.

  20. Scaffold of Selenium Nanovectors and Honey Phytochemicals for Inhibition of Pseudomonas aeruginosa Quorum Sensing and Biofilm Formation.

    PubMed

    Prateeksha; Singh, Braj R; Shoeb, M; Sharma, S; Naqvi, A H; Gupta, Vijai K; Singh, Brahma N

    2017-01-01

    Honey is an excellent source of polyphenolic compounds that are effective in attenuating quorum sensing (QS), a chemical process of cell-to-cell communication system used by the opportunistic pathogen Pseudomonas aeruginosa to regulate virulence and biofilm formation. However, lower water solubility and inadequate bioavailability remains major concerns of these therapeutic polyphenols. Its therapeutic index can be improved by using nano-carrier systems to target QS signaling potently. In the present study, we fabricated a unique drug delivery system comprising selenium nanoparticles (SeNPs; non-viral vectors) and polyphenols of honey (HP) for enhancement of anti-QS activity of HP against P. aeruginosa PAO1. The developed selenium nano-scaffold showed superior anti-QS activity, anti-biofilm efficacy, and anti-virulence potential in both in-vitro and in-vivo over its individual components, SeNPs and HP. LasR is inhibited by selenium nano-scaffold in-vitro . Using computational molecular docking studies, we have also demonstrated that the anti-virulence activity of selenium nano-scaffold is reliant on molecular binding that occurs between HP and the QS receptor LasR through hydrogen bonding and hydrophobic interactions. Our preliminary investigations with selenium-based nano-carriers hold significant promise to improve anti-virulence effectiveness of phytochemicals by enhancing effective intracellular delivery.

  1. Genotyping of Pseudomonas aeruginosa isolates from lung transplant recipients and aquatic environment-detected in-hospital transmission.

    PubMed

    Johansson, Ewa; Welinder-Olsson, Christina; Gilljam, Marita

    2014-02-01

    Lung infection with Pseudomonas aeruginosa is common in lung transplant recipients and may lead to severe complications. Bacteriological surveillance aims to detect transmission of microbes between hospital environment and patients. We sought to determine whether genotyping of P. aeruginosa isolates could improve identifications of pathways of infection. From 2004 to 2009, we performed genotyping with multiple-locus variable number of tandem repeats analysis (MLVA) and pulsed-field gel electrophoresis (PFGE) of P. aeruginosa isolates cultured from lung transplant recipients at Sahlgrenska University Hospital, Gothenburg. During a small outbreak in 2008, cultivation and genotyping of isolates from sink and drains samples from the hospital ward were performed. Pseudomona aeruginosa from 11/18 patients were genotyped to unique strains. The remaining seven patients were carriers of a P. aeruginosa strain of cluster A genotype. Pseudomona aeruginosa was isolated in 4/8 water samples, typed by MLVA also as cluster A genotype and confirmed by PFGE to be similar or identical to the isolates from four transplanted patients. In conclusion, genotyping of isolates revealed a clonal relationship between patient and water isolates, indicating in-hospital transmission of P. aeruginosa. We suggest genotyping with MLVA for rapid routine surveillance, with the PFGE method used for extended, confirmatory analyses. © 2013 APMIS. Published by John Wiley & Sons Ltd.

  2. First Survey of Metallo-β-Lactamase Producers in Clinical Isolates of Pseudomonas aeruginosa From a Referral Burn Center in Kurdistan Province.

    PubMed

    Kalantar, Enayatollah; Torabi, Vahideh; Salimizand, Heiman; Soheili, Fariborz; Beiranvand, Soheila; Soltan Dallal, Mohammad Mehdi

    2012-01-01

    Treatment of infectious diseases is becoming more challenging with each passing year. This is especially true for infections caused by Pseudomonas aeruginosa, an opportunistic pathogen with the ability to rapidly develop resistance to multiple classes of antibiotics. This study was conducted to determine the prevalence of metallo-β-lactamase (MBL)-producing strains among multidrug-resistant P. aeruginosa strains isolated from burn patients. The isolates were identified, tested for susceptibility to various antimicrobial agents, and screened for the presence of MβLs by using the double-disk synergy test. The minimal inhibitory concentration of imipenem was determined by microplate broth dilution method on Mueller-Hinton agar. To detect VIM, SIM, and GIM MBLs, the isolates were subjected to polymerase chain reaction. In this study, we identified 100 P. aeruginosa isolates from 176 clinical specimens obtained from burn patients. The isolates showed maximum resistance to ampicillin (100%), ceftazidime (94%), and ceftriaxone (89%). The CLSI-MBL phenotypic test showed that of the 100 P. aeruginosa isolates, 22 (22%) were positive for MBL production in the double-disk synergy test. Of the 22 MBL-positive P. aeruginosa isolates, 8 were resistant to imipenem. PCR analysis showed that 8 isolates were positive for blaVIM1. The other genes blaSIM1 and blaGIM1 were not detected. The study results demonstrate the serious therapeutic threat of the spread of MBL producers among P. aeruginosa populations. Metallo-β-lactamases were detected in 22% of imipenem-resistant P. aeruginosa isolates. Early detection and infection-control practices are the best antimicrobial strategies for this organism; therefore, systematic surveillance to detect MBL producers is necessary.

  3. Synergistic algicidal effect and mechanism of two diketopiperazines produced by Chryseobacterium sp. strain GLY-1106 on the harmful bloom-forming Microcystis aeruginosa

    NASA Astrophysics Data System (ADS)

    Guo, Xingliang; Liu, Xianglong; Pan, Jianliang; Yang, Hong

    2015-10-01

    A potent algicidal bacterium isolated from Lake Taihu, Chryseobacterium sp. strain GLY-1106, produces two algicidal compounds: 1106-A (cyclo(4-OH-Pro-Leu)) and 1106-B (cyclo(Pro-Leu)). Both diketopiperazines showed strong algicidal activities against Microcystis aeruginosa, the dominant bloom-forming cyanobacterium in Lake Taihu. Interestingly, these two algicidal compounds functioned synergistically. Compared with individual treatment, combined treatment with cyclo(4-OH-Pro-Leu) and cyclo(Pro-Leu) significantly enhanced algicidal activity, accelerated the increase in intracellular reactive oxygen species (ROS) levels in M. aeruginosa, and further decreased the activities of antioxidases, effective quantum yield and maximal electron transport rate of M. aeruginosa. The results also showed that the algicidal characteristics of cyclo(4-OH-Pro-Leu) are distinct from those of cyclo(Pro-Leu). Cyclo(4-OH-Pro-Leu) mainly interrupted the flux of electron transport in the cyanobacterial photosynthetic system, whereas cyclo(Pro-Leu) mainly inhibited the activity of cyanobacterial intracellular antioxidases. A possible algicidal mechanism for the synergism between cyclo(4-OH-Pro-Leu) and cyclo(Pro-Leu) is proposed, which is in accordance with their distinct algicidal characteristics in individual and combined treatment. These findings suggest that synergism between algicidal compounds might be used as an effective strategy for the future control of Microcystis blooms.

  4. Synergistic algicidal effect and mechanism of two diketopiperazines produced by Chryseobacterium sp. strain GLY-1106 on the harmful bloom-forming Microcystis aeruginosa

    PubMed Central

    Guo, Xingliang; Liu, Xianglong; Pan, Jianliang; Yang, Hong

    2015-01-01

    A potent algicidal bacterium isolated from Lake Taihu, Chryseobacterium sp. strain GLY-1106, produces two algicidal compounds: 1106-A (cyclo(4-OH-Pro-Leu)) and 1106-B (cyclo(Pro-Leu)). Both diketopiperazines showed strong algicidal activities against Microcystis aeruginosa, the dominant bloom-forming cyanobacterium in Lake Taihu. Interestingly, these two algicidal compounds functioned synergistically. Compared with individual treatment, combined treatment with cyclo(4-OH-Pro-Leu) and cyclo(Pro-Leu) significantly enhanced algicidal activity, accelerated the increase in intracellular reactive oxygen species (ROS) levels in M. aeruginosa, and further decreased the activities of antioxidases, effective quantum yield and maximal electron transport rate of M. aeruginosa. The results also showed that the algicidal characteristics of cyclo(4-OH-Pro-Leu) are distinct from those of cyclo(Pro-Leu). Cyclo(4-OH-Pro-Leu) mainly interrupted the flux of electron transport in the cyanobacterial photosynthetic system, whereas cyclo(Pro-Leu) mainly inhibited the activity of cyanobacterial intracellular antioxidases. A possible algicidal mechanism for the synergism between cyclo(4-OH-Pro-Leu) and cyclo(Pro-Leu) is proposed, which is in accordance with their distinct algicidal characteristics in individual and combined treatment. These findings suggest that synergism between algicidal compounds might be used as an effective strategy for the future control of Microcystis blooms. PMID:26423356

  5. Synergistic algicidal effect and mechanism of two diketopiperazines produced by Chryseobacterium sp. strain GLY-1106 on the harmful bloom-forming Microcystis aeruginosa.

    PubMed

    Guo, Xingliang; Liu, Xianglong; Pan, Jianliang; Yang, Hong

    2015-10-01

    A potent algicidal bacterium isolated from Lake Taihu, Chryseobacterium sp. strain GLY-1106, produces two algicidal compounds: 1106-A (cyclo(4-OH-Pro-Leu)) and 1106-B (cyclo(Pro-Leu)). Both diketopiperazines showed strong algicidal activities against Microcystis aeruginosa, the dominant bloom-forming cyanobacterium in Lake Taihu. Interestingly, these two algicidal compounds functioned synergistically. Compared with individual treatment, combined treatment with cyclo(4-OH-Pro-Leu) and cyclo(Pro-Leu) significantly enhanced algicidal activity, accelerated the increase in intracellular reactive oxygen species (ROS) levels in M. aeruginosa, and further decreased the activities of antioxidases, effective quantum yield and maximal electron transport rate of M. aeruginosa. The results also showed that the algicidal characteristics of cyclo(4-OH-Pro-Leu) are distinct from those of cyclo(Pro-Leu). Cyclo(4-OH-Pro-Leu) mainly interrupted the flux of electron transport in the cyanobacterial photosynthetic system, whereas cyclo(Pro-Leu) mainly inhibited the activity of cyanobacterial intracellular antioxidases. A possible algicidal mechanism for the synergism between cyclo(4-OH-Pro-Leu) and cyclo(Pro-Leu) is proposed, which is in accordance with their distinct algicidal characteristics in individual and combined treatment. These findings suggest that synergism between algicidal compounds might be used as an effective strategy for the future control of Microcystis blooms.

  6. Molecular Epidemiology of Mutations in Antimicrobial Resistance Loci of Pseudomonas aeruginosa Isolates from Airways of Cystic Fibrosis Patients.

    PubMed

    Greipel, Leonie; Fischer, Sebastian; Klockgether, Jens; Dorda, Marie; Mielke, Samira; Wiehlmann, Lutz; Cramer, Nina; Tümmler, Burkhard

    2016-11-01

    The chronic airway infections with Pseudomonas aeruginosa in people with cystic fibrosis (CF) are treated with aerosolized antibiotics, oral fluoroquinolones, and/or intravenous combination therapy with aminoglycosides and β-lactam antibiotics. An international strain collection of 361 P. aeruginosa isolates from 258 CF patients seen at 30 CF clinics was examined for mutations in 17 antimicrobial susceptibility and resistance loci that had been identified as hot spots of mutation by genome sequencing of serial isolates from a single CF clinic. Combinatorial amplicon sequencing of pooled PCR products identified 1,112 sequence variants that were not present in the genomes of representative strains of the 20 most common clones of the global P. aeruginosa population. A high frequency of singular coding variants was seen in spuE, mexA, gyrA, rpoB, fusA1, mexZ, mexY, oprD, ampD, parR, parS, and envZ (amgS), reflecting the pressure upon P. aeruginosa in lungs of CF patients to generate novel protein variants. The proportion of nonneutral amino acid exchanges was high. Of the 17 loci, mexA, mexZ, and pagL were most frequently affected by independent stop mutations. Private and de novo mutations seem to play a pivotal role in the response of P. aeruginosa populations to the antimicrobial load and the individual CF host. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  7. Spatial Mapping of Pyocyanin in Pseudomonas aeruginosa Bacterial Communities by Surface Enhanced Raman Scattering

    PubMed Central

    Polisetti, Sneha; Baig, Nameera F.; Morales-Soto, Nydia; Shrout, Joshua D.; Bohn, Paul W.

    2017-01-01

    Surface Enhanced Raman Spectroscopy (SERS) imaging was used in conjunction with Principal Component Analysis (PCA) for the in situ spatiotemporal mapping of the virulence factor pyocyanin, in communities of the pathogenic bacterium Pseudomonas aeruginosa. The combination of SERS imaging and PCA analysis provides a robust method for characterization of heterogeneous biological systems while circumventing issues associated with interference from sample autofluorescence and low reproducibility of SERS signals. The production of pyocyanin is found to depend both on the growth carbon source and on the specific strain of P. aeruginosa studied. A cystic fibrosis lung isolate strain of P. aeruginosa synthesizes and secretes pyocyanin when grown with glucose and glutamate, while the laboratory strain exhibits detectable production of pyocyanin only when grown with glutamate as the source of carbon. Pyocyanin production in the laboratory strain grown with glucose was below the limit of detection of SERS. In addition, the combination of SERS imaging and PCA can elucidate subtle differences in the molecular composition of biofilms. PCA loading plots from the clinical isolate exhibit features corresponding to vibrational bands of carbohydrates, which represent the mucoid biofilm matrix specific to that isolate, features that are not seen in the PCA loading plots of the laboratory strain. PMID:27354400

  8. A universal definition of ARDS: the PaO2/FiO2 ratio under a standard ventilatory setting--a prospective, multicenter validation study.

    PubMed

    Villar, Jesús; Pérez-Méndez, Lina; Blanco, Jesús; Añón, José Manuel; Blanch, Lluís; Belda, Javier; Santos-Bouza, Antonio; Fernández, Rosa Lidia; Kacmarek, Robert M

    2013-04-01

    The PaO2/FiO2 is an integral part of the assessment of patients with acute respiratory distress syndrome (ARDS). The American-European Consensus Conference definition does not mandate any standardization procedure. We hypothesized that the use of PaO2/FiO2 calculated under a standard ventilatory setting within 24 h of ARDS diagnosis allows a more clinically relevant ARDS classification. We studied 452 ARDS patients enrolled prospectively in two independent, multicenter cohorts treated with protective mechanical ventilation. At the time of ARDS diagnosis, patients had a PaO2/FiO2 ≤ 200. In the derivation cohort (n = 170), we measured PaO2/FiO2 with two levels of positive end-expiratory pressure (PEEP) (≥ 5 and ≥ 10 cmH2O) and two levels of FiO2 (≥ 0.5 and 1.0) at ARDS onset and 24 h later. Dependent upon PaO2 response, patients were reclassified into three groups: mild (PaO2/FiO2 > 200), moderate (PaO2/FiO2 101-200), and severe (PaO2/FiO2 ≤ 100) ARDS. The primary outcome measure was ICU mortality. The standard ventilatory setting that reached the highest significance difference in mortality among these categories was tested in a separate cohort (n = 282). The only standard ventilatory setting that identified the three PaO2/FiO2 risk categories in the derivation cohort was PEEP ≥ 10 cmH2O and FiO2 ≥ 0.5 at 24 h after ARDS onset (p = 0.0001). Using this ventilatory setting, patients in the validation cohort were reclassified as having mild ARDS (n = 47, mortality 17 %), moderate ARDS (n = 149, mortality 40.9 %), and severe ARDS (n = 86, mortality 58.1 %) (p = 0.00001). Our method for assessing PaO2/FiO2 greatly improved risk stratification of ARDS and could be used for enrolling appropriate ARDS patients into therapeutic clinical trials.

  9. Visualization of microbiological processes underlying stress relaxation in Pseudomonas aeruginosa biofilms.

    PubMed

    Peterson, Brandon W; Busscher, Henk J; Sharma, Prashant K; van der Mei, Henny C

    2014-06-01

    Bacterial biofilms relieve themselves from external stresses through internal rearrangement, as mathematically modeled in many studies, but never microscopically visualized for their underlying microbiological processes. The aim of this study was to visualize rearrangement processes occurring in mechanically deformed biofilms using confocal-laser-scanning-microscopy after SYTO9 (green-fluorescent) and calcofluor-white (blue-fluorescent) staining to visualize bacteria and extracellular-polymeric matrix substances, respectively. We apply 20% uniaxial deformation to Pseudomonas aeruginosa biofilms and fix deformed biofilms prior to staining, after allowing different time-periods for relaxation. Two isogenic P. aeruginosa strains with different abilities to produce extracellular polymeric substances (EPS) were used. By confocal-laser-scanning-microscopy all biofilms showed intensity distributions for fluorescence from which rearrangement of EPS and bacteria in deformed biofilms were derived. For the P. aeruginosa strain producing EPS, bacteria could not find new, stable positions within 100 s after deformation, while EPS moved toward deeper layers within 20 s. Bacterial rearrangement was not seen in P. aeruginosa biofilms deficient in production of EPS. Thus, EPS is required to stimulate bacterial rearrangement in mechanically deformed biofilms within the time-scale of our experiments, and the mere presence of water is insufficient to induce bacterial movement, likely due to its looser association with the bacteria.

  10. Rapid detection of Pseudomonas aeruginosa targeting the toxA gene in intensive care unit patients from Beijing, China

    PubMed Central

    Dong, Derong; Zou, Dayang; Liu, Hui; Yang, Zhan; Huang, Simo; Liu, Ningwei; He, Xiaoming; Liu, Wei; Huang, Liuyu

    2015-01-01

    Pseudomonas aeruginosa is a major opportunistic pathogen in hospital-acquired infections and exhibits increasing antibiotic resistance. A rapid and sensitive molecular method for its detection in clinical samples is needed to guide therapeutic treatment and to control P. aeruginosa outbreaks. In this study, we established a polymerase spiral reaction (PSR) method for rapid detection of P. aeruginosa by targeting the toxA gene, which regulates exotoxin A synthesis. Real-time turbidity monitoring and a chromogenic visualization using hydroxynaphthol blue were used to assess the reaction. All 17 non- P. aeruginosa strains tested negative, indicating the high specificity of the PSR primers. The detection limit was 2.3 pg/μl within 60 min at isothermal temperature (65°C), 10-fold more sensitive than conventional PCR. Then, the PSR assay was applied to a clinical surveillance of P. aeruginosa in three top hospitals in Beijing, China. Of the 130 sputum samples collected from ICU patients with suspected multi-resistant infections, 37 P. aeruginosa isolates were identified from the positive samples. All clinical strains belonged to 10 different P. aeruginosa multilocus sequence typing groups and exhibited high resistance to carbapenems, cephalosporins, and aminoglycosides. Interestingly, of the 33 imipenem-resistant isolates, 30 (90.9%) had lost the outer membrane porin oprD gene. Moreover, isolate SY-95, containing multiple antibiotic resistance genes, possessed the ability to hydrolyze all antibiotics used in clinic and was susceptible only to polymyxin B. Our study showed the high level of antibiotic resistance and co-occurrence of resistance genes in the clinical strains, indicating a rapid and continuing evolution of P. aeruginosa. In conclusion, we developed a P. aeruginosa PSR assay, which could be a useful tool for clinical screening, especially in case of poor resources, or for point-of-care testing. PMID:26500639

  11. First Survey of Metallo-β–Lactamase Producers in Clinical Isolates of Pseudomonas aeruginosa From a Referral Burn Center in Kurdistan Province

    PubMed Central

    Kalantar, Enayatollah; Torabi, Vahideh; Salimizand, Heiman; Soheili, Fariborz; Beiranvand, Soheila; Soltan Dallal, Mohammad Mehdi

    2012-01-01

    Background Treatment of infectious diseases is becoming more challenging with each passing year. This is especially true for infections caused by Pseudomonas aeruginosa, an opportunistic pathogen with the ability to rapidly develop resistance to multiple classes of antibiotics. Objectives This study was conducted to determine the prevalence of metallo-β-lactamase (MBL)–producing strains among multidrug-resistant P. aeruginosa strains isolated from burn patients. Materials and Methods The isolates were identified, tested for susceptibility to various antimicrobial agents, and screened for the presence of MβLs by using the double-disk synergy test. The minimal inhibitory concentration of imipenem was determined by microplate broth dilution method on Mueller-Hinton agar. To detect VIM, SIM, and GIM MBLs, the isolates were subjected to polymerase chain reaction. Results In this study, we identified 100 P. aeruginosa isolates from 176 clinical specimens obtained from burn patients. The isolates showed maximum resistance to ampicillin (100%), ceftazidime (94%), and ceftriaxone (89%). The CLSI-MBL phenotypic test showed that of the 100 P. aeruginosa isolates, 22 (22%) were positive for MBL production in the double-disk synergy test. Of the 22 MBL-positive P. aeruginosa isolates, 8 were resistant to imipenem. PCR analysis showed that 8 isolates were positive for blaVIM1. The other genes blaSIM1 and blaGIM1 were not detected. Conclusions The study results demonstrate the serious therapeutic threat of the spread of MBL producers among P. aeruginosa populations. Metallo-β-lactamases were detected in 22% of imipenem-resistant P. aeruginosa isolates. Early detection and infection-control practices are the best antimicrobial strategies for this organism; therefore, systematic surveillance to detect MBL producers is necessary. PMID:24624147

  12. Antimicrobial Resistance of Pseudomonas aeruginosa Isolated from Dogs and Cats in Primary Veterinary Hospitals in Japan.

    PubMed

    Yukawa, Shoichiro; Tsuyuki, Yuzo; Sato, Tomomi; Fukuda, Akira; Usui, Masaru; Tamura, Yutaka

    2017-07-24

    We collected 200 Pseudomonas aeruginosa isolates from dogs and cats in primary veterinary hospitals in Japan to investigate their antimicrobial resistance. Resistance rates against ciprofloxacin, cefotaxime, gentamicin, amikacin, and fosfomycin were 9%, 12.5%, 4.5%, 2.5%, and 35.5%, respectively. One strain displayed resistance (0.5%) to ceftazidime. We did not detect any imipenem-resistant or multidrug-resistant P. aeruginosa strains as defined by the Japanese Ministry of Health, Labour, and Welfare Law Concerning the Prevention of Infections and Medical Care for Patients with Infections. In addition, we did not find any P. aeruginosa isolates that produced metallo-β-lactamase, the aminoglycoside 6'-N-acetyltransferase AAC(6')-Iae, or the aminoglycoside acetyltransferase AAC(6')-Ib.

  13. Isolation and characterization of an algicidal bacterium indigenous to lake Taihu with a red pigment able to lyse microcystis aeruginosa.

    PubMed

    Yang, Fei; Wei, Hai Yan; Li, Xiao Qin; Li, Yun Hui; Li, Xiao Bo; Yin, Li Hong; Pu, Yue Pu

    2013-02-01

    To isolate and characterize indigenous algicidal bacteria and their algae-lysing compounds active against Microcystis aeruginosa, strains TH1, TH2, and FACHB 905. The bacteria were identified using the Biolog automated microbial identification system and 16S rDNA sequence analysis. The algae-lysing compounds were isolated and purified by silica gel column chromatography and reverse-phase high performance liquid chromatography. Their structures were confirmed by Nuclear Magnetic Resonance (NMR) and Fourier Transform Infrared (FT-IR) spectroscopy. Algae-lysing activity was observed using microscopy. The algae-lysing bacterium LTH-2 isolated from Lake Taihu was identified as Serratia marcescens. Strain LTH-2 secreted a red pigment identified as prodigiosin (C20H25N3O), which showed strong lytic activity with algal strains M. aeruginosa TH1, TH2, and FACHB 905 in a concentration-dependent manner. The 50% inhibitory concentration (IC50) of prodigiosin with the algal strains was 4.8 (± 0.4)× 10⁻² μg/mL, 8.9 (± 1.1)× 10⁻² μg/mL, and 1.7 (± 0.1)× 10⁻¹ μg/mL in 24 h, respectively. The bacterium LTH-2 and its pigment had strong Microcystis-lysing activity probably related to damage of cell membranes. The bacterium LTH-2 and its red pigment are potentially useful for regulating blooms of harmful M. aeruginosa. Copyright © 2013 The Editorial Board of Biomedical and Environmental Sciences. Published by China CDC. All rights reserved.

  14. Pseudomonas aeruginosa keratitis: outcomes and response to corticosteroid treatment.

    PubMed

    Sy, Aileen; Srinivasan, Muthiah; Mascarenhas, Jeena; Lalitha, Prajna; Rajaraman, Revathi; Ravindran, Meenakshi; Oldenburg, Catherine E; Ray, Kathryn J; Glidden, David; Zegans, Michael E; McLeod, Stephen D; Lietman, Thomas M; Acharya, Nisha R

    2012-01-25

    To compare the clinical course and effect of adjunctive corticosteroid therapy in Pseudomonas aeruginosa with those of all other strains of bacterial keratitis. Subanalyses were performed on data collected in the Steroids for Corneal Ulcers Trial (SCUT), a large randomized controlled trial in which patients were treated with moxifloxacin and were randomly assigned to 1 of 2 adjunctive treatment arms: corticosteroid or placebo (4 times a day with subsequent reduction). Multivariate analysis was used to determine the effect of predictors, organism, and treatment on outcomes, 3-month best-spectacle-corrected visual acuity (BSCVA), and infiltrate/scar size. The incidence of adverse events over a 3-month follow-up period was compared using Fisher's exact test. SCUT enrolled 500 patients. One hundred ten patients had P. aeruginosa ulcers; 99 of 110 (90%) enrolled patients returned for follow-up at 3 months. Patients with P. aeruginosa ulcers had significantly worse visual acuities than patients with other bacterial ulcers (P = 0.001) but showed significantly more improvement in 3-month BSCVA than those with other bacterial ulcers, adjusting for baseline characteristics (-0.14 logMAR; 95% confidence interval, -0.23 to -0.04; P = 0.004). There was no significant difference in adverse events between P. aeruginosa and other bacterial ulcers. There were no significant differences in BSCVA (P = 0.69), infiltrate/scar size (P = 0.17), and incidence of adverse events between patients with P. aeruginosa ulcers treated with adjunctive corticosteroids and patients given placebo. Although P. aeruginosa corneal ulcers have a more severe presentation, they appear to respond better to treatment than other bacterial ulcers. The authors did not find a significant benefit with corticosteroid treatment, but they also did not find any increase in adverse events. (ClinicalTrials.gov number, NCT00324168.).

  15. Pseudomonas aeruginosa Keratitis: Outcomes and Response to Corticosteroid Treatment

    PubMed Central

    Sy, Aileen; Srinivasan, Muthiah; Mascarenhas, Jeena; Lalitha, Prajna; Rajaraman, Revathi; Ravindran, Meenakshi; Oldenburg, Catherine E.; Ray, Kathryn J.; Glidden, David; Zegans, Michael E.; McLeod, Stephen D.; Lietman, Thomas M.

    2012-01-01

    Purpose. To compare the clinical course and effect of adjunctive corticosteroid therapy in Pseudomonas aeruginosa with those of all other strains of bacterial keratitis. Methods. Subanalyses were performed on data collected in the Steroids for Corneal Ulcers Trial (SCUT), a large randomized controlled trial in which patients were treated with moxifloxacin and were randomly assigned to 1 of 2 adjunctive treatment arms: corticosteroid or placebo (4 times a day with subsequent reduction). Multivariate analysis was used to determine the effect of predictors, organism, and treatment on outcomes, 3-month best-spectacle-corrected visual acuity (BSCVA), and infiltrate/scar size. The incidence of adverse events over a 3-month follow-up period was compared using Fisher's exact test. Results. SCUT enrolled 500 patients. One hundred ten patients had P. aeruginosa ulcers; 99 of 110 (90%) enrolled patients returned for follow-up at 3 months. Patients with P. aeruginosa ulcers had significantly worse visual acuities than patients with other bacterial ulcers (P = 0.001) but showed significantly more improvement in 3-month BSCVA than those with other bacterial ulcers, adjusting for baseline characteristics (−0.14 logMAR; 95% confidence interval, −0.23 to −0.04; P = 0.004). There was no significant difference in adverse events between P. aeruginosa and other bacterial ulcers. There were no significant differences in BSCVA (P = 0.69), infiltrate/scar size (P = 0.17), and incidence of adverse events between patients with P. aeruginosa ulcers treated with adjunctive corticosteroids and patients given placebo. Conclusions. Although P. aeruginosa corneal ulcers have a more severe presentation, they appear to respond better to treatment than other bacterial ulcers. The authors did not find a significant benefit with corticosteroid treatment, but they also did not find any increase in adverse events. (ClinicalTrials.gov number, NCT00324168.) PMID:22159005

  16. Australian epidemic strain pseudomonas (AES-1) declines further in a cohort segregated cystic fibrosis clinic.

    PubMed

    Griffiths, Amanda L; Wurzel, Danielle F; Robinson, Phil J; Carzino, Rosemary; Massie, John

    2012-01-01

    To evaluate changes in prevalence of an epidemic strain of Pseudomonas aeruginosa (AES-1, Australian epidemic strain, type 1) in a paediatric cystic fibrosis (CF) centre practising cohort segregation, to describe the patients' clinical characteristics at acquisition and observe mortality rates. Cohort segregation was introduced in our paediatric CF clinic January 2000. The prevalence of AES-1 was analysed in 1999, 2002 and 2007. Age at acquisition, lung function, presence of bronchiectasis, hospitalisations, prior P. aeruginosa infection and mortality rates were collected. AES-1 infection was determined by pulse-field-gel-electrophoresis (PFGE) on airway specimen cultures taken three monthly. The prevalence of AES-1 declined from 21% in 1999 to 14% in 2002 (risk difference 7% (95% CI 1,13) p=0.0256) and to 6% in 2007 (risk difference 8% (95% CI 3,13) p=0.0018). New acquisitions after the introduction of cohort segregation were uncommon (10 by 2002 and another 7 by 2007) with a declining incidence of 3.3 cases/year (1999 to 2002) compared to 1.4 cases/year (2002 to 2007). Twenty-two of 32 (69%) deaths between 1999 and 2007 occurred in patients infected with AES-1. Cohort segregation has been associated with reductions in the prevalence of AES-1 in our CF clinic. Mortality was higher in patients infected with AES-1 than other organisms. Copyright © 2011 European Cystic Fibrosis Society. Published by Elsevier B.V. All rights reserved.

  17. Identification and discrimination of Pseudomonas aeruginosa bacteria grown in blood and bile by laser-induced breakdown spectroscopy

    NASA Astrophysics Data System (ADS)

    Rehse, Steven J.; Diedrich, Jonathan; Palchaudhuri, Sunil

    2007-10-01

    Pseudomonas aeruginosa bacteria colonies have been analyzed by laser-induced breakdown spectroscopy using nanosecond laser pulses. LIBS spectra were obtained after transferring the bacteria from a nutrient-rich culture medium to a nutrient-free agar plate for laser ablation. To study the dependence of the LIBS spectrum on growth and environmental conditions, colonies were cultured on three different nutrient media: a trypticase soy agar (TSA) plate, a blood agar plate, and a medium chosen deliberately to induce bacteria membrane changes, a MacConkey agar plate containing bile salts. Nineteen atomic and ionic emission lines in the LIBS spectrum, which was dominated by inorganic elements such as calcium, magnesium and sodium, were used to identify and classify the bacteria. A discriminant function analysis was used to discriminate between the P. aeruginosa bacteria and two strains of E. coli: a non-pathogenic environmental strain and the pathogenic strain enterohemorrhagic E. coli 0157:H7 (EHEC). Nearly identical spectra were obtained from P. aeruginosa grown on the TSA plate and the blood agar plate, while the bacteria grown on the MacConkey plate exhibited easily distinguishable differences from the other two. All P. aeruginosa samples, independent of initial growth conditions, were readily discriminated from the two E. coli strains.

  18. Detection of Metallo-Beta Lactamases Among Carbapenem-Resistant Pseudomonas aeruginosa.

    PubMed

    Farajzadeh Sheikh, Ahmad; Rostami, Soodabeh; Jolodar, Abbas; Tabatabaiefar, Mohammad Amin; Khorvash, Farzin; Saki, Azadeh; Shoja, Saeed; Sheikhi, Raheleh

    2014-11-01

    Carbapenems are important drugs used for the treatment of Pseudomonas aeruginosa infections, however metallo-β-lactamases (MBL) are able to efficiently hydrolyze these classes of drugs. Immediate detection of the MBL-producing P. aeruginosa is necessary in order to accurately treat infections caused by this organism. To determine the prevalence of MBL producing P. aeruginosa in burn and non-burn patients by two phenotypic tests and polymerase chain reaction (PCR) and to compare phenotypic tests with PCR. A total of 223 non-duplicate strains of P. aeruginosa were collected from three teaching hospitals of Ahvaz, Iran. Antimicrobial susceptibility and minimum inhibitory concentrations (MICs) of carbapenems (imipenem, meropenem, doripenem and ertapenem) were determined by the Kirby-Bauer and E-test methods. Combined disk (CD) test, MBL E-test and PCR were performed for carbapenem-resistant P. aeruginosa isolates. Amongst all the P. aeruginosa isolates, 58.7% were resistant to imipenem while 31.8%, 13.5% and 74.4% were resistant to meropenem, doripenem and ertapenem, respectively. Amongst all the P. aeruginosa isolates, 44.4% were multidrug resistant and 13.45% were resistant to all of the carbapenems. The CD test with doripenem disk / 750 μg ethylene diamine tetra acetic acid (EDTA) had the highest efficiency compared to the other phenotypic tests. bla IMP and bla VIM genes were detected in 11.7% and 0.4% of isolates, respectively. bla SPM and bla NDM genes were not observed. Epidemiological and regional evaluation of MBL-producing P. aeruginosa through simple and inexpensive methods should be considered for effective treatment of carbapenem-resistant P. aeruginosa infections.

  19. Role of psl Genes in Antibiotic Tolerance of Adherent Pseudomonas aeruginosa.

    PubMed

    Murakami, Keiji; Ono, Tsuneko; Viducic, Darija; Somiya, Yoko; Kariyama, Reiko; Hori, Kenji; Amoh, Takashi; Hirota, Katsuhiko; Kumon, Hiromi; Parsek, Matthew R; Miyake, Yoichiro

    2017-07-01

    Bacteria attached to a surface are generally more tolerant to antibiotics than their planktonic counterparts, even without the formation of a biofilm. The mechanism of antibiotic tolerance in biofilm communities is multifactorial, and the genetic background underlying this antibiotic tolerance has not yet been fully elucidated. Using transposon mutagenesis, we isolated a mutant with reduced tolerance to biapenem (relative to that of the wild type) from adherent cells. Sequencing analysis revealed a mutation in the pslL gene, which is part of the polysaccharide biosynthesis operon. The Pseudomonas aeruginosa PAO1Δ pslBCD mutant demonstrated a 100-fold-lower survival rate during the exposure of planktonic and biofilm cells to biapenem; a similar phenotype was observed in a mouse infection model and in clinical strains. Transcriptional analysis of adherent cells revealed increased expression of both pslA and pelA , which are directly regulated by bis-(3',5')-cyclic dimeric GMP (c-di-GMP). Inactivation of wspF resulted in significantly increased tolerance to biapenem due to increased production of c-di-GMP. The loss of pslBCD in the Δ wspF mutant background abolished the biapenem-tolerant phenotype of the Δ wspF mutant, underscoring the importance of psl in biapenem tolerance. Overexpression of PA2133, which can catalyze the degradation of c-di-GMP, led to a significant reduction in biapenem tolerance in adherent cells, indicating that c-di-GMP is essential in mediating the tolerance effect. The effect of pslBCD on antibiotic tolerance was evident, with 50- and 200-fold-lower survival in the presence of ofloxacin and tobramycin, respectively. We speculate that the psl genes, which are activated by surface adherence through elevated intracellular c-di-GMP levels, confer tolerance to antimicrobials. Copyright © 2017 American Society for Microbiology.

  20. Mobile genetic elements of Pseudomonas aeruginosa isolates from hydrotherapy facility and respiratory infections.

    PubMed

    Pereira, S G; Cardoso, O

    2014-03-01

    The content of mobile genetic elements in Pseudomonas aeruginosa isolates of a pristine natural mineral water system associated with healthcare was compared with clinical isolates from respiratory infections. One isolate, from the therapy pool circuit, presented a class 1 integron, with 100% similarity to a class 1 integron contained in plasmid p4800 of the Klebsiella pneumoniae Kp4800 strain, which is the first time it has been reported in P. aeruginosa. Class 1 integrons were found in 25.6% of the clinical isolates. PAGI1 orf3 was more prevalent in environmental isolates, while PAGI2 c105 and PAGI3 sg100 were more prevalent in clinical isolates. Plasmids were not observed in either population. © 2013 The Authors Clinical Microbiology and Infection © 2013 European Society of Clinical Microbiology and Infectious Diseases.

  1. Dissemination of VIM-2 producing Pseudomonas aeruginosa ST233 at tertiary care hospitals in Egypt.

    PubMed

    Zafer, Mai Mahmoud; Al-Agamy, Mohamed Hamed; El-Mahallawy, Hadir Ahmed; Amin, Magdy Aly; El Din Ashour, Seif

    2015-03-12

    Pseudomonas aeruginosa is an important nosocomial pathogen, commonly causing infections in immunocompromised patients. The aim of this study was to examine the genetic relatedness of metallo-beta-lactamase (MBL) producing carbapenem resistant Pseudomonas aeruginosa clinical isolates collected from 2 tertiary hospitals in Cairo, Egypt using Multi Locus sequence typing (MLST). Phenotypic and genotypic detection of metallo-beta-lactamase for forty eight non-duplicate carbapenem resistant P. aeruginosa isolates were carried out. DNA sequencing and MLST were done. The bla VIM-2 gene was highly prevalent (28/33 strains, 85%) among 33 MBL-positive P.aeruginosa isolates. MLST revealed eleven distinct Sequence Types (STs). A unique ST233 clone producing VIM-2 was documented by MLST in P.aeruginosa strains isolated from Cairo university hospitals. The high prevalence of VIM-2 producers was not due to the spread of a single clone. The findings of the present study clearly demonstrate that clones of VIM-2 positive in our hospitals are different from those reported from European studies. Prevalence of VIM-2 producers of the same clone was detected from surgical specimens whereas oncology related specimens were showing diverse clones.

  2. Pseudomonas aeruginosa folliculitis acquired through use of a contaminated loofah sponge: an unrecognized potential public health problem.

    PubMed Central

    Bottone, E J; Perez, A A

    1993-01-01

    Pseudomonas aeruginosa folliculitis is a well-known entity that occurs among users of closed-cycle recreational water sources such as whirlpools, swimming pools, and hot tubs. In the absence of this epidemiologic link, isolated cases are difficult to diagnose. We encountered a patient who developed P. aeruginosa folliculitis subsequent to the use of a loofah sponge grossly contaminated with the same P. aeruginosa strain (serotype 10; pyocin type 1/a 4,b) that was recovered from her skin lesions. Furthermore, we demonstrated that sterile unused loofah sponges can serve as the sole growth-promoting substrate for P. aeruginosa. To obviate the potential public health problem of contaminated loofah sponges, it is strongly recommended that manufacturers append, and consumers adhere to, instructions as to the care of loofah sponges, which includes allowing the sponge to dry after use. Images PMID:8458939

  3. Environment and Colonisation Sequence Are Key Parameters Driving Cooperation and Competition between Pseudomonas aeruginosa Cystic Fibrosis Strains and Oral Commensal Streptococci

    PubMed Central

    Whiley, Robert A.; Fleming, Emily V.; Makhija, Ridhima; Waite, Richard D.

    2015-01-01

    Cystic fibrosis (CF) patient airways harbour diverse microbial consortia that, in addition to the recognized principal pathogen Pseudomonas aeruginosa, include other bacteria commonly regarded as commensals. The latter include the oral (viridans) streptococci, which recent evidence indicates play an active role during infection of this environmentally diverse niche. As the interactions between inhabitants of the CF airway can potentially alter disease progression, it is important to identify key cooperators/competitors and environmental influences if therapeutic intervention is to be improved and pulmonary decline arrested. Importantly, we recently showed that virulence of the P. aeruginosa Liverpool Epidemic Strain (LES) could be potentiated by the Anginosus-group of streptococci (AGS). In the present study we explored the relationships between other viridans streptococci (Streptococcus oralis, Streptococcus mitis, Streptococcus gordonii and Streptococcus sanguinis) and the LES and observed that co-culture outcome was dependent upon inoculation sequence and environment. All four streptococcal species were shown to potentiate LES virulence factor production in co-culture biofilms. However, in the case of S. oralis interactions were environmentally determined; in air cooperation within a high cell density co-culture biofilm occurred together with stimulation of LES virulence factor production, while in an atmosphere containing added CO2 this species became a competitor antagonising LES growth through hydrogen peroxide (H2O2) production, significantly altering biofilm population dynamics and appearance. Streptococcus mitis, S. gordonii and S. sanguinis were also capable of H2O2 mediated inhibition of P. aeruginosa growth, but this was only visible when inoculated as a primary coloniser prior to introduction of the LES. Therefore, these observations, which are made in conditions relevant to the biology of CF disease pathogenesis, show that the pathogenic and

  4. Enhanced rhamnolipid production in Burkholderia thailandensis transposon knockout strains deficient in polyhydroxyalkanoate (PHA) synthesis.

    PubMed

    Funston, Scott J; Tsaousi, Konstantina; Smyth, Thomas J; Twigg, Matthew S; Marchant, Roger; Banat, Ibrahim M

    2017-12-01

    Microbially produced rhamnolipids have significant commercial potential; however, the main bacterial producer, Pseudomonas aeruginosa, is an opportunistic human pathogen, which limits biotechnological exploitation. The non-pathogenic species Burkholderia thailandensis produces rhamnolipids; however, yield is relatively low. The aim of this study was to determine whether rhamnolipid production could be increased in Burkholderia thailandensis through mutation of genes responsible for the synthesis of the storage material polyhydroxyalkanoate (PHA), thereby increasing cellular resources for the production of rhamnolipids. Potential PHA target genes were identified in B. thailandensis through comparison with known function genes in Pseudomonas aeruginosa. Multiple knockout strains for the phbA, phbB and phbC genes were obtained and their growth characteristics and rhamnolipid and PHA production determined. The wild-type strain and an rhamnolipid (RL)-deficient strain were used as controls. Three knockout strains (ΔphbA1, ΔphbB1 and ΔphbC1) with the best enhancement of rhamnolipid production were selected for detailed study. ΔphbB1 produced the highest level of purified RL (3.78 g l -1 ) compared to the wild-type strain (1.28 g l -1 ). In ΔphbB1, the proportion of mono-rhamnolipid was also increased compared to the wild-type strain. The production of PHA was reduced by at least 80% in all three phb mutant strains, although never completely eliminated. These results suggest that, in contrast to Pseudomonas aeruginosa, knockout of the PHA synthesis pathway in Burkholderia thailandensis could be used to increase rhamnolipid production. The evidence of residual PHA production in the phb mutant strains suggests B. thailandensis possesses a secondary unelucidated PHA synthesis pathway.

  5. Catalase (KatA) Plays a Role in Protection against Anaerobic Nitric Oxide in Pseudomonas aeruginosa

    PubMed Central

    Su, Shengchang; Panmanee, Warunya; Wilson, Jeffrey J.; Mahtani, Harry K.; Li, Qian; VanderWielen, Bradley D.; Makris, Thomas M.; Rogers, Melanie; McDaniel, Cameron; Lipscomb, John D.; Irvin, Randall T.; Schurr, Michael J.; Lancaster, Jack R.; Kovall, Rhett A.; Hassett, Daniel J.

    2014-01-01

    Pseudomonas aeruginosa (PA) is a common bacterial pathogen, responsible for a high incidence of nosocomial and respiratory infections. KatA is the major catalase of PA that detoxifies hydrogen peroxide (H2O2), a reactive oxygen intermediate generated during aerobic respiration. Paradoxically, PA displays elevated KatA activity under anaerobic growth conditions where the substrate of KatA, H2O2, is not produced. The aim of the present study is to elucidate the mechanism underlying this phenomenon and define the role of KatA in PA during anaerobiosis using genetic, biochemical and biophysical approaches. We demonstrated that anaerobic wild-type PAO1 cells yielded higher levels of katA transcription and expression than aerobic cells, whereas a nitrite reductase mutant ΔnirS produced ∼50% the KatA activity of PAO1, suggesting that a basal NO level was required for the increased KatA activity. We also found that transcription of the katA gene was controlled, in part, by the master anaerobic regulator, ANR. A ΔkatA mutant and a mucoid mucA22 ΔkatA bacteria demonstrated increased sensitivity to acidified nitrite (an NO generator) in anaerobic planktonic and biofilm cultures. EPR spectra of anaerobic bacteria showed that levels of dinitrosyl iron complexes (DNIC), indicators of NO stress, were increased significantly in the ΔkatA mutant, and dramatically in a ΔnorCB mutant compared to basal levels of DNIC in PAO1 and ΔnirS mutant. Expression of KatA dramatically reduced the DNIC levels in ΔnorCB mutant. We further revealed direct NO-KatA interactions in vitro using EPR, optical spectroscopy and X-ray crystallography. KatA has a 5-coordinate high spin ferric heme that binds NO without prior reduction of the heme iron (K d ∼6 μM). Collectively, we conclude that KatA is expressed to protect PA against NO generated during anaerobic respiration. We proposed that such protective effects of KatA may involve buffering of free NO when potentially toxic concentrations of

  6. Pseudomonas Aeruginosa: Resistance to the Max

    PubMed Central

    Poole, Keith

    2011-01-01

    Pseudomonas aeruginosa is intrinsically resistant to a variety of antimicrobials and can develop resistance during anti-pseudomonal chemotherapy both of which compromise treatment of infections caused by this organism. Resistance to multiple classes of antimicrobials (multidrug resistance) in particular is increasingly common in P. aeruginosa, with a number of reports of pan-resistant isolates treatable with a single agent, colistin. Acquired resistance in this organism is multifactorial and attributable to chromosomal mutations and the acquisition of resistance genes via horizontal gene transfer. Mutational changes impacting resistance include upregulation of multidrug efflux systems to promote antimicrobial expulsion, derepression of ampC, AmpC alterations that expand the enzyme's substrate specificity (i.e., extended-spectrum AmpC), alterations to outer membrane permeability to limit antimicrobial entry and alterations to antimicrobial targets. Acquired mechanisms contributing to resistance in P. aeruginosa include β-lactamases, notably the extended-spectrum β-lactamases and the carbapenemases that hydrolyze most β-lactams, aminoglycoside-modifying enzymes, and 16S rRNA methylases that provide high-level pan-aminoglycoside resistance. The organism's propensity to grow in vivo as antimicrobial-tolerant biofilms and the occurrence of hypermutator strains that yield antimicrobial resistant mutants at higher frequency also compromise anti-pseudomonal chemotherapy. With limited therapeutic options and increasing resistance will the untreatable P. aeruginosa infection soon be upon us? PMID:21747788

  7. CsPAO4 of Citrus sinensis functions in polyamine terminal catabolism and inhibits plant growth under salt stress.

    PubMed

    Wang, Wei; Liu, Ji-Hong

    2016-08-18

    Polyamine oxidase (PAO) is a key enzyme catalyzing polyamine catabolism leading to H2O2 production. We previously demonstrated that Citrus sinensis contains six putative PAO genes, but their functions are not well understood. In this work, we reported functional elucidation of CsPAO4 in polyamine catabolism and salt stress response. CsPAO4 was localized to the apoplast and used both spermidine (Spd) and spermine (Spm) as substrates for terminal catabolism. Transgenic plants overexpressing CsPAO4 displayed prominent increase in PAO activity, concurrent with marked decrease of Spm and Spd and elevation of H2O2. Seeds of transgenic lines displayed better germination when compared with wild type (WT) under salt stress. However, both vegetative growth and root elongation of the transgenic lines were prominently inhibited under salt stress, accompanied by higher level of H2O2 and more conspicuous programmed cell death (PCD). Exogenous supply of catalase (CAT), a H2O2 scavenger, partially recovered the vegetative growth and root elongation. In addition, spermine inhibited root growth of transgenic plants. Taken together, these data demonstrated that CsPAO4 accounts for production of H2O2 causing oxidative damages under salt stress and that down-regulation of a PAO gene involved in polyamine terminal catabolism may be an alternative approach for improving salt stress tolerance.

  8. Prevalence and antibiotic resistance of Pseudomonas aeruginosa in water samples in central Italy and molecular characterization of oprD in imipenem resistant isolates

    PubMed Central

    Carloni, Elisa; Andreoni, Francesca; Magi, Silvia; Chironna, Maria; Brandi, Giorgio; Amagliani, Giulia

    2017-01-01

    Scope This study aimed to analyse the prevalence, antibiotic resistance and genetic relatedness of P. aeruginosa isolates obtained from potable and recreational water samples (n. 8,351) collected from different settings (swimming pools, n. 207; healthcare facilities, n 1,684; accommodation facilities, n. 1,518; municipal waterworks, n. 4,500; residential buildings, n. 235). Possible mechanisms underlying resistance to imipenem, with particular focus on those involving oprD-based uptake, were also explored. Methods and results Isolation and identification of Pseudomonas aeruginosa was performed according to the standardized procedure UNI EN ISO 16266:2008 followed by PCR confirmation. Antibiotic Susceptibility testing was conducted according to EUCAST standardized disk diffusion method. Genetic relatedness of strains was carried out by RAPD. The sequence of the oprD gene was analyzed by standard method. Fifty-three samples (0.63%) were positive for P. aeruginosa, of which 10/207 (4.83%) were from swimming pools. Five isolates (9.43%) were resistant to imipenem, one to Ticarcillin + Clavulanate, one to both Piperacillin and Ticarcillin + Clavulanate. The highest isolation rate of imipenem resistant P. aeruginosa was observed in swimming pool water. Identical RAPD profiles were found in isolates from the same location in the same year or even in different years. Conclusions Imipenem resistant strains were identified as carbapenemase-negative and resistance has been associated with inactivating mutations within the oprD gene, with a concomitant loss of porin. RAPD results proved that a water system can remain colonized by one strain for long periods and the contamination may be difficult to eradicate. This study has revealed the presence of P. aeruginosa in different water samples, including resistant strains, especially in swimming pools, and confirmed the role of porins as a contributing factor in carbapenem resistance in Gram-negative bacteria. PMID:29211780

  9. Prevalence and antibiotic resistance of Pseudomonas aeruginosa in water samples in central Italy and molecular characterization of oprD in imipenem resistant isolates.

    PubMed

    Schiavano, Giuditta Fiorella; Carloni, Elisa; Andreoni, Francesca; Magi, Silvia; Chironna, Maria; Brandi, Giorgio; Amagliani, Giulia

    2017-01-01

    This study aimed to analyse the prevalence, antibiotic resistance and genetic relatedness of P. aeruginosa isolates obtained from potable and recreational water samples (n. 8,351) collected from different settings (swimming pools, n. 207; healthcare facilities, n 1,684; accommodation facilities, n. 1,518; municipal waterworks, n. 4,500; residential buildings, n. 235). Possible mechanisms underlying resistance to imipenem, with particular focus on those involving oprD-based uptake, were also explored. Isolation and identification of Pseudomonas aeruginosa was performed according to the standardized procedure UNI EN ISO 16266:2008 followed by PCR confirmation. Antibiotic Susceptibility testing was conducted according to EUCAST standardized disk diffusion method. Genetic relatedness of strains was carried out by RAPD. The sequence of the oprD gene was analyzed by standard method. Fifty-three samples (0.63%) were positive for P. aeruginosa, of which 10/207 (4.83%) were from swimming pools. Five isolates (9.43%) were resistant to imipenem, one to Ticarcillin + Clavulanate, one to both Piperacillin and Ticarcillin + Clavulanate. The highest isolation rate of imipenem resistant P. aeruginosa was observed in swimming pool water. Identical RAPD profiles were found in isolates from the same location in the same year or even in different years. Imipenem resistant strains were identified as carbapenemase-negative and resistance has been associated with inactivating mutations within the oprD gene, with a concomitant loss of porin. RAPD results proved that a water system can remain colonized by one strain for long periods and the contamination may be difficult to eradicate. This study has revealed the presence of P. aeruginosa in different water samples, including resistant strains, especially in swimming pools, and confirmed the role of porins as a contributing factor in carbapenem resistance in Gram-negative bacteria.

  10. Stability, frequency and multiplicity of transposon insertions in the pyoverdine region in the chromosomes of different fluorescent pseudomonads.

    PubMed

    Cornelis, P; Anjaiah, V; Koedam, N; Delfosse, P; Jacques, P; Thonart, P; Neirinckx, L

    1992-07-01

    Tn5 mutagenesis of different fluorescent pseudomonads was achieved by conjugational transfer of the suicide vector pSUP 10141. Pyoverdine negative (Pvd-) mutants were detected by the absence of fluorescence on King's B medium and by their inability to grow in the presence of the iron chelator EDDHA [ethylenediamine di(o-hydroxyphenylacetic acid)]. In P. fluorescens ATCC 17400 and three rhizosphere isolates (one P. putida and two P. fluorescens), the percentage of Pvd- mutants ranged between 0 and 0.54%. In a P. chlororaphis rhizosphere isolate, this percentage was higher (4%). In these mutants both of the Tn5 antibiotic resistances (Km and Tc) were stable and the transposon could be detected by hybridization. In Pvd- mutants of P. fluorescens ATCC 17400, the transposon was found to be inserted twice in the chromosome while single insertions were detected in the DNA of other, randomly tested mutants. In P. aeruginosa PAO1, where 13.1% of the mutants were Pvd-, both antibiotic resistances were rapidly lost and accordingly no transposon insertion could be detected by hybridization. However, the Pvd- phenotype was generally stable in these mutants. The plasmid pNK862 containing a mini-Tn10 transposon was introduced by electroporation into P. aeruginosa PAO1 and Kmr mutants were recovered, 89% of which were Pvd- and confirmed to be P. aeruginosa by PCR amplification of the P. aeruginosa lipoprotein gene. The mini-Tn10 insertions were also found to be unstable in PAO1.

  11. Bioactive Organocopper Compound from Pseudomonas aeruginosa Inhibits the Growth of Xanthomonas citri subsp. citri.

    PubMed

    de Oliveira, Admilton G; Spago, Flavia R; Simionato, Ane S; Navarro, Miguel O P; da Silva, Caroline S; Barazetti, André R; Cely, Martha V T; Tischer, Cesar A; San Martin, Juca A B; de Jesus Andrade, Célia G T; Novello, Cláudio R; Mello, João C P; Andrade, Galdino

    2016-01-01

    Citrus canker is a very destructive disease of citrus species. The challenge is to find new compounds that show strong antibiotic activity and low toxicity to plants and the environment. The objectives of the present study were (1) to extract, purify and evaluate the secondary metabolites with antibiotic activity produced by Pseudomonas aeruginosa LV strain in vitro against Xanthomonas citri subsp. citri (strain 306), (2) to determine the potential of semi-purified secondary metabolites in foliar application to control citrus canker under greenhouse conditions, and (3) to identify antibiotic activity in orange leaf mesophyll infected with strain 306, by electron microscopy. Two pure bioactive compounds were isolated, an organocopper antibiotic compound (OAC) and phenazine-1-carboxamide. Phenazine-1-carboxamide did not show any antibiotic activity under the experimental conditions used in this study. The OAC showed a high level of antibiotic activity with a minimum inhibitory concentration of 0.12 μg mL(-1). In greenhouse tests for control of citrus canker in orange trees, the semi-purified fraction F3d reduced lesion formation by about 97%. The concentration used was 500 times lower than that for the recommended commercial copper-based product. Electron microscopy showed that F3d altered the exopolysaccharide matrix and caused cell lysis of the pathogen inside the citrus canker lesions. These results suggest that secondary metabolites produced by inducing P. aeruginosa LV strain have a high potential to be used as a bioproduct to control citrus canker.

  12. Bioactive Organocopper Compound from Pseudomonas aeruginosa Inhibits the Growth of Xanthomonas citri subsp. citri

    PubMed Central

    de Oliveira, Admilton G.; Spago, Flavia R.; Simionato, Ane S.; Navarro, Miguel O. P.; da Silva, Caroline S.; Barazetti, André R.; Cely, Martha V. T.; Tischer, Cesar A.; San Martin, Juca A. B.; de Jesus Andrade, Célia G. T.; Novello, Cláudio R.; Mello, João C. P.; Andrade, Galdino

    2016-01-01

    Citrus canker is a very destructive disease of citrus species. The challenge is to find new compounds that show strong antibiotic activity and low toxicity to plants and the environment. The objectives of the present study were (1) to extract, purify and evaluate the secondary metabolites with antibiotic activity produced by Pseudomonas aeruginosa LV strain in vitro against Xanthomonas citri subsp. citri (strain 306), (2) to determine the potential of semi-purified secondary metabolites in foliar application to control citrus canker under greenhouse conditions, and (3) to identify antibiotic activity in orange leaf mesophyll infected with strain 306, by electron microscopy. Two pure bioactive compounds were isolated, an organocopper antibiotic compound (OAC) and phenazine-1-carboxamide. Phenazine-1-carboxamide did not show any antibiotic activity under the experimental conditions used in this study. The OAC showed a high level of antibiotic activity with a minimum inhibitory concentration of 0.12 μg mL-1. In greenhouse tests for control of citrus canker in orange trees, the semi-purified fraction F3d reduced lesion formation by about 97%. The concentration used was 500 times lower than that for the recommended commercial copper-based product. Electron microscopy showed that F3d altered the exopolysaccharide matrix and caused cell lysis of the pathogen inside the citrus canker lesions. These results suggest that secondary metabolites produced by inducing P. aeruginosa LV strain have a high potential to be used as a bioproduct to control citrus canker. PMID:26903992

  13. Relevance of multidrug-resistant Pseudomonas aeruginosa infections in cystic fibrosis.

    PubMed

    Stefani, S; Campana, S; Cariani, L; Carnovale, V; Colombo, C; Lleo, M M; Iula, V D; Minicucci, L; Morelli, P; Pizzamiglio, G; Taccetti, G

    2017-09-01

    Multidrug-resistant (MDR) Pseudomonas aeruginosa is an important issue for physicians who take care of patients with cystic fibrosis (CF). Here, we review the latest research on how P. aeruginosa infection causes lung function to decline and how several factors contribute to the emergence of antibiotic resistance in P. aeruginosa strains and influence the course of the infection course. However, many aspects of the practical management of patients with CF infected with MDR P. aeruginosa are still to be established. Less is known about the exact role of susceptibility testing in clinical strategies for dealing with resistant infections, and there is an urgent need to find a tool to assist in choosing the best therapeutic strategy for MDR P. aeruginosa infection. One current perception is that the selection of antibiotic therapy according to antibiogram results is an important component of the decision-making process, but other patient factors, such as previous infection history and antibiotic courses, also need to be evaluated. On the basis of the known issues and the best current data on respiratory infections caused by MDR P. aeruginosa, this review provides practical suggestions to optimize the diagnostic and therapeutic management of patients with CF who are infected with these pathogens. Copyright © 2017 Elsevier GmbH. All rights reserved.

  14. Activity and interactions of antibiotic and phytochemical combinations against Pseudomonas aeruginosa in vitro

    PubMed Central

    Jayaraman, Premkumar; Sakharkar, Meena K; Lim, Chu Sing; Tang, Thean Hock; Sakharkar, Kishore R.

    2010-01-01

    In this study the in vitro activities of seven antibiotics (ciprofloxacin, ceftazidime, tetracycline, trimethoprim, sulfamethoxazole, polymyxin B and piperacillin) and six phytochemicals (protocatechuic acid, gallic acid, ellagic acid, rutin, berberine and myricetin) against five P. aeruginosa isolates, alone and in combination are evaluated. All the phytochemicals under investigation demonstrate potential inhibitory activity against P. aeruginosa. The combinations of sulfamethoxazole plus protocatechuic acid, sulfamethoxazole plus ellagic acid, sulfamethoxazole plus gallic acid and tetracycline plus gallic acid show synergistic mode of interaction. However, the combinations of sulfamethoxazole plus myricetin shows synergism for three strains (PA01, DB5218 and DR3062). The synergistic combinations are further evaluated for their bactericidal activity against P. aeruginosa ATCC strain using time-kill method. Sub-inhibitory dose responses of antibiotics and phytochemicals individually and in combination are presented along with their interaction network to suggest on the mechanism of action and potential targets for the phytochemicals under investigation. The identified synergistic combinations can be of potent therapeutic value against P. aeruginosa infections. These findings have potential implications in delaying the development of resistance as the antibacterial effect is achieved with lower concentrations of both drugs (antibiotics and phytochemicals). PMID:20941374

  15. Application of bioflocculating property of Pseudomonas aeruginosa strain IASST201 in treatment of oil-field formation water.

    PubMed

    Pathak, Mihirjyoti; Devi, Arundhuti; Sarma, Hridip Kumar; Lal, Banwari

    2014-07-01

    A bioflocculating activity of 89.8% was depicted by an activated sludge-borne bacteria Pseudomonas aeruginosa strain IASST201 with a yield of bioflocculant of 2.68 g L(-1) obtained from production media broth after optimization of different parameters. The highest bioflocculation efficiency was found at the pre-stationary phase of the bacterial growth period in the production media broth at 96th hour examined from a growth-flocculation kinetics study. 85.67% of bioflocculation was observed in oil-field formation water, with a separation of 68.7% of aliphatic hydrocarbon contents of the formation water after the application of the bacterial bioflocculant by entrapment mechanism with formation of flocs which was analyzed and examined comparatively through gas-chromatography. Extensive removal of heavy metal contents of the oil-field formation water due to bioflocculation was estimated by Atomic Absorption Spectrophotometer (AAS). The SEM and AFM studies declare the extracellular polymeric nature of the bioflocculant produced by this bacterium clumped within bacterial biofilm supported with FTIR study of the extracted bioflocculant. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  16. IMP-29, a Novel IMP-Type Metallo-β-Lactamase in Pseudomonas aeruginosa

    PubMed Central

    Jeannot, Katy; Poirel, Laurent; Robert-Nicoud, Marjorie; Cholley, Pascal; Nordmann, Patrice

    2012-01-01

    Analysis of two clonally related multiresistant Pseudomonas aeruginosa isolates led to the identification of a novel IMP-type metallo-β-lactamase. IMP-29 was significantly different from the other IMP variants (the closest variant being IMP-5 with 93% amino acid identity). The blaIMP-29 gene cassette was carried by a class 1 integron in strain 10.298, while in strain 10.266 it was located in a rearranged DNA region on a 30-kb conjugative plasmid. Biochemical analysis confirmed that IMP-29 efficiently hydrolyzed carbapenems. PMID:22290960

  17. The Pseudomonas aeruginosa ribbon-helix-helix DNA-binding protein AlgZ (AmrZ) controls twitching motility and biogenesis of type IV pili.

    PubMed

    Baynham, Patricia J; Ramsey, Deborah M; Gvozdyev, Borys V; Cordonnier, Ellen M; Wozniak, Daniel J

    2006-01-01

    Pseudomonas aeruginosa is an opportunistic pathogen that is commonly found in water and soil. In order to colonize surfaces with low water content, P. aeruginosa utilizes a flagellum-independent form of locomotion called twitching motility, which is dependent upon the extension and retraction of type IV pili. This study demonstrates that AlgZ, previously identified as a DNA-binding protein absolutely required for transcription of the alginate biosynthetic operon, is required for twitching motility. AlgZ may be required for the biogenesis or function of type IV pili in twitching motility. Transmission electron microscopy analysis of an algZ deletion in nonmucoid PAO1 failed to detect surface pili. To examine expression and localization of PilA (the major pilin subunit), whole-cell extracts and cell surface pilin preparations were analyzed by Western blotting. While the PilA levels present in whole-cell extracts were similar for wild-type P. aeruginosa and P. aeruginosa with the algZ deletion, the amount of PilA on the surface of the cells was drastically reduced in the algZ mutant. Analysis of algZ and algD mutants indicates that the DNA-binding activity of AlgZ is essential for the regulation of twitching motility and that this is independent of the role of AlgZ in alginate expression. These data show that AlgZ DNA-binding activity is required for twitching motility independently of its role in alginate production and that this involves the surface localization of type IV pili. Given this new role in twitching motility, we propose that algZ (PA3385) be designated amrZ (alginate and motility regulator Z).

  18. Isolation and determination of four potential antimicrobial components from Pseudomonas aeruginosa extracts

    PubMed Central

    Xu, Ling-Qing; Zeng, Jian-Wen; Jiang, Chong-He; Wang, Huan; Li, Yu-Zhen; Wen, Wei-Hong; Li, Jie-Hua; Wang, Feng; Ting, Wei-Jen; Sun, Zi-Yong; Huang, Chih-Yang

    2017-01-01

    Background: Pseudomonas aeruginosa can cause disease and also can be isolated from the skin of healthy people. Additionally, it exhibits certain antimicrobial effects against other microorganisms. Methods: We collected 60 strains of P. aeruginosa and screened their antimicrobial effects against Staphylococcus aureus (ATCC 25923) using the filter paper-disk method, the cross-streaking method and the co-culture method and then evaluated the antimicrobial activity of the chloroform-isolated S. aureus extracts against methicillin-resistant S. aureus (MRSA, Gram-positive cocci), vancomycin intermediate-resistant S. aureus (VISA, Gram-positive cocci), Corynebacterium spp. (CS, Gram-positive bacilli), Acinetobacter baumannii (AB, Gram-negative bacilli), Moraxella catarrhalis (MC, Gram-negative diplococcus), Candida albicans (CA, fungi), Candida tropicalis (CT, fungi), Candida glabrata (CG, fungi) and Candida parapsilosis (CP, fungi). Results: The PA06 and PA46 strains have strong antimicrobial effects. High-performance liquid chromatography (HPLC) analysis revealed that the major components of PA06 and PA46 that exhibit antimicrobial activity are functionally similar to phenazine-1-carboxylic acid (PCA) and pyocyanin. Preparative HPLC was performed to separate and isolate the 4 major potential antimicrobial components: PA06ER10, PA06ER16, PA06ER23 and PA06ER31. Further, the molecular masses of PA06ER10 (260.1), PA06ER16 (274.1), PA06ER23 (286.1) and PA06ER31 (318.2) were determined by electrospray ionization (ESI) mass spectrometry. Conclusion: P. aeruginosa can produce small molecules with potential antimicrobial activities against MRSA, VISA, CS, MC, CA, CT, CG and CP but not against AB. PMID:29200950

  19. Phylogenetic Distribution of CRISPR-Cas Systems in Antibiotic-Resistant Pseudomonas aeruginosa.

    PubMed

    van Belkum, Alex; Soriaga, Leah B; LaFave, Matthew C; Akella, Srividya; Veyrieras, Jean-Baptiste; Barbu, E Magda; Shortridge, Dee; Blanc, Bernadette; Hannum, Gregory; Zambardi, Gilles; Miller, Kristofer; Enright, Mark C; Mugnier, Nathalie; Brami, Daniel; Schicklin, Stéphane; Felderman, Martina; Schwartz, Ariel S; Richardson, Toby H; Peterson, Todd C; Hubby, Bolyn; Cady, Kyle C

    2015-11-24

    Pseudomonas aeruginosa is an antibiotic-refractory pathogen with a large genome and extensive genotypic diversity. Historically, P. aeruginosa has been a major model system for understanding the molecular mechanisms underlying type I clustered regularly interspaced short palindromic repeat (CRISPR) and CRISPR-associated protein (CRISPR-Cas)-based bacterial immune system function. However, little information on the phylogenetic distribution and potential role of these CRISPR-Cas systems in molding the P. aeruginosa accessory genome and antibiotic resistance elements is known. Computational approaches were used to identify and characterize CRISPR-Cas systems within 672 genomes, and in the process, we identified a previously unreported and putatively mobile type I-C P. aeruginosa CRISPR-Cas system. Furthermore, genomes harboring noninhibited type I-F and I-E CRISPR-Cas systems were on average ~300 kb smaller than those without a CRISPR-Cas system. In silico analysis demonstrated that the accessory genome (n = 22,036 genes) harbored the majority of identified CRISPR-Cas targets. We also assembled a global spacer library that aided the identification of difficult-to-characterize mobile genetic elements within next-generation sequencing (NGS) data and allowed CRISPR typing of a majority of P. aeruginosa strains. In summary, our analysis demonstrated that CRISPR-Cas systems play an important role in shaping the accessory genomes of globally distributed P. aeruginosa isolates. P. aeruginosa is both an antibiotic-refractory pathogen and an important model system for type I CRISPR-Cas bacterial immune systems. By combining the genome sequences of 672 newly and previously sequenced genomes, we were able to provide a global view of the phylogenetic distribution, conservation, and potential targets of these systems. This analysis identified a new and putatively mobile P. aeruginosa CRISPR-Cas subtype, characterized the diverse distribution of known CRISPR-inhibiting genes, and

  20. Levofloxacin-imipenem combination prevents the emergence of resistance among clinical isolates of Pseudomonas aeruginosa.

    PubMed

    Lister, Philip D; Wolter, Daniel J

    2005-02-15

    A 2-compartment in vitro pharmacokinetic model (IVPM) was used to assess the potential of a levofloxacin-imipenem combination to prevent the emergence of resistance during treatment of Pseudomonas aeruginosa infection. Log-phase cultures (10(8) cfu/mL) of 3 clinical isolates were inoculated into the peripheral compartment of the IVPMs and were treated with simulated human doses of levofloxacin (750 mg) and imipenem (250 mg). Pharmacodynamics and the emergence of resistance were evaluated over the course of 24 h. Resistant mutants were evaluated for transcriptional expression of specific efflux pumps. Initially, rapid killing was observed in association with each regimen. However, with levofloxacin and imipenem alone, rapid regrowth was observed as a result of the selection of resistant subpopulations. Analysis of mutants selected by levofloxacin demonstrated that mexEF-oprN-overexpressing subpopulations resistant to both levofloxacin and imipenem were selected from cultures of all 3 strains. Nevertheless, the levofloxacin-imipenem combination rapidly eradicated all 3 P. aeruginosa strains. These data suggest that levofloxacin-imipenem may be an effective combination for preventing the emergence of resistance among P. aeruginosa strains, even when subpopulations resistant to both drugs are present. Further studies are warranted to evaluate the use of this combination against strains with established resistance to either or both drugs.

  1. Structural basis for effectiveness of siderophore-conjugated monocarbams against clinically relevant strains of Pseudomonas aeruginosa

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Han, Seungil; Zaniewski, Richard P.; Marr, Eric S.

    Pseudomonas aeruginosa is an opportunistic Gram-negative pathogen that causes nosocomial infections for which there are limited treatment options. Penicillin-binding protein PBP3, a key therapeutic target, is an essential enzyme responsible for the final steps of peptidoglycan synthesis and is covalently inactivated by {beta}-lactam antibiotics. Here we disclose the first high resolution cocrystal structures of the P. aeruginosa PBP3 with both novel and marketed {beta}-lactams. These structures reveal a conformational rearrangement of Tyr532 and Phe533 and a ligand-induced conformational change of Tyr409 and Arg489. The well-known affinity of the monobactam aztreonam for P. aeruginosa PBP3 is due to a distinct hydrophobicmore » aromatic wall composed of Tyr503, Tyr532, and Phe533 interacting with the gem-dimethyl group. The structure of MC-1, a new siderophore-conjugated monocarbam complexed with PBP3 provides molecular insights for lead optimization. Importantly, we have identified a novel conformation that is distinct to the high-molecular-weight class B PBP subfamily, which is identifiable by common features such as a hydrophobic aromatic wall formed by Tyr503, Tyr532, and Phe533 and the structural flexibility of Tyr409 flanked by two glycine residues. This is also the first example of a siderophore-conjugated triazolone-linked monocarbam complexed with any PBP. Energetic analysis of tightly and loosely held computed hydration sites indicates protein desolvation effects contribute significantly to PBP3 binding, and analysis of hydration site energies allows rank ordering of the second-order acylation rate constants. Taken together, these structural, biochemical, and computational studies provide a molecular basis for recognition of P. aeruginosa PBP3 and open avenues for future design of inhibitors of this class of PBPs.« less

  2. Profile of Virulence Factors in the Multi-Drug Resistant Pseudomonas aeruginosa Strains of Human Urinary Tract Infections (UTI).

    PubMed

    Habibi, Asghar; Honarmand, Ramin

    2015-12-01

    Putative virulence factors are responsible for the pathogenicity of UTIs caused by Pseudomonas aeruginosa (P. aeruginosa). Resistance of P. aeruginosa to commonly used antibiotics is caused by the extreme overprescription of those antibiotics. The goal of the present study was to investigate the prevalence of virulence factors and the antibiotic resistance patterns of P. aeruginosa isolates in UTI cases in Iran. Two hundred and fifty urine samples were collected from patients who suffered from UTIs. Samples were cultured immediately, and those that were P. aeruginosa-positive were analyzed for the presence of virulence genes using polymerase chain reaction (PCR) testing. Antimicrobial susceptibility testing (AST) was performed using the disk diffusion method. Of the 250 urine samples analyzed, 8 samples (3.2%) were positive for P. aeruginosa. The prevalence of P. aeruginosa in male and female patients was 2.7% and 3.5%, respectively, (P = 0.035). In patients less than 10 years old, it was 4.2%, and in patients more than 55 years old, it was 4.2%. These were the most commonly infected groups. The highest levels of resistance were seen against ampicillin (87.5%), norfloxacin (62.5%), gentamycin (62.5%), amikacin (62.5%), and aztreonam (62.5%), while the lowest were seen for meropenem (0%), imipenem (12.5%), and polymyxin B (12.5%). LasB (87.5%), pclH (75%), pilB (75%), and exoS (75%) were the most commonly detected virulence factors in the P. aeruginosa isolates. It is logical to first prescribe meropenem, imipenem, and polymyxin B in cases of UTIs caused by P. aeruginosa. Medical practitioners should be aware of the presence of levels of antibiotic resistance in hospitalized UTI patients in Iran.

  3. Oxygen-dependent regulation of c-di-GMP synthesis by SadC controls alginate production in Pseudomonas aeruginosa.

    PubMed

    Schmidt, Annika; Hammerbacher, Anna Silke; Bastian, Mike; Nieken, Karen Jule; Klockgether, Jens; Merighi, Massimo; Lapouge, Karine; Poschgan, Claudia; Kölle, Julia; Acharya, K Ravi; Ulrich, Martina; Tümmler, Burkhard; Unden, Gottfried; Kaever, Volkhard; Lory, Stephen; Haas, Dieter; Schwarz, Sandra; Döring, Gerd

    2016-10-01

    Pseudomonas aeruginosa produces increased levels of alginate in response to oxygen-deprived conditions. The regulatory pathway(s) that links oxygen limitation to increased synthesis of alginate has remained elusive. In the present study, using immunofluorescence microscopy, we show that anaerobiosis-induced alginate production by planktonic PAO1 requires the diguanylate cyclase (DGC) SadC, previously identified as a regulator of surface-associated lifestyles. Furthermore, we found that the gene products of PA4330 and PA4331, located in a predicted operon with sadC, have a major impact on alginate production: deletion of PA4330 (odaA, for oxygen-dependent alginate synthesis activator) caused an alginate production defect under anaerobic conditions, whereas a PA4331 (odaI, for oxygen-dependent alginate synthesis inhibitor) deletion mutant produced alginate also in the presence of oxygen, which would normally inhibit alginate synthesis. Based on their sequence, OdaA and OdaI have predicted hydratase and dioxygenase reductase activities, respectively. Enzymatic assays using purified protein showed that unlike OdaA, which did not significantly affect DGC activity of SadC, OdaI inhibited c-di-GMP production by SadC. Our data indicate that SadC, OdaA and OdaI are components of a novel response pathway of P. aeruginosa that regulates alginate synthesis in an oxygen-dependent manner. © 2016 Society for Applied Microbiology and John Wiley & Sons Ltd.

  4. Bactericidal Activity of Ceragenin CSA-13 in Cell Culture and in an Animal Model of Peritoneal Infection.

    PubMed

    Bucki, Robert; Niemirowicz, Katarzyna; Wnorowska, Urszula; Byfield, Fitzroy J; Piktel, Ewelina; Wątek, Marzena; Janmey, Paul A; Savage, Paul B

    2015-10-01

    Ceragenins constitute a novel family of cationic antibiotics characterized by a broad spectrum of antimicrobial activities, which have mostly been assessed in vitro. Using a polarized human lung epithelial cell culture system, we evaluated the antibacterial activities of the ceragenin CSA-13 against two strains of Pseudomonas aeruginosa (PAO1 and Xen5). Additionally, the biodistribution and bactericidal activity of a CSA-13-IRDye 800CW derivate were assessed using an animal model of peritoneal infection after PAO1 challenge. In cell culture, CSA-13 bactericidal activities against PAO1 and Xen5 were higher than the activities of the human cathelicidin peptide LL-37. Increased CSA-13 activity was observed in polarized human lung epithelial cell cultures subjected to butyric acid treatment, which is known to increase endogenous LL-37 production. Eight hours after intravenous or intraperitoneal injection, the greatest CSA-13-IRDye 800CW accumulation was observed in mouse liver and kidneys. CSA-13-IRDye 800CW administration resulted in decreased bacterial outgrowth from abdominal fluid collected from animals subjected to intraperitoneal PAO1 infection. These observations indicate that CSA-13 may synergistically interact with antibacterial factors that are naturally present at mucosal surfaces and it maintains its antibacterial activity in the infected abdominal cavity. Cationic lipids such as CSA-13 represent excellent candidates for the development of new antibacterial compounds. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  5. Isolation and characterization of novel strains of Pseudomonas aeruginosa and Serratia marcescens possessing high efficiency to degrade gasoline, kerosene, diesel oil, and lubricating oil.

    PubMed

    Wongsa, Patcharaporn; Tanaka, Makiko; Ueno, Akio; Hasanuzzaman, Mohammad; Yumoto, Isao; Okuyama, Hidetoshi

    2004-12-01

    Bacteria possessing high capacity to degrade gasoline, kerosene, diesel oil, and lubricating oil were screened from several areas of Hokkaido, Japan. Among isolates, two strains, WatG and HokM, which were identified as new strains of Pseudomonas aeruginosa and Serratia marcescens species, respectively, showed relatively high capacity and wide spectrum to degrade the hydrocarbons in gasoline, kerosene, diesel, and lubricating oil. About 90-95% of excess amount of total diesel oil and kerosene added to mineral salts media as a sole carbon source could be degraded by WatG within 2 and 3 weeks, respectively. The same amount of lubricating oil was 60% degraded within 2 weeks. Strain HokM was more capable than WatG in degrading aromatic compounds in gasoline. This strain could also degrade kerosene, diesel, and lubricating oil with a capacity of 50-60%. Thus, these two isolates have potential to be useful for bioremediation of sites highly contaminated with petroleum hydrocarbons.

  6. Pseudomonas aeruginosa ventilator-associated pneumonia management.

    PubMed

    Ramírez-Estrada, Sergio; Borgatta, Bárbara; Rello, Jordi

    2016-01-01

    Ventilator-associated pneumonia is the most common infection in intensive care unit patients associated with high morbidity rates and elevated economic costs; Pseudomonas aeruginosa is one of the most frequent bacteria linked with this entity, with a high attributable mortality despite adequate treatment that is increased in the presence of multiresistant strains, a situation that is becoming more common in intensive care units. In this manuscript, we review the current management of ventilator-associated pneumonia due to P. aeruginosa, the most recent antipseudomonal agents, and new adjunctive therapies that are shifting the way we treat these infections. We support early initiation of broad-spectrum antipseudomonal antibiotics in present, followed by culture-guided monotherapy de-escalation when susceptibilities are available. Future management should be directed at blocking virulence; the role of alternative strategies such as new antibiotics, nebulized treatments, and vaccines is promising.

  7. Quorum sensing systems differentially regulate the production of phenazine-1-carboxylic acid in the rhizobacterium Pseudomonas aeruginosa PA1201

    PubMed Central

    Sun, Shuang; Zhou, Lian; Jin, Kaiming; Jiang, Haixia; He, Ya-Wen

    2016-01-01

    Pseudomonas aeruginosa strain PA1201 is a newly identified rhizobacterium that produces high levels of the secondary metabolite phenazine-1-carboxylic acid (PCA), the newly registered biopesticide Shenqinmycin. PCA production in liquid batch cultures utilizing a specialized PCA-promoting medium (PPM) typically occurs after the period of most rapid growth, and production is regulated in a quorum sensing (QS)-dependent manner. PA1201 contains two PCA biosynthetic gene clusters phz1 and phz2; both clusters contribute to PCA production, with phz2 making a greater contribution. PA1201 also contains a complete set of genes for four QS systems (LasI/LasR, RhlI/RhlR, PQS/MvfR, and IQS). By using several methods including gene deletion, the construction of promoter-lacZ fusion reporter strains, and RNA-Seq analysis, this study investigated the effects of the four QS systems on bacterial growth, QS signal production, the expression of phz1 and phz2, and PCA production. The possible mechanisms for the strain- and condition-dependent expression of phz1 and phz2 were discussed, and a schematic model was proposed. These findings provide a basis for further genetic engineering of the QS systems to improve PCA production. PMID:27456813

  8. Epidemic Pseudomonas aeruginosa infection in patients with cystic fibrosis is not a risk factor for poor clinical Outcomes following lung transplantation.

    PubMed

    Pritchard, Julia; Thakrar, Mitesh V; Somayaji, Ranjani; Surette, Michael G; Rabin, Harvey R; Helmersen, Doug; Lien, Dale; Purighalla, Swathi; Waddell, Barbara; Parkins, Michael D

    2016-05-01

    Epidemic strains of Pseudomonas aeruginosa (ePA) causing infection in cystic fibrosis (CF) have been commonly identified from clinics around the world. ePA disproportionally impacts CF patient pre-transplant outcomes manifesting in increased exacerbation frequency, worsened treatment burden and increased rate of lung function decline, and disproportionally leads to death and/or transplantation. As other CF factors such as pre-transplant infection with multi-resistant organisms, and isolation of P. aeruginosa in the post transplant graft, may impact post-transplant outcomes, we sought to determine if infection with ePA similarly adversely impact post-transplant outcomes. Between 1991-2014, 53 CF patients from our center received lung transplants. Bacterial strain typing was performed retrospectively on isolates collected prior to transplantation. Comprehensive chart reviews were performed to obtain baseline patient characteristics and post-transplant outcomes. Of the 53 transplanted patients, 57% of patients were infected with ePA prior to transplant; the other 43% of patients had unique strains of P. aeruginosa. Mean age at transplant was 29.0years for ePA and 33.3years for unique (p=0.04). There were no differences in overall survival (HR=0.75, 95% CI 0.31-1.79), bronchiolitis obliterans syndrome (BOS) free survival (HR 1.43, 95% CI 0.54-4.84) or all other assessed outcomes including exacerbation frequency, chronic renal failure, acute cellular rejections, Aspergillus infection, airway stenosis, and post-transplant lymphoproliferative disorder. Unlike pre-transplant outcomes, CF patients infected with ePA do not experience worse post-transplant outcomes than those infected with unique strains. Therefore, lung transplantation should be considered for all patients with P. aeruginosa infection and end stage lung disease, irrespective of infection with ePA. Copyright © 2015 European Cystic Fibrosis Society. Published by Elsevier B.V. All rights reserved.

  9. Investigation of the enhanced antimicrobial activity of combination dry powder inhaler formulations of lactoferrin.

    PubMed

    Marshall, Lindsay J; Oguejiofor, Wilson; Price, Robert; Shur, Jagdeep

    2016-12-05

    The airways of most people with cystic fibrosis are colonized with biofilms of the Gram-negative, opportunistic pathogen Pseudomonas aeruginosa. Delivery of antibiotics directly to the lung in the form of dry powder aerosols offers the potential to achieve high local concentrations directly to the biofilms. Unfortunately, current aerosolised antibiotic regimes are unable to efficiently eradicate these biofilms from the airways. We investigated the ability of the innate antimicrobial, lactoferrin, to enhance the activity of two aminoglycoside antibiotics (tobramycin and gentamicin) against biofilms of P. aeruginosa strain PAO1. Biofilms were prepared in 96 well polystyrene plates. Combinations of the antibiotics and various lactoferrin preparations were spray dried. The bacterial cell viability of the various spray dried combinations was determined. Iron-free lactoferrin (apo lactoferrin) induced a 3-log reduction in the killing of planktonic cell by the aminoglycoside antibiotics (p<0.01) and also reduced both the formation and persistence of P. aeruginosa biofilms (p<0.01). Combinations of lactoferrin and an aminoglycoside displays potential as an effective new therapeutic strategy in the treatment of P. aeruginosa biofilms infections such as those typical of the CF lungs. Copyright © 2016 Elsevier B.V. All rights reserved.

  10. Anti-biofilm activity of biogenic selenium nanoparticles and selenium dioxide against clinical isolates of Staphylococcus aureus, Pseudomonas aeruginosa, and Proteus mirabilis.

    PubMed

    Shakibaie, Mojtaba; Forootanfar, Hamid; Golkari, Yaser; Mohammadi-Khorsand, Tayebe; Shakibaie, Mohammad Reza

    2015-01-01

    The aim of the present study was to investigate the anti-biofilm activity of biologically synthesized selenium nanoparticles (Se NPs) against the biofilm produced by clinically isolated bacterial strains compared to that of selenium dioxide. Thirty strains of Staphylococcus aureus, Pseudomonas aeruginosa, and Proteus mirabilis were isolated from various specimens of the patients hospitalized in different hospitals (Kerman, Iran). Quantification of the biofilm using microtiter plate assay method introduced 30% of S. aureus, 13% of P. aeruginosa and 17% of P. mirabilis isolates as severely adherent strains. Transmission electron micrograph (TEM) of the purified Se NPs (produced by Bacillus sp. MSh-1) showed individual and spherical nano-structure in the size range of 80-220nm. Obtained results of the biofilm formation revealed that selenium nanoparticles inhibited the biofilm of S. aureus, P. aeruginosa, and P. mirabilis by 42%, 34.3%, and 53.4%, respectively, compared to that of the non-treated samples. Effect of temperature and pH on the biofilm formation in the presence of Se NPs and SeO2 was also evaluated. Copyright © 2014 Elsevier GmbH. All rights reserved.

  11. Dynamics of the Action of Biocides in Pseudomonas aeruginosa Biofilms▿†

    PubMed Central

    Bridier, A.; Dubois-Brissonnet, F.; Greub, G.; Thomas, V.; Briandet, R.

    2011-01-01

    The biocidal activity of peracetic acid (PAA) and benzalkonium chloride (BAC) on Pseudomonas aeruginosa biofilms was investigated by using a recently developed confocal laser scanning microscopy (CLSM) method that enables the direct and real-time visualization of cell inactivation within the structure. This technique is based on monitoring the loss of fluorescence that corresponds to the leakage of a fluorophore out of cells due to membrane permeabilization by the biocides. Although this approach has previously been used with success with various Gram-positive species, it is not directly applicable to the visualization of Gram-negative strains such as P. aeruginosa, particularly because of limitations regarding fluorescence staining. After adapting the staining procedure to P. aeruginosa, the action of PAA and BAC on the biofilm formed by strain ATCC 15442 was investigated. The results revealed specific inactivation patterns as a function of the mode of action of the biocides. While PAA treatment triggered a uniform loss of fluorescence in the structure, the action of BAC was first localized at the periphery of cell clusters and then gradually spread throughout the biofilm. Visualization of the action of BAC in biofilms formed by three clinical isolates then confirmed the presence of a delay in penetration, showing that diffusion-reaction limitations could provide a major explanation for the resistance of P. aeruginosa biofilms to this biocide. Biochemical analysis suggested a key role for extracellular matrix characteristics in these processes. PMID:21422224

  12. Cryo-transmission electron microscopy of frozen-hydrated sections of Escherichia coli and Pseudomonas aeruginosa.

    PubMed

    Matias, Valério R F; Al-Amoudi, Ashraf; Dubochet, Jacques; Beveridge, Terry J

    2003-10-01

    High-pressure freezing of Escherichia coli K-12 and Pseudomonas aeruginosa PAO1 in the presence of cryoprotectants provided consistent vitrification of cells so that frozen-hydrated sections could be cut, providing approximately 2-nm resolution of structure. The size and shape of the bacteria, as well as their surface and cytoplasmic constituents, were nicely preserved and compared well with other published high-resolution techniques. Cells possessed a rich cytoplasm containing a diffuse dispersion of ribosomes and genetic material. Close examination of cells revealed that the periplasmic space was compressed during cryosectioning, a finding which provided supporting evidence that this space is filled by a compressible gel. Since the outer membrane and peptidoglycan layer are bonded together via lipoproteins, the space between them (although still part of the periplasmic space) was not as compacted. Even when this cryosectioning compression was taken into account, there was still substantial variability in the width of the periplasmic space. It is possible that the protoplast has some capacity to float freely within the periplasm.

  13. Preliminary study : optimization of pH and salinity for biosurfactant production from Pseudomonas aeruginosa in diesel fuel and crude oil medium

    NASA Astrophysics Data System (ADS)

    Ikhwani, A. Z. N.; Nurlaila, H. S.; Ferdinand, F. D. K.; Fachria, R.; Hasan, A. E. Z.; Yani, M.; Setyawati, I.; Suryani

    2017-03-01

    Biosurfactant is secondary metabolite surface active compound produced by microorganisms which is nontoxic and eco-friendly. Microorganism producing biosurfactant that is quite potential to use in many applications is from Pseudomonas aeruginosa strains. Good quality of biosurfactant production from microbes is supported by the suitable nutrients and environmental factors. The aim of this research was to obtain preliminary o data upon the optimum pH and salinity for the production of biosurfactant from Pseudomonas aeruginosa ATCC 15442 in diesel fuel and crude oil medium. P. aeruginosa ATCC 15442 cultured in diesel fuel and crude oil as carbon source showed biosurfactant activity. P.aeruginosa-derived biosurfactant was capable to form stable emulsion for 24 hours (EI24) in hydrocarbons n-hexane solutions. The particular biosurfactant showed EI24 highest value at pH 7 (31.02%) and 1% NaCl (24.00%) when P. aeruginosa was grown in 10% diesel fuel medium in mineral salt solution. As for the media crude oil, the highest EI24 value was at pH 6 (52.16%) and 1% NaCl (33.30%).

  14. Establishing quality control ranges for antimicrobial susceptibility testing of Escherichia coli, Pseudomonas aeruginosa, and Staphylococcus aureus: a cornerstone to develop reference strains for Korean clinical microbiology laboratories.

    PubMed

    Hong, Sung Kuk; Choi, Seung Jun; Shin, Saeam; Lee, Wonmok; Pinto, Naina; Shin, Nari; Lee, Kwangjun; Hong, Seong Geun; Kim, Young Ah; Lee, Hyukmin; Kim, Heejung; Song, Wonkeun; Lee, Sun Hwa; Yong, Dongeun; Lee, Kyungwon; Chong, Yunsop

    2015-11-01

    Quality control (QC) processes are being performed in the majority of clinical microbiology laboratories to ensure the performance of microbial identification and antimicrobial susceptibility testing by using ATCC strains. To obtain these ATCC strains, some inconveniences are encountered concerning the purchase cost of the strains and the shipping time required. This study was focused on constructing a database of reference strains for QC processes using domestic bacterial strains, concentrating primarily on antimicrobial susceptibility testing. Three strains (Escherichia coli, Pseudomonas aeruginosa, and Staphylococcus aureus) that showed legible results in preliminary testing were selected. The minimal inhibitory concentrations (MICs) and zone diameters (ZDs) of eight antimicrobials for each strain were determined according to the CLSI M23. All resulting MIC and ZD ranges included at least 95% of the data. The ZD QC ranges obtained by using the CLSI method were less than 12 mm, and the MIC QC ranges extended no more than five dilutions. This study is a preliminary attempt to construct a bank of Korean QC strains. With further studies, a positive outcome toward cost and time reduction can be anticipated.

  15. Intricate interactions between the bloom-forming cyanobacterium Microcystis aeruginosa and foreign genetic elements, revealed by diversified clustered regularly interspaced short palindromic repeat (CRISPR) signatures.

    PubMed

    Kuno, Sotaro; Yoshida, Takashi; Kaneko, Takakazu; Sako, Yoshihiko

    2012-08-01

    Clustered regularly interspaced short palindromic repeats (CRISPR) confer sequence-dependent, adaptive resistance in prokaryotes against viruses and plasmids via incorporation of short sequences, called spacers, derived from foreign genetic elements. CRISPR loci are thus considered to provide records of past infections. To describe the host-parasite (i.e., cyanophages and plasmids) interactions involving the bloom-forming freshwater cyanobacterium Microcystis aeruginosa, we investigated CRISPR in four M. aeruginosa strains and in two previously sequenced genomes. The number of spacers in each locus was larger than the average among prokaryotes. All spacers were strain specific, except for a string of 11 spacers shared in two closely related strains, suggesting diversification of the loci. Using CRISPR repeat-based PCR, 24 CRISPR genotypes were identified in a natural cyanobacterial community. Among 995 unique spacers obtained, only 10 sequences showed similarity to M. aeruginosa phage Ma-LMM01. Of these, six spacers showed only silent or conservative nucleotide mutations compared to Ma-LMM01 sequences, suggesting a strategy by the cyanophage to avert CRISPR immunity dependent on nucleotide identity. These results imply that host-phage interactions can be divided into M. aeruginosa-cyanophage combinations rather than pandemics of population-wide infectious cyanophages. Spacer similarity also showed frequent exposure of M. aeruginosa to small cryptic plasmids that were observed only in a few strains. Thus, the diversification of CRISPR implies that M. aeruginosa has been challenged by diverse communities (almost entirely uncharacterized) of cyanophages and plasmids.

  16. Intricate Interactions between the Bloom-Forming Cyanobacterium Microcystis aeruginosa and Foreign Genetic Elements, Revealed by Diversified Clustered Regularly Interspaced Short Palindromic Repeat (CRISPR) Signatures

    PubMed Central

    Kuno, Sotaro; Kaneko, Takakazu; Sako, Yoshihiko

    2012-01-01

    Clustered regularly interspaced short palindromic repeats (CRISPR) confer sequence-dependent, adaptive resistance in prokaryotes against viruses and plasmids via incorporation of short sequences, called spacers, derived from foreign genetic elements. CRISPR loci are thus considered to provide records of past infections. To describe the host-parasite (i.e., cyanophages and plasmids) interactions involving the bloom-forming freshwater cyanobacterium Microcystis aeruginosa, we investigated CRISPR in four M. aeruginosa strains and in two previously sequenced genomes. The number of spacers in each locus was larger than the average among prokaryotes. All spacers were strain specific, except for a string of 11 spacers shared in two closely related strains, suggesting diversification of the loci. Using CRISPR repeat-based PCR, 24 CRISPR genotypes were identified in a natural cyanobacterial community. Among 995 unique spacers obtained, only 10 sequences showed similarity to M. aeruginosa phage Ma-LMM01. Of these, six spacers showed only silent or conservative nucleotide mutations compared to Ma-LMM01 sequences, suggesting a strategy by the cyanophage to avert CRISPR immunity dependent on nucleotide identity. These results imply that host-phage interactions can be divided into M. aeruginosa-cyanophage combinations rather than pandemics of population-wide infectious cyanophages. Spacer similarity also showed frequent exposure of M. aeruginosa to small cryptic plasmids that were observed only in a few strains. Thus, the diversification of CRISPR implies that M. aeruginosa has been challenged by diverse communities (almost entirely uncharacterized) of cyanophages and plasmids. PMID:22636003

  17. Molecular Epidemiology and Mechanisms of Carbapenem Resistance in Pseudomonas aeruginosa Isolates from Spanish Hospitals▿

    PubMed Central

    Gutiérrez, O.; Juan, C.; Cercenado, E.; Navarro, F.; Bouza, E.; Coll, P.; Pérez, J. L.; Oliver, A.

    2007-01-01

    All (236) Pseudomonas aeruginosa isolates resistant to imipenem and/or meropenem collected during a multicenter (127-hospital) study in Spain were analyzed. Carbapenem-resistant isolates were found to be more frequently resistant to all β-lactams and non-β-lactam antibiotics than carbapenem-susceptible isolates (P < 0.001), and up to 46% of the carbapenem-resistant isolates met the criteria used to define multidrug resistance (MDR). Pulsed-field gel electrophoresis revealed remarkable clonal diversity (165 different clones were identified), and with few exceptions, the levels of intra- and interhospital dissemination of clones were found to be low. Carbapenem resistance was driven mainly by the mutational inactivation of OprD, accompanied or not by the hyperexpression of AmpC or MexAB-OprM. Class B carbapenemases (metallo-β-lactamases [MBLs]) were detected in a single isolate, although interestingly, this isolate belonged to one of the few epidemic clones documented. The MBL-encoding gene (blaVIM-2), along with the aminoglycoside resistance determinants, was transferred to strain PAO1 by electroporation, demonstrating its plasmid location. The class 1 integron harboring blaVIM-2 was characterized as well, and two interesting features were revealed: intI1 was found to be disrupted by a 1.1-kb insertion sequence, and a previously undescribed aminoglycoside acetyltransferase-encoding gene [designated aac(6′)-32] preceded blaVIM-2. AAC(6′)-32 showed 80% identity to AAC(6′)-Ib′ and the recently described AAC(6′)-31, and when aac(6′)-32 was cloned into Escherichia coli, it conferred resistance to tobramycin and reduced susceptibility to gentamicin and amikacin. Despite the currently low prevalence of epidemic clones with MDR, active surveillance is needed to detect and prevent the dissemination of these clones, particularly those producing integron- and plasmid-encoded MBLs, given their additional capacity for the intra- and interspecies spread of MDR

  18. Leaf Extracts of Mangifera indica L. Inhibit Quorum Sensing – Regulated Production of Virulence Factors and Biofilm in Test Bacteria

    PubMed Central

    Husain, Fohad M.; Ahmad, Iqbal; Al-thubiani, Abdullah S.; Abulreesh, Hussein H.; AlHazza, Ibrahim M.; Aqil, Farrukh

    2017-01-01

    Quorum sensing (QS) is a global gene regulatory mechanism in bacteria for various traits including virulence factors. Disabling QS system with anti-infective agent is considered as a potential strategy to prevent bacterial infection. Mangifera indica L. (mango) has been shown to possess various biological activities including anti-QS. This study investigates the efficacy of leaf extracts on QS-regulated virulence factors and biofilm formation in Gram negative pathogens. Mango leaf (ML) extract was tested for QS inhibition and QS-regulated virulence factors using various indicator strains. It was further correlated with the biofilm inhibition and confirmed by electron microscopy. Phytochemical analysis was carried out using ultra performance liquid chromatography (UPLC) and gas chromatography–mass spectrometry (GC-MS) analysis. In vitro evaluation of anti-QS activity of ML extracts against Chromobacterium violaceum revealed promising dose-dependent interference in violacein production, by methanol extract. QS inhibitory activity is also demonstrated by reduction in elastase (76%), total protease (56%), pyocyanin (89%), chitinase (55%), exopolysaccharide production (58%) and swarming motility (74%) in Pseudomonas aeruginosa PAO1 at 800 μg/ml concentration. Biofilm formation by P. aeruginosa PAO1 and Aeromonas hydrophila WAF38 was reduced considerably (36–82%) over control. The inhibition of biofilm was also observed by scanning electron microscopy. Moreover, ML extracts significantly reduced mortality of Caenorhabditis elegans pre-infected with PAO1 at the tested concentration. Phytochemical analysis of active extracts revealed very high content of phenolics in methanol extract and a total of 14 compounds were detected by GC-MS and UPLC. These findings suggest that phytochemicals from the ML could provide bioactive anti-infective and needs further investigation to isolate and uncover their therapeutic efficacy. PMID:28484444

  19. The OprB porin plays a central role in carbohydrate uptake in Pseudomonas aeruginosa.

    PubMed

    Wylie, J L; Worobec, E A

    1995-06-01

    Using interposon mutagenesis, we have generated strains of Pseudomonas aeruginosa which lack or overexpress the substrate-selective OprB porin of this species. A marked decrease or increase in the initial uptake of glucose by these strains verified the role of OprB in facilitating the diffusion of glucose across the outer membrane of P. aeruginosa. However, we also demonstrated that the loss or overexpression of OprB had a similar effect on the uptake of three other sugars able to support the growth of this bacterium (mannitol, glycerol, and fructose). This effect was restricted to carbohydrate transport; arginine uptake was identical in mutant and wild-type strains. These results indicated that OprB cannot be considered strictly a glucose-selective porin; rather, it acts as a central component of carbohydrate transport and is more accurately described as a carbohydrate-selective porin.

  20. Innovative material containing the natural product curcumin, with enhanced antimicrobial properties for active packaging.

    PubMed

    Papadimitriou, A; Ketikidis, I; Stathopoulou, M-E K; Banti, C N; Papachristodoulou, C; Zoumpoulakis, L; Agathopoulos, S; Vagenas, G V; Hadjikakou, S K

    2018-03-01

    Curcumin (Curc) reacts with zinc di‑iodine (ZnI 2 ) in 2:1molar ratio in the presence of an excess of a base triethylamine ((CH 3 CH 2 ) 3 N) in methanol (CH 3 OH) solution towards the amorphous solid material of formula [ZnI 2 (Curc) 2 ] (1). The complex was characterized by melting point (m.p.), Fourier Transform-Infra Red (FT-IR) and Nuclear Magnetic Resonance of hydrogen nucleus ( 1 H NMR) spectroscopy. The formula of 1 was determined by X-ray fluorescence (XRF) analysis. The retention of the structure in solution was confirmed by 1 H NMR spectroscopy. The antimicrobial activity of the complex has been studied against the bacteria Pseudomonas aeruginosa (PAO1). The Minimum Inhibitory Concentrations (MIC) of the compounds 1 and Curc against P. aeruginosa (PAO1) are: 71.3μΜ (75.3μg/mL) for [ZnI 2 (Curc) 2 ] and 339μM (125μg/mL) for Curc, respectively. Moreover, the antimicrobial activity of the new material which was diffused in polystyrene against biofilm formed by PAO1 was also calculated. Copyright © 2017 Elsevier B.V. All rights reserved.

  1. Pseudomonas aeruginosa ventilator-associated pneumonia management

    PubMed Central

    Ramírez-Estrada, Sergio; Borgatta, Bárbara; Rello, Jordi

    2016-01-01

    Ventilator-associated pneumonia is the most common infection in intensive care unit patients associated with high morbidity rates and elevated economic costs; Pseudomonas aeruginosa is one of the most frequent bacteria linked with this entity, with a high attributable mortality despite adequate treatment that is increased in the presence of multiresistant strains, a situation that is becoming more common in intensive care units. In this manuscript, we review the current management of ventilator-associated pneumonia due to P. aeruginosa, the most recent antipseudomonal agents, and new adjunctive therapies that are shifting the way we treat these infections. We support early initiation of broad-spectrum antipseudomonal antibiotics in present, followed by culture-guided monotherapy de-escalation when susceptibilities are available. Future management should be directed at blocking virulence; the role of alternative strategies such as new antibiotics, nebulized treatments, and vaccines is promising. PMID:26855594

  2. Identification of the putrescine biosynthetic genes in Pseudomonas aeruginosa and characterization of agmatine deiminase and N-carbamoylputrescine amidohydrolase of the arginine decarboxylase pathway.

    PubMed

    Nakada, Yuji; Itoh, Yoshifumi

    2003-03-01

    Putrescine can be synthesized either directly from ornithine by ornithine decarboxylase (ODC; the speC product) or indirectly from arginine via arginine decarboxylase (ADC; the speA product). The authors identified the speA and speC genes in Pseudomonas aeruginosa PAO1. The activities of the two decarboxylases were similar and each enzyme alone appeared to direct sufficient formation of the polyamine for normal growth. A mutant defective in both speA and speC was a putrescine auxotroph. In this strain, agmatine deiminase (the aguA product) and N-carbamoylputrescine amidohydrolase (the aguB product), which were initially identified as the catabolic enzymes of agmatine, biosynthetically convert agmatine to putrescine in the ADC pathway: a double mutant of aguAB and speC was a putrescine auxotroph. AguA was purified as a homodimer of 43 kDa subunits and AguB as a homohexamer of 33 kDa subunits. AguA specifically deiminated agmatine with K(m) and K(cat) values of 0.6 mM and 4.2 s(-1), respectively. AguB was specific to N-carbamoylputrescine and the K(m) and K(cat) values of the enzyme for the substrate were 0.5 mM and 3.3 s(-1), respectively. Whereas AguA has no structural relationship to any known C-N hydrolases, AguB is a protein of the nitrilase family that performs thiol-assisted catalysis. Inhibition by SH reagents and the conserved cysteine residue in AguA and its homologues suggested that this enzyme is also involved in thiol-mediated catalysis.

  3. Activity of MK-7655 combined with imipenem against Enterobacteriaceae and Pseudomonas aeruginosa.

    PubMed

    Livermore, David M; Warner, Marina; Mushtaq, Shazad

    2013-10-01

    MK-7655 is a novel inhibitor of class A and C β-lactamases. We investigated its potential to protect imipenem. Chequerboard MICs were determined by CLSI agar dilution: (i) for Enterobacteriaceae with carbapenemases; (ii) for Enterobacteriaceae with carbapenem resistance contingent on combinations of impermeability together with an extended-spectrum β-lactamase or AmpC enzyme; and (iii) for Pseudomonas aeruginosa and other non-fermenters. At a concentration of 4 mg/L, MK-7655 reduced imipenem MICs for Enterobacteriaceae with KPC carbapenemases from 16-64 mg/L to 0.12-1 mg/L. Synergy also was seen for Enterobacteriaceae with impermeability-mediated carbapenem resistance, with weaker synergy seen for isolates with the OXA-48 enzyme. On the other hand, MK-7655 failed to potentiate imipenem against Enterobacteriaceae with metallo-carbapenemases. In the case of P. aeruginosa, where endogenous AmpC confers slight protection versus imipenem, 4 mg/L MK-7655 reduced the MIC of imipenem for all isolates, except those with metallo-carbapenemases: the MICs of imipenem fell from 1-2 mg/L to 0.25-0.5 mg/L for imipenem-susceptible P. aeruginosa and from 16-64 mg/L to 1-4 mg/L for OprD-deficient strains. No potentiation was seen for chryseobacteria or for Stenotrophomonas maltophilia. MK-7655 potentiated imipenem against Enterobacteriaceae with KPC carbapenemases or combinations of β-lactamase and impermeability, but not those with metallo-carbapenemases. It augmented the activity of imipenem against P. aeruginosa in general and OprD mutants in particular.

  4. Isolation and characterization of T7-like lytic bacteriophages infecting multidrug resistant Pseudomonas aeruginosa isolated from Egypt.

    PubMed

    El Didamony, Gamal; Askora, Ahmed; Shehata, Aya A

    2015-06-01

    In this study, two lytic phages designated as ϕPSZ1 and ϕPSZ2 infecting multidrug resistant Pseudomonas aeruginosa were isolated from sewage samples collected in Zagazig, Egypt. Morphological analysis by transmission electron microscopy revealed that both phages belong to the podoviridae family and resembles typical T7-like phages. ϕPSZ1 has a head of about 60 ± 5 nm in diameter with a short tail of 19 ± 2 nm in length, while ϕPSZ2 has a head of about 57 ± 5 nm in diameter with a short tail of 14 ± 2 nm in length. Both phages were shown to be able to infect 13 different P. aeruginosa strains and has no effect on other tested bacteria. In spite of morphological similarity, these phages showed diverged genomic sequences revealed by restriction enzyme digestion analysis. One-step growth curves of bacteriophages revealed eclipse and latent periods of 12 min for ϕPSZ1 and 15 min for ϕPSZ2, respectively, with burst sizes of about 100 per infected cell. Phage treatment prevented the growth of P. aeruginosa for up to 18 h with multiplicity of infection ratios of 1. These results suggest that both phages have a high potential for phage application to control P. aeruginosa.

  5. Engineered probiotic Escherichia coli can eliminate and prevent Pseudomonas aeruginosa gut infection in animal models

    PubMed Central

    Hwang, In Young; Koh, Elvin; Wong, Adison; March, John C.; Bentley, William E.; Lee, Yung Seng; Chang, Matthew Wook

    2017-01-01

    Bacteria can be genetically engineered to kill specific pathogens or inhibit their virulence. We previously developed a synthetic genetic system that allows a laboratory strain of Escherichia coli to sense and kill Pseudomonas aeruginosa in vitro. Here, we generate a modified version of the system, including a gene encoding an anti-biofilm enzyme, and use the probiotic strain Escherichia coli Nissle 1917 as host. The engineered probiotic shows in vivo prophylactic and therapeutic activity against P. aeruginosa during gut infection in two animal models (Caenorhabditis elegans and mice). These findings support the further development of engineered microorganisms with potential prophylactic and therapeutic activities against gut infections. PMID:28398304

  6. Effect of Catechins, Green tea Extract and Methylxanthines in Combination with Gentamicin Against Staphylococcus aureus and Pseudomonas aeruginosa

    PubMed Central

    Fazly Bazzaz, Bibi Sedigheh; Sarabandi, Sahar; Khameneh, Bahman; Hosseinzadeh, Hossein

    2016-01-01

    Objectives: Bacterial resistant infections have become a global health challenge and threaten the society’s health. Thus, an urgent need exists to find ways to combat resistant pathogens. One promising approach to overcoming bacterial resistance is the use of herbal products. Green tea catechins, the major green tea polyphenols, show antimicrobial activity against resistant pathogens. The present study aimed to investigate the effect of catechins, green tea extract, and methylxanthines in combination with gentamicin against standard and clinical isolates of Staphylococcus aureus (S. aureus) and the standard strain of Pseudomonas aeruginosa (P. aeruginosa). Methods: The minimum inhibitory concentration (MIC) and the minimum bactericidal concentration (MBC) values of different agents against bacterial strains were determined. The interactions of green tea extract, epigallate catechin, epigallocatechin gallate, two types of methylxanthine, caffeine, and theophylline with gentamicin were studied in vitro by using a checkerboard method and calculating the fraction inhibitory concentration index (FICI). Results: The MICs of gentamicin against bacterial strains were in the range of 0.312 - 320 μg/mL. The MIC values of both types of catechins were 62.5 - 250 μg/ mL. Green tea extract showed insufficient antibacterial activity when used alone. Methylxanthines had no intrinsic inhibitory activity against any of the bacterial strains tested. When green tea extract and catechins were combined with gentamicin, the MIC values of gentamicin against the standard strains and a clinical isolate were reduced, and synergistic activities were observed (FICI < 1). A combination of caffeine with gentamicin did not alter the MIC values of gentamicin. Conclusion: The results of the present study revealed that green tea extract and catechins potentiated the antimicrobial action of gentamicin against some clinical isolates of S. aureus and standard P. aeruginosa strains. Therefore

  7. Assessment of PaO2/FiO2 for stratification of patients with moderate and severe acute respiratory distress syndrome

    PubMed Central

    Villar, Jesús; Blanco, Jesús; del Campo, Rafael; Andaluz-Ojeda, David; Díaz-Domínguez, Francisco J; Muriel, Arturo; Córcoles, Virgilio; Suárez-Sipmann, Fernando; Tarancón, Concepción; González-Higueras, Elena; López, Julia; Blanch, Lluis; Pérez-Méndez, Lina; Fernández, Rosa Lidia; Kacmarek, Robert M

    2015-01-01

    Objectives A recent update of the definition of acute respiratory distress syndrome (ARDS) proposed an empirical classification based on ratio of arterial partial pressure of oxygen to fraction of inspired oxygen (PaO2/FiO2) at ARDS onset. Since the proposal did not mandate PaO2/FiO2 calculation under standardised ventilator settings (SVS), we hypothesised that a stratification based on baseline PaO2/FiO2 would not provide accurate assessment of lung injury severity. Design A prospective, multicentre, observational study. Setting A network of teaching hospitals. Participants 478 patients with eligible criteria for moderate (100<PaO2/FiO2≤200) and severe (PaO2/FiO2≤100) ARDS and followed until hospital discharge. Interventions We examined physiological and ventilator parameters in association with the PaO2/FiO2 at ARDS onset, after 24 h of usual care and at 24 h under a SVS. At 24 h, patients were reclassified as severe, moderate, mild (200<PaO2/FiO2≤300) ARDS and non-ARDS (PaO2/FiO2>300). Primary and secondary outcomes Group severity and hospital mortality. Results At ARDS onset, 173 patients had a PaO2/FiO2≤100 but only 38.7% met criteria for severe ARDS at 24 h under SVS. When assessed under SVS, 61.3% of patients with severe ARDS were reclassified as moderate, mild and non-ARDS, while lung severity and hospital mortality changed markedly with every PaO2/FiO2 category (p<0.000001). Our model of risk stratification outperformed the stratification using baseline PaO2/FiO2 and non-standardised PaO2/FiO2 at 24 h, when analysed by the predictive receiver operating characteristic (ROC) curve: area under the ROC curve for stratification at baseline was 0.583 (95% CI 0.525 to 0.636), 0.605 (95% CI 0.552 to 0.658) at 24 h without SVS and 0.693 (95% CI 0.645 to 0.742) at 24 h under SVS (p<0.000001). Conclusions Our findings support the need for patient assessment under SVS at 24 h after ARDS onset to assess disease severity, and have implications for the

  8. The effect of in silico targeting Pseudomonas aeruginosa patatin-like protein D, for immunogenic administration.

    PubMed

    Chirani, Alireza Salimi; Majidzadeh, Robabeh; Pouriran, Ramin; Heidary, Mohsen; Nasiri, Mohammad Javad; Gholami, Mehrdad; Goudarzi, Mehdi; Omrani, Vahid Fallah

    2018-02-05

    The vaccine candidates that have been introduced for immunization against Pseudomonas aeruginosa (P. aeruginosa) strains are quite diverse. In fact, there has been no proper antigen to act as an effective immunogenic substance against this ubiquitous pathogen in the market as yet. The complications caused by this bacterium due to the rapid development of multiple drug resistant strains have led to clinical problems worldwide. P. aeruginosa encodes many specific virulence elements that could be used as appropriate vaccine candidates. Type Vd secretion system, also known as patatin-like protein D, is a novel P. aeruginosa auto-transporter system. It is known that cellular or humoral immune responses could be elevated by chimeric proteins carrying epitopes. It has been recognized that in silico tools are essential for the evaluation of new chimeric antigens. In this study, we have considered the patatin-like protein D (PlpD) molecule from P. aeruginosa and predicted some immunogenic properties of this strong cytotoxic phospholipase A2 with the use of in-depth computational and immunoinformatics assessment methods The novelty of our in silico study is the modeling and assessment of both humoral and cellular immune potential against the PlpD molecule. The molecule was considered by multiple sequence alignment and homology valuation. The extremely conserved regions in the PlpD were predicted. The allergenic and physicochemical property predictions on the PlpD state that the molecule is a non-allergic and stable molecule. High-resolution secondary and tertiary conformations were created. Indeed, the B-cell and T-cell epitope mapping on the chimeric target protein confirmed that the engineered protein contained a tremendous number of both B-cell and T-cell corresponding epitopes. This investigation magnificently attained the chimeric molecule as being a potent lipolytic enzyme composed of numerous B-cell and T-cell restricted epitopes and could induce both humoral and

  9. Characterization of elastase-deficient clinical isolates of Pseudomonas aeruginosa.

    PubMed Central

    Hamood, A N; Griswold, J; Colmer, J

    1996-01-01

    Elastase production in Pseudomonas aeruginosa is regulated by the lasR, lasI, rhlR, and rhlI genes. Recently, we have analyzed several clinical isolates of P. aeruginosa for the production of elastase and other extracellular virulence factors. Four of these isolates (CIT1, CIW5, CIW7, and CIW8) produced no elastolytic activity. We have characterized these isolates with respect to their elastase-deficient phenotype. Elastase was detected by immunoblotting experiments using elastase-specific antiserum. We also determined the presence of IasB and IasR mRNAs by Northern (RNA) blot hybridization experiments using lasB and lasR internal probes, respectively. None of the four elastase-deficient strains produced either the elastase protein or the lasB mRNA. Complementation experiments (using plasmids carrying either the lasB or the lasR gene) were conducted to determine if the isolates carry defective lasB or lasR genes. The presence of either a lasB or a lasR plasmid in CIW7 and CIW8 resulted in the production of very low levels of elastase and lasB mRNA. Neither elastase nor lasB mRNA was detected in CIT1 and CIW5 carrying the lasB plasmid. The presence of the lasR plasmid in CIT1 and CIW5 resulted in the production of lasB mRNA and elastase protein in CIW5 only. All elastase-deficient strains produced detectable levels of lasR mRNA which were enhanced in the presence of the lasR plasmid. The Pseudomonas autoinducer (which is encoded by lasI) was also produced by all strains. CIT1 produced both hemolysin and alkaline protease but was defective in pyocyanin production. These results suggest that (i) CIT1 may contain a defect in a lasB-regulatory gene, (ii) CIW5 carries a defect within lasR, and (iii) the defect in isolates CIW7 and CIW8 affects the efficiency of lasB transcription. PMID:8757847

  10. Study on the resistance mechanism via outer membrane protein OprD2 and metal β-lactamase expression in the cell wall of Pseudomonas aeruginosa.

    PubMed

    Cai, Shuangqi; Chen, Yiqiang; Song, Dezhi; Kong, Jinliang; Wu, Yanbin; Lu, Huasong

    2016-11-01

    The aim of the present study was to evaluate the imipenem-resistant mechanism via the outer membrane protein (OMP) OprD2 and metal β-lactamase expression in the cell wall of Pseudomonas aeruginosa . The Pseudomonas aeruginosa was clinically separated and validated by VITEK-2 full-automatic bacteria analyzer. Drug resistance, sensitive antibiotics and minimum inhibitory concentration (MIC) were tested using the drug sensitivity analysis system. The phenotype positive strains of MBL genes were screened using the Kirby-Bauer diffusion method by adding metal ion-chelating agent EDTA on the imipenem susceptibility paper. IMP-1, VIM-1 and SPM metaloenzyme genes were tested by polymerase chain reaction (PCR)-telomeric repeat amplification protocol (TRAP). The OMP OprD2 genes were tested by PCR-TRAP, and the protein expression was tested using western blot analysis. The location of OMP OprD2 was confirmed using the sodium salicylate inhibition test. The results showed that 80 portions (40%) of MBL-positive strains were screened out of 200 specimens. Imipenem-resistant Pseudomonas aeruginosa (IRPA) and MIC values were significantly higher than quality control bacteria and control bacteria (P<0.05). A total of 35 cases with IMP-1 positive, 20 with VIM-1 positive, 16 with SPM positive, 5 with 2 positive genes and 4 with 3 positive genes were screened among MBL positive strains. A total of 150 portions (75%) of OprD2 deficiencies were screened from 200 specimens. The standard strains and sensitive strains showed OprD2 protein bands at 45 kDa while no OprD2 protein bands appeared in OprD2 deficiency strains. It was in accordance with gene detection. In conclusion, OMP OprD2 deficiency and MBL phenotype positivity may be important mechanisms of IRPA.

  11. Comprehensive MALDI-TOF biotyping of the non-redundant Harvard Pseudomonas aeruginosa PA14 transposon insertion mutant library.

    PubMed

    Oumeraci, Tonio; Jensen, Vanessa; Talbot, Steven R; Hofmann, Winfried; Kostrzewa, Markus; Schlegelberger, Brigitte; von Neuhoff, Nils; Häussler, Susanne

    2015-01-01

    Pseudomonas aeruginosa is a gram-negative bacterium that is ubiquitously present in the aerobic biosphere. As an antibiotic-resistant facultative pathogen, it is a major cause of hospital-acquired infections. Its rapid and accurate identification is crucial in clinical and therapeutic environments. In a large-scale MALDI-TOF mass spectrometry-based screen of the Harvard transposon insertion mutant library of P. aeruginosa strain PA14, intact-cell proteome profile spectra of 5547 PA14 transposon mutants exhibiting a plethora of different phenotypes were acquired and analyzed. Of all P. aeruginosa PA14 mutant profiles 99.7% were correctly identified as P. aeruginosa with the Biotyper software on the species level. On the strain level, 99.99% of the profiles were mapped to five different individual P. aeruginosa Biotyper database entries. A principal component analysis-based approach was used to determine the most important discriminatory mass features between these Biotyper groups. Although technical replicas were consistently categorized to specific Biotyper groups in 94.2% of the mutant profiles, biological replicas were not, indicating that the distinct proteotypes are affected by growth conditions. The PA14 mutant profile collection presented here constitutes the largest coherent P. aeruginosa MALDI-TOF spectral dataset publicly available today. Transposon insertions in thousands of different P. aeruginosa genes did not affect species identification from MALDI-TOF mass spectra, clearly demonstrating the robustness of the approach. However, the assignment of the individual spectra to sub-groups proved to be non-consistent in biological replicas, indicating that the differentiation between biotyper groups in this nosocomial pathogen is unassured.

  12. Catalytic activity of the two-component flavin-dependent monooxygenase from Pseudomonas aeruginosa toward cinnamic acid derivatives.

    PubMed

    Furuya, Toshiki; Kino, Kuniki

    2014-02-01

    4-Hydroxyphenylacetate 3-hydroxylases (HPAHs) of the two-component flavin-dependent monooxygenase family are attractive enzymes that possess the catalytic potential to synthesize valuable ortho-diphenol compounds from simple monophenol compounds. In this study, we investigated the catalytic activity of HPAH from Pseudomonas aeruginosa strain PAO1 toward cinnamic acid derivatives. We prepared Escherichia coli cells expressing the hpaB gene encoding the monooxygenase component and the hpaC gene encoding the oxidoreductase component. E. coli cells expressing HpaBC exhibited no or very low oxidation activity toward cinnamic acid, o-coumaric acid, and m-coumaric acid, whereas they rapidly oxidized p-coumaric acid to caffeic acid. Interestingly, after p-coumaric acid was almost completely consumed, the resulting caffeic acid was further oxidized to 3,4,5-trihydroxycinnamic acid. In addition, HpaBC exhibited oxidation activity toward 3-(4-hydroxyphenyl)propanoic acid, ferulic acid, and coniferaldehyde to produce the corresponding ortho-diphenols. We also investigated a flask-scale production of caffeic acid from p-coumaric acid as the model reaction for HpaBC-catalyzed syntheses of hydroxycinnamic acids. Since the initial concentrations of the substrate p-coumaric acid higher than 40 mM markedly inhibited its HpaBC-catalyzed oxidation, the reaction was carried out by repeatedly adding 20 mM of this substrate to the reaction mixture. Furthermore, by using the HpaBC whole-cell catalyst in the presence of glycerol, our experimental setup achieved the high-yield production of caffeic acid, i.e., 56.6 mM (10.2 g/L) within 24 h. These catalytic activities of HpaBC will provide an easy and environment-friendly synthetic approach to hydroxycinnamic acids.

  13. Pseudomonas aeruginosa prevalence, antibiotic resistance and antimicrobial use in Chinese burn wards from 2007 to 2014

    PubMed Central

    Dou, Yi; Guo, Feng; Zhou, Zengding; Shi, Yan

    2017-01-01

    Objective To assess the application of antibacterial agents, alongside pathogen prevalence and Pseudomonas aeruginosa drug resistance, with the aim of understanding the impact of inappropriate antibacterial use. Methods This retrospective study assessed bacteria from wounds, catheters, blood, faeces, urine and sputum of hospitalized patients in burn wards between 2007 and 2014. The intensity of use of antibacterial agents and resistance of P. aeruginosa to common anti-Gram-negative antibiotics were measured. Results Annual detection rates of Staphylococcus aureus were significantly decreased, whereas annual detection rates of P. aeruginosa and Klebsiella pneumoniae were significantly increased. Multidrug-resistant strains of P. aeruginosa were increased. The intensity of use of some anti-Gramnegative antibiotics positively correlated with resistance rates of P. aeruginosa to similar antimicrobials. Conclusion In burn wards, more attention should be paid to P. aeruginosa and K. pneumoniae. The use of ciprofloxacin, ceftazidime and cefoperazone/sulbactam should be limited to counter the related increase in resistance levels. PMID:28443385

  14. Requirements for Pseudomonas aeruginosa Type I-F CRISPR-Cas Adaptation Determined Using a Biofilm Enrichment Assay.

    PubMed

    Heussler, Gary E; Miller, Jon L; Price, Courtney E; Collins, Alan J; O'Toole, George A

    2016-11-15

    CRISPR (clustered regularly interspaced short palindromic repeat)-Cas (CRISPR-associated protein) systems are diverse and found in many archaea and bacteria. These systems have mainly been characterized as adaptive immune systems able to protect against invading mobile genetic elements, including viruses. The first step in this protection is acquisition of spacer sequences from the invader DNA and incorporation of those sequences into the CRISPR array, termed CRISPR adaptation. Progress in understanding the mechanisms and requirements of CRISPR adaptation has largely been accomplished using overexpression of cas genes or plasmid loss assays; little work has focused on endogenous CRISPR-acquired immunity from viral predation. Here, we developed a new biofilm-based assay system to enrich for Pseudomonas aeruginosa strains with new spacer acquisition. We used this assay to demonstrate that P. aeruginosa rapidly acquires spacers protective against DMS3vir, an engineered lytic variant of the Mu-like bacteriophage DMS3, through primed CRISPR adaptation from spacers present in the native CRISPR2 array. We found that for the P. aeruginosa type I-F system, the cas1 gene is required for CRISPR adaptation, recG contributes to (but is not required for) primed CRISPR adaptation, recD is dispensable for primed CRISPR adaptation, and finally, the ability of a putative priming spacer to prime can vary considerably depending on the specific sequences of the spacer. Our understanding of CRISPR adaptation has expanded largely through experiments in type I CRISPR systems using plasmid loss assays, mutants of Escherichia coli, or cas1-cas2 overexpression systems, but there has been little focus on studying the adaptation of endogenous systems protecting against a lytic bacteriophage. Here we describe a biofilm system that allows P. aeruginosa to rapidly gain spacers protective against a lytic bacteriophage. This approach has allowed us to probe the requirements for CRISPR adaptation in

  15. Diguanylate cyclase activity of the Mycobacterium leprae T cell antigen ML1419c

    PubMed Central

    Rotcheewaphan, Suwatchareeporn; Belisle, John T.; Webb, Kristofor J.; Kim, Hee-Jin; Spencer, John S.

    2016-01-01

    The second messenger, bis-(3′,5′)-cyclic dimeric guanosine monophosphate (cyclic di-GMP), is involved in the control of multiple bacterial phenotypes, including those that impact host–pathogen interactions. Bioinformatics analyses predicted that Mycobacterium leprae, an obligate intracellular bacterium and the causative agent of leprosy, encodes three active diguanylate cyclases. In contrast, the related pathogen Mycobacterium tuberculosis encodes only a single diguanylate cyclase. One of the M. leprae unique diguanylate cyclases (ML1419c) was previously shown to be produced early during the course of leprosy. Thus, functional analysis of ML1419c was performed. The gene encoding ML1419c was cloned and expressed in Pseudomonas aeruginosa PAO1 to allow for assessment of cyclic di-GMP production and cyclic di-GMP-mediated phenotypes. Phenotypic studies revealed that ml1419c expression altered colony morphology, motility and biofilm formation of P. aeruginosa PAO1 in a manner consistent with increased cyclic di-GMP production. Direct measurement of cyclic di-GMP levels by liquid chromatography–mass spectrometry confirmed that ml1419c expression increased cyclic di-GMP production in P. aeruginosa PAO1 cultures in comparison to the vector control. The observed phenotypes and increased levels of cyclic di-GMP detected in P. aeruginosa expressing ml1419c could be abrogated by mutation of the active site in ML1419c. These studies demonstrated that ML1419c of M. leprae functions as diguanylate cyclase to synthesize cyclic di-GMP. Thus, this protein was renamed DgcA (Diguanylate cyclase A). These results also demonstrated the ability to use P. aeruginosa as a heterologous host for characterizing the function of proteins involved in the cyclic di-GMP pathway of a pathogen refractory to in vitro growth, M. leprae. PMID:27450520

  16. Diguanylate cyclase activity of the Mycobacterium leprae T cell antigen ML1419c.

    PubMed

    Rotcheewaphan, Suwatchareeporn; Belisle, John T; Webb, Kristofor J; Kim, Hee-Jin; Spencer, John S; Borlee, Bradley R

    2016-09-01

    The second messenger, bis-(3',5')-cyclic dimeric guanosine monophosphate (cyclic di-GMP), is involved in the control of multiple bacterial phenotypes, including those that impact host-pathogen interactions. Bioinformatics analyses predicted that Mycobacterium leprae, an obligate intracellular bacterium and the causative agent of leprosy, encodes three active diguanylate cyclases. In contrast, the related pathogen Mycobacterium tuberculosis encodes only a single diguanylate cyclase. One of the M. leprae unique diguanylate cyclases (ML1419c) was previously shown to be produced early during the course of leprosy. Thus, functional analysis of ML1419c was performed. The gene encoding ML1419c was cloned and expressed in Pseudomonas aeruginosa PAO1 to allow for assessment of cyclic di-GMP production and cyclic di-GMP-mediated phenotypes. Phenotypic studies revealed that ml1419c expression altered colony morphology, motility and biofilm formation of P. aeruginosa PAO1 in a manner consistent with increased cyclic di-GMP production. Direct measurement of cyclic di-GMP levels by liquid chromatography-mass spectrometry confirmed that ml1419c expression increased cyclic di-GMP production in P. aeruginosa PAO1 cultures in comparison to the vector control. The observed phenotypes and increased levels of cyclic di-GMP detected in P. aeruginosa expressing ml1419c could be abrogated by mutation of the active site in ML1419c. These studies demonstrated that ML1419c of M. leprae functions as diguanylate cyclase to synthesize cyclic di-GMP. Thus, this protein was renamed DgcA (Diguanylate cyclase A). These results also demonstrated the ability to use P. aeruginosa as a heterologous host for characterizing the function of proteins involved in the cyclic di-GMP pathway of a pathogen refractory to in vitro growth, M. leprae.

  17. Isolation and Molecular Characterization of a Model Antagonistic Pseudomonas aeruginosa Divulging In Vitro Plant Growth Promoting Characteristics.

    PubMed

    Uzair, Bushra; Kausar, Rehana; Bano, Syeda Asma; Fatima, Sammer; Badshah, Malik; Habiba, Ume; Fasim, Fehmida

    2018-01-01

    The use of microbial technologies in agriculture is currently expanding quite rapidly with the identification of new bacterial strains, which are more effective in promoting plant growth. In the present study 18 strains of Pseudomonas were isolated from soil sample of Balochistan coastline. Among isolated Pseudomonas strains four designated as SP19, SP22, PS24, and SP25 exhibited biocontrol activities against phytopathogenic fungi, that is, Rhizopus microsporus, Fusarium oxysporum, Aspergillus niger, Alternaria alternata, and Penicillium digitatum ; PS24 identified as Pseudomonas aeruginosa by 16srRNA gene bank accession number EU081518 was selected on the basis of its antifungal activity to explore its potential as plant growth promotion. PS24 showed multiple plant growth promoting attributes such as phosphate solubilization activity, indole acetic acid (IAA), siderophore, and HCN production. In order to determine the basis for antifungal properties, antibiotics were extracted from King B broth of PS24 and analyzed by TLC. Pyrrolnitrin antibiotic was detected in the culture of strain PS24. PS24 exhibited antifungal activities found to be positive for hydrogen cyanide synthase Hcn BC gene. Sequencing of gene of Hcn BC gene of strain PS24 revealed 99% homology with the Pseudomonas aeruginosa strain PA01 . The sequence of PS24 had been submitted in gene bank accession number KR605499. Ps. aeruginosa PS24 with its multifunctional biocontrol possessions can be used to bioprotect the crop plants from phytopathogens.

  18. Gallium induces the production of virulence factors in Pseudomonas aeruginosa.

    PubMed

    García-Contreras, Rodolfo; Pérez-Eretza, Berenice; Lira-Silva, Elizabeth; Jasso-Chávez, Ricardo; Coria-Jiménez, Rafael; Rangel-Vega, Adrián; Maeda, Toshinari; Wood, Thomas K

    2014-02-01

    The novel antimicrobial gallium is a nonredox iron III analogue with bacteriostatic and bactericidal properties, effective for the treatment of Pseudomonas aeruginosa in vitro and in vivo in mouse and rabbit infection models. It interferes with iron metabolism, transport, and presumably its homeostasis. As gallium exerts its antimicrobial effects by competing with iron, we hypothesized that it ultimately will lead cells to an iron deficiency status. As iron deficiency promotes the expression of virulence factors in vitro and promotes the pathogenicity of P. aeruginosa in animal models, it is anticipated that treatment with gallium will also promote the production of virulence factors. To test this hypothesis, the reference strain PA14 and two clinical isolates from patients with cystic fibrosis were exposed to gallium, and their production of pyocyanin, rhamnolipids, elastase, alkaline protease, alginate, pyoverdine, and biofilm was determined. Gallium treatment induced the production of all the virulence factors tested in the three strains except for pyoverdine. In addition, as the Ga-induced virulence factors are quorum sensing controlled, co-administration of Ga and the quorum quencher brominated furanone C-30 was assayed, and it was found that C-30 alleviated growth inhibition from gallium. Hence, adding both C-30 and gallium may be more effective in the treatment of P. aeruginosa infections. © 2013 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.

  19. [Prevalence of Acinetobacter baumannii and Pseudomonas aeruginosa isolates resistant to imipenem by production of metallo-β-lactamases in Rabat Military Teaching Hospital Mohammed V].

    PubMed

    Gildas Comlan Zohoun, Alban; Moket, Danièle; El Hamzaoui, Sakina

    2013-01-01

    We studied the production of metallo-β-lactamases (MBL) in Acinetobacter baumannii and Pseudomonas aeruginosa strains resistant to imipenem at the Rabat Mohammed V military teaching hospital, according to Yong et al.'s method, using a sterilized solution of EDTA 0.5 M pH 8. One hundred and five bacterial strains (48 A. baumannii and 57 P. aeruginosa) were identified. 45 (42.9%) with 34 A. baumannii and 11 P. aeruginosa were resistant to imipenem. The prevalence of MBL producing strains was 22.2% (10/45). The existence of this isolates resistant to imipenem by producing metallo-β-lactamases is an emerging public health problem. It is necessary to implemente infection control programs to avoid spreading of multidrug resistant bacteria.

  20. The OprB porin plays a central role in carbohydrate uptake in Pseudomonas aeruginosa.

    PubMed Central

    Wylie, J L; Worobec, E A

    1995-01-01

    Using interposon mutagenesis, we have generated strains of Pseudomonas aeruginosa which lack or overexpress the substrate-selective OprB porin of this species. A marked decrease or increase in the initial uptake of glucose by these strains verified the role of OprB in facilitating the diffusion of glucose across the outer membrane of P. aeruginosa. However, we also demonstrated that the loss or overexpression of OprB had a similar effect on the uptake of three other sugars able to support the growth of this bacterium (mannitol, glycerol, and fructose). This effect was restricted to carbohydrate transport; arginine uptake was identical in mutant and wild-type strains. These results indicated that OprB cannot be considered strictly a glucose-selective porin; rather, it acts as a central component of carbohydrate transport and is more accurately described as a carbohydrate-selective porin. PMID:7768797