Occurrence of the Plasmid-Mediated Fluoroquinolone Resistance qepA1 Gene in Two Clonal Clinical Isolates of CTX-M-15-Producing Escherichia coli from Algeria.

PubMed

Yanat, Betitera; Dali Yahia, Radia; Yazi, Leila; Machuca, Jesús; Díaz-De-Alba, Paula; Touati, Abdelaziz; Pascual, Álvaro; Rodríguez-Martínez, José-Manuel

2017-06-01

QepA is a plasmid-mediated quinolone resistance determinant of low prevalence described worldwide, mainly in Enterobacteriaceae. This study describes, for the first time in Algeria, two clonally related, QepA-producing Escherichia coli clinical isolates positive for CTX-M-15. The clonal spread of these multidrug-resistant isolates is a major public health concern.

Clinical and biochemical significance of toxin production by Aeromonas hydrophila.

PubMed Central

Kindschuh, M; Pickering, L K; Cleary, T G; Ruiz-Palacios, G

1987-01-01

Production of cytotoxin and enterotoxin by Aeromonas strains obtained from stools of 50 children in Mexico and Texas and from blood of 9 children with sepsis was determined. Results were correlated with clinical features of infected children as well as with biochemical traits of Aeromonas strains. Cytotoxin was produced by 40 of 42 Aeromonas strains (95%) isolated from stools of children with diarrhea, by all 8 isolates from stools of well children, and by all 9 isolates from children with sepsis. There was no difference in the quantities (amount of cytotoxin per milligram of protein required to kill 50% of the cells) of cytotoxin produced and in clinical manifestations among the groups. None of the isolates produced a toxin that could be neutralized by antiserum raised against Shiga toxin produced by Shigella dysenteriae 1 60R. Heat-labile-like enterotoxin (LT) was produced by 26 of 42 stool isolates (62%), while only 1 of the 42 isolates (2%) produced enterotoxinlike activity in suckling mice; 65% of the cytotoxin-producing strains also produced an LT-like material. All strains from blood produced LT-like material, and 2 of 6 (33%) produced activity in suckling mice. All strains produced hemolysin; 37 of 57 (65%) were Voges-Proskauer positive; 27 of 57 (47%) were lysine decarboxylase positive by API 20E strips, none were positive for lysine decarboxylose production by lysin-iron agar slants at 24 h, but 17 of 54 (31%) were positive at 48 h. There was no correlation between biochemical reactions and enterotoxin or cytotoxin production. There appears to be no correlation between toxin production by Aeromonas spp. and gastroenteritis. PMID:3584426

Uropathogenic Escherichia coli ST131 in urinary tract infections in children.

PubMed

Yun, Ki Wook; Lee, Mi-Kyung; Kim, Wonyong; Lim, In Seok

2017-07-01

Escherichia coli sequence type (ST) 131, a multidrug-resistant clone causing extraintestinal infections, has rapidly become prevalent worldwide. However, the epidemiological and clinical features of pediatric infections are poorly understood. We aimed to explore the characteristics of ST131 Escherichia coli isolated from Korean children with urinary tract infections. We examined 114 uropathogenic E. coli (UPEC) isolates from children hospitalized at Chung-Ang University Hospital between 2011 and 2014. Bacterial strains were classified into STs by partial sequencing of seven housekeeping genes ( adk , fumC , gyrB , icd , mdh , purA , and recA ). Clinical characteristics and antimicrobial susceptibility were compared between ST131 and non-ST131 UPEC isolates. Sixteen UPEC isolates (14.0%) were extended-spectrum β-lactamase (ESBL)-producers; 50.0% of ESBL-producers were ST131 isolates. Of all the isolates tested, 13.2% (15 of 114) were classified as ST131. There were no statistically significant associations between ST131 and age, sex, or clinical characteristics, including fever, white blood cell counts in urine and serum, C-reactive protein, radiologic abnormalities, and clinical outcome. However, ST131 isolates showed significantly lower rates of susceptibility to cefazolin (26.7%), cefotaxime (40.0%), cefepime (40.0%), and ciprofloxacin (53.3%) than non-ST131 isolates (65.7%, 91.9%, 92.9%, and 87.9%, respectively; P <0.001 for all). ESBL was more frequently produced in ST131 (53.3%) than in non-ST131 (8.1%) isolates ( P <0.01). ST131 E. coli isolates were prevalent uropathogens in children at a single medical center in Korea between 2011 and 2014. Although ST131 isolates showed higher rates of antimicrobial resistance, clinical presentation and outcomes of patients were similar to those of patients infected with non-ST131 isolates.

Molecular and epidemiological characterization of IMP-type metallo-β-lactamase-producing Enterobacter cloacae in a Large tertiary care hospital in Japan.

PubMed

Hayakawa, Kayoko; Miyoshi-Akiyama, Tohru; Kirikae, Teruo; Nagamatsu, Maki; Shimada, Kayo; Mezaki, Kazuhisa; Sugiki, Yuko; Kuroda, Emi; Kubota, Shiho; Takeshita, Nozomi; Kutsuna, Satoshi; Tojo, Masayoshi; Ohmagari, Norio

2014-06-01

IMP-type metallo-β-lactamase enzymes have been reported in different geographical areas and in various Gram-negative bacteria. However, the risk factors and epidemiology pertaining to IMP-type metallo-β-lactamase-producing Enterobacter cloacae (IMP-producing E. cloacae) have not been systematically evaluated. We conducted a retrospective, matched case-control study of patients from whom IMP-producing E. cloacae isolates were obtained, in addition to performing thorough molecular analyses of the clinically obtained IMP-producing E. cloacae isolates. Unique cases with IMP-producing E. cloacae isolation were included. Patients with IMP-producing E. cloacae were matched to uninfected controls at a ratio of 1 to 3. Fifteen IMP-producing E. cloacae cases were identified, with five of the isolates being obtained from blood, and they were matched to 45 uninfected controls. All (100%) patients from whom IMP-producing E. cloacae isolates were obtained had indwelling devices at the time of isolation, compared with one (2.2%) uninfected control. Independent predictors for isolation of IMP-producing E. cloacae were identified as cephalosporin exposure and invasive procedures within 3 months. Although in-hospital mortality rates were similar between cases and controls (14.3% versus 13.3%), the in-hospital mortality of patients with IMP-producing E. cloacae-caused bacteremia was significantly higher (40%) than the rate in controls. IMP-producing E. cloacae isolates were frequently positive for other resistance determinants. The MICs of meropenem and imipenem were not elevated; 10 (67%) and 12 (80%) of the 15 IMP-producing E. cloacae isolates had a MIC of ≤ 1 μg/ml. A phylogenetic tree showed a close relationship among the IMP-producing E. cloacae samples. Indwelling devices, exposure to cephalosporin, and a history of invasive procedures were associated with isolation of IMP-producing E. cloacae. Screening for carbapenemase production is important in order to apply appropriate clinical management and infection control measures. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

First outbreak of VIM-2 metallo-β-lactamase-producing Pseudomonas aeruginosa in The Netherlands: microbiology, epidemiology and clinical outcomes.

PubMed

Van der Bij, A K; Van Mansfeld, R; Peirano, G; Goessens, W H F; Severin, J A; Pitout, J D D; Willems, R; Van Westreenen, M

2011-06-01

This study was designed to investigate the prevalence and characteristics of metallo-β-lactamase (MBL)-producing Pseudomonas aeruginosa in a tertiary care centre in The Netherlands, a country that is considered to have a low prevalence of antibiotic-resistant bacteria. Imipenem-resistant P. aeruginosa isolates cultured from clinical specimens during 2008-2009 were analysed phenotypically and molecularly by polymerase chain reaction (PCR) with sequencing. Genotyping was performed by multiple-locus variable-number tandem repeat (VNTR) analysis (MLVA). Clinical information was obtained by electronic chart review for all patients infected or colonised with an imipenem-resistant P. aeruginosa isolate that was included in the study. In total, 106 imipenem-resistant P. aeruginosa isolates were included. The bla(VIM-2) gene was detected in 35/106 isolates (33%) and was associated with integrons. Compared with non-MBL-producing imipenem-resistant P. aeruginosa, VIM-2 MBL-producing isolates showed higher rates of multidrug resistance. Patients with VIM-2 MBL-producing isolates were more likely to be admitted to the Intensive Care Unit (ICU) and had a higher risk of invasive infection, including development of bacteraemia. MLVA identified two separate VIM-2 MBL-producing clones, responsible for outbreaks in the ICU but also affecting 10 other departments. This is the first reported outbreak of VIM-2 MBL-producing P. aeruginosa in The Netherlands. Once introduced, VIM-2 MBL-producing P. aeruginosa cause significant infections and are easily spread within the hospital setting. Copyright © 2011 Elsevier B.V. and the International Society of Chemotherapy. All rights reserved.

Incidence of temonera, sulphuhydryl variables and cefotaximase genes associated with β-lactamase producing escherichia coli in clinical isolates

PubMed Central

Isaiah, Ibeh Nnana; Nche, Bikwe Thomas; Nwagu, Ibeh Georgina; Nwagu, Ibeh Isaiah

2011-01-01

Background: the occurrence of the different types of Extended spectrum beta Lactamase producing Escherichia coli with the, Sulphurhydryl variable, Temonera and the Cefotaximase have been on the rise Aim: The study was to determine the prevalence of extended spectrum beta lactamase gene resistance across the clinical isolates of hospitalized patients. Materials and Method: Three hundred and fifty isolates of Escherichia coli were received from different clinical specimens. The susceptibility profile of the isolates against 10 different antibiotics was examined, the MICs (Minimum Inhibitory Concentration) for ceftazidime were also determined using micro-broth dilution assay. Isolates showing MIC ≥ 6 μg/ml for ceftazidime were screened for ESBL (PCT)phenotypic confirmatory test and subjected to PCR (polymerase chain reaction) to further. Results: By disk diffusion test, there was resistance to ceftazidime and cefotaxime were 180(51.4%) and 120 (34.2%) respectively. However, all strains were susceptible to imipenem. 250 isolates showed MICs≥ 6 μg/ml for ceftazidime of which 180 (72%) were positive for extended spectrum beta lactamase. The prevalence of Sulphurhydryl variable, Temonera and the Cefotaximase among these isolates were 17.1%, 6.6% and 17%, respectively. Conclusion: For the identification of extended spectrum beta lactamase producing isolates it is recommended that clinical laboratories adopt simple test based on Cinical laboratory standard institute recommendation for confirming extended spectrum beta lactamase production in enterobacteriacea species. PMID:22363078

Escherichia coli Sequence Type 131 H30 Is the Main Driver of Emerging Extended-Spectrum-β-Lactamase-Producing E. coli at a Tertiary Care Center.

PubMed

Johnson, James R; Johnston, Brian; Thuras, Paul; Launer, Bryn; Sokurenko, Evgeni V; Miller, Loren G

2016-01-01

The H 30 strain of Escherichia coli sequence type 131 (ST131- H 30) is a recently emerged, globally disseminated lineage associated with fluoroquinolone resistance and, via its H 30Rx subclone, the CTX-M-15 extended-spectrum beta-lactamase (ESBL). Here, we studied the clonal background and resistance characteristics of 109 consecutive recent E. coli clinical isolates (2015) and 41 historical ESBL-producing E. coli blood isolates (2004 to 2011) from a public tertiary care center in California with a rising prevalence of ESBL-producing E. coli isolates. Among the 2015 isolates, ST131, which was represented mainly by ST131- H 30, was the most common clonal lineage (23% overall). ST131- H 30 accounted for 47% (8/17) of ESBL-producing, 47% (14/30) of fluoroquinolone-resistant, and 33% (11/33) of multidrug-resistant isolates. ST131- H 30 also accounted for 53% (8/14) of dually fluoroquinolone-resistant, ESBL-producing isolates, with the remaining 47% comprised of diverse clonal groups that contributed a single isolate each. ST131- H 30Rx, with CTX-M-15, was the major ESBL producer (6/8) among ST131- H 30 isolates. ST131- H 30 and H 30Rx also dominated (46% and 37%, respectively) among the historical ESBL-producing isolates (2004 to 2011), without significant temporal shifts in relative prevalence. Thus, this medical center's recently emerging ESBL-producing E. coli strains, although multiclonal, are dominated by ST131- H 30 and H 30Rx, which are the only clonally expanded fluoroquinolone-resistant, ESBL-producing lineages. Measures to rapidly and effectively detect, treat, and control these highly successful lineages are needed. IMPORTANCE The ever-rising prevalence of resistance to first-line antibiotics among clinical Escherichia coli isolates leads to worse clinical outcomes and higher health care costs, thereby creating a need to discover its basis so that effective interventions can be developed. We found that the H 30 subset within E. coli sequence type 131 (ST131- H 30) is currently, and has been since at least 2004, the main E. coli lineage contributing to key resistance phenotypes-including extended-spectrum-beta-lactamase (ESBL) production, fluoroquinolone resistance, multidrug resistance, and dual ESBL production-plus-fluoroquinolone resistance-at a United States tertiary care center with a rising prevalence of ESBL-producing E. coli isolates. This identifies ST131- H 30 as a target for diagnostic tests and preventive measures designed to curb the emergence of multidrug-resistant E. coli isolates and/or to blunt its clinical impact.

Indistinguishable NDM-producing Escherichia coli isolated from recreational waters, sewage, and a clinical specimen in Ireland, 2016 to 2017

PubMed Central

Mahon, Bláthnaid M; Brehony, Carina; McGrath, Elaine; Killeen, James; Cormican, Martin; Hickey, Paul; Keane, Shane; Hanahoe, Belinda; Dolan, Ann; Morris, Dearbháile

2017-01-01

In this study, New Delhi metallo-beta-lactamase (NDM)-producing Enterobacteriaceae were identified in Irish recreational waters and sewage. Indistinguishable NDM-producing Escherichia coli by pulsed-field gel electrophoresis were isolated from sewage, a fresh water stream and a human source. NDM-producing Klebsiella pneumoniae isolated from sewage and seawater in the same area were closely related to each other and to a human isolate. This raises concerns regarding the potential for sewage discharges to contribute to the spread of carbapenemase-producing Enterobacteriaceae. PMID:28449738

Extended-Spectrum beta (β)-Lactamases and Antibiogram in Enterobacteriaceae from Clinical and Drinking Water Sources from Bahir Dar City, Ethiopia

PubMed Central

Abera, Bayeh; Kibret, Mulugeta; Mulu, Wondemagegn

2016-01-01

Background The spread of Extended-Spectrum beta (β)-Lactamases (ESBL)-producing Enterobacteriaceae has become a serious global problem. ESBL-producing Enterobacteriaceae vary based on differences in antibiotic use, nature of patients and hospital settings. This study was aimed at determining ESBL and antibiogram in Enterobacteriaceae isolates from clinical and drinking water sources in Bahir Dar City, Northwest Ethiopia. Methods Enterobacteriaceae species were isolated from clinical materials and tap water using standard culturing procedures from September 2013 to March 2015. ESBL-producing-Enterobacteriaceae were detected using double-disk method by E-test Cefotaxim/cefotaxim+ clavulanic acid and Ceftazidime/ceftazidime+ clavulanic acid (BioMerieux SA, France) on Mueller Hinton agar (Oxoid, UK). Results Overall, 274 Enterobacteriaceae were isolated. Of these, 210 (44%) were from patients and 64 (17.1%) were from drinking water. The median age of the patients was 28 years. Urinary tract infection and blood stream infection accounted for 60% and 21.9% of Enterobacteriaceae isolates, respectively. Klebsiella pneumoniae was isolated from 9 (75%) of neonatal sepsis. The overall prevalence of ESBL-producing Enterobacteriaceae in clinical and drinking water samples were 57.6% and 9.4%, respectively. The predominant ESBL-producers were K. pneumoniae 34 (69.4%) and Escherichia coli 71 (58.2%). Statistically significant associations were noted between ESBL-producing and non- producing Enterobacteriaceae with regard to age of patients, infected body sites and patient settings (P = 0.001). ESBL-producing Enterobacteriaceae showed higher levels of resistance against chloramphenicol, ciprofloxacin and cotrimoxazole than non-ESBL producers (P = 0.001) Conclusions ESBL-producing Enterobacteriaceae coupled with high levels of other antimicrobials become a major concern for treatment of patients with invasive infections such as blood stream infections, neonatal sepsis and urinary tract infections. ESBL-producing Enterobacteriaceae were also detected in drinking water sources. PMID:27846254

Study on biofilm-forming properties of clinical isolates of Staphylococcus aureus.

PubMed

Taj, Yasmeen; Essa, Farhan; Aziz, Faisal; Kazmi, Shahana Urooj

2012-05-14

The purpose of this study was to observe the formation of biofilm, an important virulence factor, by isolates of Staphylococcus aureus (S. aureus) in Pakistan by different conventional methods and through electron microscopy. We screened 115 strains of S. aureus isolated from different clinical specimens by tube method (TM), air-liquid interface coverslip assay method, Congo red agar (CRA) method, and scanning electron microscopy (SEM). Out of 115 S. aureus isolates, 63 (54.78%) showed biofilm formation by tube method. Biofilm forming bacteria were further categorized as high producers (n = 23, 20%) and moderate producers (n = 40, 34.78%). TM coordinated well with the coverslip assay for strong biofilm-producing strains in 19 (16.5%) isolates. By coverslip method, weak producers were difficult to differentiate from biofilm negative isolates. Screening on CRA showed biofilm formation only in four (3.47%) strains. Scanning electron micrographs showed the biofilm-forming strains of S. aureus arranged in a matrix on the propylene surface and correlated well with the TM. Biofilm production is a marker of virulence for clinically relevant staphylococcal infections. It can be studied by various methods but screening on CRA is not recommended for investigation of biofilm formation in Staphylococcus aureus. Electron micrograph images correlate well with the biofilm production as observed by TM.

Dissemination of IMP-6 metallo-β-lactamase-producing Pseudomonas aeruginosa sequence type 235 in Korea.

PubMed

Seok, Yoonmi; Bae, Il Kwon; Jeong, Seok Hoon; Kim, Soo Hyun; Lee, Hyukmin; Lee, Kyungwon

2011-12-01

To investigate the epidemiological traits of Pseudomonas aeruginosa clinical isolates producing metallo-β-lactamases (MBLs) in Korea. A total of 386 non-duplicate P. aeruginosa clinical isolates were collected from Korea in 2009. Detection of MBL genes was performed by PCR. The genetic organization of class 1 integrons carrying the MBL gene cassette was investigated by PCR mapping and sequencing. The epidemiological relationships of the isolates were investigated by multilocus sequence typing and PFGE. Of 386 P. aeruginosa isolates, 30 (7.8%) isolates carried the bla(IMP-6) gene and 1 (0.3%) isolate carried the bla(VIM-2) gene. A probe specific for the bla(IMP-6) gene was hybridized to an ∼950 kbp I-CeuI-macrorestriction fragment from all 30 isolates and a probe specific for the bla(VIM-2) gene also hybridized to an ∼500 kbp I-CeuI-macrorestriction fragment from 1 isolate (BDC10). All 31 MBL-producing isolates shared an identical sequence type (ST), ST235, and they carried the same bla(OXA-50) allelic type, bla(OXA-50g). All MBL-producing isolates showed similar XbaI-macrorestriction patterns (similarity >85%), irrespective of MBL genotype. P. aeruginosa ST235 carrying the chromosomally located bla(IMP-6) gene is widely disseminated in Korea.

Mass Spectrometry and Multiplex Antigen Assays to Assess Microbial Quality and Toxin Production of Staphylococcus aureus Strains Isolated from Clinical and Food Samples

PubMed Central

Attien, Paul; Sina, Haziz; Moussaoui, Wardi; Zimmermann-Meisse, Gaëlle; Dadié, Thomas; Keller, Daniel; Riegel, Philippe; Edoh, Vincent; Kotchoni, Simeon O.; Djè, Marcellin; Prévost, Gilles

2014-01-01

The aim of our study was to investigate the microbial quality of meat products and on some clinical samples in Abidjan focused on Staphylococcus genus and the toxin production profile of Staphylococcus aureus (S. aureus) isolated. Bacteria were collected from 240 samples of three meat products sold in Abidjan and 180 samples issued from clinical infections. The strains were identified by both microbiological and MALDI-TOF-MS methods. The susceptibility to antibiotics was determined by the disc diffusion method. The production of Panton-Valentine Leukocidin, LukE/D, and epidermolysins was screened using radial gel immunodiffusion. The production of staphylococcal enterotoxins and TSST-1 was screened by a Bio-Plex Assay. We observed that 96/240 of meat samples and 32/180 of clinical samples were contaminated by Staphylococcus. Eleven species were isolated from meats and 4 from clinical samples. Forty-two S. aureus strains were isolated from ours samples. Variability of resistance was observed for most of the tested antibiotics but none of the strains displays a resistance to imipenem and quinolones. We observed that 89% of clinical S. aureus were resistant to methicillin against 58% for those issued from meat products. All S. aureus isolates issued from meat products produce epidermolysins whereas none of the clinical strains produced these toxins. The enterotoxins were variably produced by both clinical and meat product samples. PMID:24987686

The Role of OmpK35, OmpK36 Porins, and Production of β-Lactamases on Imipenem Susceptibility in Klebsiella pneumoniae Clinical Isolates, Cairo, Egypt.

PubMed

Wassef, Mona; Abdelhaleim, Mona; AbdulRahman, Eiman; Ghaith, Doaa

2015-12-01

OmpK35 and OmpK36 are the major outer membrane porins of Klebsiella pneumoniae. We aimed to study the effect of combined porin loss and production of extended-spectrum β-lactamases (ESBLs) on imipenem susceptibility among K. pneumoniae clinical isolates. This study included 91 suspected ESBL-producing K. pneumoniae clinical isolates, isolated from different patient specimens at the Cairo University hospital from January to June 2010. All isolates were subjected to genotypic analysis of the outer membrane protein gene expression using reverse transcription-PCR (RT-PCR) and analysis of OmpK35/36 of 38 isolates by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). By RT-PCR, loss of Omp35 was detected in 78 (85.7%) isolates, loss of Omp36 was detected in 64 (70.32%), and loss of both porins was detected in 62 (68.1%). Out of 91 isolates, 45 (49.5%) were resistant to cefoxitin, and 17 (18.7%) were confirmed as derepressed AmpC producers. Omp35 was lost in all FOX-resistant isolates, whereas Omp36 was lost in 42 (93.3%) (p-value 0.002). The mean of ceftazidime inhibition zone diameter was significantly decreased among ESBL-producing isolates with loss of Omp35/36 (p-value 0.041 and 0.006), respectively. The mean of imipenem minimal inhibitory concentration (MIC) was markedly increased to 8.55 μg/ml among AmpC-producing isolates with Omp35/36 loss, while the mean of imipenem MIC among the 66 confirmed ESBL producers was 0.32 μg/ml. Imipenem MIC was markedly increased among K. pneumoniae isolates showing AmpC production with loss of both porins OmpK35/36. Meanwhile, the association of porin OmpK35/36 loss with ESBL production was not a direct cause of resistance to imipenem.

Whole genome sequencing of ESBL-producing Escherichia coli isolated from patients, farm waste and canals in Thailand.

PubMed

Runcharoen, Chakkaphan; Raven, Kathy E; Reuter, Sandra; Kallonen, Teemu; Paksanont, Suporn; Thammachote, Jeeranan; Anun, Suthatip; Blane, Beth; Parkhill, Julian; Peacock, Sharon J; Chantratita, Narisara

2017-09-06

Tackling multidrug-resistant Escherichia coli requires evidence from One Health studies that capture numerous potential reservoirs in circumscribed geographic areas. We conducted a survey of extended β-lactamase (ESBL)-producing E. coli isolated from patients, canals and livestock wastewater in eastern Thailand between 2014 and 2015, and analyzed isolates using whole genome sequencing. The bacterial collection of 149 isolates consisted of 84 isolates from a single hospital and 65 from the hospital sewer, canals and farm wastewater within a 20 km radius. E. coli ST131 predominated the clinical collection (28.6%), but was uncommon in the environment. Genome-based comparison of E. coli from infected patients and their immediate environment indicated low genetic similarity overall between the two, although three clinical-environmental isolate pairs differed by ≤ 5 single nucleotide polymorphisms. Thai E. coli isolates were dispersed throughout a phylogenetic tree containing a global E. coli collection. All Thai ESBL-positive E. coli isolates were multidrug resistant, including high rates of resistance to tobramycin (77.2%), gentamicin (77.2%), ciprofloxacin (67.8%) and trimethoprim (68.5%). ESBL was encoded by six different CTX-M elements and SHV-12. Three isolates from clinical samples (n = 2) or a hospital sewer (n = 1) were resistant to the carbapenem drugs (encoded by NDM-1, NDM-5 or GES-5), and three isolates (clinical (n = 1) and canal water (n = 2)) were resistant to colistin (encoded by mcr-1); no isolates were resistant to both carbapenems and colistin. Tackling ESBL-producing E. coli in this setting will be challenging based on widespread distribution, but the low prevalence of resistance to carbapenems and colistin suggests that efforts are now required to prevent these from becoming ubiquitous.

Metallo-β-lactamase-producing Pseudomonas aeruginosa in the Netherlands: the nationwide emergence of a single sequence type.

PubMed

Van der Bij, A K; Van der Zwan, D; Peirano, G; Severin, J A; Pitout, J D D; Van Westreenen, M; Goessens, W H F

2012-09-01

Recently, the first outbreak of clonally related VIM-2 metallo-β-lactamase (MBL)-producing Pseudomonas aeruginosa in a Dutch tertiary-care centre was described. Subsequently, a nationwide surveillance study was performed in 2010-2011, which identified the presence of VIM-2 MBL-producing P. aeruginosa in 11 different hospitals. Genotyping by multiple-locus variable-number tandem-repeat analysis (MLVA) showed that the majority of the 82 MBL-producing isolates found belonged to a single MLVA type (n = 70, 85%), identified as ST111 by multilocus sequence typing (MLST). As MBL-producing isolates cause serious infections that are difficult to treat, the presence of clonally related isolates in various hospitals throughout the Netherlands is of nationwide concern. © 2012 The Authors. Clinical Microbiology and Infection © 2012 European Society of Clinical Microbiology and Infectious Diseases.

Metallo-Beta-Lactamase Producing Pseudomonas aeruginosa in a Healthcare Setting in Alexandria, Egypt.

PubMed

Abaza, Amani F; El Shazly, Soraya A; Selim, Heba S A; Aly, Gehan S A

2017-09-27

Pseudomonas aeruginosa has emerged as a major healthcare associated pathogen that creates a serious public health disaster in both developing and developed countries. In this work we aimed at studying the occurrence of metallo-beta-lactamase (MBL) producing P. aeruginosa in a healthcare setting in Alexandria, Egypt. This cross sectional study included 1583 clinical samples that were collected from patients admitted to Alexandria University Students' Hospital. P. aeruginosa isolates were identified using standard microbiological methods and were tested for their antimicrobial susceptibility patterns using single disc diffusion method according to the Clinical and Laboratory Standards Institute recommendations. Thirty P. aeruginosa isolates were randomly selected and tested for their MBL production by both phenotypic and genotypic methods. Diagnostic Epsilometer test was done to detect metallo-beta-lactamase enzyme producers and polymerase chain reaction test was done to detect imipenemase (IMP), Verona integron-encoded (VIM) and Sao Paulo metallo-beta-lactamase (IMP) encoding genes. Of the 1583 clinical samples, 175 (11.3%) P. aeruginosa isolates were identified. All the 30 (100%) selected P. aeruginosa isolates that were tested for MBL production by Epsilometer test were found to be positive; where 19 (63.3%) revealed blaSPM gene and 11 (36.7%) had blaIMP gene. blaVIM gene was not detected in any of the tested isolates. Isolates of MBL producing P. aeruginosa were highly susceptible to polymyxin B 26 (86.7%) and highly resistant to amikacin 26 (86.7%). MBL producers were detected phenotypically by Epsilometer test in both carbapenem susceptible and resistant P. aeruginosa isolates. blaSPM was the most commonly detected MBL gene in P. aeruginosa isolates.

Molecular characteristics of extended-spectrum beta-lactamases and qnr determinants in Enterobacter species from Japan.

PubMed

Kanamori, Hajime; Yano, Hisakazu; Hirakata, Yoichi; Hirotani, Ayako; Arai, Kazuaki; Endo, Shiro; Ichimura, Sadahiro; Ogawa, Miho; Shimojima, Masahiro; Aoyagi, Tetsuji; Hatta, Masumitsu; Yamada, Mitsuhiro; Gu, Yoshiaki; Tokuda, Koichi; Kunishima, Hiroyuki; Kitagawa, Miho; Kaku, Mitsuo

2012-01-01

The incidence of extended-spectrum β-lactamases (ESBLs) has been increasing worldwide, but screening criteria for detection of ESBLs are not standardized for AmpC-producing Enterobacteriaceae such as Enterobacter species. In this study, we investigated the prevalence of ESBLs and/or AmpC β-lactamases in Japanese clinical isolates of Enterobacter spp. and the association of plasmid-mediated quinolone resistance (PMQR) determinants with ESBL producers. A total of 364 clinical isolates of Enterobacter spp. collected throughout Japan between November 2009 and January 2010 were studied. ESBL-producing strains were assessed by the CLSI confirmatory test and the boronic acid disk test. PCR and sequencing were performed to detect CTX-M, TEM, and SHV type ESBLs and PMQR determinants. For ESBL-producing Enterobacter spp., pulsed-field gel electrophoresis (PFGE) was performed using XbaI restriction enzyme. Of the 364 isolates, 22 (6.0%) were ESBL producers. Seven isolates of Enterobacter cloacae produced CTX-M-3, followed by two isolates producing SHV-12. Two isolates of Enterobacter aerogenes produced CTX-M-2. Of the 22 ESBL producers, 21 had the AmpC enzyme, and six met the criteria for ESBL production in the boronic acid test. We found a significant association of qnrS with CTX-M-3-producing E. cloacae. The 11 ESBL-producing Enterobacter spp. possessing bla(CTX-M), bla(SHV), or bla(TEM) were divided into six unique PFGE types. This is the first report about the prevalence of qnr determinants among ESBL-producing Enterobacter spp. from Japan. Our results suggest that ESBL-producing Enterobacter spp. with qnr determinants are spreading in Japan.

Molecular Characteristics of Extended-Spectrum Beta-Lactamases and qnr Determinants in Enterobacter Species from Japan

PubMed Central

Hirakata, Yoichi; Hirotani, Ayako; Arai, Kazuaki; Endo, Shiro; Ichimura, Sadahiro; Ogawa, Miho; Shimojima, Masahiro; Aoyagi, Tetsuji; Hatta, Masumitsu; Yamada, Mitsuhiro; Gu, Yoshiaki; Tokuda, Koichi; Kunishima, Hiroyuki; Kitagawa, Miho; Kaku, Mitsuo

2012-01-01

The incidence of extended-spectrum β-lactamases (ESBLs) has been increasing worldwide, but screening criteria for detection of ESBLs are not standardized for AmpC-producing Enterobacteriaceae such as Enterobacter species. In this study, we investigated the prevalence of ESBLs and/or AmpC β-lactamases in Japanese clinical isolates of Enterobacter spp. and the association of plasmid-mediated quinolone resistance (PMQR) determinants with ESBL producers. A total of 364 clinical isolates of Enterobacter spp. collected throughout Japan between November 2009 and January 2010 were studied. ESBL-producing strains were assessed by the CLSI confirmatory test and the boronic acid disk test. PCR and sequencing were performed to detect CTX-M, TEM, and SHV type ESBLs and PMQR determinants. For ESBL-producing Enterobacter spp., pulsed-field gel electrophoresis (PFGE) was performed using XbaI restriction enzyme. Of the 364 isolates, 22 (6.0%) were ESBL producers. Seven isolates of Enterobacter cloacae produced CTX-M-3, followed by two isolates producing SHV-12. Two isolates of Enterobacter aerogenes produced CTX-M-2. Of the 22 ESBL producers, 21 had the AmpC enzyme, and six met the criteria for ESBL production in the boronic acid test. We found a significant association of qnrS with CTX-M-3-producing E. cloacae. The 11 ESBL-producing Enterobacter spp. possessing bla CTX-M, bla SHV, or bla TEM were divided into six unique PFGE types. This is the first report about the prevalence of qnr determinants among ESBL-producing Enterobacter spp. from Japan. Our results suggest that ESBL-producing Enterobacter spp. with qnr determinants are spreading in Japan. PMID:22719857

Utility of Combining Whole Genome Sequencing with Traditional Investigational Methods To Solve Foodborne Outbreaks of Salmonella Infections Associated with Chicken: A New Tool for Tackling This Challenging Food Vehicle.

PubMed

Crowe, Samuel J; Green, Alice; Hernandez, Kimberly; Peralta, Vi; Bottichio, Lyndsay; Defibaugh-Chavez, Stephanie; Douris, Aphrodite; Gieraltowski, Laura; Hise, Kelley; La-Pham, Karen; Neil, Karen P; Simmons, Mustafa; Tillman, Glenn; Tolar, Beth; Wagner, Darlene; Wasilenko, Jamie; Holt, Kristin; Trees, Eija; Wise, Matthew E

2017-04-01

High consumption rates and a multitude of brands make multistate foodborne outbreaks of Salmonella infections associated with chicken challenging to investigate, but whole genome sequencing is a powerful tool that can be used to assist investigators. Whole genome sequencing of pathogens isolated from clinical, environmental, and food samples is increasingly being used in multistate foodborne outbreak investigations to determine with unprecedented resolution how closely related these isolates are to one another genetically. In 2014, federal and state health officials investigated an outbreak of 146 Salmonella Heidelberg infections in 24 states. A follow-up analysis was conducted after the conclusion of the investigation in which 27 clinical and 24 food isolates from the outbreak underwent whole genome sequencing. These isolates formed seven clades, the largest of which contained clinical isolates from a subcluster of case patients who attended a catered party. One isolate from a chicken processed by a large producer was closely related genetically (zero to three single-nucleotide polymorphism differences) to the clinical isolates from these subcluster case patients. Chicken from this large producer was also present in the kitchen of the caterer on the day before the event, thus providing additional evidence that the chicken from this producer was the outbreak source. This investigation highlights how whole genome sequencing can be used with epidemiologic and traceback evidence to identify chicken sources of foodborne outbreaks.

Utility of Combining Whole Genome Sequencing with Traditional Investigational Methods To Solve Foodborne Outbreaks of Salmonella Infections Associated with Chicken: A New Tool for Tackling This Challenging Food Vehicle

PubMed Central

Crowe, Samuel J.; Green, Alice; Hernandez, Kimberly; Peralta, Vi; Bottichio, Lyndsay; Defibaugh-Chavez, Stephanie; Douris, Aphrodite; Gieraltowski, Laura; Hise, Kelley; La-Pham, Karen; Neil, Karen P.; Simmons, Mustafa; Tillman, Glenn; Tolar, Beth; Wagner, Darlene; Wasilenko, Jamie; Holt, Kristin; Trees, Eija; Wise, Matthew E.

2017-01-01

High consumption rates and a multitude of brands make multistate foodborne outbreaks of Salmonella infections associated with chicken challenging to investigate, but whole genome sequencing is a powerful tool that can be used to assist investigators. Whole genome sequencing of pathogens isolated from clinical, environmental, and food samples is increasingly being used in multistate foodborne outbreak investigations to determine with unprecedented resolution how closely related these isolates are to one another genetically. In 2014, federal and state health officials investigated an outbreak of 146 Salmonella Heidelberg infections in 24 states. A follow-up analysis was conducted after the conclusion of the investigation in which 27 clinical and 24 food isolates from the outbreak underwent whole genome sequencing. These isolates formed seven clades, the largest of which contained clinical isolates from a subcluster of case patients who attended a catered party. One isolate from a chicken processed by a large producer was closely related genetically (zero to three single-nucleotide polymorphism differences) to the clinical isolates from these subcluster case patients. Chicken from this large producer was also present in the kitchen of the caterer on the day before the event, thus providing additional evidence that the chicken from this producer was the outbreak source. This investigation highlights how whole genome sequencing can be used with epidemiologic and traceback evidence to identify chicken sources of foodborne outbreaks. PMID:28294686

Isolation, genotyping, and antimicrobial resistance of zoonotic shiga toxin-producing escherichia coli

USDA-ARS?s Scientific Manuscript database

Shiga toxin-producing Escherichia coli (STEC) is an enteric pathogen linked to outbreaks of human gastroenteritis with diverse clinical spectra. Traditional culture and isolation methods, including selective enrichment and differential plating, have enabled the effective recovery of STEC. Ruminants ...

Medicinal Plants Used by a Mbyá-Guarani Tribe Against Infections: Activity on KPC-Producing Isolates and Biofilm-Forming Bacteria.

PubMed

Brandelli, Clara Lia Costa; Ribeiro, Vanessa Bley; Zimmer, Karine Rigon; Barth, Afonso Luís; Tasca, Tiana; Macedo, Alexandre José

2015-11-01

The traditional use of medicinal plants for treatment of infectious diseases by an indigenous Mbyá-Guarani tribe from South Brazil was assessed by evaluating the antibiotic and antibiofilm activities against relevant bacterial pathogens. Aqueous extracts from 10 medicinal plants were prepared according to indigenous Mbyá-Guarani traditional uses. To evaluate antibiotic (OD600) and antibiofilm (crystal violet method) activities, Pseudomonas aeruginosa ATCC 27853, Staphylococcus epidermidis ATCC 35984 and seven multi-drug resistant Klebsiella pneumoniae carbapenemase (KPC)-producing bacterial clinical isolates were challenged with the extracts. Furthermore, the susceptibility profile of KPC-producing bacteria and the ability of these isolates to form biofilm were evaluated. The plants Campomanesia xanthocarpa, Maytenus ilicifolia, Bidens pilosa and Verbena sp. showed the best activity against bacterial growth and biofilm formation. The majority of KPC-producing isolates, which showed strong ability to form biofilm and a multidrug resistance profile, was inhibited by more than 50% by some extracts. The Enterobacter cloacae (KPC 05) clinical isolate was the only one resistant to all extracts. This study confirms the importance of indigenous traditional medicinal knowledge and describes for the first time the ability of these plants to inhibit biofilm formation and/or bacterial growth of multi-drug resistant KPC-producing isolates.

Prevalence and Molecular Characterization of Plasmid-mediated Extended-Spectrum β-Lactamase Genes (balaTEM, blaCTX and blASHV) Among Urinary Escherichia coli Clinical Isolates in Mashhad, Iran

PubMed Central

Nakhaei Moghaddam, Mahboobeh; Forghanifard, Mohammad Mahdi; Moshrefi, Sheila

2012-01-01

Objective(s) Extended-spectrum beta-lactamase (ESBL) producing bacteria have an important role in nosocomial infections. Due to the limited availability of information about the molecular epidemiology of ESBL producing bacteria in Mashhad, we decided to investigate about TEM, CTX and SHV ESBLs among urinary Escherichia coli isolates in Mashhad, a city in northeast Iran. Materials and Methods One hundred and eleven clinical isolates of E. coli were diagnosed from hospitalized patients in 2009. After performing antibiogram and phenotypic confirmation test, polymerase chain reaction (PCR) was performed by blaTEM, blaSHV and blaCTX primers and restriction digestion was carried out using PstI and TaqI (Fermentas-Lithuania) for confirmation. Results ESBL producers of E. coli isolates were 33.3%. Among 37 ESBL-producing isolates, 35 (94.6%), 21 (56.8%) and 5 (13.5%) were shown to have blaCTX, blaTEM and blaSHV, genes respectively. Co-resistance to non-beta lactam antibiotics was observed more with ESBL producers (P < 0.05). Conclusion The results showed that the studied ESBL genes are found with high prevalence and among them blaCTX is more widespread in urine E. coli isolates in Mashhad. PMID:23493415

Negative correlation between phospholipase and esterase activity produced by Fusarium isolates.

PubMed

Ishida, K; Alviano, D S; Silva, B G; Guerra, C R; Costa, A S; Nucci, M; Alviano, C S; Rozental, S

2012-05-01

Fusarium species have emerged as one of the more outstanding groups of clinically important filamentous fungi, causing localized and life-threatening invasive infections with high morbidity and mortality. The ability to produce different types of hydrolytic enzymes is thought to be an important virulence mechanism of fungal pathogens and could be associated with the environment of the microorganism. Here, we have measured the production of two distinct lipolytic enzymes, phospholipase and esterase, by sixteen Fusarium isolates recovered from the hospital environment, immunocompromised patients' blood cultures, foot interdigital space scrapings from immunocompromised patients, and foot interdigital space scrapings from immunocompetent patients (4 isolates each). Fourteen of these 16 isolates were identified as Fusarium solani species complex (FSSC) and two were identified as F. oxysporum species complex (FOSC). Some relevant genus characteristics were visualized by light and electron microscopy such as curved and multicelled macroconidia with 3 or 4 septa, microconidia, phialides, and abundant chlamydospores. All Fusarium isolates were able to produce esterase and phospholipase under the experimental conditions. However, a negative correlation was observed between these two enzymes, indicating that a Fusarium isolate with high phospholipase activity has low esterase activity and vice versa. In addition, Fusarium isolated from clinical material produced more phospholipases, while environmental strains produced more esterases. These observations may be correlated with the different types of substrates that these fungi need to degrade during their nutrition processes.

Negative correlation between phospholipase and esterase activity produced by Fusarium isolates

PubMed Central

Ishida, K.; Alviano, D.S.; Silva, B.G.; Guerra, C.R.; Costa, A.S.; Nucci, M.; Alviano, C.S.; Rozental, S.

2012-01-01

Fusarium species have emerged as one of the more outstanding groups of clinically important filamentous fungi, causing localized and life-threatening invasive infections with high morbidity and mortality. The ability to produce different types of hydrolytic enzymes is thought to be an important virulence mechanism of fungal pathogens and could be associated with the environment of the microorganism. Here, we have measured the production of two distinct lipolytic enzymes, phospholipase and esterase, by sixteen Fusarium isolates recovered from the hospital environment, immunocompromised patients' blood cultures, foot interdigital space scrapings from immunocompromised patients, and foot interdigital space scrapings from immunocompetent patients (4 isolates each). Fourteen of these 16 isolates were identified as Fusarium solani species complex (FSSC) and two were identified as F. oxysporum species complex (FOSC). Some relevant genus characteristics were visualized by light and electron microscopy such as curved and multicelled macroconidia with 3 or 4 septa, microconidia, phialides, and abundant chlamydospores. All Fusarium isolates were able to produce esterase and phospholipase under the experimental conditions. However, a negative correlation was observed between these two enzymes, indicating that a Fusarium isolate with high phospholipase activity has low esterase activity and vice versa. In addition, Fusarium isolated from clinical material produced more phospholipases, while environmental strains produced more esterases. These observations may be correlated with the different types of substrates that these fungi need to degrade during their nutrition processes. PMID:22415116

Structural study of the exopolysaccharide produced by a clinical isolate of Burkholderia cepacia.

PubMed

Cescutti, P; Bosco, M; Picotti, F; Impallomeni, G; Leitão, J H; Richau, J A; Sá-Correia, I

2000-07-14

The primary structure of the exopolysaccharide produced by a clinical isolate of the bacterium Burkholderia cepacia was studied by means of methylation analysis, selective degradation, NMR spectroscopy, and electrospray mass spectrometry. The resulting data showed that the parent repeating unit of the exopolysaccharide is a highly branched heptasaccharide with the following structure: Two acetyl groups are present per repeating unit, as noncarbohydrate substituents. Copyright 2000 Academic Press.

Molecular and clinical characterization of plasmid-mediated AmpC β-lactamase-producing Escherichia coli bacteraemia: a comparison with extended-spectrum β-lactamase-producing and non-resistant E. coli bacteraemia.

PubMed

Matsumura, Y; Nagao, M; Iguchi, M; Yagi, T; Komori, T; Fujita, N; Yamamoto, M; Matsushima, A; Takakura, S; Ichiyama, S

2013-02-01

Plasmid-mediated AmpC β-lactamase-producing Escherichia coli (AmpC-E) bacteraemia was characterized by comparison with bacteraemia caused by extended-spectrum β-lactamase (ESBL)-producing E. coli (ESBL-E) and non-resistant E. coli (NR-E) in the era of the worldwide spread of the CTX-M-15-producing O25b-ST131-B2 clone. Of 706 bloodstream E. coli isolates collected between 2005 and 2010 in three Japanese university hospitals, 111 ESBL screening-positive isolates were analysed for AmpC and ESBL genes by PCR. A case-control study was performed in which the cases consisted of all of the patients with AmpC-E bacteraemia. Phylogenetic groups, sequence types and O25b serotype were determined. Twenty-seven AmpC-E isolates (26 of which were of the CMY-2 type) were identified, and 54 ESBL-E and 54 NR-E isolates were selected for the controls. Nineteen AmpC-E isolates were also positive for ESBL. CTX-M-14 was the most prevalent ESBL type among both the AmpC-E and ESBL-E isolates. The O25b-ST131-B2 clone was the most prevalent among the ESBL-E isolates (26%) and the second most prevalent among the NR-E isolates (13%), but only one O25b-ST131-B2 clone was found among the AmpC-E isolates. Twenty-three different sequence types were identified among the AmpC-E isolates. When compared with bacteraemia with ESBL-E, previous isolation of multidrug-resistant bacteria and intravascular catheterization were independently associated with a lower risk for AmpC-E. When compared with NR-E bacteraemia, prior use of antibiotics was the only significant risk factor for AmpC-E. Unlike the spread of the O25b-ST131-B2 clone between ESBL-E and NR-E, the AmpC-E isolates were not dominated by any specific clone. © 2012 The Authors. Clinical Microbiology and Infection © 2012 European Society of Clinical Microbiology and Infectious Diseases.

Comparison of atypical Brachyspira spp. clinical isolates and classic strains in a mouse model of swine dysentery.

PubMed

Burrough, Eric; Strait, Erin; Kinyon, Joann; Bower, Leslie; Madson, Darin; Schwartz, Kent; Frana, Timothy; Songer, J Glenn

2012-12-07

Multiple Brachyspira spp. can colonize the porcine colon, and the presence of the strongly beta-hemolytic Brachyspira hyodysenteriae is typically associated with clinical swine dysentery. Recently, several Brachyspira spp. have been isolated from the feces of pigs with clinical disease suggestive of swine dysentery, yet these isolates were not identified as B. hyodysenteriae by genotypic or phenotypic methods. This study used a mouse model of swine dysentery to compare the pathogenic potential of seventeen different Brachyspira isolates including eight atypical clinical isolates, six typical clinical isolates, the standard strain of B. hyodysenteriae (B204), and reference strains of Brachyspira intermedia and Brachyspira innocens. Results revealed that strongly beta-hemolytic isolates induced significantly greater cecal inflammation than weakly beta-hemolytic isolates regardless of the genetic identification of the isolate, and that strongly beta-hemolytic isolates identified as 'Brachyspira sp. SASK30446' and B. intermedia by PCR produced lesions indistinguishable from those caused by B. hyodysenteriae in this model. Copyright © 2012 Elsevier B.V. All rights reserved.

Isolation of NDM-1-producing multidrug-resistant Pseudomonas putida from a paediatric case of acute gastroenteritis, India.

PubMed

Bhattacharya, D; Dey, S; Kadam, S; Kalal, S; Jali, S; Koley, H; Sinha, R; Nag, D; Kholkute, S D; Roy, S

2015-05-01

Pseudomonas putida is an uncommon opportunistic pathogen, usually susceptible to antimicrobial agents. Data concerning resistance to antimicrobial agents in clinical P. putida isolates are limited. To the best of our knowledge we report for the first time the isolation of NDM-1-producing multidrug-resistant P. putida from a case of acute gastroenteritis. The isolate showed resistance to a wide range of antimicrobials, including fluoroquinolones, third-generation cephalosporins and carbapenems. The isolate also exhibited multiple mutations in the quinolone resistance determining region and showed the presence of qepA, bla TEM , bla OXA1 and bla OXA7 genes. The present study highlights the importance of looking for the relatively rare aetiological agents in clinical samples that do not yield common pathogens.

Isolation of NDM-1-producing multidrug-resistant Pseudomonas putida from a paediatric case of acute gastroenteritis, India

PubMed Central

Bhattacharya, D.; Dey, S.; Kadam, S.; Kalal, S.; Jali, S.; Koley, H.; Sinha, R.; Nag, D.; Kholkute, S.D.; Roy, S.

2015-01-01

Pseudomonas putida is an uncommon opportunistic pathogen, usually susceptible to antimicrobial agents. Data concerning resistance to antimicrobial agents in clinical P. putida isolates are limited. To the best of our knowledge we report for the first time the isolation of NDM-1-producing multidrug-resistant P. putida from a case of acute gastroenteritis. The isolate showed resistance to a wide range of antimicrobials, including fluoroquinolones, third-generation cephalosporins and carbapenems. The isolate also exhibited multiple mutations in the quinolone resistance determining region and showed the presence of qepA, blaTEM, blaOXA1 and blaOXA7 genes. The present study highlights the importance of looking for the relatively rare aetiological agents in clinical samples that do not yield common pathogens. PMID:25893095

Emergence of VIM-4 metallo-β-lactamase-producing Klebsiella pneumoniae ST15 clone in the Clinical Centre University of Pécs, Hungary.

PubMed

Melegh, S; Kovács, K; Gám, T; Nyul, A; Patkó, B; Tóth, A; Damjanova, I; Mestyán, G

2014-01-01

Since November 2009 carbapenemase-producing Klebsiella pneumoniae isolates have been detected in increasing numbers at the Clinical Centre University of Pécs. Molecular typing was performed for 102 clinical isolates originating from different time periods and various departments of the Clinical Centre. Pulsed-field gel electrophoresis revealed the predominance of a single clone (101/102), identified as sequence type ST15. PCR and sequencing showed the presence of blaCTX-M-15 and blaVIM-4 genes. The blaVIM-4 was located on a class 1 integron designated In238b. To our knowledge, this is the first description of a blaVIM-4 gene in the predominant CTX-M-15 extended spectrum β-lactamase-producing Hungarian Epidemic Clone/ST15. © 2013 The Authors Clinical Microbiology and Infection © 2013 European Society of Clinical Microbiology and Infectious Diseases.

Phenotypic Characterization of Multidrug-resistant Escherichia Coli with Special Reference to Extended-spectrum-beta-lactamases and Metallo-beta-lactamases in a Tertiary Care Center.

PubMed

Shrestha, B; Shrestha, S; Mishra, S K; Kattel, H P; Tada, T; Ohara, H; Kirikae, T; Rijal, B P; Sherchand, J B; Pokhrel, B M

2015-01-01

The increasing reports on extended-spectrum-beta-lactamase and metallo-beta-lactamase producing Escherichia coli have addressed a potential threat to global health since it is found to be highly resistance to most of the currently available antibiotics including carbapenems. The present study was aimed to determine the antibiogram of extended-spectrum-beta-lactamase and metallo-beta-lactamase producing MDR E. coli isolates from various clinical samples. This was a cross-sectional study conducted over a period of seven months from December 2013 to July 2014 at bacteriology laboratory of Tribhuvan University Teaching Hospital. A total of 250 clinical specimens (urine, pus, sputum, blood, body fluid, bile, tissue and central venous pressure line tip) were processed from inpatients, with multidrug-resistant Escherichia coli infections. Standard microbiological techniques were used for isolation and identification of the isolates. The presence of extended-spectrum-beta-lactamase was detected by phenotypic confirmatory test recommended by Clinical and Laboratory Standards Institute and imipenem (IMP) /EDTA combined disc method was performed to detect metallo-beta-lactamase mediated resistance mechanism. We found high level of beta lactamase mediated resistance mechanism as part of multidrug resistance. Among 250 MDR isolates, 60% isolates were extended-spectrum-beta-lactamase producers and 17.2% isolates were metallo-beta-lactamase producers. Co-existence of extended-spectrum-beta-lactamase and metallo-beta-lactamase identified in 6.8% isolates. Beta-lactamase mediated resistance mechanisms are accounting very high in the multidrug resistant isolates of E. coli. Therefore, early detection of beta lactamase mediated resistant strains and their current antibiotic susceptibility pattern is necessary to avoid treatment failure and prevent the spread of MDR.

A study on the characterization of Propionibacterium acnes isolated from ocular clinical specimens.

PubMed

Sowmiya, Murali; Malathi, Jambulingam; Swarnali, Sen; Priya, Jeyavel Padma; Therese, Kulandai Lily; Madhavan, Hajib N

2015-10-01

There are only a few reports available on characterization of Propionibacterium acnes isolated from various ocular clinical specimens. We undertook this study to evaluate the role of P. acnes in ocular infections and biofilm production, and also do the phylogenetic analysis of the bacilli. One hundred isolates of P. acnes collected prospectively from ocular clinical specimens at a tertiary care eye hospital between January 2010 and December 2011, were studied for their association with various ocular disease conditions. The isolates were also subjected to genotyping and phylogenetic analysis, and were also tested for their ability to produce biofilms. Among preoperative conjunctival swabs, P. acnes was a probably significant pathogen in one case; a possibly significant pathogen in two cases. In other clinical conditions, 13 per cent isolates were probably significant pathogens and 38 per cent as possibly significant pathogens. The analysis of 16S rRNA gene revealed four different phylogenies whereas analysis of recA gene showed two phylogenies confirming that recA gene was more reliable than 16S rRNA with less sequence variation. Results of polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) had 100 per cent concordance with phylogenetic results. No association was seen between P. acnes subtypes and biofilm production. RecA gene phylogenetic studies revealed two different phylogenies. RFLP technique was found to be cost-effective with high sensitivity and specificity in phylogenetic analysis. No association between P. acnes subtypes and pathogenetic ability was observed. Biofilm producing isolates showed increased antibiotic resistance compared with non-biofilm producing isolates.

Prevalence and Molecular Characteristics of Carbapenemase-Producing Enterobacteriaceae From Five Hospitals in Korea.

PubMed

Jeong, Seok Hoon; Kim, Han Sung; Kim, Jae Seok; Shin, Dong Hoon; Kim, Hyun Soo; Park, Min Jeong; Shin, Saeam; Hong, Jun Sung; Lee, Seung Soon; Song, Wonkeun

2016-11-01

The emergence of carbapenemase-producing Enterobacteriaceae (CPE) represents a major clinical problem because these bacteria are resistant to most antibiotics. CPE remain relatively uncommon in Korea. We report the prevalence, clinical characteristics, and molecular epidemiology of CPE isolates collected from five university hospitals in Korea. Between January and December 2015, 393 non-duplicated isolates that were nonsusceptible to ertapenem were analyzed. Production of carbapenemase, extended-spectrum β-lactamase, and AmpC β-lactamase was determined by genotypic tests. Antimicrobial susceptibility profiles were determined by using an Etest. Clonality of Klebsiella pneumoniae carbapenemase (KPC)-2-producing and oxacillinase (OXA)-232-producing Klebsiella pneumoniae isolates was determined by pulsed-field gel electrophoresis (PFGE). Of the 393 isolates tested, 79 (20.1%) were CPE. Of these 79 isolates, 47 (59.5%) harbored the bla(OXA-232) gene while the remaining isolates carried genes bla(KPC-2) (n=27), bla(IMP-1) (n=4), and bla(NDM-1) (n=1). Among the 24 KPC-2 K. pneumoniae isolates from hospital B, 100% were resistant to carbapenems, 8% to colistin, and 0% to tigecycline. Among the 45 OXA-232 K. pneumoniae at hospital C, 95% were resistant to ertapenem, 68% to imipenem, 95% to meropenem, 10% to colistin, and 24% to tigecycline. PFGE analysis revealed a unique pattern for KPC-2 K. pneumoniae and identified 30 isolates belonging to the dominant pulsotypes (PT)1 and PT2 among 41 OXA-232 K. pneumoniae isolates. CPE strains are present in Korea, with the majority of K. pneumoniae isolates producing OXA-232 and KPC-2. The prevalence and predominant genotypes of CPE show hospital-specific differences.

ESBL and MBL in Cefepime Resistant Pseudomonas aeruginosa: An Update from a Rural Area in Northern India

PubMed Central

Biswas, Debasis; Kakati, Barnali; Singh, Malvika

2016-01-01

Introduction Cefepime, a fourth generation cephalosporin, is widely used for the empirical treatment of serious infections in critically ill hospitalized patients. Pseudomonas aeruginosa (P. aeruginosa), one of the commonest bacteria causing nosocomial infections has a propensity to develop antibiotic resistance quite promptly. Aim We undertook this study to assess the efficacy of cefepime against current clinical isolates of P. aeruginosa and to study existence of different beta-lactamase enzymes among cefepime resistant P. aeruginosa isolates. Materials and Methods Total of 618 isolates of P. aeruginosa recovered consecutively from various clinical samples of a tertiary care hospital were analysed. Their Antimicrobial sensitivity profile against piperacilin (100μg), piperacillin/tazobactam (100μg/10μg), ceftazidime (30μg), cefoperazone (75μg), cefepime (30μg), ciprofloxacin (5μg), gentamycin (10μg), amikacin (30μg) and imipenem (10μg) (Himedia) was tested by Kirby-Bauer disc diffusion method (Clinical and Laboratory Standards Institute guidelines). We further looked for ESBL, MBL and ESBL + MBL co producers among the cefepime resistant isolates by two different methods (combined double disc synergy test, imipenem-EDTA combined disc test and vitek2). Results Among 618 consecutive clinical isolates of P. aeruginosa, we observed resistance to cefepime in 457 (74%) isolates. We observed resistance to ciprofloxacin (n=506, 82%) in maximum number of isolates followed by that to Gentamycin (n=475, 77%), amikacin (n=366, 60%), and cefoperazone (n=350, 56.6%). Among all our cefepime resistant P. aeruginosa isolates only 27(6%) were ESBL producers, 18(4%) MBL producers and 2(0.4%) were ESBL+ MBL co-producers. All the ESBL and MBL isolates were also tested by VITEK 2 advanced expert system (bioMırieux Vitek Systems Inc, Hazelwood, MO, France) which revealed a 100% concordance with the phenotypic method tested. Conclusion This paper highlights the need to reconsider prescribing empirical antibiotics for Pseudomonas infections in this region and formulate a strong antibiotic policy to curb the menace of spread of multidrug resistant strains. PMID:27190800

ESBL and MBL in Cefepime Resistant Pseudomonas aeruginosa: An Update from a Rural Area in Northern India.

PubMed

Kotwal, Aarti; Biswas, Debasis; Kakati, Barnali; Singh, Malvika

2016-04-01

Cefepime, a fourth generation cephalosporin, is widely used for the empirical treatment of serious infections in critically ill hospitalized patients. Pseudomonas aeruginosa (P. aeruginosa), one of the commonest bacteria causing nosocomial infections has a propensity to develop antibiotic resistance quite promptly. We undertook this study to assess the efficacy of cefepime against current clinical isolates of P. aeruginosa and to study existence of different beta-lactamase enzymes among cefepime resistant P. aeruginosa isolates. Total of 618 isolates of P. aeruginosa recovered consecutively from various clinical samples of a tertiary care hospital were analysed. Their Antimicrobial sensitivity profile against piperacilin (100μg), piperacillin/tazobactam (100μg/10μg), ceftazidime (30μg), cefoperazone (75μg), cefepime (30μg), ciprofloxacin (5μg), gentamycin (10μg), amikacin (30μg) and imipenem (10μg) (Himedia) was tested by Kirby-Bauer disc diffusion method (Clinical and Laboratory Standards Institute guidelines). We further looked for ESBL, MBL and ESBL + MBL co producers among the cefepime resistant isolates by two different methods (combined double disc synergy test, imipenem-EDTA combined disc test and vitek2). Among 618 consecutive clinical isolates of P. aeruginosa, we observed resistance to cefepime in 457 (74%) isolates. We observed resistance to ciprofloxacin (n=506, 82%) in maximum number of isolates followed by that to Gentamycin (n=475, 77%), amikacin (n=366, 60%), and cefoperazone (n=350, 56.6%). Among all our cefepime resistant P. aeruginosa isolates only 27(6%) were ESBL producers, 18(4%) MBL producers and 2(0.4%) were ESBL+ MBL co-producers. All the ESBL and MBL isolates were also tested by VITEK 2 advanced expert system (bioMırieux Vitek Systems Inc, Hazelwood, MO, France) which revealed a 100% concordance with the phenotypic method tested. This paper highlights the need to reconsider prescribing empirical antibiotics for Pseudomonas infections in this region and formulate a strong antibiotic policy to curb the menace of spread of multidrug resistant strains.

Characterization of carbapenem nonsusceptible Pseudomonas aeruginosa in Denmark: a nationwide, prospective study.

PubMed

Hansen, Frank; Johansen, Helle Krogh; Østergaard, Claus; Arpi, Magnus; Hansen, Dennis Schrøder; Littauer, Pia; Holm, Anette; Heltberg, Ole; Schumacher, Helga; Fuursted, Kurt; Lykke, Mari-Ann Domar; Tønning, Birgitte; Hammerum, Anette M; Justesen, Ulrik Stenz

2014-02-01

From January 1st 2011 through June 30th 2011, 116 nonreplicate, noncystic fibrosis-related Pseudomonas aeruginosa isolates with reduced carbapenem susceptibility were collected from 12 out of 13 Danish departments of clinical microbiology. The presence of acquired β-lactamases was assessed with combination tablet-diffusion methodology and polymerase chain reaction. In addition, antimicrobial susceptibility testing, an efflux pump inhibitor assay, and pulsed-field gel electrophoresis (PFGE) were performed. Isolates producing acquired β-lactamases were further investigated by serotyping and multi locus sequence typing. Eight isolates produced the metallo-β-lactamase (MBL) VIM-2, and one isolate produced OXA-10 and VEB-1-like extended-spectrum beta-lactamase (ESBL). Phenotypic indications of derepressed AmpC and efflux pump were seen in 56 and 43 isolates, respectively. Overall, the results indicate that mutational factors related to permeability--often combined with derepressed, chromosomal AmpC--is the main factor behind carbapenem nonsusceptibility in Danish P. aeruginosa isolates. The ESBL producer and all the VIM producers belonged to international clones. PFGE revealed that most of the isolates were unrelated, but clonal spread was seen; the 116 isolates distributed in 97 PFGE types, with the largest cluster consisting of 4 isolates (including three isolates from the same hospital with 100% similarity). Thirty-two isolates were pair-wise related, while the remaining isolates were clonally unrelated, as were all nine ESBL/MBL producers.

Meropenem-Vaborbactam Tested against Contemporary Gram-Negative Isolates Collected Worldwide during 2014, Including Carbapenem-Resistant, KPC-Producing, Multidrug-Resistant, and Extensively Drug-Resistant Enterobacteriaceae.

PubMed

Castanheira, Mariana; Huband, Michael D; Mendes, Rodrigo E; Flamm, Robert K

2017-09-01

We evaluated the activity of meropenem-vaborbactam against contemporary nonfastidious Gram-negative clinical isolates, including Enterobacteriaceae isolates with resistance phenotypes and carbapenemase genotypes. Meropenem-vaborbactam (inhibitor at 8 μg/ml) and comparators were susceptibility tested by reference broth microdilution methods against 14,304 Gram-negative clinical isolates collected worldwide during 2014. Carbapenemase-encoding genes were screened by PCR and sequencing. Meropenem-vaborbactam (MIC 50/90 , ≤0.015/0.06 μg/ml) inhibited 99.1 and 99.3% of the 10,426 Enterobacteriaceae isolates tested at ≤1 and ≤2 μg/ml, respectively. Meropenem inhibited 97.3 and 97.7% of these isolates at the same concentrations. Against Enterobacteriaceae isolates displaying carbapenem-resistant Enterobacteriaceae (CRE) ( n = 265), multidrug-resistant (MDR) ( n = 1,210), and extensively drug-resistant (XDR) ( n = 161) phenotypes, meropenem-vaborbactam displayed MIC 50/90 values of 0.5/32, 0.03/1, and 0.5/32 μg/ml, respectively, whereas meropenem activities were 16/>32, 0.06/32, and 0.5/32 μg/ml, respectively. Among all geographic regions, the highest meropenem-vaborbactam activities were observed for CRE and MDR isolates from the United States (MIC 50/90 , 0.03/1 and 0.03/0.12 μg/ml, respectively). Meropenem-vaborbactam was very active against 135 KPC producers, and all isolates were inhibited by concentrations of ≤8 μg/ml (133 isolates by concentrations of ≤2 μg/ml). This combination had limited activity against isolates producing metallo-β-lactamases (including 25 NDM-1 and 16 VIM producers) and/or oxacillinases (27 OXA-48/OXA-163 producers) that were detected mainly in Asia-Pacific and some European countries. The activity of meropenem-vaborbactam was similar to that of meropenem alone against Pseudomonas aeruginosa , Acinetobacter spp., and Stenotrophomonas maltophilia Meropenem-vaborbactam was active against contemporary Enterobacteriaceae isolates collected worldwide, and this combination demonstrated enhanced activity compared to those of meropenem and most comparator agents against CRE isolates and KPC producers, the latter of which are often MDR. Copyright © 2017 American Society for Microbiology.

Carbapenemase-producing Enterobacteriaceae: a 2-year surveillance in a hospital in Iaşi, Romania.

PubMed

Braun, Sascha D; Dorneanu, Olivia S; Vremeră, Teodora; Reißig, Annett; Monecke, Stefan; Ehricht, Ralf

2016-01-01

Limited information is currently available about the prevalence of carbapenemase-producing Enterobacteriaceae (CPE) in Romania. Routine tests of 1,993 clinical isolates at a hospital in Iaşi yielded 46 isolates that were resistant to carbapenems. All 46 isolates were phenotypically and genotypically analyzed using VITEK-2 and DNA microarray-based assays. Isolates were assigned to Klebsiella pneumoniae and Enterobacter cloacae. For 39 isolates, carbapenem resistance was confirmed and 37 harbored at least one carbapenem resistance gene. Two isolates were probably resistant due to AmpC β-lactamases in combination with a porin loss. The overall concordance between detected phenotype and genotype was 95%. Our data show that carbapenemase-producing isolates with different underlying resistance mechanisms are still rare in Iaşi, but the global rise of CPE warrants intensified surveillance.

Spread of multidrug-resistant Acinetobacter baumannii and Pseudomonas aeruginosa clones in patients with ventilator-associated pneumonia in an adult intensive care unit at a university hospital.

PubMed

Royer, Sabrina; Faria, Ana Luiza Souza; Seki, Liliane Miyuki; Chagas, Thiago Pavoni Gomes; Campos, Paola Amaral de; Batistão, Deivid William da Fonseca; Asensi, Marise Dutra; Gontijo Filho, Paulo P; Ribas, Rosineide Marques

2015-01-01

In Brazil, ventilator-associated pneumonia (VAP) caused by carbapenem resistant Acinetobacter baumannii and Pseudomonas aeruginosa isolates are associated with significant mortality, morbidity and costs. Studies on the clonal relatedness of these isolates could lay the foundation for effective infection prevention and control programs. We sought to study the epidemiological and molecular characteristics of A. baumannii vs. P. aeruginosa VAP in an adult intensive care unit (ICU). It was conducted a cohort study of patients with VAP caused by carbapenem resistant A. baumannii and P. aeruginosa during 14 months in an adult ICU. Genomic studies were used to investigate the clonal relatedness of carbapenem resistant OXA-23-producing A. baumannii and P. aeruginosa clinical isolates. The risk factors for acquisition of VAP were also evaluated. Clinical isolates were collected for analysis as were samples from the environment and were typed using pulsed field gel electrophoresis. Multivariate logistic regression analysis identified trauma diagnosed at admission and inappropriate antimicrobial therapy as independent variables associated with the development of A. baumannii VAP and hemodialysis as independent variable associated with P. aeruginosa VAP. All carbapenem resistant clinical and environmental isolates of A. baumannii were OXA-23 producers. No MBL-producer P. aeruginosa was detected. Molecular typing revealed a polyclonal pattern; however, clone A (clinical) and H (surface) were the most frequent among isolates of A. baumannii tested, with a greater pattern of resistance than other isolates. In P. aeruginosa the most frequent clone I was multi-sensitive. These findings suggest the requirement of constant monitoring of these microorganisms in order to control the spread of these clones in the hospital environment. Copyright © 2015 Elsevier Editora Ltda. All rights reserved.

Identification of pathogenic factors potentially involved in Staphylococcus aureus keratitis using proteomics.

PubMed

Khan, Shamila; Cole, Nerida; Hume, Emma B H; Garthwaite, Linda L; Nguyen-Khuong, Terry; Walsh, Bradley J; Willcox, Mark D P

2016-10-01

Staphylococcus is a leading cause of microbial keratitis, characterized by destruction of the cornea by bacterial exoproteins and host-associated factors. The aim of this study was to compare extracellular and cell-associated proteins produced by two different isolates of S. aureus, a virulent clinical isolate (Staph 38) and a laboratory strain (Staphylococcus aureus 8325-4) of weaker virulence in the mouse keratitis model. Proteins were analyzed using 2D polyacrylamide gel electrophoresis and identified by subsequent mass spectrometry. Activity of staphylococcal adhesins was assessed by allowing strains to bind to various proteins adsorbed onto polymethylmethacrylate squares. Thirteen proteins in the extracellular fraction and eight proteins in the cell-associated fractions after bacterial growth were produced in increased amounts in the clinical isolate Staph 38. Four of these proteins were S. aureus virulence factor adhesins, fibronectin binding protein A, staphopain, glyceraldehyde-3-phosphate dehydrogenase 2 and extracellular adherence protein. The clinical isolate Staph 38 adhered to a greater extent to all mammalian proteins tested, indicating the potential of the adhesins to be active on its surface. Other proteins with increased expression in Staph 38 included potential moonlighting proteins and proteins involved in transcription or translation. This is the first demonstration of the proteome of S. aureus isolates from keratitis. These results indicate that the virulent clinical isolate produces more potentially important virulence factors compared to the less virulent laboratory strain and these may be associated with the ability of a S. aureus strain to cause more severe keratitis. Copyright © 2016 Elsevier Ltd. All rights reserved.

Isolation of Shiga toxin-producing Escherichia coli from fresh produce using STEC heart infusion washed blood agar with mitomycin-C.

PubMed

Lin, Andrew; Nguyen, Lam; Clotilde, Laurie M; Kase, Julie A; Son, Insook; Lauzon, Carol R

2012-11-01

The ability to detect and isolate Shiga toxin-producing Escherichia coli (STEC) remains a major challenge for food microbiologists. Although methods based on nucleic acids and antibodies have improved detection of STECs in foods, isolation of these bacteria remains arduous. STEC isolation is necessary for matching food, environmental, and clinical isolates during outbreak investigations and for distinguishing between pathogenic and nonpathogenic organisms. STEC heart infusion washed blood agar with mitomycin-C (SHIBAM) is a modification of washed sheep blood agar prepared by adding mitomycin-C and optimizing both the washed blood and base agar to better isolate STECs. Most STEC isolates produce a zone of hemolysis on SHIBAM plates and are easily distinguishable from background microbiota. Here, we present data supporting the use of SHIBAM to isolate STECs from fresh produce. SHIBAM was tested for accuracy in identifying STECs (365 of 410 STEC strains were hemolytic, and 63 of 73 E. coli strains that did not produce Shiga toxin were not hemolytic) and for recovery from artificially inoculated fresh produce (11 of 24 romaine lettuce samples and 6 of 24 tomato samples). STEC recovery with SHIBAM agar was greatly improved when compared with recovery on Levine's eosin-methylene blue agar as a reference method.

Detection of Different β-Lactamases and their Co-existence by Using Various Discs Combination Methods in Clinical Isolates of Enterobacteriaceae and Pseudomonas spp.

PubMed Central

Rawat, Vinita; Singhai, Monil; Verma, Pankaj Kumar

2013-01-01

Background: Resistance to broad spectrum beta-lactams mediated by extended spectrum β-lactamase (ESBL), AmpC, and metallobetalactamase (MBLs) enzymes are an increasing problem worldwide. The study was aimed to detect occurrence rate and to evaluate different substrates and inhibitors by disc combination method for detecting varying degree of β-lactamase enzymes and their co-production. Materials and Methods: A disc panel containing imipenem (IMP), IMP/EDTA, ceftazidime (CA), ceftazidime-tazobactum (CAT), CAT/cloxacillin (CLOX), ceftazidime-clavulanic acid (CAC), CAC/CLOX, cefoxitin (CN), and CN/CLOX in a single plate was used to detect presence of ESBLs, AmpC, and MBLs and/or their co-existence in 184 consecutive, nonrepetitive, clinical isolates of Enterobacteriace (n = 96) and Pseudomonas spp. (n = 88) from pus samples of hospitalized patients, resistant to 3rd generation cephalosporins. Results: Out of a total of 96 clinical isolates of Enterobacteriaceae, 18.7, 20.8, and 27% were pure ESBL, AmpC, and MBL producers, respectively. ESBL and AmpC were co-produced by 25% isolates. Among 88 Pseudomonas spp. 38.6, 13, and 6% were pure MBL, ESBL, and AmpC producers, respectively. ESBL/AmpC and MBL/AmpC co-production was seen in 20% and 18% isolates, respectively. Among ESBL and AmpC co-producers, CA/CAC/CLOX disc combination (DC) missed 7 of the 24 ESBL producers in Enterobacteriace and 4 of the 18 ESBL in Pseudomonas spp., which were detected by CA/CAT/CLOX DC. No mechanism was detected among 8.3% Enterobacteriaceae and 2.3% Pseudomonas isolates. Conclusion: Diagnostic problems posed by co-existence of different classes of β-lactamases in a single isolate could be solved by disc combination method by using simple panel of discs containing CA, CAT, CAT/CLOX, IMP, and IMP/EDTA. PMID:24014963

Dissemination of metallo-β-lactamase-producing Pseudomonas aeruginosa of sequence type 235 in Asian countries.

PubMed

Kim, Moon Jung; Bae, Il Kwon; Jeong, Seok Hoon; Kim, So Hyun; Song, Jae Hoon; Choi, Jae Young; Yoon, Sang Sun; Thamlikitkul, Visanu; Hsueh, Po-Ren; Yasin, Rohani Md; Lalitha, M K; Lee, Kyungwon

2013-12-01

To investigate the epidemiological traits of metallo-β-lactamase (MBL)-producing Pseudomonas aeruginosa (MPPA) clinical isolates collected by the Asian Network for Surveillance of Resistant Pathogens (ANSORP). A total of 16 MPPA clinical isolates were collected from six Asian countries in 2000 to 2009 by ANSORP. The MBL gene was detected by PCR amplification. The genetic organization of the class 1 integron carrying the MBL gene cassette was investigated by PCR mapping and sequencing. Southern blotting, repetitive sequence-based PCR and multilocus sequence typing (MLST) experiments were performed to characterize the isolates. PCR and sequencing experiments detected the blaVIM-2 (n = 12), blaVIM-3 (n = 1), blaIMP-6 (n = 2) and blaIMP-26 (n = 1) genes. The MBL genes were located on the chromosome in all isolates except one. Furthermore, all the MBL genes were located in a class 1 integron. All the MPPA isolates from Malaysia, Thailand, Sri Lanka and Korea were identified as sequence type (ST) 235 by MLST. Three VIM-2-producing isolates from India were identified as ST773, and one isolate harbouring VIM-3 from Taiwan was identified as ST298. P. aeruginosa ST235 might play a role in dissemination of MBL genes in Asian countries.

Does Extracellular DNA Production Vary in Staphylococcal Biofilms Isolated From Infected Implants versus Controls?

PubMed

Zatorska, Beata; Groger, Marion; Moser, Doris; Diab-Elschahawi, Magda; Lusignani, Luigi Segagni; Presterl, Elisabeth

2017-08-01

Prosthetic implant infections caused by Staphylococcus aureus and epidermidis are major challenges for early diagnosis and treatment owing to biofilm formation on the implant surface. Extracellular DNA (eDNA) is actively excreted from bacterial cells in biofilms, contributing to biofilm stability, and may offer promise in the detection or treatment of such infections. (1) Does DNA structure change during biofilm formation? (2) Are there time-dependent differences in eDNA production during biofilm formation? (3) Is there differential eDNA production between clinical and control Staphylococcal isolates? (4) Is eDNA production correlated to biofilm thickness? We investigated eDNA presence during biofilm formation in 60 clinical and 30 control isolates of S aureus and S epidermidis. The clinical isolates were isolated from patients with infections of orthopaedic prostheses and implants: 30 from infected hip prostheses and 30 from infected knee prostheses. The control isolates were taken from healthy volunteers who had not been exposed to antibiotics and a hospital environment during the previous 3 and 12 months, respectively. Control S epidermidis was isolated from the skin of the antecubital fossa, and control S aureus was isolated from the nares. For the biofilm experiments the following methods were used to detect eDNA: (1) fluorescent staining with 4',6-diamidino-2-phenylindole (DAPI), (2) eDNA extraction using a commercial kit, and (3) confocal laser scanning microscopy for 24-hour biofilm observation using propidium iodide and concanavalin-A staining; TOTO ® -1 and SYTO ® 60 staining were used for observation and quantification of eDNA after 6 and 24 hours of biofilm formation. Additionally antibiotic resistance was described. eDNA production as observed by confocal laser scanning microscopy was greater in clinical isolates than controls (clinical isolates mean ± SD: 1.84% ± 1.31%; control mean ± SD: 1.17% ± 1.37%; p < 0.005) after 6 hours of biofilm formation. After 24 hours, the amount of eDNA was greater in biofilms of S epidermidis than in biofilms of S aureus (S aureus mean ± SD: 1.35% ± 2.0%; S epidermidis mean ± SD: 6.42% ± 10.6%; p < 0.05). Clinical isolates of S aureus and S epidermidis produced more eDNA than control isolates at 6 hours of biofilm formation. The extraction method also showed that clinical isolates produced substantially greater amounts of eDNA than controls. S aureus and S epidermidis exhibit a differential production of DNA with time. Clinical isolates associated with implant infections produce greater amounts of eDNA than controls. Future research might focus on the diagnostic value of eDNA as a surrogate laboratory marker for biofilm formation in implant infections. eDNA should be considered as a potential future diagnostic tool or even a possible target to modify biofilms for successful treatment of biofilm-associated infections.

Clinical Saccharomyces cerevisiae isolates cannot cross the epithelial barrier in vitro.

PubMed

Pérez-Torrado, Roberto; Llopis, Silvia; Jespersen, Lene; Fernández-Espinar, Teresa; Querol, Amparo

2012-06-15

Saccharomyces cerevisiae is generally considered to be a safe organism and is essential to produce many different kinds of foods as well as being widely used as a dietary supplement. However, several isolates, which are genetically related to brewing and baking yeasts, have shown virulent traits, being able to produce human infections in immunodeficient patients. Previously it has been shown that the administration of S. cerevisiae clinical isolates can lead to systemic infections, reaching several organs in murine systems. In this work, we studied S. cerevisiae clinical isolates in an in vitro intestinal epithelial barrier model, comparing their behaviour with that of several strains of the related pathogens Candida glabrata and Candida albicans. The results showed that, in contrast to C. glabrata and C. albicans, S. cerevisiae was not able to cross the intestinal barrier. We concluded that S. cerevisiae can only perform opportunistic or passive crossings when epithelial barrier integrity is previously compromised. Copyright © 2012 Elsevier B.V. All rights reserved.

Clinical and molecular epidemiology of chromosome-mediated resistance to third-generation cephalosporins in Enterobacter isolates in eastern France.

PubMed

Talon, D; Bailly, P; Bertrand, X; Thouverez, M; Mulin, B

2000-07-01

To determine both the prevalence of group I beta-lactamase-related resistance and the clinical setting in which resistance to expanded-spectrum cephalosporins occurs. Isolates of Enterobacter spp. were sensitivity tested to a range of antibiotics, and selected isolates were DNA fingerprinted by pulsed-field gel electrophoresis. The medical records of all patients with positive cultures for Enterobacter spp. were reviewed to determine the effect of previous antibiotic treatment on the susceptibility profile of these organisms. The crude incidence of colonization/infection (n = 315) was 0.51 per 100 patients and 0.73 per 1000 days of hospitalization. The 4-day and 7-day Kaplan-Meier rates of colonization/infection with Enterobacter were estimated to be 7.57% (standard deviation (SD = 3.26%) and 4.16% (SD = 2.88%)), respectively. The time lag to colonization/infection with isolates producing large amounts of Bush group 1 beta-lactamase (HLBL) (27.35 +/- 27.30 days) was significantly different from that to colonization/infection with wild-type isolates (13.59 +/- 17.93 days) (P = 0.036). Ninety-six isolates (30.5%) demonstrated acquired resistance to expanded-spectrum cephalosporins: 34 isolates (10.8%) produced extended-spectrum beta-lactamase, and 62 isolates (19.7%) produced HLBL. The 89 Enterobacter isolates susceptible to third-generation cephalosporins yielded 84 major DNA patterns, and the 45 HLBL isolates yielded 38 major DNA patterns. The risk of colonization/infection with HLBL-producing Enterobacter was higher in cases of antimicrobial treatment with third-generation cephalosporins or a fluoroquinolone, and in cases of urinary tract colonization/infection. The judicious use in hospitals of both expanded-spectrum cephalosporins and other antibiotics such as fluoroquinolones is necessary to curtail the emergence of resistance in Enterobacter spp.

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.

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

Occurrence of Staphylococcal Ocular Infections of Food Producing Animals in Nsukka Southeast, Nigeria

PubMed Central

Udegbunam, Sunday Ositadinma; Udegbunam, Rita Ijeoma; Anyanwu, Madubuike Umunna

2014-01-01

Staphylococcal ocular infections of food animals have been somewhat under diagnosed probably due to the ubiquitous nature of staphylococcal organisms. This study was undertaken to determine the occurrence of staphylococcal ocular infections of food producing animals in Nsukka Southeast, Nigeria, and to determine the antibiogram of the isolated staphylococci. A total of 5,635 food producing animals were externally examined for signs of clinical ocular conditions. Animals that showed clinical eye lesions were further examined using pen light to assess the entire globe and the pupillary reflex. Blindness was assessed using menace blink reflex, palpebral reflex and obstacle methods. Isolation and identification of staphylococcal isolates from ocular swabs were done by standard methods. Antibiogram of the isolates was determined by disc diffusion method. Sixty-three (1.1%) of the examined animals showed signs of ocular condition. Thirty-one (49.2%) of the cultured swabs yielded Staphylococcus aureus (S. aureus). Isolation rates from different animal species were caprine (60%), ovine (33.3%), bovine (12.5%), and porcine (0%). Resistance of the isolates was 100% to ampicillin/cloxacillin, 90% to tetracycline, 80% to streptomycin, 71% to chloramphenicol, 20% to erythromycin, 16% to gentamicin, and 0% to ciprofloxacin and norfloxacin. Twenty-five (81%) of the isolates were multi-drug resistant. This study has shown that antibiotic-resistant staphylococci are associated with a sizeable percentage of ocular infections of food producing animals and should be considered during diagnosis and treatment. PMID:24693461

Detection of Extended-Spectrum β-Lactamases in Clinical Isolates of Enterobacter cloacae and Enterobacter aerogenes

PubMed Central

Tzelepi, Eva; Giakkoupi, Panagiota; Sofianou, Danai; Loukova, Veneta; Kemeroglou, Anastassia; Tsakris, Athanassios

2000-01-01

The aim of the present study was to investigate the frequency of extended-spectrum β-lactamases (ESBLs) in a consecutive collection of clinical isolates of Enterobacter spp. The abilities of various screening methods to detect ESBLs in enterobacters were simultaneously tested. Among the 68 consecutive isolates (56 Enterobacter cloacae and 12 Enterobacter aerogenes isolates) that were analyzed for β-lactamase content, 21 (25 and 58%, respectively) possessed transferable ESBLs with pIs of 8.2 and phenotypic characteristics of SHV-type enzymes, 8 (14.3%) of the E. cloacae isolates produced a previously nondescribed, clavulanate-susceptible ESBL that exhibited a pI of 6.9 and that conferred a ceftazidime resistance phenotype on Escherichia coli transconjugants, and 2 E. cloacae isolates produced both of these enzymes. Among the total of 31 isolates that were considered ESBL producers, the Vitek ESBL detection test was positive for 2 (6.5%) strains, and the conventional double-disk synergy test (DDST) with amoxicillin-clavulanate and with expanded-spectrum cephalosporins and aztreonam was positive for 5 (16%) strains. Modifications of the DDST consisting of closer application of the disks (at 20 instead of 30 mm), the use of cefepime, and the use of both modifications increased the sensitivity of this test to 71, 61, and 90%, respectively. Of the 37 isolates for which isoelectric focusing failed to determine ESBLs, the Vitek test was false positive for 1 isolate and the various forms of DDSTs were false-positive for 3 isolates. PMID:10655342

Clonal distribution and associated characteristics of Escherichia coli clinical and surveillance isolates from a military medical center.

PubMed

Manges, Amee R; Mende, Katrin; Murray, Clinton K; Johnston, Brian D; Sokurenko, Evgeni V; Tchesnokova, Veronika; Johnson, James R

2017-04-01

Antimicrobial-resistant Escherichia coli are a concern for military health services. We studied 100 extended-spectrum beta-lactamase (ESBL)-producing and non-producing E. coli clinical and surveillance isolates from military personnel and civilians at Brooke Army Medical Center (2007-2011). Major E. coli lineages, most prominently ST10 (24%), ST131 (16%), and ST648 (8%), were distributed much as reported for other North American populations. ST131, represented mainly by its resistance-associated ST131-H30 clonal subset, was uniquely associated with a clinical origin, regardless of ESBL status. Thus, clonal background predicted resistance phenotype and clinical versus surveillance origin, and these findings could assist military clinicians and epidemiologists. Published by Elsevier Inc.

Molecular typing of environmental and clinical strains of Vibrio vulnificus isolated in the northeastern USA.

PubMed

Reynaud, Yann; Pitchford, Steven; De Decker, Sophie; Wikfors, Gary H; Brown, Christopher L

2013-01-01

Vibrio vulnificus is a ubiquitous marine bacterium that is responsible for infections and some seafood-related illnesses and deaths in the United States, mainly in individuals with compromised health status in the Gulf of Mexico region. Most phylogenetic studies focus on V. vulnificus strains isolated in the southern United States, but almost no genetic data are available on northeastern bacterial isolates of clinical or environmental origin. Our goal in this study was to examine the genetic diversity of environmental strains isolated from commercially-produced oysters and in clinical strains of known pathogenicity in northeastern United States. We conducted analyses of a total of eighty-three strains of V. vulnificus, including 18 clinical strains known to be pathogenic. A polyphasic, molecular-typing approach was carried out, based upon established biotypes, vcg, CPS, 16S rRNA types and three other genes possibly associated with virulence (arylsulfatase A, mtlABC, and nanA). An established Multi Locus Sequence Typing (MLST) method was also performed. Phylogenetic analyses of these markers and MLST results produced similar patterns of clustering of strains into two main lineages (we categorized as 'LI' and 'LII'), with clinical and environmental strains clustering together in both lineages. Lineage LII was comprised primarily but not entirely of clinical bacterial isolates. Putative virulence markers were present in both clinical and environmental strains. These results suggest that some northeastern environmental strains of V. vulnificus are phylogenetically close to clinical strains and probably are capable of virulence. Further studies are necessary to assess the risk of human illness from consuming raw oysters harvested in the northeastern US.

Clinical MRSA isolates from skin and soft tissue infections show increased in vitro production of phenol soluble modulins

PubMed Central

Berlon, Nicholas R.; Qi, Robert; Sharma-Kuinkel, Batu K.; Joo, Hwang-Soo; Park, Lawrence P.; George, Dennis; Thaden, Joshua T.; Messina, Julia A.; Maskarinec, Stacey A.; Mueller-Premru, Manica; Athan, Eugene; Tattevin, Pierre; Pericas, Juan M.; Woods, Christopher W.; Otto, Michael; Fowler, Vance G.

2016-01-01

Summary Background Phenol-soluble modulins (PSMs) are amphipathic, pro-inflammatory proteins secreted by most Staphylococcus aureus isolates. This study tested the hypothesis that in vitro PSM production levels are associated with specific clinical phenotypes. Methods 177 methicillin-resistant S. aureus (MRSA) isolates from infective endocarditis (IE), skin and soft tissue infection (SSTI), and hospital-acquired/ventilator-associated pneumonia (HAP) were matched by geographic origin, then genotyped using spa-typing. In vitro PSM production was measured by high performance liquid chromatography/mass spectrometry. Statistical analysis was performed using Chi-squared or Kruskal–Wallis tests as appropriate. Results Spa type 1 was significantly more common in SSTI isolates (62.7% SSTI; 1.7% IE; 16.9% HAP; p < 0.0001) while HAP and IE isolates were more commonly spa type 2 (0% SSTI; 37.3% IE; 40.7% HAP; p < 0.0001). USA300 isolates produced the highest levels of PSMs in vitro. SSTI isolates produced significantly higher quantities of PSMα1-4, PSMβ1, and δ-toxin than other isolates (p < 0.001). These findings persisted when USA300 isolates were excluded from analysis. PMID:26079275

Distribution and antimicrobial susceptibility profile of extended-spectrum β-lactamase-producing Proteus mirabilis strains recently isolated in Japan.

PubMed

Kanayama, Akiko; Kobayashi, Intetsu; Shibuya, Kazutoshi

2015-02-01

Here we report on the prevalence of extended-spectrum β-lactamase (ESBL)-producing Proteus mirabilis from a nationwide antimicrobial resistance survey in different geographical regions of Japan. A total of 799 P. mirabilis isolates recovered between July 2009 and June 2010 from 314 healthcare facilities were characterised according to ESBL production, source, location and antimicrobial susceptibility pattern. ESBL production was found in 364 (45.6%) of the isolates, among which 354 (97.3%) produced CTX-M-2 group β-lactamases. Of the 349 ESBL-producing isolates in which the inpatient or outpatient status of the source was known, 324 (92.8%) were from inpatients and 25 (7.2%) were from outpatients (P<0.05). Results of pulsed-field gel electrophoresis (PFGE) analysis performed on 66 of the ESBL-producers generated a distribution of PFGE patterns into 21 groups. Genetic relatedness was seen among isolates within a region, which is consistent with horizontal transmission. With respect to the frequency of ESBL-producers by specimen source, 12/14 (85.7%) central venous catheter specimens yielded ESBL-producing P. mirabilis compared with 159/405 (39.3%), 119/209 (56.9%), 42/77 (54.5%) and 20/49 (40.8%), respectively, for isolates from urine, sputum, decubitus ulcer and wound specimens. Among the ESBL-producers, non-susceptibility to ciprofloxacin was found in 74.2% of the ESBL-producing isolates compared with 17.7% of the ESBL-non-producing isolates. These results show that approximately one-half of the P. mirabilis isolates from clinical specimens in Japan are ESBL-producers and that the potential for concomitant fluoroquinolone resistance must also be considered. Copyright © 2014 Elsevier B.V. and the International Society of Chemotherapy. All rights reserved.

Neutralizing Activity of Broadly Neutralizing Anti-HIV-1 Antibodies against Clade B Clinical Isolates Produced in Peripheral Blood Mononuclear Cells.

PubMed

Cohen, Yehuda Z; Lorenzi, Julio C C; Seaman, Michael S; Nogueira, Lilian; Schoofs, Till; Krassnig, Lisa; Butler, Allison; Millard, Katrina; Fitzsimons, Tomas; Daniell, Xiaoju; Dizon, Juan P; Shimeliovich, Irina; Montefiori, David C; Caskey, Marina; Nussenzweig, Michel C

2018-03-01

Recently discovered broadly neutralizing antibodies (bNAbs) against HIV-1 demonstrate extensive breadth and potency against diverse HIV-1 strains and represent a promising approach for the treatment and prevention of HIV-1 infection. The breadth and potency of these antibodies have primarily been evaluated by using panels of HIV-1 Env-pseudotyped viruses produced in 293T cells expressing molecularly cloned Env proteins. Here we report on the ability of five bNAbs currently in clinical development to neutralize circulating primary HIV-1 isolates derived from peripheral blood mononuclear cells (PBMCs) and compare the results to those obtained with the pseudovirus panels used to characterize the bNAbs. The five bNAbs demonstrated significantly less breadth and potency against clinical isolates produced in PBMCs than against Env-pseudotyped viruses. The magnitude of this difference in neutralizing activity varied, depending on the antibody epitope. Glycan-targeting antibodies showed differences of only 3- to 4-fold, while antibody 10E8, which targets the membrane-proximal external region, showed a nearly 100-fold decrease in activity between published Env-pseudotyped virus panels and PBMC-derived primary isolates. Utilizing clonal PBMC-derived primary isolates and molecular clones, we determined that the observed discrepancy in bNAb performance is due to the increased sensitivity to neutralization exhibited by 293T-produced Env-pseudotyped viruses. We also found that while full-length molecularly cloned viruses produced in 293T cells exhibit greater sensitivity to neutralization than PBMC-derived viruses do, Env-pseudotyped viruses produced in 293T cells generally exhibit even greater sensitivity to neutralization. As the clinical development of bNAbs progresses, it will be critical to determine the relevance of each of these in vitro neutralization assays to in vivo antibody performance. IMPORTANCE Novel therapeutic and preventive strategies are needed to contain the HIV-1 epidemic. Antibodies with exceptional neutralizing activity against HIV-1 may provide several advantages to traditional HIV drugs, including an improved side-effect profile, a reduced dosing frequency, and immune enhancement. The activity of these antibodies has been established in vitro by utilizing HIV-1 Env-pseudotyped viruses derived from circulating viruses but produced in 293T cells by pairing Env proteins with a backbone vector. We tested PBMC-produced circulating viruses against five anti-HIV-1 antibodies currently in clinical development. We found that the activity of these antibodies against PBMC isolates is significantly less than that against 293T Env-pseudotyped viruses. This decline varied among the antibodies tested, with some demonstrating moderate reductions in activity and others showing an almost 100-fold reduction. As the development of these antibodies progresses, it will be critical to determine how the results of different in vitro tests correspond to performance in the clinic. Copyright © 2018 Cohen et al.

Evaluation of several phenotypic methods for the detection of carbapenemase-producing Pseudomonas aeruginosa.

PubMed

Heinrichs, A; Huang, T D; Berhin, C; Bogaerts, P; Glupczynski, Y

2015-07-01

The purpose of this investigation was to compare several phenotypic methods, including combined disk tests (CDT) containing metallo-β-lactamase (MBL) inhibitors or cloxacillin, and the Carba NP test for the detection of carbapenemase-producing Pseudomonas aeruginosa (CPPA). A new CDT using imipenem (10 μg) ± cloxacillin 4,000 μg and the Carba NP test were evaluated to detect CPPA. In addition, four commercially available combined disks containing a carbapenem and ethylene-diamine-tetra-acetic acid (EDTA) or dipicolinic acid (DPA) as the inhibitor were tested in order to detect MBL-positive P. aeruginosa. All these phenotypic methods were evaluated on 188 imipenem non-susceptible P. aeruginosa (CPPA, n = 75) isolates divided into 26 well-characterized collection strains and 162 non-duplicate clinical isolates referred to the national reference laboratory in 2013. For the total of 188 isolates tested, CDT containing EDTA or DPA displayed high sensitivities (99%) and specificities (95%) for detecting MBL-producing isolates. CDT with cloxacillin showed a sensitivity and specificity of 97%/96% compared to 88%/99% for the Carba NP test in order to detect CPPA. For the 162 clinical isolates, CDT containing EDTA or DPA displayed a high negative predictive value (NPV) (99%) for detecting MBL-producing isolates. CDT with cloxacillin showed an NPV of 98%, compared to 95% for the Carba NP test in order to detect CPPA. In our setting, CDT associating imipenem ± EDTA or ± DPA performed best for the detection of MBL-producing P. aeruginosa. Imipenem/imipenem-cloxacillin test yielded good NPV to exclude the presence of MBL in imipenem non-susceptible isolates.

Dissemination of multidrug-resistant blaCTX-M-15/IncFIIk plasmids in Klebsiella pneumoniae isolates from hospital- and community-acquired human infections in Tunisia.

PubMed

Mansour, Wejdene; Grami, Raoudha; Ben Haj Khalifa, Anis; Dahmen, Safia; Châtre, Pierre; Haenni, Marisa; Aouni, Mahjoub; Madec, Jean-Yves

2015-11-01

This study investigated the molecular features of extended-spectrum β-lactamase (ESBL)-producing Klebsiella pneumoniae from hospital- and community-acquired (HA/CA) infections in the region of Mahdia, Tunisia. Among 336 K. pneumoniae isolates recovered from both clinical contexts between July 2009 and December 2011, 49 and 15 were ESBL producers and originated from clinical and community sources, respectively. All isolates produced the CTX-M-15 enzyme. As shown by Southern blot on S1 nuclease treatment followed by pulsed-field gel electrophoresis (PFGE) gels, the blaCTX-M-15 gene was carried on IncFII (n=4), IncFIIk (n=25), IncL/M (n=4), IncK (n=1), or untypeable (n=15) plasmids in HA isolates. In CA isolates, the blaCTX-M-15 gene was carried on IncFIIk (n=6), IncFII (n=1), IncHI1 (n=1), or untypeable (n=7) plasmids. In all, 23 and 11 PFGE types were found among the HA and CA isolates. Multilocus sequence typing on representative isolates shows diverse sequence types (STs), such as ST307, ST101, ST39, ST4, ST140, ST15, and ST307 in HA isolates and ST101, ST664, and ST323 in CA isolates. This study is the first comprehensive report of ESBL plasmids in K. pneumoniae from HA and CA infections in Tunisia. Copyright © 2015 Elsevier Inc. All rights reserved.

Emergence of Multidrug Resistance and Metallo-beta-lactamase Producing Acinetobacter baumannii Isolated from Patients in Shiraz, Iran

PubMed Central

Moghadam, MN; Motamedifar, M; Sarvari, J; Sedigh, Ebrahim-Saraie H; Mousavi, Same M; Moghadam, FN

2016-01-01

Background: Metallo-beta-lactamase (MβL) enzymes production is one of the most important resistance mechanisms against carbapenems in some bacteria including Acinetobacter baumannii. Aims: This study was aimed to determine the antimicrobial susceptibility and the prevalence of MβL among carbapenem-resistant isolates of A. baumannii. Materials and Methods: In this cross-sectional study from October 2012 to April 2013, 98 isolates were identified as A. baumannii using Microgen™ kits and confirmed by molecular method. These isolates were tested for antimicrobial susceptibilities by disk diffusion method according to the Clinical and Laboratory Standards Institute guidelines. Carbapenem-resistant isolates were further detected phenotypically by MβL minimal inhibitory concentration (MIC)-test strips, and subsequently positive MβL isolates were confirmed by polymerase chain reaction (PCR). Results: Overall, 98% (96/98) of A. baumannii isolates were detected as carbapenem-resistant by MIC test. Highest sensitivity to the tested antibiotic with 42.9% (42/98) was observed to colistin. Of 96 carbapenem-resistant isolates, 43 were phenotypically positive for MβL; out of 43 isolates, 37 were confirmed for the presence of MβL genes by PCR. Conclusion: The frequency of drug resistance among the clinical samples of A. baumannii isolated in our study against most of the antibiotics was very high. Moreover, all MβL producing isolates were multidrug resistance. Therefore, systematic surveillance to detect MβL producing bacteria and rational prescription and use of carbapenems could be helpful to prevent the spread of carbapenem resistance. PMID:27398247

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.

Genotypic and phenotypic profiles of Escherichia coli isolates belonging to clinical sequence type 131 (ST131), clinical non-ST131, and fecal non-ST131 lineages from India.

PubMed

Hussain, Arif; Ranjan, Amit; Nandanwar, Nishant; Babbar, Anshu; Jadhav, Savita; Ahmed, Niyaz

2014-12-01

In view of the epidemiological success of CTX-M-15-producing lineages of Escherichia coli and particularly of sequence type 131 (ST131), it is of significant interest to explore its prevalence in countries such as India and to determine if antibiotic resistance, virulence, metabolic potential, and/or the genetic architecture of the ST131 isolates differ from those of non-ST131 isolates. A collection of 126 E. coli isolates comprising 43 ST131 E. coli, 40 non-ST131 E. coli, and 43 fecal E. coli isolates collected from a tertiary care hospital in India was analyzed. These isolates were subjected to enterobacterial repetitive intergenic consensus (ERIC)-based fingerprinting, O typing, phylogenetic grouping, antibiotic sensitivity testing, and virulence and antimicrobial resistance gene (VAG) detection. Representative isolates from this collection were also analyzed by multilocus sequence typing (MLST), conjugation, metabolic profiling, biofilm production assay, and zebra fish lethality assay. All of the 43 ST131 E. coli isolates were exclusively associated with phylogenetic group B2 (100%), while most of the clinical non-ST131 and stool non-ST131 E. coli isolates were affiliated with the B2 (38%) and A (58%) phylogenetic groups, respectively. Significantly greater proportions of ST131 isolates (58%) than non-ST131 isolates (clinical and stool E. coli isolates, 5% each) were technically identified to be extraintestinal pathogenic E. coli (ExPEC). The clinical ST131, clinical non-ST131, and stool non-ST131 E. coli isolates exhibited high rates of multidrug resistance (95%, 91%, and 91%, respectively), extended-spectrum-β-lactamase (ESBL) production (86%, 83%, and 91%, respectively), and metallo-β-lactamase (MBL) production (28%, 33%, and 0%, respectively). CTX-M-15 was strongly linked with ESBL production in ST131 isolates (93%), whereas CTX-M-15 plus TEM were present in clinical and stool non-ST131 E. coli isolates. Using MLST, we confirmed the presence of two NDM-1-positive ST131 E. coli isolates. The aggregate bioscores (metabolite utilization) for ST131, clinical non-ST131, and stool non-ST131 E. coli isolates were 53%, 52%, and 49%, respectively. The ST131 isolates were moderate biofilm producers and were more highly virulent in zebra fish than non-ST131 isolates. According to ERIC-based fingerprinting, the ST131 strains were more genetically similar, and this was subsequently followed by the genetic similarity of clinical non-ST131 and stool non-ST131 E. coli strains. In conclusion, our data provide novel insights into aspects of the fitness advantage of E. coli lineage ST131 and suggest that a number of factors are likely involved in the worldwide dissemination of and infections due to ST131 E. coli isolates. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

Occurrence of qnr-positive clinical isolates in Klebsiella pneumoniae producing ESBL or AmpC-type beta-lactamase from five pediatric hospitals in China.

PubMed

Wang, Aihua; Yang, Yonghong; Lu, Quan; Wang, Yi; Chen, Yuan; Deng, Li; Ding, Hui; Deng, Qiulian; Wang, Li; Shen, Xuzhuang

2008-06-01

The plasmid-mediated quinolone resistance qnr genes in clinical isolates in adults have been described in different countries; however, the frequency of their occurrence has not been detected in pediatric patients. A total of 410 clinical isolates of Klebsiella pneumoniae, identified as producers of an extended-spectrum beta-lactamase (ESBL), or AmpC beta-lactamase, were collected from five children's hospitals in China during 2005-2006. The isolates were screened for the presence of the qnrA, qnrB, and qnrS genes, and then the harboring qnr gene isolates were detected for a bla gene coding for the TEM, SHV, CTX-M, and plasmid-mediated ampC gene by a PCR experiment. Ninety-two isolates (22.7%) were positive for the qnr gene, including 10 of qnrA (2.4%), 25 of qnrB (6.1%), and 62 of qnrS (15.1%). Eighty-one of the 92 (88.0%) qnr-positive isolates carried at least one bla gene for TEM, SHV, CTX-M, or DHA-1. The ciprofloxacin resistance increased 16-256-fold and oflaxacin resistance increased 2-32-fold in transconjugants, respectively. These results indicated that the plasmid-mediated qnr quinolone resistance gene was qnrS, followed by qnrB and qnrA. Most of the isolates also carried a bla gene coding ESBL or ampC gene coding DHA-1 among Klebsiella pneumoniae isolated from Chinese pediatric patients.

Characterization of Enterobacteriaceae isolates obtained from a tertiary care hospital in Mexico, which produces extended-spectrum β-lactamase.

PubMed

Morfín-Otero, Rayo; Mendoza-Olazarán, Soraya; Silva-Sánchez, Jesús; Rodríguez-Noriega, Eduardo; Laca-Díaz, Jorge; Tinoco-Carrillo, Perla; Petersen, Luis; López, Perla; Reyna-Flores, Fernando; Alcantar-Curiel, Dolores; Garza-Ramos, Ulises; Garza-González, Elvira

2013-10-01

The prevalence and genetic characteristics of Escherichia coli and Klebsiella pneumoniae clinical isolates producing extended-spectrum β-lactamase (ESBL) were examined. Between October 2010 and March 2011, E. coli (n=460) and K. pneumoniae (n=78) isolates were collected at a tertiary care hospital in Guadalajara, Mexico. The minimum inhibitory concentration (MIC) for each isolate was determined using a broth microdilution method, and ESBL production was assayed. The presence of β-lactamase genes, blaSHV, blaCTX-M, and blaTLA-1, was detected by PCR and confirmed with sequencing. Only ESBL-producing isolates were further subjected to pulsed-field gel electrophoresis (PFGE) and plasmid profiling. All of the ESBL isolates were multidrug resistant and 75/460 (16.3%) E. coli isolates and 21/78 (26.9%) K. pneumoniae isolates were found to produce ESBL. For the E. coli isolates, >95% susceptibility to amikacin, meropenem, fosfomycin, imipenem, and nitrofurantoin was observed. For K. pneumoniae, similar results were obtained, with discrepancies observed for gentamicin and nitrofurantoin. PFGE further identified eleven pulsotypes for E. coli and three clusters of K. pneumoniae. CTX-M-15 was detected in 85% of ESBL-producing E. coli and in 76% of ESBL-producing K. pneumoniae. In contrast, SHV-5 ESBL was identified in 17% of E. coli isolates and in 86% of K. pneumoniae isolates. The bla-TLA-1 gene was not detected in any of the 96 isolates analyzed. Overall, CTX-M-15 and SHV-5 were found to have a high rate of spread throughout the hospital and were associated with strong multidrug resistance.

Hospital clonal dissemination of Enterobacter aerogenes producing carbapenemase KPC-2 in a Chinese teaching hospital.

PubMed

Qin, Xiaohua; Yang, Yang; Hu, Fupin; Zhu, Demei

2014-02-01

Carbapenems are first-line agents for the treatment of serious nosocomial infections caused by multidrug-resistant Enterobacteriaceae. However, resistance to carbapenems has increased dramatically among Enterobacteriaceae in our hospital. In this study, we report clonal dissemination caused by carbapenem-resistant Enterobacter aerogenes (CREA). In 2011, CREA was identified from 12 patients admitted to the neurosurgical ward. All 12 clinical isolates were non-susceptible to cefotaxime, ceftazidime, cefoxitin, ertapenem, imipenem or meropenem. All isolates carried the gene encoding Klebsiella pneumoniae carbapenemase-2 (KPC-2), except for the isolate E4. However, a remarkably lower expression level of the porin OmpF was detected in the non-KPC-2-producing isolate E4 on SDS-PAGE compared with the carbapenem-susceptible isolate. Epidemiological and molecular investigations showed that a single E. aerogenes strain (PFGE type A), including seven KPC-2-producing clinical isolates, was primarily responsible for the first isolation and subsequent dissemination. In a case-control study, we identified risk factors for infection/colonization with CREA. Mechanical ventilation, the changing of sickbeds and previous use of broad-spectrum antibiotics were identified as potential risk factors. Our findings suggest that further studies should focus on judicious use of available antibiotics, implementation of active antibiotic resistance surveillance and strict implementation of infection-control measures to avoid the rapid spread or clonal dissemination caused by carbapenem-resistant Enterobacteriaceae in healthcare facilities.

Regional Spread of CTX-M-2-Producing Proteus mirabilis with the Identical Genetic Structure in Japan.

PubMed

Kato, Karin; Matsumura, Yasufumi; Yamamoto, Masaki; Nagao, Miki; Takakura, Shunji; Ichiyama, Satoshi

2017-07-01

In this study, we analyzed the molecular epidemiology of extended-spectrum β-lactamase (ESBL)-producing Proteus mirabilis isolates collected from the central region of Japan. Between 2005 and 2012, 820 clinical P. mirabilis isolates were obtained from ten acute care hospitals in Japan. We characterized ESBL confirmatory test-positive isolates by sequencing the ESBL genes and their flanking regions, detecting plasmid replicons, and performing pulsed-field gel electrophoresis (PFGE). Ninety-six isolates (12%) were positive according to the ESBL confirmatory test; all these isolates possessed bla CTX-M-2 with the same flanking structure of upstream ΔISEcp1 and a downstream region identical to downstream bla KLUA-1 . IncT was the prevalent, and only, replicon found in 63 isolates. PFGE analysis detected eight clusters with more than one isolate, among which three included 56 isolates and six included isolates from multiple hospitals. CTX-M-2-producing P. mirabilis with an identical genetic structure flanking bla CTX-M-2 is dominant in this Japanese region, and there is evidence for the clonal spread of isolates.

Isolation and characterization of an atypical Siberian sturgeon herpesvirus strain in Russia: novel North American Acipenserid herpesvirus 2 strain in Europe?

PubMed

Doszpoly, A; Kalabekov, I M; Breyta, R; Shchelkunov, I S

2017-10-01

Siberian sturgeon herpesvirus (SbSHV) was isolated in Russia for the first time in 2006. Nine SbSHV isolates were recovered from different fish hatcheries producing the same cytopathic effect in cell cultures, the same clinical signs and mortality kinetics in virus-infected fish and the same virus neutralization pattern and shared identical nucleotide sequences. In 2011, a new isolate was recovered from juvenile sturgeon, which caused completely different cytopathic effect. That isolate was not readily neutralized by Siberian sturgeon hyperimmune antisera, and its DNA was not recognized by the routine PCR developed for SbSHV detection. Molecular study of the novel isolate revealed that it was more closely related to North American Acipenserid herpesvirus 2 (AciHV-2) isolates from white sturgeon, while the genome sequences of the former SbSHV isolates showed high similarity to the AciHV-2 isolated from shortnose sturgeon. While clinical signs and mortality caused by the novel isolate in infected Siberian sturgeon were similar to those of the formerly described SbSHV isolates, the incubation period and mean time to death produced by the novel isolate were twice as long. The differences between the former isolates and the recent one suggest that a novel SbSHV strain emerged in Europe and the molecular findings imply its North American origin. © 2017 John Wiley & Sons Ltd.

Epidemiology of Extended-Spectrum β-Lactamase-Producing Enterobacter Isolates in a Spanish Hospital during a 12-Year Period

PubMed Central

Cantón, Rafael; Oliver, Antonio; Coque, Teresa M.; Varela, María del Carmen; Pérez-Díaz, José Claudio; Baquero, Fernando

2002-01-01

Fifteen Enterobacter clinical isolates (11 Enterobacter cloacae isolates, 3 Enterobacter aerogenes isolates, and 1 Enterobacter gergoviae isolate), representing 0.4% of all Enterobacter isolates recovered in our hospital from 1989 to 2000, were suspected of harboring an extended-spectrum β-lactamase (ESBL). These isolates were recovered from 14 different patients. ESBLs were transferred by conjugation into an Escherichia coli recipient strain. Pulsed-field gel electrophoresis (PFGE) revealed a single clone of E. aerogenes and six different clones of E. cloacae. Four of these E. cloacae clonal types were represented by only one isolate each, but the other two were represented by three and four isolates, respectively. Isoelectric focusing, susceptibility phenotyping, PCR analysis, and sequencing demonstrated the presence of three different ESBLs. The most frequent was the recently characterized CTX-M-10 ESBL, which was found in the E. gergoviae isolate and in all but one of the E. cloacae isolates. The remaining E. cloacae isolate harbored a TEM-27 ESBL, and the three E. aerogenes isolates harbored a TEM-24 ESBL. PFGE revealed that our E. aerogenes strain was indistinguishable from the French TEM-24-producing E. aerogenes endemic clone. Although a low prevalence of ESBL-producing Enterobacter isolates was found in our institution over a 12-year period, a diversity of nonepidemic E. cloacae clones was detected, as was the persistence of the CTX-M-10 β-lactamase. The presence of the TEM-24-producing E. aerogenes French clone in our institution also demonstrates the intercountry dissemination of ESBL-producing isolates. PMID:11923338

In vivo efficacy of human simulated regimens of carbapenems and comparator agents against NDM-1-producing Enterobacteriaceae.

PubMed

Wiskirchen, Dora E; Nordmann, Patrice; Crandon, Jared L; Nicolau, David P

2014-01-01

Doripenem and ertapenem have demonstrated efficacy against several NDM-1-producing isolates in vivo, despite having high MICs. In this study, we sought to further characterize the efficacy profiles of humanized regimens of standard (500 mg given every 8 h) and high-dose, prolonged infusion of doripenem (2 g given every 8 h, 4-h infusion) and 1 g of ertapenem given intravenously every 24 h and the comparator regimens of ceftazidime at 2 g given every 8 h (2-h infusion), levofloxacin at 500 mg every 24 h, and aztreonam at 2 g every 6 h (1-h infusion) against a wider range of isolates in a murine thigh infection model. An isogenic wild-type strain and NDM-1-producing Klebsiella pneumoniae and eight clinical NDM-1-producing members of the family Enterobacteriaceae were tested in immunocompetent- and neutropenic-mouse models. The wild-type strain was susceptible to all of the agents, while the isogenic NDM-1-producing strain was resistant to ceftazidime, doripenem, and ertapenem. Clinical NDM-1-producing strains were resistant to nearly all five of the agents (two were susceptible to levofloxacin). In immunocompetent mice, all of the agents produced ≥1-log10 CFU reductions of the isogenic wild-type and NDM-1-producing strains after 24 h. Minimal efficacy of ceftazidime, aztreonam, and levofloxacin against the clinical NDM-1-producing strains was observed. However, despite in vitro resistance, ≥1-log10 CFU reductions of six of eight clinical strains were achieved with high-dose, prolonged infusion of doripenem and ertapenem. Slight enhancements of doripenem activity over the standard doses were obtained with high-dose, prolonged infusion for three of the four isolates tested. Similar efficacy observations were noted in neutropenic mice. These data suggest that carbapenems are a viable treatment option for infections caused by NDM-1-producing Enterobacteriaceae.

Phenotypic characterization of enzymatic activity of clinical dermatophyte isolates from animals with and without skin lesions and humans.

PubMed

Gnat, Sebastian; Łagowski, Dominik; Nowakiewicz, Aneta; Zięba, Przemysław

2018-05-20

The pathogenesis of dermatophytoses is associated with the secretion of enzymes degrading the infected tissue components. Although many studies on enzymatic activity of dermatophytes have been conducted over the years, there have been no concrete proposals on construction of the profile of enzymes characteristic of individual species, genus, or ecological types of dermatophytes. The aim of this study was to assess the capability of clinical dermatophyte isolates from both symptomatic and asymptomatic animals and humans to produce different enzymes. Clinical isolates of 234 dermatophyte strains collected during routine examination of animal health were used in this study. The enzymatic production of keratinase, elastase, phospholipase, lipase, protease, DNase, and gelatinase as well as the haemolytic activity were evaluated using specific test media. The overall degree of enzymatic activity of the analysed clinical isolates of the dermatophytes was 67%. All tested clinical isolates of different species of dermatophytes showed keratinase activity and 96% additionally exhibited phospholipase activity. The weakest activity among the tested enzymes was demonstrated for elastase and gelatinase. 83% of the isolates of the dermatophytes showed haemolytic activity. Our data indicate that clinical isolates of dermatophytes from different species produce enzymes with different levels of activities. Profile of enzymes characteristic of individual species, genus, or ecological types of dermatophytes is possibly dependent upon factors related to the host. The relationship between each enzyme and the occurrence of skin lesions in animals and humans or asymptomatic animal carriers varies on whether the infection is caused by T. mentagrophytes, T. verrucosum, or M. canis. Interestingly, only keratinase seems to be correlated with the appearance of dermatophyte infections, irrespective of the pathogen species, and elastase is a characteristic enzyme for dermatophyte strains infecting humans. Haemolysis seems to be dependent on host factors and is more common in the case of human dermatophyte isolates. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

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.

Genotypic and phenotypic characterization of invasive neonatal Escherichia coli clinical isolates.

PubMed

Shakir, Salika Mehreen; Goldbeck, Jessica Marie; Robison, Denise; Eckerd, Annette Marie; Chavez-Bueno, Susana

2014-11-01

The objective of this study was to describe the clinical characteristics of neonates with Escherichia coli bacteremia and the antibiotic resistance pattern of the bacterial isolates. We assessed the isolates' genetic relatedness and virulence phenotypic characteristics in vitro. A total of 24 neonates with E. coli bacteremia were identified prospectively in a tertiary-care hospital. Clinical and antibiotic resistance data were investigated. The E. coli isolates were analyzed by multilocus sequence typing (MLST); the presence of the K1 capsule and their ability to invade intestinal epithelial cells were also assessed. Most newborns were very low birth weight infants. Overall, 75% of the isolates were ampicillin resistant and 17% were gentamicin and tobramycin nonsusceptible. MLST determined sequence types 95 and 131 (ST95 and ST131) predominated, with ST131 becoming significantly more prevalent recently. The K1 capsule was present in 50% of the isolates. ST131 isolates and those producing bacteremia in newborns younger than 7 days showed a highly invasive phenotype. Resistance to antibiotics currently used empirically to treat newborns is present in bacteremia-producing E. coli. Clonal spread among newborns of multidrug-resistant E. coli is possible; therefore, continued surveillance is needed. Identification of additional virulence factors associated with increased invasion in neonatal E. coli strains is important and further studies are warranted. Thieme Medical Publishers 333 Seventh Avenue, New York, NY 10001, USA.

Unusual Outbreak of Clinical Mastitis in Dairy Sheep Caused by Streptococcus equi subsp. zooepidemicus

PubMed Central

Las Heras, Alfonso; Vela, Ana I.; Fernández, Elena; Legaz, Emilio; Domínguez, Lucas; Fernández-Garayzábal, Jose F.

2002-01-01

This work describes an outbreak of clinical mastitis affecting 13 of 58 lactating ewes due to Streptococcus equi subsp. zooepidemicus. S. equi subsp. zooepidemicus was isolated in pure culture from all milk samples. All the clinical isolates had identical biochemical profiles and antimicrobial susceptibility patterns and also exhibited indistinguishable macrorestriction patterns by pulsed-field gel electrophoresis, indicating that all cases of mastitis were produced by a single strain. PMID:11880454

The mazEF toxin-antitoxin system as a novel antibacterial target in Acinetobacter baumannii.

PubMed

Ghafourian, Sobhan; Good, Liam; Sekawi, Zamberi; Hamat, Rukman Awang; Soheili, Sara; Sadeghifard, Nourkhoda; Neela, Vasanthakumari

2014-07-01

Although analysis of toxin-antitoxin (TA) systems can be instructive, to date, there is no information on the prevalence and identity of TA systems based on a large panel of Acinetobacter baumannii clinical isolates. The aim of the current study was to screen for functional TA systems among clinical isolates of A. baumannii and to identify the systems' locations. For this purpose, we screened 85 A. baumannii isolates collected from different clinical sources for the presence of the mazEF, relBE and higBA TA genes. The results revealed that the genes coding for the mazEF TA system were commonly present in all clinical isolates of A. baumannii. Reverse transcriptase-polymerase chain reaction analysis showed that transcripts were produced in the clinical isolates. Our findings showed that TA genes are prevalent, harboured by chromosomes and transcribed within A. baumannii. Hence, activation of the toxin proteins in the mazEF TA system should be investigated further as an effective antibacterial strategy against this bacterium.

Prevalence of Class D Carbapenemases among Extended-Spectrum β-Lactamases Producing Escherichia coli Isolates from Educational Hospitals in Shahrekord

PubMed Central

Damavandi, Mohammad-Sadegh; Latif Pour, Mohammad

2016-01-01

Introduction Extended-spectrum β-lactamases (ESBLs) are a set of plasmid-borne, various and quickly evolving enzymes that are a main therapeutic issue now-a-days for inpatient and outpatient treatment. Aim The aim of this study was to determine multi-drug resistance (MDR) and ESBLs producing E. coli strains, prevalence of class D Carbapenemases among ESBLs producing Escherichia coli isolates from educational hospitals in Shahrekord, Iran. Materials and Methods Uropathogenic Escherichia coli strains were isolated from patients with Urinary Tract Infections (UTIs). The agar disc diffusion test was used to characterize the antimicrobial sensitivity of the E. coli isolates. The ESBL positive strains were identified by phenotypic double-disk synergy test, by third-generation cephalosporin in combination with or without clavulanic acid. Multiplex PCR was carried out for detection of the three families of OXA-type carbapenamases including OXA-23, OXA-24, and OXA-48 in E. coli strains. Results All bacterial isolates were susceptible to meropenem. Ninety isolates produced ESBL, 55 E. coli isolates from inpatients, and 35 isolates from outpatients, with a significant association (p< 0.05). The prevalence of OXA-23, OXA-24, and OXA-48 in the ESBLs producing isolates was respectively 21%, 18%, and 11% for inpatients, and 10%, 8%, and 6% for outpatients. Conclusion ESBL-producing E. coli isolates are also a major threat in the clinical setting. The findings of this study indicated the high occurrence of ESBLs and multiple antibiotic resistance in E. coli isolates. PMID:27462579

In vitro activity of flomoxef and comparators against Escherichia coli, Klebsiella pneumoniae and Proteus mirabilis producing extended-spectrum β-lactamases in China.

PubMed

Yang, Qiwen; Zhang, Hui; Cheng, Jingwei; Xu, Zhipeng; Xu, Yingchun; Cao, Bin; Kong, Haishen; Ni, Yuxing; Yu, Yunsong; Sun, Ziyong; Hu, Bijie; Huang, Wenxiang; Wang, Yong; Wu, Anhua; Feng, Xianju; Liao, Kang; Shen, Dingxia; Hu, Zhidong; Chu, Yunzhuo; Lu, Juan; Su, Jianrong; Gui, Bingdong; Duan, Qiong; Zhang, Shufang; Shao, Haifeng

2015-05-01

The objective of this study was to better understand the in vitro activity of flomoxef against clinical extended-spectrum β-lactamase (ESBL)-producing Enterobacteriaceae. A total of 401 ESBL-producing isolates, including 196 Escherichia coli, 124 Klebsiella pneumoniae and 81 Proteus mirabilis, were collected consecutively from 21 hospitals in China in 2013. Minimum inhibitory concentrations (MICs) were determined by broth microdilution methods. Phenotypic identification of ESBL production was detected as recommended by the Clinical and Laboratory Standards Institute (CLSI). ESBL genes were detected by PCR and sequencing. Flomoxef, doripenem, meropenem, ertapenem, cefmetazole and piperacillin/tazobactam exhibited good activity against ESBL-producing isolates, with susceptibility rates >90%. Tigecycline showed good activity against E. coli and K. pneumoniae (100% and 97.6%, respectively). Cefotaxime and cefepime showed very low activities against ESBL-producing isolates, with susceptibility rates of 0-0.8% and 1.0-13.6%, respectively. blaCTX-M were the major ESBL genes, with occurrence in 99.5% of E. coli, 91.1% of K. pneumoniae and 97.5% of P. mirabilis. blaCTX-M-14 was the predominant ESBL gene, detected in 46.9% (188/401) of the isolates, followed by blaCTX-M-15 (21.4%), blaCTX-M-55 (17.2%), blaCTX-M-65 (12.7%) and blaCTX-M-3 (6.7%). Flomoxef exhibited excellent activity against the different CTX-M-type ESBL-producing isolates, with MIC50 and MIC90 values of 0.064-0.125μg/mL and 0.25-0.5μg/mL, respectively. Against the isolates solely producing CTX-M-14, -15, -55, -3 or -65, flomoxef showed susceptibility rates of 98.6%, 98.0%, 98.1%, 100.0% and 97.4%, respectively. In conclusion, flomoxef showed good activity against ESBL-producing Enterobacteriaceae and may be a choice to treat infections caused by these isolates in China. Copyright © 2015 Elsevier B.V. and the International Society of Chemotherapy. All rights reserved.

Prevalence of 16S rRNA methylase genes among β-lactamase-producing Enterobacteriaceae clinical isolates in Saudi Arabia

PubMed Central

Al Sheikh, Yazeed A.; Marie, Mohammed Ali M.; John, James; Krishnappa, Lakshmana Gowda; Dabwab, Khaled Homoud M.

2014-01-01

Background Co production of 16S rRNA methylases gene and β-Lactamase gene among Enterobacteriaceae isolates conferring resistance to both therapeutic options has serious implications for clinicians worldwide. Methods To study co existence of 16S rRNA methylases (armA, rmtA, rmtB, rmtC, rmtD, and npmA) and β-Lactamase (blaTEM-1, blaSHV-12, blaCTX-M-14) genes, we screened all phenotypic positive β-Lactamase producing enterobacteriaceae by polymerase chain reaction (PCR) targeting above genes. A total of 330 enterobacteriaceae strains were collected during study period out of that 218 isolates were identified phenotypically as β-Lactamase producers, which include 50 (22.9%) Escherichia coli; 92 (42.2%) Klebsiella pneumoniae, 44 (20.2%), Citrobactor freundii and 32 (14.7%) Enterobacter spp. Results Among this 218, only 188 isolates harbored the resistant gene for β-Lactamase production. Major β-Lactamase producing isolates were bla TEM-1 type. 122 (56 %) isolates were found to produce any one of the 16S rRNA methylase genes. A total of 116 isolates co produced β-Lactamase and at least one 16S rRNA methylases gene Co production of armA gene was found in 26 isolates with rmtB and in 4 isolates with rmtC. The rmtA and rmtD genes were not detected in any of the tested isolates. Six isolates were positive for a 16S rRNA methylase gene alone. Conclusion β-Lactamase producing isolates appears to coexist with 16S rRNA methylase predominantly armA and rmtB genes in the same isolate. We conclude the major β-Lactamase and 16S rRNA methylases co-producer was K. pneumoniae followed by E. coli. We suggest further work on evaluating other β-lactamases types and novel antibiotic resistance mechanisms among Enterobacteriaceae. PMID:25005152

Clinical, microbiological, and experimental animal studies of Candida lipolytica.

PubMed Central

Walsh, T J; Salkin, I F; Dixon, D M; Hurd, N J

1989-01-01

Candida lipolytica was recovered from six patients in three different clinical centers. The index isolate caused a persistent fungemia with catheter-associated Candida thrombophlebitis, the second isolate was from a polymicrobial sinusitis, and the remaining four isolates were involved in tissue colonization. These and 20 other isolates were consistent in their morphological and physiological characteristics. All formed true hyphae and blastoconidia on cornmeal-Tween 80 agar and all assimilated glucose, glycerol, and erythritol. In a murine model of disseminated candidiasis, the index isolate that caused clinical fungemia caused no mortality and produced only two lesions on a kidney, as determined at necropsy. The nine isolates selected for in vitro antifungal susceptibility studies had intermediate susceptibilities to amphotericin B but were susceptible to ketoconazole. We conclude that C. lipolytica is a weakly virulent pathogen which may require an intravascular foreign body to cause fungemia. Images PMID:2745702

Putative Virulence Factor Expression by Clinical and Food Isolates of Bacillus spp. after Growth in Reconstituted Infant Milk Formulae

PubMed Central

Rowan, Neil J.; Deans, Karen; Anderson, John G.; Gemmell, Curtis G.; Hunter, Iain S.; Chaithong, Thararat

2001-01-01

Forty-seven strains representing 14 different Bacillus species isolated from clinical and food samples were grown in reconstituted infant milk formulae (IMF) and subsequently assessed for adherence to, invasion of, and cytotoxicity toward HEp-2 and Caco-2 cells. Cell-free supernatant fluids from 38 strains (81%) were shown to be cytotoxic, 43 strains (91%) adhered to the test cell lines, and 23 strains (49%) demonstrated various levels of invasion. Of the 21 Bacillus cereus strains examined, 5 (24%) were invasive. A larger percentage of clinically derived Bacillus species (20%) than of similar species tested from the food environment were invasive. Increased invasion occurred after growth of selected Bacillus species in reconstituted IMF containing glucose. While PCR primer studies revealed that many different Bacillus species contained DNA sequences encoding the hemolysin BL (HBL) enterotoxin complex and B. cereus enterotoxin T, not all of these isolates expressed these diarrheagenic genes after growth in reconstituted IMF. Of the 47 Bacillus isolates examined, 3 isolates of B. cereus and 1 isolate of B. subtilis produced the HBL enterotoxin after 18 h of growth in brain heart infusion broth. However, eight isolates belonging to the species B. cereus, B. licheniformis, B. circulans, and B. megaterium were found to produce this enterotoxin after growth in reconstituted IMF when assessed with the B. cereus enterotoxin (diarrheal type) reversed passive latex agglutination (RPLA) kit. It is concluded that several Bacillus species occurring occasionally in clinical specimens and food samples are of potential medical significance due to the expression of putative virulence factors. PMID:11525980

Molecular epidemiology of Pseudomonas aeruginosa clinical isolates from Korea producing β-lactamases with extended-spectrum activity.

PubMed

Bae, Il Kwon; Suh, Borum; Jeong, Seok Hoon; Wang, Kang-Kyun; Kim, Yong-Rok; Yong, Dongeun; Lee, Kyungwon

2014-07-01

This study was performed to investigate the prevalence and molecular epidemiology of Pseudomonas aeruginosa isolates from Korea that produce enzymes with extended-spectrum (ES) activity to β-lactams. A total of 205 non-duplicate P. aeruginosa clinical isolates were collected from 18 university hospitals in Korea. PCR and sequencing experiments were performed to identify genes encoding β-lactamases. PCR mapping and sequencing of the regions surrounding the β-lactamase genes were performed. Multilocus sequence typing experiments were performed. The most common sequence type (ST) was ST235 (n = 96), and 2 single-locus variants of ST235, ST1015 (n = 1) and ST1162 (n = 1), were also identified. These 3 STs were grouped as a clonal complex (CC), CC235. The remaining 107 isolates were identified as 59 different STs. Isolates belonging to CC235 showed higher rates of non-susceptibility to imipenem (85.4% versus 47.7%) and meropenem (92.7% versus 52.3%) compared to non-CC235 isolates. All the metallo-β-lactamase (MBL)-producing isolates were identified as CC235, except for 1 ST591. Genes encoding OXA-17 and OXA-142 were detected in 1 isolate and 4 isolates of CC235, respectively; while the bla(SHV-12) gene was detected in 4 non-CC235 isolates. Class A and D β-lactamases with ES activity play a role in acquiring ceftazidime resistance in P. aeruginosa in Korea. Production of IMP-6 and VIM-2 MBLs is the main mechanisms in acquiring resistance to ceftazidime and carbapenems in P. aeruginosa isolates in Korea. Clonal spread of P. aeruginosa CC235 may be an important conduit for the dissemination of MBL genes in Korea. Copyright © 2014 Elsevier Inc. All rights reserved.

Development and Validation of a Lateral Flow Immunoassay for Rapid Detection of NDM-Producing Enterobacteriaceae

PubMed Central

Boutal, Hervé; Naas, Thierry; Devilliers, Karine; Oueslati, Saoussen; Bernabeu, Sandrine; Simon, Stéphanie

2017-01-01

ABSTRACT The global spread of carbapenemase-producing Enterobacteriaceae (CPE) that are often resistant to most, if not all, classes of antibiotics is a major public health concern. The NDM-1 carbapenemase is among the most worrisome carbapenemases given its rapid worldwide spread. We have developed and evaluated a lateral flow immunoassay (LFIA) (called the NDM LFIA) for the rapid and reliable detection of NDM-like carbapenemase-producing Enterobacteriaceae from culture colonies. We evaluated the NDM LFIA using 175 reference enterobacterial isolates with characterized β-lactamase gene content and 74 nonduplicate consecutive carbapenem-resistant clinical isolates referred for expertise to the French National Reference Center (NRC) for Antibiotic Resistance during a 1-week period (in June 2016). The reference collection included 55 non-carbapenemase producers and 120 carbapenemase producers, including 27 NDM producers. All 27 NDM-like carbapenemase producers of the reference collection were correctly detected in less than 15 min by the NDM LFIA, including 22 strains producing NDM-1, 2 producing NDM-4, 1 producing NDM-5, 1 producing NDM-7, and 1 producing NDM-9. All non-NDM-1 producers gave a negative result with the NDM LFIA. No cross-reaction was observed with carbapenemases (VIM, IMP, NDM, KPC, and OXA-48-like), extended-spectrum β-lactamases (ESBLs) (TEM, SHV, and CTX-M), AmpCs (CMY-2, DHA-2, and ACC-1), and oxacillinases (OXA-1, -2, -9, and -10). Similarly, among the 74 referred nonduplicate consecutive clinical isolates, all 7 NDM-like producers were identified. Overall, the sensitivity and specificity of the assay were 100% for NDM-like carbapenemase detection with strains cultured on agar. The NDM LFIA was efficient, rapid, and easy to implement in the routine workflow of a clinical microbiology laboratory for the confirmation of NDM-like carbapenemase-producing Enterobacteriaceae. PMID:28404680

Route of infection alters virulence of neonatal septicemia Escherichia coli clinical isolates

PubMed Central

Cole, Bryan K.; Scott, Edgar; Ilikj, Marko; Bard, David; Akins, Darrin R.; Dyer, David W.

2017-01-01

Escherichia coli is the leading cause of Gram-negative neonatal septicemia in the United States. Invasion and passage across the neonatal gut after ingestion of maternal E. coli strains produce bacteremia. In this study, we compared the virulence properties of the neonatal E. coli bacteremia clinical isolate SCB34 with the archetypal neonatal E. coli meningitis strain RS218. Whole-genome sequencing data was used to compare the protein coding sequences among these clinical isolates and 33 other representative E. coli strains. Oral inoculation of newborn animals with either strain produced septicemia, whereas intraperitoneal injection caused septicemia only in pups infected with RS218 but not in those injected with SCB34. In addition to being virulent only through the oral route, SCB34 demonstrated significantly greater invasion and transcytosis of polarized intestinal epithelial cells in vitro as compared to RS218. Protein coding sequences comparisons highlighted the presence of known virulence factors that are shared among several of these isolates, and revealed the existence of proteins exclusively encoded in SCB34, many of which remain uncharacterized. Our study demonstrates that oral acquisition is crucial for the virulence properties of the neonatal bacteremia clinical isolate SCB34. This characteristic, along with its enhanced ability to invade and transcytose intestinal epithelium are likely determined by the specific virulence factors that predominate in this strain. PMID:29236742

Characterization of AmpC, CTX-M and MBLs types of β-lactamases in clinical isolates of Klebsiella pneumoniae and Escherichia coli producing Extended Spectrum β-lactamases in Kerman, Iran

PubMed Central

Mansouri, Shahla; Kalantar Neyestanaki, Davood; Shokoohi, Mostafa; Halimi, Shahnaz; Beigverdi, Reza; Rezagholezadeh, Fereshteh; Hashemi, Ali

2014-01-01

Background: Extended spectrum β-lactamases (ESBLs) and AmpC β-lactamases enzyme are major sources of resistance to β-lactam antibiotics especially in Enterobacteriaceae such as Escherichia coli and Klebsiella pneumoniae. Increasing frequency of the co-existence of ESBLs with AmpC-β-lactamases in bacteria is a serious threat for treating bacterial infections. Objectives: The aim of this study was to determine the presence of AmpC and CTX-M types of β-lactamases in clinical isolates of E. coli and K. pneumoniae producing ESBLs. Materials and Methods: Resistance to different antibiotics was determined using the standard disk diffusion method. ESBLs, MBLs and AmpC-β-lactamases were detected by the combination double disk test (CDDT) method and polymerase chain reaction (PCR) was used to determine blaCTX-M genes in the ESBLs and AmpC positive isolates. Results: The prevalence of ESBLs and AmpC-β-lactamase producer isolates was 181 (43.8%) and 133 (37.2%), respectively. The prevalence of blaCTX-M among isolates was 61 (14.7%). Conclusions: Outbreak of isolates co-expressing AmpC-β-lactamases and ESBLs can cause serious problems in the future, regarding the treatment of infections caused by these common enteric pathogens. PMID:25147671

Bacterial and fungal keratitis in Upper Egypt: In vitro screening of enzymes, toxins and antifungal activity

PubMed Central

Gharamah, Abdullah A; Moharram, Ahmed M; Ismail, Mady A; AL-Hussaini, Ashraf K

2014-01-01

Purpose: This work was conducted to study the ability of bacterial and fungal isolates from keratitis cases in Upper Egypt to produce enzymes, toxins, and to test the isolated fungal species sensitivity to some therapeutic agents. Materials and Methods: One hundred and fifteen patients clinically diagnosed to have microbial keratitis were investigated. From these cases, 37 bacterial isolates and 25 fungal isolates were screened for their ability to produce extra-cellular enzymes in solid media. In addition, the ability of fungal isolates to produce mycotoxins and their sensitivity to 4 antifungal agents were tested. Results: Protease, lipase, hemolysins, urease, phosphatase, and catalase were detected respectively in 48.65%, 37.84%, 59.46%, 43.24%, 67.57%, and 100% out of 37 bacterial isolates tested. Out of 25 fungal isolates tested during the present study, 80% were positive for protease, 84% for lipase and urease, 28% for blood hemolysis, and 100% for phosphatase and catalase enzymes. Thirteen fungal isolates were able to produce detectable amounts of 7 mycotoxins in culture medium (aflatoxins (B1, B2, G1, and G2), sterigmatocystin, fumagillin, diacetoxyscirpenol, zearalenone, T-2 toxin, and trichodermin). Among the antifungal agents tested in this study, terbinafine showed the highest effect against most isolates in vitro. Conclusion: In conclusion, the ability of bacterial and fungal isolates to produce extracellular enzymes and toxins may be aid in the invasion and destruction of eye tissues, which, in turn, lead to vision loss. PMID:24008795

Increasing proportion of carbapenemase-producing Enterobacteriaceae and emergence of a MCR-1 producer through a multicentric study among hospital-based and private laboratories in Belgium from September to November 2015.

PubMed

Huang, Te Din; Bogaerts, Pierre; Berhin, Catherine; Hoebeke, Martin; Bauraing, Caroline; Glupczynski, Youri

2017-05-11

Carbapenemase-producing Enterobacteriaceae (CPE) strains have been increasingly reported in Belgium. We aimed to determine the proportion of CPE among Enterobacteriaceae isolated from hospitalised patients and community outpatients in Belgium in 2015. For the hospitalised patients, the results were compared to a previous similar survey performed in the same hospitals in 2012. Twenty-four hospital-based and 10 private laboratories collected prospectively 200 non-duplicated Enterobacteriaceae isolates from clinical specimens. All isolates were screened locally by carbapenem disk diffusion using European Committee on Antimicrobial Susceptibility Testing methodology. Putative CPE strains with inhibition zone diameters below the screening breakpoints were referred centrally for confirmation of carbapenemase production. From September to November 2015, we found a proportion of clinical CPE of 0.55% (26/4,705) and of 0.60% (12/1,991) among hospitalised patients and among ambulatory outpatients respectively. Klebsiella pneumoniae (26/38) and OXA-48-like carbapenemase (28/38) were the predominant species and enzyme among CPE. One OXA-48-producing Escherichia coli isolated from a hospital was found carrying plasmid-mediated MCR-1 colistin resistance. Compared with the 2012 survey, we found a significant increased proportion of clinical CPE (0.55% in 2015 vs 0.25% in 2012; p = 0.02) and an increased proportion of hospitals (13/24 in 2015 vs 8/24 in 2012) with at least one CPE detected. The study results confirmed the concerning spread of CPE including a colistin-resistant MCR-1 producer in hospitals and the establishment of CPE in the community in Belgium. This article is copyright of The Authors, 2017.

Emergence of Salmonella genomic island 1 (SGI1) among Proteus mirabilis clinical isolates in Dijon, France.

PubMed

Siebor, Eliane; Neuwirth, Catherine

2013-08-01

Salmonella genomic island 1 (SGI1) is often encountered in antibiotic-resistant Salmonella enterica and exceptionally in Proteus mirabilis. We investigated the prevalence of SGI1-producing clinical isolates of P. mirabilis in our hospital (Dijon, France). A total of 57 strains of P. mirabilis resistant to amoxicillin and/or gentamicin and/or trimethoprim/sulfamethoxazole isolated from August 2011 to February 2012 as well as 9 extended-spectrum β-lactamase (ESBL)-producing P. mirabilis from our collection were tested for the presence of SGI1 by PCR. The complete SGI1 structure from positive isolates [backbone and multidrug resistance (MDR) region] was sequenced. SGI1 was detected in 7 isolates; 5 out of the 57 isolates collected during the study period (9%) and 2 out of the 9 ESBL-producing strains of our collection. The structures of the seven SGI1s were distinct. Three different backbones were identified: one identical to the SGI1 backbone from the epidemic Salmonella Typhimurium DT104, one with variations already described in SGI1-K from Salmonella Kentucky (deletion and insertion of IS1359 in the region spanning from S005 to S009) and one with a variation never detected before (deletion from S005 to S009). Six different MDR regions were identified: four simple variants containing resistance genes already described and two variants harbouring a very complex structure including regions derived from several transposons and IS26 elements with aphA1a never reported to date in SGI1. SGI1 variants are widely distributed among P. mirabilis clinical strains and might spread to other commensal Enterobacteriaceae. This would become a serious public health problem.

Identification and characterization of metallo-β-lactamases producing Pseudomonas aeruginosa clinical isolates in University Hospital from Zanjan Province, Iran.

PubMed

Doosti, Masoumeh; Ramazani, Ali; Garshasbi, Maryam

2013-01-01

Infectious by Pseudomonas aeruginosa has spread worldwide and metallo-beta-lactamases (MBL) are being reported with increasing frequency. The aim of this study was to investigate the antibiotic susceptibility and distribution of blaVIM and blaIMP genes in P. aeruginosa isolates from Zanjan Province of Iran. A total of 70 P. aeruginosa isolates were identified from patients admitted at intensive care units. The antimicrobial susceptibility was tested by disk diffusion (Kirby-Bauer) method and for production of MBL using double-disk synergy test (DDST). After DNA extraction, the presence of blaVIM and blaIMP genes and class 1 integron were detected by PCR. Most of the isolates were resistant to meropenem, cefotaxime and imipenem (IPM). Also, 44/70 (62.85%) IPM resistant isolates were confirmed by DDST. Of the 44 clinical isolates, 41 (93%) isolates showed MIC≥4 µg/ml for IPM. Based on the DDST results, 36 (87.8%) were confirmed to be MBL producers. PCR amplification showed that 23/41 (56%) carried blaVIM and 10/41 (24.3%) possessed blaIMP gene. Also, 31/44 (70.5%) isolates contained class 1 integron gene. Our results highlight that the genes for Verona integron-encoded metallo-β-lactamase, IPM β-lactamases and class 1 integrons were predominantly present among the IPM-resistant P. aeruginosa tested in our province and also the frequency of blaVIM type is higher than blaIMP. This is the first report of P. aeruginosa strains producing blaIMP with high frequency from Zanjan province of Iran.

Efficacy of non-carbapenem antibiotics for pediatric patients with first febrile urinary tract infection due to extended-spectrum beta-lactamase-producing Escherichia coli.

PubMed

Abe, Yoshifusa; Inan-Erdogan, Işil; Fukuchi, Kunihiko; Wakabayashi, Hitomi; Ogawa, Yasuha; Hibino, Satoshi; Sakurai, Shunsuke; Matsuhashi, Kazuhiko; Watanabe, Yoshitaka; Hashimoto, Kaori; Ugajin, Kazuhisa; Itabashi, Kazuo

2017-08-01

Although carbapenem is the recommended for urinary tract infection (UTI) caused by extended-spectrum beta-lactamase (ESBL)-producing organisms, non-carbapenems have been reported to be effective for adult patients with UTI caused by ESBL-producing organisms. The purpose of this study was to evaluate the efficacy of non-carbapenems for pediatric patients with UTI due to ESBL-producing Escherichia coli (E. coli) based on the microbiologic and clinical outcomes. Fifteen children, who were treated for first febrile UTI caused by ESBL-producing E. coli were enrolled in this study. Antimicrobial susceptibilities and ESBL production were determined according to the Clinical and Laboratory Standards Institute guidelines. To detect CTX-M genes, polymerase chain reaction was performed with specific primers for bla CTX-M detection. Of the 15 enrolled patients, 10 (66.7%) were boys and 5 (33.3%) were girls, with a median age of four months. VUR was detected in six patients (40%). For detection of bla CTX-M by PCR, CTX-M-3, CTX-M-8, CTX-M-14, and CTX-M-15 were detected in five, one, eight, and one patient, respectively. Overall, 14 of the 15 isolates (93.3%) were susceptible for fosfomycin (FOM), and all isolates were susceptible for cefmetazole (CMZ), flomoxef (FMOX), and imipenem/cilastatin (IPM/CS). Of the 15 patients, 12 (80%) clinically improved without the use of carbapenems. In conclusion, even if isolates of ESBL-producing E. coli are multidrug resistant based on MIC assessment, clinical susceptibility to non-carbapenems, such as CMZ, FMOX, and FOM, is possible. Accordingly, carbapenems may not be required all the time for treatment of pediatric UTI in clinical practice. Copyright © 2017 Japanese Society of Chemotherapy and The Japanese Association for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

Draft Genome Sequence of the First New Delhi Metallo-β-Lactamase (NDM-1)-Producing Escherichia coli Strain Isolated in Peru.

PubMed

Tamariz, Jesus; Llanos, Carlos; Seas, Carlos; Montenegro, Paola; Lagos, Jose; Fernandes, Miriam R; Cerdeira, Louise; Lincopan, Nilton

2018-03-29

We present here the draft genome sequence of the first New Delhi metallo-β-lactamase (NDM-1)-producing Escherichia coli strain, belonging to sequence type 155 (ST155), isolated in Peru. Assembly of this draft genome resulted in 5,061,184 bp, revealing a clinically significant resistome for β-lactams, aminoglycosides, tetracyclines, phenicols, sulfonamides, trimethoprim, and fluoroquinolones. Copyright © 2018 Tamariz et al.

Functional Characterization of Bacteria Isolated from Ancient Arctic Soil Exposes Diverse Resistance Mechanisms to Modern Antibiotics

PubMed Central

Perron, Gabriel G.; Whyte, Lyle; Turnbaugh, Peter J.; Goordial, Jacqueline; Hanage, William P.; Dantas, Gautam; Desai, Michael M.

2015-01-01

Using functional metagenomics to study the resistomes of bacterial communities isolated from different layers of the Canadian high Arctic permafrost, we show that microbial communities harbored diverse resistance mechanisms at least 5,000 years ago. Among bacteria sampled from the ancient layers of a permafrost core, we isolated eight genes conferring clinical levels of resistance against aminoglycoside, β-lactam and tetracycline antibiotics that are naturally produced by microorganisms. Among these resistance genes, four also conferred resistance against amikacin, a modern semi-synthetic antibiotic that does not naturally occur in microorganisms. In bacteria sampled from the overlaying active layer, we isolated ten different genes conferring resistance to all six antibiotics tested in this study, including aminoglycoside, β-lactam and tetracycline variants that are naturally produced by microorganisms as well as semi-synthetic variants produced in the laboratory. On average, we found that resistance genes found in permafrost bacteria conferred lower levels of resistance against clinically relevant antibiotics than resistance genes sampled from the active layer. Our results demonstrate that antibiotic resistance genes were functionally diverse prior to the anthropogenic use of antibiotics, contributing to the evolution of natural reservoirs of resistance genes. PMID:25807523

CTX-M-15 and OXA-10 beta lactamases in multi drug resistant Pseudomonas aeruginosa: First report from Pakistan.

PubMed

Ullah, Waheed; Qasim, Muhammad; Rahman, Hazir; Khan, Saadullah; Rehman, Zia Ur; Ali, Nawab; Muhammad, Noor

2017-04-01

Pseudomonas aeruginosa is an emerging threat to public health worldwide due to their rapid development of drug resistance including beta-lactamases. The present study was designed to investigate the incidence of β-lactamases and genotypic pattern of CTX and OXA in the clinical isolate of multidrug resistant P. aeruginosa. In this study a total of 102 MDR P. aeruginosa isolates obtained from Lady Reading Hospital, Peshawar, Pakistan were subjected to extended spectrum beta lactamase (ESBL), metallo beta lactamase (MBL) and plasmid mediated β-lactamase (AmpC) detection using phenotypic and molecular methods. Furthermore, sequencing of CTX and OXA gene was performed. Out of 102 MDR P. aeruginosa isolates, 71 (69.6%) were beta lactamase producers. The incidence of ESBL, MBL and AmpC in clinical isolates of P. aeruginosa was found to be 23.94%, 40.84% and 35.21% respectively. Co-production of ESBL and AmpC were also observed in some isolates. There were 14 (19.71%) CTX-M-15 harboring isolates which were ESBL (64.28%), MBL (21.42%) and AmpC (14.28%) producer. Co-production of ESBL/MBL (14.28%), ESBL/AmpC (14.28%) and MBL/AmpC (14.28%) were also observed in the CTX M-15 harboring isolates while 12.28% isolates were not ESBL, MBL or AmpC producer. OXA-10 was detected in 8 (11.26%) isolates which were ESBL (12.5%), MBL (37.5%) and AmpC (12.5%) producer. OXA 10 isolates also exhibit co-production of ESBL/AmpC (12.5%) and MBL/AmpC (12.5%). All CTX-M-15 carried the class A β-lactamase conserved domain while OXA-10 harbored conserved domain of class D β-lactamase. The current study for the first time reported and characterized the CTX-M-15 and OXA-10 among MDR P. aeruginosa isolates from Pakistan. Further efforts are needed to understand the molecular mechanism of drug resistance with CTX and OXA harboring P. aeruginosa isolates. Copyright © 2017 Elsevier Ltd. All rights reserved.

Bacteraemia due to non-ESBL-producing Escherichia coli O25b:H4 sequence type 131: insights into risk factors, clinical features and outcomes.

PubMed

Morales-Barroso, Isabel; López-Cerero, Lorena; Molina, José; Bellido, Mar; Navarro, María Dolores; Serrano, Lara; González-Galán, Verónica; Praena, Julia; Pascual, Alvaro; Rodríguez-Baño, Jesús

2017-04-01

The epidemiology and outcomes of bloodstream infections (BSIs) caused by Escherichia coli ST131 isolates not producing extended-spectrum β-lactamases (ESBLs) are not well defined despite being more prevalent than ESBL-producers. In this study, risk factors and the impact on outcome of BSIs caused by non-ESBL-producing ST131 E. coli versus non-ST131 E. coli were investigated. A case-control study was performed in two tertiary centres to identify risk factors for ST131. Molecular methods were used to investigate all E. coli isolates from blood cultures for those belonging to O25b:H4-ST131 clonal group. fimH alleles were characterised in ST131 isolates. Multivariate analysis was performed by logistic regression or Cox regression as appropriate. A total of 33 ST131 E. coli cases and 56 controls were studied. ST131 isolates showed higher rates of resistance to ampicillin and ciprofloxacin; fimH alleles were H30 in 14 isolates (42.4%) and H22 in 12 isolates (36.4%). Only recent surgery (OR = 7.03, 95% CI 1.71-28.84; P = 0.007) and unknown source of bacteraemia (OR = 5.37, 95% CI 0.93-30.81; P = 0.05) were associated with ST131. ST131 isolates showed no association with 30-day mortality, therapeutic failure, presentation with severe sepsis/shock or length of stay. Bacteraemia due to non-ESBL-producing O25b:H4-ST131 E. coli showed few differences in terms of risk factors as well as similar outcome to non-ST131 E. coli. These data support the notion that ST131 strains are not less clinically virulent despite showing increased antimicrobial resistance, but also that they are not more virulent than other clonal groups causing BSI. Copyright © 2017 Elsevier B.V. and International Society of Chemotherapy. All rights reserved.

Detection of carbapenemases and other mechanisms of enzymatic resistance to β-lactams in Enterobacteriaceae with diminished susceptibility to carbapenems in a tertiary care hospital.

PubMed

Gómara, Marta; López-Calleja, Ana Isabel; Iglesia, Berta María Pilar Vela; Cerón, Isabel Ferrer; López, Antonio Rezusta; Pinilla, María José Revillo

2018-05-01

Our objective was to characterize the enzymatic β-lactam resistance in clinical Enterobacteriaceae isolates with diminished susceptibility to carbapenems from 2013 to 2014 at Hospital Universitario Miguel Servet. A total of 63 clinical isolates were analyzed for the presence of carbapenemases (KPC, OXA-48 and MBL), ESBLs and AmpC enzymes by combined disk methods and PCR detection of carbapenemase-encoding and beta-lactamase-encoding genes. Fifteen isolates had a phenotypic test compatible with carbapenemase production; two of these were confirmed by PCR as OXA-48 producers. ESBL detection was positive in 27 isolates (43%); plasmid-mediated AmpC was detected in nine isolates (14.2%) and derepressed AmpC β-lactamase was present in 18 isolates (28%). During the study period, the decreased susceptibility to carbapenems in Enterobacteriaceae in our area was not due to true carbapenemases but rather to β-lactamase activity (82.5% were ESBL or AmpC producers), probably in combination with decreased permeability of the outer membrane. Copyright © 2017 Elsevier España, S.L.U. and Sociedad Española de Enfermedades Infecciosas y Microbiología Clínica. All rights reserved.

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

A simple screening test for the detection of metallo-β-lactamase-producing Pseudomonas aeruginosa and Acinetobacter in a tertiary care hospital.

PubMed

Wan Nor Amilah, W A W; Noor Izani, N J; Ng, W K; Ashraful Haq, J

2012-12-01

Clinical utilization of carbapenems remains under threat with the emergence of acquired carbapenemase-producing bacteria, particularly metallo-β-lactamases (MBL). Rapid detection of MBL-producing Gram-negative bacilli is essential to prevent their widespread dissemination. However, no standardized detection method is available for routine laboratory use. The purpose of the study was to evaluate a chelating-agent based double disk synergic test and disk potentiation test for MBL-producing strain detection and to determine the isolation rate of MBL-producing Pseudomonas aeruginosa and Acinetobacter from clinical samples in our tertiary teaching hospital. A total of 22 and 66 imipenem-resistant P. aeruginosa and Acinetobacter isolates respectively were tested with ceftazidime (CAZ) disk by modified double disk synergic test and disk potentiation test using ethylenediaminetetraacetic acid (EDTA) and 2-mercaptopropionic acid (as chelating agents) to detect MBL production. The tests were compared with EDTA-phenanthroline-imipenem (EPI) microdilution MIC test as gold standard. MBL positive strains were detected in 17 (77.3%) P. aeruginosa and 2 (3.5%) Acinetobacter isolates. The disk potentiation test with 2-mercaptopropionic acid (2-MPA) dilution of 1:12 provided the most acceptable sensitivities and specificities (88.2% sensitivity and 100% specificity in P. aeruginosa; 100% sensitivity and specificity in Acinetobacter) compared to other screening methods used in this study. This study provided useful information on the local prevalence of MBL-producing P. aeruginosa and Acinetobacter in our hospital. Disc potentiation test with CAZ/2-MPA disc appears to be reliable and convenient MBL detection method in the routine clinical laboratory.

beta-lactamase production by oral pigmented Prevotella species isolated from young children.

PubMed

Könönen, E; Nyfors, S; Mättö, J; Asikainen, S; Jousimies-Somer, H

1997-09-01

The frequency of beta-lactamase production by oral pigmented Prevotella species isolated from 23 healthy young children and the minimal inhibitory concentrations (MICs) for 186 available beta-lactamase-positive isolates were examined by using the chromogenic cephalosporin disk test (AB BIODISK, Solna, Sweden) and the Etest (AB BIODISK) and/or the agar dilution method of the National Committee for Clinical Laboratory Standards (Villanova, PA, USA), respectively. beta-Lactamase-positive Prevotella melaninogenica strains were isolated from all children, and more than two-thirds of the Prevotella denticola and Prevotella loescheii strains isolated from the children were beta-lactamase-positive. The beta-lactamase-producing Prevotella intermedia group consisted of Prevotella nigrescens and the P. intermedia/ P. nigrescens-like organism (PINLO); P. intermedia was not found. Only two P. nigrescens isolates but most of the PINLO isolates produced beta-lactamase. The MICs for beta-lactamase-producing strains varied between 0.38 and 64 micrograms/mL. beta-Lactamase production by oral pigmented Prevotella species colonizing young children is already frequent. The phenomenon should be taken into account in the treatment of pediatric anaerobic infections of oral origin.

Eliciting antibiotics active against the ESKAPE pathogens in a collection of actinomycetes isolated from mountain soils.

PubMed

Zhu, Hua; Swierstra, Jasper; Wu, Changsheng; Girard, Geneviève; Choi, Young Hae; van Wamel, Willem; Sandiford, Stephanie K; van Wezel, Gilles P

2014-08-01

The rapid emergence of multidrug-resistant (MDR) bacterial pathogens poses a major threat for human health. In recent years, genome sequencing has unveiled many poorly expressed antibiotic clusters in actinomycetes. Here, we report a well-defined ecological collection of >800 actinomycetes obtained from sites in the Himalaya and Qinling mountains, and we used these in a concept study to see how efficiently antibiotics can be elicited against MDR pathogens isolated recently from the clinic. Using 40 different growth conditions, 96 actinomycetes were identified - predominantly Streptomyces - that produced antibiotics with efficacy against the MDR clinical isolates referred to as ESKAPE pathogens: Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa and/or Enterobacter cloacae. Antimicrobial activities that fluctuated strongly with growth conditions were correlated with specific compounds, including borrelidin, resistomycin, carbomethoxy-phenazine, and 6,7,8- and 5,6,8-trimethoxy-3-methylisocoumarin, of which the latter was not described previously. Our work provided insights into the potential of actinomycetes as producers of drugs with efficacy against clinical isolates that have emerged recently and also underlined the importance of targeting a specific pathogen. © 2014 The Authors.

Comparison of the MUREX C. albicans, Albicans-Sure, and BactiCard Candida test kits with the germ tube test for presumptive identification of Candida albicans.

PubMed Central

Crist, A E; Dietz, T J; Kampschroer, K

1996-01-01

The MUREX C. albicans (MC)(Murex Diagnostics), Albicans-Sure (AS) (Clinical Standards Laboratories), and BactiCard Candida (BC) (Remel) test kits were compared with the germ tube (GT) test for the rapid, presumptive identification of Candida albicans. All three test kits detect the enzymes L-proline aminopeptidase and beta-galactosaminidase in yeast cells grown on culture media and are based on the principle that C. albicans produces both enzymes whereas other yeasts produce only one or neither of the enzymes. The organisms evaluated were fresh clinical isolates identified by methods routinely used in our laboratory (API 20C system and conventional methods) and included 303 C. albicans isolates, 153 Candida glabrata isolates, 70 Candida tropicalis isolates, 36 Candida parapsilosis isolates, 13 isolates of other Candida spp., 5 Cryptococcus neoformans isolates, and 3 Saccharomyces cerevisiae isolates. The MC, AS, BC, and GT tests detected 299 (98.7%), 300 (99.0%), 301 (99.3%), and 287 (94.7%) C. albicans isolates, respectively. There was one false-positive result with both the MC and BC kits and two false-positive results with the GT test. The enzymatic methods evaluated in this study provide rapid and accurate alternatives to the GT test for the presumptive identification of C. albicans. PMID:8880535

Extended-spectrum β-lactamases and plasmid-mediated quinolone resistance in enterobacterial clinical isolates from neonates in Tunisia.

PubMed

Charfi, Karama; Grami, Raoudha; Ben Jeddou, Abir; Messaoudi, Aziza; Mani, Yosra; Bouallegue, Olfa; Boujaafar, Noureddine; Aouni, Mahjoub; Mammeri, Hedi; Mansour, Wejdène

2017-09-01

This study was conducted to investigate extended-spectrum-β-lactamase (ESBL) producing Enterobacteriaceae isolates from the Center of Maternity and Neonatology of Monastir, Tunisia. Fourty-six strains out of 283 were found to produce ESBL: Klebsiella pneumoniae (n = 37), Escherichia coli (n = 6), Enterobacter cloacae (n = 2), and Citrobacter freundi (n = 1). Genotyping analysis, using ERIC2 and RAPD, showed that strains were clonally unrelated. PCR amplification followed by sequencing revealed that all strains produced CTX-M-15. This enzyme was co-produced with TEM and SHV determinants in 34 and 36 strains respectively. The bla CTXM-15 gene was bracked by ISEcp1 and/or IS26 in 42 out of the 46 ESBL positive strains. The quinolone resistance determinants were associated to the ESBL producing isolates: we identified the qnrB1 gene in six isolates and the aac(6')-Ib-cr gene in five isolates. This epidemiological study shows the widespread of CTX-M-15 and qnr determinants among enterobacterial isolates from neonates hospitalized at the center of Maternity and Neonatology of Monastir suggesting either mother portage or horizontal transmission. Copyright © 2017 Elsevier Ltd. All rights reserved.

Comparison of Predictors and Mortality Between Bloodstream Infections Caused by ESBL-Producing Escherichia coli and ESBL-Producing Klebsiella pneumoniae.

PubMed

Scheuerman, Oded; Schechner, Vered; Carmeli, Yehuda; Gutiérrez-Gutiérrez, Belen; Calbo, Esther; Almirante, Benito; Viale, Pier-Luigy; Oliver, Antonio; Ruiz-Garbajosa, Patricia; Gasch, Oriol; Gozalo, Monica; Pitout, Johann; Akova, Murat; Peña, Carmen; Molina, Jose; Hernández-Torres, Alicia; Venditti, Mario; Prim, Nuria; Origüen, Julia; Bou, German; Tacconelli, Evelina; Tumbarello, Maria; Hamprecht, Axel; Karaiskos, Ilias; de la Calle, Cristina; Pérez, Federico; Schwaber, Mitchell J; Bermejo, Joaquin; Lowman, Warren; Hsueh, Po-Ren; Navarro-San Francisco, Carolina; Bonomo, Robert A; Paterson, David L; Pascual, Alvaro; Rodríguez-Baño, Jesus

2018-06-01

OBJECTIVETo compare the epidemiology, clinical characteristics, and mortality of patients with bloodstream infections (BSI) caused by extended-spectrum β-lactamase (ESBL)-producing Escherichia coli (ESBL-EC) versus ESBL-producing Klebsiella pneumoniae (ESBL-KP) and to examine the differences in clinical characteristics and outcome between BSIs caused by isolates with CTX-M versus other ESBL genotypesMETHODSAs part of the INCREMENT project, 33 tertiary hospitals in 12 countries retrospectively collected data on adult patients diagnosed with ESBL-EC BSI or ESBL-KP BSI between 2004 and 2013. Risk factors for ESBL-EC versus ESBL-KP BSI and for 30-day mortality were examined by bivariate analysis followed by multivariable logistic regression.RESULTSThe study included 909 patients: 687 with ESBL-EC BSI and 222 with ESBL-KP BSI. ESBL genotype by polymerase chain reaction amplification of 286 isolates was available. ESBL-KP BSI was associated with intensive care unit admission, cardiovascular and neurological comorbidities, length of stay to bacteremia >14 days from admission, and a nonurinary source. Overall, 30-day mortality was significantly higher in patients with ESBL-KP BSI than ESBL-EC BSI (33.7% vs 17.4%; odds ratio, 1.64; P=.016). CTX-M was the most prevalent ESBL subtype identified (218 of 286 polymerase chain reaction-tested isolates, 76%). No differences in clinical characteristics or in mortality between CTX-M and non-CTX-M ESBLs were detected.CONCLUSIONSClinical characteristics and risk of mortality differ significantly between ESBL-EC and ESBL-KP BSI. Therefore, all ESBL-producing Enterobacteriaceae should not be considered a homogeneous group. No differences in outcomes between genotypes were detected.CLINICAL TRIALS IDENTIFIERClinicalTrials.gov. Identifier: NCT01764490.Infect Control Hosp Epidemiol 2018;39:660-667.

The isolation of Cryptococcus neoformans from pigeon droppings and serotyping of naturally and clinically sourced isolates in China.

PubMed

Li, A; Nishimura, K; Taguchi, H; Tanaka, R; Wu, S; Miyaji, M

1993-10-01

This is the first report on the isolation of Cryptococcus neoformans from pigeon droppings in China and their serotypes. C. neoformans colonies which produced brown colonies on caffeic acid-cornmeal agar were found in Twenty-five out of thirty-six samples of pigeon droppings. Fifty-one colonies randomly picked from the positive samples were identified as C. neoformans by a commercially available kit for carbon source assimilation test and Christensen's urea agar. Forty (78%) out of the 51 strains were serotyped as A and 11 (22%) as AD. At the same time, seventeen out of nineteen clinical isolates were serotyped as A and 2 as B. There are three findings in our results. One is that only C. neoformans var. neoformans strains could be isolated from pigeon droppings, although the variety gattii strains were found in the clinical isolates obtained in the same geographic site in China. The second is that serotype A strains were most frequently seen in natural and clinical materials in the southeast part of China, and serotype AD strains were isolated in pigeon droppings but not in clinical materials. The third is that the coexistence of serotype A and AD cells of C. neoformans strains in same samples of pigeon droppings were observed.

Clonal dissemination of multilocus sequence type ST15 KPC-2-producing Klebsiella pneumoniae in Bulgaria.

PubMed

Markovska, Rumyana; Stoeva, Temenuga; Schneider, Ines; Boyanova, Lyudmila; Popova, Valentina; Dacheva, Daniela; Kaneva, Radka; Bauernfeind, Adolf; Mitev, Vanyo; Mitov, Ivan

2015-10-01

A total of 36 consecutive clinical and two fecal-screening carbapenem-resistant Klebsiella pneumoniae isolates from two Bulgarian university hospitals (Varna and Pleven) were investigated. Susceptibility testing, conjugation experiments, and plasmid replicon typing were carried out. Beta-lactamases were characterized by isoelectric focusing, PCR, and sequencing. Clonal relatedness was investigated by RAPD and multilocus sequence typing (MLST). Most of the isolates demonstrated multidrug resistance profile. Amikacin and tigecycline retained good activity with susceptibility rates of 95 and 87%, respectively. The resistance rate to colistin was 63%. Six RAPD- and MLST-types were identified: the dominating MLST-type was ST15 (27 isolates), followed by ST76 (six isolates), and ST1350 (two isolates). ST101, ST258, and ST151 were detected once. All except one of the K. pneumoniae produced KPC-2, mostly in combination with CTX-M-15, while for one isolate (ST101) the enzymes OXA-48 and CTX-M-14 were found. All KPC-2-producing transconjugants revealed the presence of IncFII plasmid. The OXA-48- and CTX-M-14-producing isolate showed the presence of L/M replicon type. The dissemination of KPC-2-producing K.pneumoniae in Bulgaria is mainly due to the sustained spread of successful ST15 clone and to a lesser extent of ST76 clone. This is the first report of OXA-48 producing ST101 K. pneumoniae in Bulgaria. © 2015 APMIS. Published by John Wiley & Sons Ltd.

In Vitro Efficacy of Nonantibiotic Treatments on Biofilm Disruption of Gram-Negative Pathogens and an In Vivo Model of Infectious Endometritis Utilizing Isolates from the Equine Uterus

PubMed Central

McCue, Patrick M.; Borlee, Grace I.; Loncar, Kristen D.; Hennet, Margo L.

2015-01-01

In this study, we evaluated the ability of the equine clinical treatments N-acetylcysteine, EDTA, and hydrogen peroxide to disrupt in vitro biofilms and kill equine reproductive pathogens (Escherichia coli, Pseudomonas aeruginosa, or Klebsiella pneumoniae) isolated from clinical cases. N-acetylcysteine (3.3%) decreased biofilm biomass and killed bacteria within the biofilms of E. coli isolates. The CFU of recoverable P. aeruginosa and K. pneumoniae isolates were decreased, but the biofilm biomass was unchanged. Exposure to hydrogen peroxide (1%) decreased the biofilm biomass and reduced the CFU of E. coli isolates, K. pneumoniae isolates were observed to have a reduction in CFU, and minimal effects were observed for P. aeruginosa isolates. Chelating agents (EDTA formulations) reduced E. coli CFU but were ineffective at disrupting preformed biofilms or decreasing the CFU of P. aeruginosa and K. pneumoniae within a biofilm. No single nonantibiotic treatment commonly used in equine veterinary practice was able to reduce the CFU and biofilm biomass of all three Gram-negative species of bacteria evaluated. An in vivo equine model of infectious endometritis was also developed to monitor biofilm formation, utilizing bioluminescence imaging with equine P. aeruginosa isolates from this study. Following infection, the endometrial surface contained focal areas of bacterial growth encased in a strongly adherent “biofilm-like” matrix, suggesting that biofilms are present during clinical cases of infectious equine endometritis. Our results indicate that Gram-negative bacteria isolated from the equine uterus are capable of producing a biofilm in vitro, and P. aeruginosa is capable of producing biofilm-like material in vivo. PMID:26719448

In Vitro Efficacy of Nonantibiotic Treatments on Biofilm Disruption of Gram-Negative Pathogens and an In Vivo Model of Infectious Endometritis Utilizing Isolates from the Equine Uterus.

PubMed

Ferris, Ryan A; McCue, Patrick M; Borlee, Grace I; Loncar, Kristen D; Hennet, Margo L; Borlee, Bradley R

2016-03-01

In this study, we evaluated the ability of the equine clinical treatments N-acetylcysteine, EDTA, and hydrogen peroxide to disrupt in vitro biofilms and kill equine reproductive pathogens (Escherichia coli, Pseudomonas aeruginosa, or Klebsiella pneumoniae) isolated from clinical cases. N-acetylcysteine (3.3%) decreased biofilm biomass and killed bacteria within the biofilms of E. coli isolates. The CFU of recoverable P. aeruginosa and K. pneumoniae isolates were decreased, but the biofilm biomass was unchanged. Exposure to hydrogen peroxide (1%) decreased the biofilm biomass and reduced the CFU of E. coli isolates, K. pneumoniae isolates were observed to have a reduction in CFU, and minimal effects were observed for P. aeruginosa isolates. Chelating agents (EDTA formulations) reduced E. coli CFU but were ineffective at disrupting preformed biofilms or decreasing the CFU of P. aeruginosa and K. pneumoniae within a biofilm. No single nonantibiotic treatment commonly used in equine veterinary practice was able to reduce the CFU and biofilm biomass of all three Gram-negative species of bacteria evaluated. An in vivo equine model of infectious endometritis was also developed to monitor biofilm formation, utilizing bioluminescence imaging with equine P. aeruginosa isolates from this study. Following infection, the endometrial surface contained focal areas of bacterial growth encased in a strongly adherent "biofilm-like" matrix, suggesting that biofilms are present during clinical cases of infectious equine endometritis. Our results indicate that Gram-negative bacteria isolated from the equine uterus are capable of producing a biofilm in vitro, and P. aeruginosa is capable of producing biofilm-like material in vivo. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

Carbapenemase-Producing Klebsiella pneumoniae in Romania: A Six-Month Survey

PubMed Central

Straut, Monica; Usein, Codruta Romanita; Cristea, Dana; Ciontea, Simona; Codita, Irina; Rafila, Alexandru; Nica, Maria; Buzea, Mariana; Baicus, Anda; Ghita, Mihaela Camelia; Nistor, Irina; Tuchiluş, Cristina; Indreas, Marina; Antohe, Felicia; Glasner, Corinna; Grundmann, Hajo; Jasir, Aftab; Damian, Maria

2015-01-01

This study presents the first characterization of carbapenem-non-susceptible Klebsiella pneumoniae isolates by means of a structured six-month survey performed in Romania as part of an Europe-wide investigation. Klebsiella pneumoniae clinical isolates from different anatomical sites were tested for antibiotic susceptibility by phenotypic methods and confirmed by PCR for the presence of four carbapenemase genes. Genome macrorestriction fingerprinting with XbaI was used to analyze the relatedness of carbapenemase-producing Klebsiella pneumoniae isolates collected from eight hospitals. Among 75 non-susceptible isolates, 65 were carbapenemase producers. The most frequently identified genotype was OXA-48 (n = 51 isolates), eight isolates were positive for bla NDM-1 gene, four had the bla KPC-2 gene, whereas two were positive for bla VIM-1. The analysis of PFGE profiles of OXA-48 and NDM-1 producing K. pneumoniae suggests inter-hospitals and regional transmission of epidemic clones. This study presents the first description of K. pneumoniae strains harbouring bla KPC-2 and bla VIM-1 genes in Romania. The results of this study highlight the urgent need for the strengthening of hospital infection control measures in Romania in order to curb the further spread of the antibiotic resistance. PMID:26599338

Carbapenemase-Producing Klebsiella pneumoniae in Romania: A Six-Month Survey.

PubMed

Lixandru, Brandusa Elena; Cotar, Ani Ioana; Straut, Monica; Usein, Codruta Romanita; Cristea, Dana; Ciontea, Simona; Tatu-Chitoiu, Dorina; Codita, Irina; Rafila, Alexandru; Nica, Maria; Buzea, Mariana; Baicus, Anda; Ghita, Mihaela Camelia; Nistor, Irina; Tuchiluş, Cristina; Indreas, Marina; Antohe, Felicia; Glasner, Corinna; Grundmann, Hajo; Jasir, Aftab; Damian, Maria

2015-01-01

This study presents the first characterization of carbapenem-non-susceptible Klebsiella pneumoniae isolates by means of a structured six-month survey performed in Romania as part of an Europe-wide investigation. Klebsiella pneumoniae clinical isolates from different anatomical sites were tested for antibiotic susceptibility by phenotypic methods and confirmed by PCR for the presence of four carbapenemase genes. Genome macrorestriction fingerprinting with XbaI was used to analyze the relatedness of carbapenemase-producing Klebsiella pneumoniae isolates collected from eight hospitals. Among 75 non-susceptible isolates, 65 were carbapenemase producers. The most frequently identified genotype was OXA-48 (n = 51 isolates), eight isolates were positive for blaNDM-1 gene, four had the blaKPC-2 gene, whereas two were positive for blaVIM-1. The analysis of PFGE profiles of OXA-48 and NDM-1 producing K. pneumoniae suggests inter-hospitals and regional transmission of epidemic clones. This study presents the first description of K. pneumoniae strains harbouring blaKPC-2 and blaVIM-1 genes in Romania. The results of this study highlight the urgent need for the strengthening of hospital infection control measures in Romania in order to curb the further spread of the antibiotic resistance.

Bacterial and fungal endophthalmitis in upper Egypt: related species and risk factors.

PubMed

Gharamah, A A; Moharram, A M; Ismail, M A; Al-Hussaini, A K

2012-08-01

To study risk factors, contributing factors of bacterial and fungal endophthalmitis in Upper Egypt, test the isolated species sensitive to some therapeutic agents, and to investigate the air-borne bacteria and fungi in opthalmology operating rooms. Thirty one cases of endophthalmitis were clinically diagnosed and microbiologically studied. Indoor air-borne bacteria and fungi inside four air-conditioned operating rooms in the Ophthalmology Department at Assiut University Hospitals were also investigated. The isolated microbes from endophthalmitis cases were tested for their ability to produce some extracellular enzymes including protease, lipase, urease, phosphatase and catalase. Also the ability of 5 fungal isolates from endophthalmitis origin to produce mycotoxins and their sensitivity to some therapeutic agents were studied. Results showed that bacteria and fungi were responsihle for infection in 10 and 6 cases of endophthalmitis, respectively and only 2 cases produced a mixture of bacteria and fungi. Trauma was the most prevalent risk factor of endophthalmitis where 58.1% of the 31 cases were due to trauma. In ophthalmology operating rooms, different bacterial and fungal species were isolated. 8 bacterial and 5 fungal isolates showed their ability to produce enzymes while only 3 fungal isolates were able to produce mycotoxins. Terbinafine showed the highest effect against most isolates in vitro. The ability of bacterial and fungal isolates to produce extracellular enzymes and mycotoxins may be aid in the invasion and destruction of eye tissues. Microbial contamination of operating rooms with air-borne bacteria and fungi in the present work may be a source of postoperative endophthalmitis.

Dissemination of IMP-6-producing Pseudomonas aeruginosa ST244 in multiple cities in China.

PubMed

Chen, Y; Sun, M; Wang, M; Lu, Y; Yan, Z

2014-07-01

Pseudomonas aeruginosa is an important opportunistic pathogen responsible for nosocomial infections and is currently reported to be a worldwide nosocomial menace. The aim of this study was to investigate the epidemiological traits and the distribution of metallo-β-lactamases (MBLs)-producing P. aeruginosa clinical isolates in ten cities in China between January 2010 and May 2012. Antimicrobial susceptibility was determined by disc diffusion assay and the minimum inhibitory concentrations (MICs) of imipenem and meropenem were also determined by the Etest according to Clinical and Laboratory Standards Institute (CLSI) guidelines. In addition, polymerase chain reaction (PCR) and DNA sequencing were applied to detect bla MBL genes, and their epidemiological relationships were investigated by multilocus sequence typing (MLST). Of 368 P. aeruginosa isolates, MLST analysis identified 138 sequence types (STs), including 122 known and 16 novel STs, and the most frequently detected clone was ST244, followed by ST235. Besides, our study revealed that 25 isolates carried the bla IMP-6 gene and three isolates carried the bla VIM-2 gene, and a probe specific for both genes could be hybridised to an ~1,125-kb fragment in all isolates. Interestingly, all of the bla IMP-6-producing isolates shared an identical ST, ST244, and exhibited a higher level of resistance to several antibiotics. Overall, these observations suggest that P. aeruginosa ST244 carrying the chromosomally located bla IMP-6 gene is widely disseminated in multiple cites in China.

Molecular epidemiology of infant botulism in California and elsewhere, 1976-2010.

PubMed

Dabritz, Haydee A; Hill, Karen K; Barash, Jason R; Ticknor, Lawrence O; Helma, Charles H; Dover, Nir; Payne, Jessica R; Arnon, Stephen S

2014-12-01

Infant botulism (IB), first identified in California in 1976, results from Clostridium botulinum spores that germinate, multiply, and produce botulinum neurotoxin (BoNT) in the immature intestine. From 1976 to 2010 we created an archive of 1090 BoNT-producing isolates consisting of 1012 IB patient (10 outpatient, 985 hospitalized, 17 sudden death), 25 food, 18 dust/soils, and 35 other strains. The mouse neutralization assay determined isolate toxin type (56% BoNT/A, 32% BoNT/B). Amplified fragment-length polymorphism (AFLP) analysis of the isolates was combined with epidemiologic information. The AFLP dendrogram, the largest to date, contained 154 clades; 52% of isolates clustered in just 2 clades, 1 BoNT/A (n=418) and 1 BoNT/B (n=145). These clades constituted an endemic C. botulinum population that produced the entire clinical spectrum of IB. Isolates from the patient's home environment (dust/soil, honey) usually located to the same AFLP clade as the patient's isolate, thereby identifying the likely source of infective spores. C. botulinum A(B) strains were identified in California for the first time. Combining molecular methods and epidemiological data created an effective tool that yielded novel insights into the genetic diversity of C. botulinum and the clinical spectrum, occurrence, and distribution of IB in California. © The Author 2014. Published by Oxford University Press on behalf of the Infectious Diseases Society of America. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Plasmid-Mediated Resistance to Expanded-Spectrum Cephalosporins among Enterobacter aerogenes Strains

PubMed Central

Pitout, Johann D. D.; Thomson, Kenneth S.; Hanson, Nancy D.; Ehrhardt, Anton F.; Coudron, Philip; Sanders, Christine C.

1998-01-01

Resistance to expanded-spectrum cephalosporins commonly develops in Enterobacter aerogenes during therapy due to selection of mutants producing high levels of the chromosomal Bush group 1 β-lactamase. Recently, resistant strains producing plasmid-mediated extended-spectrum β-lactamases (ESBLs) have been isolated as well. A study was designed to investigate ESBL production among 31 clinical isolates of E. aerogenes from Richmond, Va., with decreased susceptibility to expanded-spectrum cephalosporins and a positive double-disk potentiation test. Antibiotic susceptibility was determined by standard disk diffusion and agar dilution procedures. β-Lactamases were investigated by an isoelectric focusing overlay technique which simultaneously determined isoelectric points (pIs) and substrate or inhibitor profiles. Decreased susceptibility to cefotaxime, ceftazidime, and aztreonam (MIC range, 1 to 64 μg/ml) was detected and associated with resistance to gentamicin and trimethoprim-sulfamethoxazole. All strains produced an inducible Bush group 1 β-lactamase (pI 8.3). Twenty-nine of the 31 isolates also produced an enzyme similar to SHV-4 (pI 7.8), while 1 isolate each produced an enzyme similar to SHV-3 (pI 6.9) and to SHV-5 (pI 8.2). The three different SHV-derived ESBLs were transferred by transconjugation to Escherichia coli C600N and amplified by PCR. Plasmid profiles of the clinical isolates showed a variety of different large plasmids. Because of the linkage of resistance to aminoglycosides and trimethoprim-sulfamethoxazole with ESBL production, it is possible that the usage of these drugs was responsible for selecting plasmid-mediated resistance to extended-spectrum cephalosporins in E. aerogenes. Furthermore, it is important that strains such as these be recognized, because they can be responsible for institutional spread of resistance genes. PMID:9517938

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.

In vitro activities and detection performances of cefmetazole and flomoxef for extended-spectrum β-lactamase and plasmid-mediated AmpC β-lactamase-producing Enterobacteriaceae.

PubMed

Matsumura, Yasufumi; Yamamoto, Masaki; Nagao, Miki; Tanaka, Michio; Takakura, Shunji; Ichiyama, Satoshi

2016-04-01

To investigate the in vitro activities of cephamycins (cefmetazole and flomoxef) for extended-spectrum β-lactamase (ESBL)- and plasmid-mediated AmpC β-lactamase (pAmpC)-producing Enterobacteriaceae, a total of 574 third-generation cephalosporin-resistant clinical isolates were collected at a Japanese multicenter study. PCR and sequencing identified 394 isolates with only ESBL genes, 63 isolates with only pAmpC genes, and 6 isolates with both ESBL and pAmpC genes. blaCTX-M types predominated 95.5% of the ESBL genes, and blaCMY-2 predominated 91.3% of the pAmpC genes. The MIC50/90 values of cefmetazole and flomoxef were ≤ 1/4 and ≤ 1/≤ 1 μg/mL for isolates with only ESBL genes, respectively, and 16/>16 and 8/16 μg/mL for isolates with only pAmpC genes, respectively. Flomoxef ≥ 4 μg/mL had the best screening performance for the detection of isolates with pAmpC genes. Flomoxef had better in vitro activities against ESBL-producing Enterobacteriaceae and provided a clearer distinction between ESBL and pAmpC-producing Enterobacteriaceae compared to cefmetazole. Copyright © 2016 Elsevier Inc. All rights reserved.

Artemisinin-resistant Plasmodium falciparum clinical isolates can infect diverse mosquito vectors of Southeast Asia and Africa

PubMed Central

St. Laurent, Brandyce; Miller, Becky; Burton, Timothy A.; Amaratunga, Chanaki; Men, Sary; Sovannaroth, Siv; Fay, Michael P.; Miotto, Olivo; Gwadz, Robert W.; Anderson, Jennifer M.; Fairhurst, Rick M.

2015-01-01

Artemisinin-resistant Plasmodium falciparum parasites are rapidly spreading in Southeast Asia, yet nothing is known about their transmission. This knowledge gap and the possibility that these parasites will spread to Africa endanger global efforts to eliminate malaria. Here we produce gametocytes from parasite clinical isolates that displayed artemisinin resistance in patients and in vitro, and use them to infect native and non-native mosquito vectors. We show that contemporary artemisinin-resistant isolates from Cambodia develop and produce sporozoites in two Southeast Asian vectors, Anopheles dirus and Anopheles minimus, and the major African vector, Anopheles coluzzii (formerly Anopheles gambiae M). The ability of artemisinin-resistant parasites to infect such highly diverse Anopheles species, combined with their higher gametocyte prevalence in patients, may explain the rapid expansion of these parasites in Cambodia and neighbouring countries, and further compromise efforts to prevent their global spread. PMID:26485448

Artemisinin-resistant Plasmodium falciparum clinical isolates can infect diverse mosquito vectors of Southeast Asia and Africa.

PubMed

St Laurent, Brandyce; Miller, Becky; Burton, Timothy A; Amaratunga, Chanaki; Men, Sary; Sovannaroth, Siv; Fay, Michael P; Miotto, Olivo; Gwadz, Robert W; Anderson, Jennifer M; Fairhurst, Rick M

2015-10-20

Artemisinin-resistant Plasmodium falciparum parasites are rapidly spreading in Southeast Asia, yet nothing is known about their transmission. This knowledge gap and the possibility that these parasites will spread to Africa endanger global efforts to eliminate malaria. Here we produce gametocytes from parasite clinical isolates that displayed artemisinin resistance in patients and in vitro, and use them to infect native and non-native mosquito vectors. We show that contemporary artemisinin-resistant isolates from Cambodia develop and produce sporozoites in two Southeast Asian vectors, Anopheles dirus and Anopheles minimus, and the major African vector, Anopheles coluzzii (formerly Anopheles gambiae M). The ability of artemisinin-resistant parasites to infect such highly diverse Anopheles species, combined with their higher gametocyte prevalence in patients, may explain the rapid expansion of these parasites in Cambodia and neighbouring countries, and further compromise efforts to prevent their global spread.

Antifungal activity of the lipopeptides produced by Bacillus amyloliquefaciens anti-CA against Candida albicans isolated from clinic.

PubMed

Song, Bo; Rong, Yan-Jun; Zhao, Ming-Xin; Chi, Zhen-Ming

2013-08-01

The bacterium Bacillus amyloliquefaciens anti-CA isolated from mangrove system was found to be able to actively kill Candida albicans isolated from clinic. The bacterial strain anti-CA could produce high level of bioactive substance, amylase and protease in the cheap medium containing 2.0 % soybean meal, 2.0 % wheat flour, pH 6.5 within 26 h. After purification, the main bioactive substance was confirmed to be a cyclic lipopeptide containing a heptapeptide, L-Asp→L-Leu→L-Leu→L-Val→L-Val→L-Glu→L-Leu and a 3-OH fatty acid (15 carbons). In addition to C. albicans, the purified lipopeptide can also kill many yeast strains including Metschnikowia bicuspidata, Candida tropicalis, Yarrowia lipolytica and Saccharomyces cerevisiae. After treated by the purified lipopeptide, both the whole cells and protoplasts of C. albicans were destroyed.

Evaluation of Etest MBL for Detection of blaIMP-1 and blaVIM-2 Allele-Positive Clinical Isolates of Pseudomonas spp. and Acinetobacter spp.

PubMed Central

Lee, Kyungwon; Yong, Dongeun; Yum, Jong Hwa; Lim, Yong Sik; Bolmström, Anne; Qwärnström, Anette; Karlsson, Åsa; Chong, Yunsop

2005-01-01

The Etest MBL (AB BIODISK, Solna, Sweden) correctly differentiated all 57 isolates of Acinetobacter spp. and Pseudomonas aeruginosa with the blaIMP-1 allele and 135 of 137 (98.5%) Acinetobacter spp. and Pseudomonas spp. isolates with the blaVIM-2 allele. The Etest MBL was reliable for detecting the IMP-1- and VIM-2-producing Pseudomonas and Acinetobacter isolates. PMID:15695713

Relationship between clinical site of isolation and ability to form biofilms in vitro in nontypeable Haemophilus influenzae.

PubMed

Obaid, Najla A; Jacobson, Glenn A; Tristram, Stephen

2015-03-01

Nontypeable Haemophilus influenzae (NTHi) is an opportunistic pathogen associated with a range of infections, including various lower respiratory infections, otitis media, and conjunctivitis. There is some debate as to whether or not NTHi produces biofilms and, if so, whether or not this is relevant to pathogenesis. Although many studies have examined the association between in vitro biofilm formation and isolates from a specific infection type, few have made comparisons from isolates from a broad range of isolates grouped by clinical source. In our study 50 NTHi from different clinical sources, otitis media, conjunctivitis, lower respiratory tract infections in both cystic fibrosis and non-cystic fibrosis patients, and nasopharyngeal carriage, plus 10 nasopharyngeal isolates of the commensal Haemophilus haemolyticus were tested for the ability to form biofilm by using a static microtitre plate crystal violet assay. A high degree of variation in biofilm forming ability was observed across all isolates, with no statistically significant differences observed between the groups, with the exception of the isolates from conjunctivitis. These isolates had uniformly lower biofilm forming ability compared with isolates from the other groups (p < 0.005).

Carbapenem-resistant Citrobacter spp. isolated in Spain from 2013 to 2015 produced a variety of carbapenemases including VIM-1, OXA-48, KPC-2, NDM-1 and VIM-2.

PubMed

Arana, David M; Ortega, Adriana; González-Barberá, Eva; Lara, Noelia; Bautista, Verónica; Gómez-Ruíz, Dolores; Sáez, David; Fernández-Romero, Sara; Aracil, Belén; Pérez-Vázquez, María; Campos, José; Oteo, Jesús

2017-12-01

There is little information about carbapenemase-producing (CP) Citrobacter spp. We studied the molecular epidemiology and microbiological features of CP Citrobacter spp. isolates collected in Spain (2013-15). In total, 119 isolates suspected of being CP by the EUCAST screening cut-off values were analysed. Carbapenemases and ESBLs were characterized using PCR and sequencing. The genetic relationship among Citrobacter freundii isolates was studied by PFGE. Of the 119 isolates, 63 (52.9%) produced carbapenemases, of which 37 (58.7%) produced VIM-1, 20 (31.7%) produced OXA-48, 12 (19%) produced KPC-2, 2 (3.2%) produced NDM-1 and 1 (1.6%) produced VIM-2; 9 C. freundii isolates co-produced VIM-1 plus OXA-48. Fourteen isolates (22.2%) also carried ESBLs: 8 CTX-M-9 plus SHV-12, 2 CTX-M-9, 2 SHV-12 and 2 CTX-M-15. Fifty-seven isolates (90.5%) were C. freundii, 4 (6.3%) were Citrobacter koseri, 1 (1.6%) was Citrobacter amalonaticus and 1 (1.6%) was Citrobacter braakii. By EUCAST breakpoints, eight (12.7%) of the CP isolates were susceptible to the four carbapenems tested. In the 53 CP C. freundii analysed by PFGE, a total of 44 different band patterns were observed. Four PFGE clusters were identified: cluster 1 included eight isolates co-producing VIM-1 and OXA-48; blaVIM-1 was carried in a class 1 integron (intI-blaVIM-1-aacA4-dfrB1-aadA1-catB2-qacEΔ1/sul1) and blaOXA-48 was carried in a Tn1999.2 transposon. We observed the clonal and polyclonal spread of CP Citrobacter spp. across several Spanish geographical areas. Four species of Citrobacter spp. produced up to five carbapenemase types, including co-production of VIM-1 plus OXA-48. Some CP Citrobacter spp. isolates were susceptible to the four carbapenems tested, a finding with potential clinical implications. © The Author 2017. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Rapid detection of carbapenemase-producing Klebsiella pneumoniae strains derived from blood cultures by Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry (MALDI-TOF MS).

PubMed

Sakarikou, Christina; Ciotti, Marco; Dolfa, Camilla; Angeletti, Silvia; Favalli, Cartesio

2017-03-08

Carbapenemase-producing Enterobacteriaceae (CPE), particularly carbapenemase-producing Klebsiella pneumoniae isolates, are important causative agents of nosocomial infections associated with significant mortality rates mostly in critical wards. The rapid detection and typing of these strains is critical either for surveillance purposes and to prevent outbreaks and optimize antibiotic therapy. In this study, the MALDI-TOF MS method was used to detect rapidly these isolates from blood cultures (BCs) and to obtain proteomic profiles enable to discriminate between carbapenemase-producing and non-carbapenemase-producing strains. Fifty-five K. pneumoniae strains were tested. Identification and carbapenemase-production detection assay using Ertapenem were performed both from bacterial pellets extracted directly from BCs flasks and from subcultures of these strains. For all isolates, a complete antimicrobial susceptibility testing and a genotypic characterization were performed. We found 100% agreement between the carbapenemase-producing profile generated by MALDI TOF MS and that obtained using conventional methods. The assay detected and discriminated different carbapenemase-producing K. pneumoniae isolates within 30 min to 3 h after incubation with Ertapenem. MALDI-TOF MS is a promising, rapid and economical method for the detection of carbapenemase-producing K. pneumoniae strains that could be successfully introduced into the routine diagnostic workflow of clinical microbiology laboratories.

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.

Serogroup, virulence, and molecular traits of Vibrio parahaemolyticus isolated from clinical and cockle sources in northeastern Thailand.

PubMed

Mala, Wanida; Alam, Munirul; Angkititrakul, Sunpetch; Wongwajana, Suwin; Lulitanond, Viraphong; Huttayananont, Sriwanna; Kaewkes, Wanlop; Faksri, Kiatichai; Chomvarin, Chariya

2016-04-01

Vibrio parahaemolyticus is responsible for seafood-borne gastroenteritis worldwide. Isolates of V. parahaemolyticus from clinical samples (n=74) and cockles (Anadara granosa) (n=74) in Thailand were analyzed by serotyping, determination of virulence and related marker genes present, response to antimicrobial agents, and genetic relatedness. Serological analysis revealed 31 different serotypes, 10 of which occurred among both clinical and cockle samples. The clinical isolates commonly included the pandemic serogroup O3:K6, while a few of the cockle isolates exhibited likely pandemic serovariants such as O3:KUT and O4:KUT, but not O3:K6. The pandemic (orf8 gene-positive) strains were more frequently found among clinical isolates (78.4%) than cockle isolates (28.4%) (p<0.001). Likewise, the virulence and related marker genes were more commonly detected among clinical than cockle isolates; i.e., tdh gene (93.2% versus 29.7%), vcrD2 (97.3% versus 23.0%), vopB2 (89.2% versus 13.5%), vopT (98.6% versus 36.5%) (all p<0.001) and trh (10.8% versus 1.4%) (p<0.05). Pulsed-field gel electrophoresis of NotI-digested genomic DNA of 41 randomly selected V. parahaemolyticus isolates representing different serotypes produced 33 pulsotypes that formed 5 different clusters (clonal complexes) (A-E) in a dendrogram. Vibrio parahaemolyticus O3:K6 and likely related pandemic serotypes were especially common among the numerous clinical isolates in cluster C, suggesting a close clonal link among many of these isolates. Most clinical and cockle isolates were resistant to ampicillin. This study indicates that O3:K6 and its likely serovariants based on the PFGE clusters, are causative agents. Seafoods such as cockles potentially serve as a source of virulent V. parahaemolyticus, but further work is required to identify possible additional sources. Copyright © 2016. Published by Elsevier B.V.

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

Clinical and molecular epidemiology of hospital Enterococcus faecalis isolates in eastern France.

PubMed

Mulin, Blandine; Bailly, Pascale; Thouverez, Michelle; Cailleaux, Vincent; Cornette, Christian; Dupont, Marie-Jeanne; Talon, Daniel

1999-03-01

OBJECTIVE: To report on the occurrence of Enterococcus faecalis hospital isolates obtained during 1 year in hospitals in the Franche-Comté region of France. METHODS: Clinical isolates of E. faecalis of different antibiotic susceptibility phenotypes from hospitalized patients were characterized by pulsed-field gel electrophoresis. Patients with positive cultures were investigated by three case-control studies to identify risk factors for colonization/infection. RESULTS: The crude incidence of colonization/infection was 2.37%, and 4-day and 7-day colonization rates after admission were 10.0% and 6.36%, respectively. The rates of high-level resistance to kanamycin (HLKR) and to gentamicin (HLGR) were 47.1% and 7.1%, respectively. No isolate was resistant to glycopeptides or produced beta-lactamase. The 209 hospital isolates obtained during the study yielded 98 major DNA patterns, of which two were major epidemic patterns including HLKR isolates. No single factor was significantly associated with colonization/infection by HLKR isolates. The length of hospitalization before isolation was associated with colonization by HLGR isolates. CONCLUSIONS: The isolation frequency of E. faecalis strains with acquired resistance to aminoglycoside antibiotics, and the wide dissemination of resistant strains with characteristics that allow them to persist and spread, argue for further large prospective surveys of clinical isolates of E. faecalis in hospitals.

AmpC β-lactamases in nosocomial isolates of Klebsiella pneumoniae from India

PubMed Central

Gupta, Varsha; Kumarasamy, Karthikeyan; Gulati, Neelam; Garg, Ritu; Krishnan, Padma; Chander, Jagdish

2012-01-01

Background & objectives: AmpC β-lactamases are clinically significant since these confer resistance to cephalosporins in the oxyimino group, 7-α methoxycephalosporins and are not affected by available β-lactamase inhibitors. In this study we looked for both extended spectrum β-lactamases (ESBL) and AmpC β-lactamases in Klebsiella pneumoniae clinical isolates. Methods: One hundred consecutive, non-duplicate clinical isolates of K. pneumoniae collected over a period of one year (June 2008 - June 2009) were included in the study. An antibiotic susceptibility method was used with 10 antibiotics for Gram-negative infections which helped in screening for ESBL and AmpC β-lactamases and also in confirmation of ESBL production. The detection of AmpC β-lactamases was done based on screening and confirmatory tests. For screening, disc diffusion zones of cefoxitin <18 mm was taken as cefoxitin resistant. All cefoxitin resistant isolates were tested further by AmpC disk test and modified three dimensional test. Multiplex-PCR was performed for screening the presence of plasmid-mediated AmpC genes. Results: Of the 100 isolates of K. pneumoniae studied, 48 were resistant to cefoxitin on screening. AmpC disk test was positive in 32 (32%) isolates. This was also confirmed with modified three dimensional test. Indentation indicating strong AmpC producer was observed in 25 isolates whereas little distortion (weak AmpC) was observed in 7 isolates. ESBL detection was confirmed by a modification of double disk synergy test in 56 isolates. Cefepime was the best cephalosporin in synergy with tazobactam for detecting ESBL production in isolates co-producing AmpC β-lactamases. The subsets of isolates phenotypically AmpC β-lactamase positive were subjected to amplification of six different families of AmpC gene using multiplex PCR. The sequence analysis revealed 12 CMY-2 and eight DHA-1 types. Interpretation & conclusions: Tazobactam was the best β-lactamase inhibitor for detecting ESBL in presence of AmpC β-lactamase as this is a very poor inducer of AmpC gene. Amongst cephalosporins, cefepime was the best cephalosporin in detecting ESBL in presence of AmpC β-lactamase as it is least hydrolyzed by AmpC enzymes. Cefepime-tazobactam combination disk test would be a simple and best method in detection of ESBLs in Enterobacteriaceae co-producing AmpC β-lactamase in the routine diagnostic microbiology laboratories. PMID:22960890

Draft genome sequence of Escherichia coli ST977: A clinical multidrug-resistant strain harbouring blaNDM-3 isolated from a bloodstream infection.

PubMed

Li, Xi; Sun, Long; Zhu, Yongze; Shen, Mengyuan; Tu, Yuexing

2018-04-14

The emergence of carbapenem-resistant Escherichia coli has become a serious challenge to manage in the clinic because of multidrug resistance. Here we report the draft genome sequence of NDM-3-producing E. coli strain NT1 isolated from a bloodstream infection in China. Whole genomic DNA of E. coli strain NT1 was extracted and was sequenced using an Illumina HiSeq™ X Ten platform. The generated sequence reads were assembled using CLC Genomics Workbench. The draft genome was annotated using Rapid Annotation using Subsystem Technology (RAST). Bioinformatics analysis was further performed. The genome size was calculated at 5,353 620bp, with 5297 protein-coding sequences and the presence of genes conferring resistance to aminoglycosides, β-lactams, quinolones, macrolides, phenicols, sulphonamides, tetracycline and trimethoprim. In addition, genes encoding virulence factors were also identified. To our knowledge, this is the first report of an E. coli strain producing NDM-3 isolated from a human bloodstream infection. The genome sequence will provide valuable information to understand antibiotic resistance mechanisms and pathogenic mechanisms in this strain. Close surveillance is urgently needed to monitor the spread of NDM-3-producing isolates. Copyright © 2018 International Society for Chemotherapy of Infection and Cancer. Published by Elsevier Ltd. All rights reserved.

A multiplex lateral flow immunoassay for the rapid identification of NDM-, KPC-, IMP- and VIM-type and OXA-48-like carbapenemase-producing Enterobacteriaceae

PubMed Central

Boutal, Hervé; Vogel, Anaïs; Bernabeu, Sandrine; Devilliers, Karine; Creton, Elodie; Cotellon, Garence; Plaisance, Marc; Oueslati, Saoussen; Dortet, Laurent; Jousset, Agnès; Simon, Stéphanie; Naas, Thierry; Volland, Hervé

2018-01-01

Abstract Objectives The global spread of carbapenemase-producing Enterobacteriaceae represents a substantial challenge in clinical practice and rapid and reliable detection of these organisms is essential. The aim of this study was to develop and validate a lateral flow immunoassay (Carba5) for the detection of the five main carbapenemases (KPC-, NDM-, VIM- and IMP-type and OXA-48-like). Methods Carba5 was retrospectively and prospectively evaluated using 296 enterobacterial isolates from agar culture. An isolated colony was suspended in extraction buffer and then loaded on the manufactured Carba5. Results All 185 isolates expressing a carbapenemase related to one of the Carba5 targets were correctly and unambiguously detected in <15 min. All other isolates gave negative results except those producing OXA-163 and OXA-405, which are considered low-activity carbapenemases. No cross-reaction was observed with non-targeted carbapenemases, ESBLs, AmpCs or oxacillinases (OXA-1, -2, -9 and -10). Overall, this assay reached 100% sensitivity and 95.3% (retrospectively) to 100% (prospectively) specificity. Conclusions Carba5 is efficient, rapid and easy to implement in the routine workflow of a clinical microbiology laboratory for confirmation of the five main carbapenemases encountered in Enterobacteriaceae. PMID:29365094

Molecular characterization and epidemiology of cefoxitin resistance among Enterobacteriaceae lacking inducible chromosomal ampC genes from hospitalized and non-hospitalized patients in Algeria: description of new sequence type in Klebsiella pneumoniae isolates.

PubMed

Gharout-Sait, Alima; Touati, Abdelaziz; Guillard, Thomas; Brasme, Lucien; de Champs, Christophe

2015-01-01

In this study, 922 consecutive non-duplicate clinical isolates of Enterobacteriaceae obtained from hospitalized and non-hospitalized patients at Bejaia, Algeria were analyzed for AmpC-type β-lactamases production. The ampC genes and their genetic environment were characterized using polymerase chain reaction (PCR) and sequencing. Plasmid incompatibility groups were determined by using PCR-based replicon typing. Phylogenetic grouping and multilocus sequence typing were determined for molecular typing of the plasmid-mediated AmpC (pAmpC) isolates. Of the isolates, 15 (1.6%) were identified as AmpC producers including 14 CMY-4-producing isolates and one DHA-1-producing Klebsiella pneumoniae. All AmpC-producing isolates co-expressed the broad-spectrum TEM-1 β-lactamase and three of them co-produced CTX-M and/or SHV-12 ESBL. Phylogenetic grouping and virulence genotyping of the E. coli isolates revealed that most of them belonged to groups D and B1. Multilocus sequence typing analysis of K. pneumoniae isolates identified four different sequence types (STs) with two new sequences: ST1617 and ST1618. Plasmid replicon typing indicates that blaCMY-4 gene was located on broad host range A/C plasmid, while LVPK replicon was associated with blaDHA-1. All isolates carrying blaCMY-4 displayed the transposon-like structures ISEcp1/ΔISEcp1-blaCMY-blc-sugE. Our study showed that CMY-4 was the main pAmpC in the Enterobacteriaceae isolates in Algeria. Copyright © 2015 Elsevier Editora Ltda. All rights reserved.

An in vitro biofilm model to examine the effect of antibiotic ointments on biofilms produced by burn wound bacterial isolates.

PubMed

Hammond, Adrienne A; Miller, Kyle G; Kruczek, Cassandra J; Dertien, Janet; Colmer-Hamood, Jane A; Griswold, John A; Horswill, Alexander R; Hamood, Abdul N

2011-03-01

Topical treatment of burn wounds is essential as reduced blood supply in the burned tissues restricts the effect of systemic antibiotics. On the burn surface, microorganisms exist within a complex structure termed a biofilm, which enhances bacterial resistance to antimicrobial agents significantly. Since bacteria differ in their ability to develop biofilms, the susceptibility of these biofilms to topically applied antibiotics varies, making it essential to identify which topical antibiotics efficiently disrupt or prevent biofilms produced by these pathogens. Yet, a simple in vitro assay to compare the susceptibility of biofilms produced by burn wound isolates to different topical antibiotics has not been reported. Biofilms were developed by inoculating cellulose disks on agar plates with burn wound isolates and incubating for 24h. The biofilms were then covered for 24h with untreated gauze or gauze coated with antibiotic ointment and remaining microorganisms were quantified and visualized microscopically. Mupirocin and triple antibiotic ointments significantly reduced biofilms produced by the Staphylococcus aureus and Pseudomonas aeruginosa burn wound isolates tested, as did gentamicin ointment, with the exception of one P. aeruginosa clinical isolate. The described assay is a practical and reproducible approach to identify topical antibiotics most effective in eliminating biofilms produced by burn wound isolates. Copyright © 2010 Elsevier Ltd and ISBI. All rights reserved.

Antimicrobial susceptibility of Salmonella isolates from healthy pigs and chickens (2008-2011).

PubMed

de Jong, Anno; Smet, Annemieke; Ludwig, Carolin; Stephan, Bernd; De Graef, Evelyne; Vanrobaeys, Mia; Haesebrouck, Freddy

2014-07-16

Using the agar dilution method, antimicrobial susceptibility to human-use antibiotics was determined among Belgian faecal Salmonella isolates from healthy pigs and broiler chickens. Both epidemiological cut-off values and clinical breakpoints were applied for interpretation of the results. Cephalosporin-resistant isolates were examined for the presence of genes encoding CTX-M, SHV, TEM and CMY β-lactamases. All isolates with decreased quinolone susceptibility were screened for plasmid-borne genes qnr, qepA and aac(6')-Ib-cr. In all, 368 Salmonella isolates were recovered from pigs and 452 from chickens. Clinical resistance to ciprofloxacin was absent in isolates of both host species, and was 1.9 and 13.1% to cefotaxime in pig and poultry isolates, respectively. Decreased susceptibility to cefotaxime amounted to 2.2 and 0.7%, whereas for ciprofloxacin this was 3.0 and 23.0% in pig and poultry isolates, respectively. Ciprofloxacin decreased susceptibility was limited to few serovars, mainly Paratyphi B. Multidrug resistance was markedly higher for pig isolates (39.7%) than for chicken isolates (17.3%). Sixty-six cefotaxime-resistant isolates, 59 from chickens and 7 from pigs, were phenotypically determined as ESBL/AmpC producers; predominantly Paratyphi B and Typhimurium serovars. BlaCTX-M (mostly blaCTXM-1, but also blaCTXM-2 and blaCTXM-9) and blaTEM-52 were the predominant ESBL genes. Only few isolates expressed SHV-12 or an AmpC enzyme (CMY-2). Isolates of four serovars carried qnr genes: Brandenburg and Llandof from pigs, both qnrS; Indiana and Paratyphi B from chickens with qnrB and qnrA. The latter isolate carried blaCTX-M-9 and was the only strain with a plasmid-borne quinolone resistance gene among the ESBL/AmpC producers. This Salmonella survey confirms that the ESBL/AmpC producers are particularly prevalent in chickens (12.8%), and much less in pigs (1.9%). A link between plasmid-borne quinolone resistance genes and ESBLs/AmpC was uncommon. Copyright © 2014 Elsevier B.V. All rights reserved.

Multilocus variable-number tandem-repeat analysis of clinical isolates of Aspergillus flavus from Iran reveals the first cases of Aspergillus minisclerotigenes associated with human infection

PubMed Central

2014-01-01

Background Aspergillus flavus is intensively studied for its role in infecting crop plants and contaminating produce with aflatoxin, but its role as a human pathogen is less well understood. In parts of the Middle East and India, A. flavus surpasses A. fumigatus as a cause of invasive aspergillosis and is a significant cause of cutaneous, sinus, nasal and nail infections. Methods A collection of 45 clinical and 10 environmental A. flavus isolates from Iran were analysed using Variable-Number Tandem-Repeat (VNTR) markers with MICROSAT and goeBURST to determine their genetic diversity and their relatedness to clinical and environmental A. flavus isolates from Australia. Phylogeny was assessed using partial β-tubulin and calmodulin gene sequencing, and mating type was determined by PCR. Antifungal susceptibility testing was performed on selected isolates using a reference microbroth dilution method. Results There was considerable diversity in the A. flavus collection, with no segregation on goeBURST networks according to source or geographic location. Three Iranian isolates, two from sinus infections and one from a paranasal infection grouped with Aspergillus minisclerotigenes, and all produced B and G aflatoxin. Phylogenic analysis using partial β-tubulin and calmodulin sequencing confirmed two of these as A. minisclerotigenes, while the third could not be differentiated from A. flavus and related species within Aspergillus section flavi. Based on epidemiological cut-off values, the A. minisclerotigens and A. flavus isolates tested were susceptible to commonly used antifungal drugs. Conclusions This is the first report of human infection due to A. minisclerotigenes, and it raises the possiblity that other species within Aspergillus section flavi may also cause clinical disease. Clinical isolates of A. flavus from Iran are not distinct from Australian isolates, indicating local environmental, climatic or host features, rather than fungal features, govern the high incidence of A. flavus infection in this region. The results of this study have important implications for biological control strategies that aim to reduce aflatoxin by the introduction of non-toxigenic strains, as potentially any strain of A. flavus, and closely related species like A. minisclerotigenes, might be capable of human infection. PMID:24986045

Performance evaluation of three automated identification systems in detecting carbapenem-resistant Enterobacteriaceae.

PubMed

He, Qingwen; Chen, Weiyuan; Huang, Liya; Lin, Qili; Zhang, Jingling; Liu, Rui; Li, Bin

2016-06-21

Carbapenem-resistant Enterobacteriaceae (CRE) is prevalent around the world. Rapid and accurate detection of CRE is urgently needed to provide effective treatment. Automated identification systems have been widely used in clinical microbiology laboratories for rapid and high-efficient identification of pathogenic bacteria. However, critical evaluation and comparison are needed to determine the specificity and accuracy of different systems. The aim of this study was to evaluate the performance of three commonly used automated identification systems on the detection of CRE. A total of 81 non-repetitive clinical CRE isolates were collected from August 2011 to August 2012 in a Chinese university hospital, and all the isolates were confirmed to be resistant to carbapenems by the agar dilution method. The potential presence of carbapenemase genotypes of the 81 isolates was detected by PCR and sequencing. Using 81 clinical CRE isolates, we evaluated and compared the performance of three automated identification systems, MicroScan WalkAway 96 Plus, Phoenix 100, and Vitek 2 Compact, which are commonly used in China. To identify CRE, the comparator methodology was agar dilution method, while the PCR and sequencing was the comparator one to identify CPE. PCR and sequencing analysis showed that 48 of the 81 CRE isolates carried carbapenemase genes, including 23 (28.4 %) IMP-4, 14 (17.3 %) IMP-8, 5 (6.2 %) NDM-1, and 8 (9.9 %) KPC-2. Notably, one Klebsiella pneumoniae isolate produced both IMP-4 and NDM-1. One Klebsiella oxytoca isolate produced both KPC-2 and IMP-8. Of the 81 clinical CRE isolates, 56 (69.1 %), 33 (40.7 %) and 77 (95.1 %) were identified as CRE by MicroScan WalkAway 96 Plus, Phoenix 100, and Vitek 2 Compact, respectively. The sensitivities/specificities of MicroScan WalkAway, Phoenix 100 and Vitek 2 were 93.8/42.4 %, 54.2/66.7 %, and 75.0/36.4 %, respectively. The MicroScan WalkAway and Viteck2 systems are more reliable in clinical identification of CRE, whereas additional tests are required for the Pheonix 100 system. Our study provides a useful guideline for using automated identification systems for CRE identification.

Comparable High Rates of Extended-Spectrum-Beta-Lactamase-Producing Escherichia coli in Birds of Prey from Germany and Mongolia

PubMed Central

Guenther, Sebastian; Aschenbrenner, Katja; Stamm, Ivonne; Bethe, Astrid; Semmler, Torsten; Stubbe, Annegret; Stubbe, Michael; Batsajkhan, Nyamsuren; Glupczynski, Youri; Wieler, Lothar H.; Ewers, Christa

2012-01-01

Frequent contact with human waste and liquid manure from intensive livestock breeding, and the increased loads of antibiotic-resistant bacteria that result, are believed to be responsible for the high carriage rates of ESBL-producing E. coli found in birds of prey (raptors) in Central Europe. To test this hypothesis against the influence of avian migration, we initiated a comparative analysis of faecal samples from wild birds found in Saxony-Anhalt in Germany and the Gobi-Desert in Mongolia, regions of dissimilar human and livestock population characteristics and agricultural practices. We sampled a total of 281 wild birds, mostly raptors with primarily north-to-south migration routes. We determined antimicrobial resistance, focusing on ESBL production, and unravelled the phylogenetic and clonal relatedness of identified ESBL-producing E. coli isolates using multi-locus sequence typing (MLST) and macrorestriction analyses. Surprisingly, the overall carriage rates (approximately 5%) and the proportion of ESBL-producers among E. coli (Germany: 13.8%, Mongolia: 10.8%) were similar in both regions. Whereas bla CTX-M-1 predominated among German isolates (100%), bla CTX-M-9 was the most prevalent in Mongolian isolates (75%). We identified sequence types (STs) that are well known in human and veterinary clinical ESBL-producing E. coli (ST12, ST117, ST167, ST648) and observed clonal relatedness between a Mongolian avian ESBL-E. coli (ST167) and a clinical isolate of the same ST that originated in a hospitalised patient in Europe. Our data suggest the influence of avian migratory species in the transmission of ESBL-producing E. coli and challenge the prevailing assumption that reducing human influence alone invariably leads to lower rates of antimicrobial resistance. PMID:23300857

Comparable high rates of extended-spectrum-beta-lactamase-producing Escherichia coli in birds of prey from Germany and Mongolia.

PubMed

Guenther, Sebastian; Aschenbrenner, Katja; Stamm, Ivonne; Bethe, Astrid; Semmler, Torsten; Stubbe, Annegret; Stubbe, Michael; Batsajkhan, Nyamsuren; Glupczynski, Youri; Wieler, Lothar H; Ewers, Christa

2012-01-01

Frequent contact with human waste and liquid manure from intensive livestock breeding, and the increased loads of antibiotic-resistant bacteria that result, are believed to be responsible for the high carriage rates of ESBL-producing E. coli found in birds of prey (raptors) in Central Europe. To test this hypothesis against the influence of avian migration, we initiated a comparative analysis of faecal samples from wild birds found in Saxony-Anhalt in Germany and the Gobi-Desert in Mongolia, regions of dissimilar human and livestock population characteristics and agricultural practices. We sampled a total of 281 wild birds, mostly raptors with primarily north-to-south migration routes. We determined antimicrobial resistance, focusing on ESBL production, and unravelled the phylogenetic and clonal relatedness of identified ESBL-producing E. coli isolates using multi-locus sequence typing (MLST) and macrorestriction analyses. Surprisingly, the overall carriage rates (approximately 5%) and the proportion of ESBL-producers among E. coli (Germany: 13.8%, Mongolia: 10.8%) were similar in both regions. Whereas bla(CTX-M-1) predominated among German isolates (100%), bla(CTX-M-9) was the most prevalent in Mongolian isolates (75%). We identified sequence types (STs) that are well known in human and veterinary clinical ESBL-producing E. coli (ST12, ST117, ST167, ST648) and observed clonal relatedness between a Mongolian avian ESBL-E. coli (ST167) and a clinical isolate of the same ST that originated in a hospitalised patient in Europe. Our data suggest the influence of avian migratory species in the transmission of ESBL-producing E. coli and challenge the prevailing assumption that reducing human influence alone invariably leads to lower rates of antimicrobial resistance.

Antimicrobial susceptibility and mechanisms of fosfomycin resistance in extended-spectrum β-lactamase-producing Escherichia coli strains from urinary tract infections in Wenzhou, China.

PubMed

Bi, Wenzi; Li, Bin; Song, Jiangning; Hong, Youliang; Zhang, Xiaoxiao; Liu, Haiyang; Lu, Hong; Zhou, Tieli; Cao, Jianming

2017-07-01

Fosfomycin in combination with various antibiotics represents an excellent clinically efficacious regimen for the treatment of urinary tract infections (UTIs) caused by extended-spectrum β-lactamase (ESBL)-producing Escherichia coli. Underlying mechanisms of fosfomycin resistance remain largely uncharacterised. To investigate the antibacterial efficacy of fosfomycin against ESBL-producing E. coli, 356 non-repetitive ESBL-producing E. coli clinical isolates were collected from urine specimens from patients with UTI in Wenzhou, China, from January 2011 to December 2015. Antimicrobial sensitivity testing indicated that 6.7% (24/356) of the ESBL-producing E. coli strains were resistant to fosfomycin. The fosA3 gene encoding a fosfomycin-modifying enzyme was detected in 20 isolates by PCR and sequencing, alone or in combination with other ESBL determinants. Conjugation experiments and Southern blotting demonstrated that 70% (14/20) of the fosA3-positive isolates possessed transferable plasmids (ca. 54.2 kb) co-harbouring the ESBL resistance gene bla CTX-M and the fosfomycin resistance gene fosA3. Among the four fosfomycin-resistant fosA3-negative E. coli isolates, three contained amino acid substitutions (Ile28Asn and Phe30Leu in MurA and Leu297Phe in GlpT). The results indicate that presence of the fosA3 gene is the primary mechanism of fosfomycin resistance in ESBL-producing E. coli isolates in Wenzhou, China. In addition, a plasmid (ca. 54.2 kb) co-harbouring fosA3 and bla CTX-M genes is horizontally transferable. Furthermore, a low degree of homology in the fosfomycin-resistant E. coli was confirmed using multilocus sequence typing (MLST), suggesting that there is no obvious phenomenon of clonal dissemination. Copyright © 2017 Elsevier B.V. and International Society of Chemotherapy. All rights reserved.

Carbapenems and piperacillin/tazobactam for the treatment of bacteremia caused by extended-spectrum β-lactamase-producing Proteus mirabilis.

PubMed

Tsai, Hsih-Yeh; Chen, Yen-Hsu; Tang, Hung-Jen; Huang, Chi-Chang; Liao, Chun-Hsing; Chu, Fang-Yeh; Chuang, Yin-Ching; Sheng, Wang-Huei; Ko, Wen-Chien; Hsueh, Po-Ren

2014-11-01

This study was intended to delineate the role of carbapenems and piperacillin/tazobactam in treating bacteremia caused by extended-spectrum β-lactamase (ESBL)-producing Proteus mirabilis. We performed a multicenter and retrospective study of the patients with ESBL-producing P. mirabilis bacteremia. The outcomes of the patients treated by piperacillin/tazobactam or a carbapenem for at least 48 hours and the MICs of the prescribed drugs for these isolates were analyzed. Forty-seven patients with available clinical data were included. The overall 30-day mortality rate was 29.8%. All available isolates (n = 44) were susceptible to ertapenem, meropenem, and doripenem, and 95.6% were susceptible to piperacillin/tazobactam; however, only 11.4% of the isolates were susceptible to imipenem. Among the 3 patients infected with isolates exhibiting non-susceptibility to imipenem (MIC ≥2 mg/L) who were treated with imipenem, none died within 28 days. The 30-day (14.3% versus 23.1%, P = 0.65) or in-hospital (19.1% versus 30.8%, P = 0.68) mortality rate of 21 patients treated by a carbapenem was lower than that of 13 treated by piperacillin/tazobactam. However, among those treated by piperacillin/tazobactam, the mortality rate of those infected by the isolates with lower piperacillin/tazobactam MICs (≤0.5/4 mg/L) was lower than that of the isolates with MICs of ≥1/4 mg/L (0%, 0/7 versus 60%, 3/5; P = 0.045). ESBL-producing P. mirabilis bacteremia is associated with significant mortality, and carbapenem therapy could be regarded as the drugs of choice. The role of piperacillin/tazobactam, especially for the infections due to the isolates with an MIC ≤0.5/4 mg/L, warrants more clinical studies. Copyright © 2014 Elsevier Inc. All rights reserved.

Characterisation of non-autoinducing tropodithietic Acid (TDA) production from marine sponge Pseudovibrio species.

PubMed

Harrington, Catriona; Reen, F Jerry; Mooij, Marlies J; Stewart, Fiona A; Chabot, Jean-Baptiste; Guerra, Antonio F; Glöckner, Frank O; Nielsen, Kristian F; Gram, Lone; Dobson, Alan D W; Adams, Claire; O'Gara, Fergal

2014-12-10

The search for new antimicrobial compounds has gained added momentum in recent years, paralleled by the exponential rise in resistance to most known classes of current antibiotics. While modifications of existing drugs have brought some limited clinical success, there remains a critical need for new classes of antimicrobial compound to which key clinical pathogens will be naive. This has provided the context and impetus to marine biodiscovery programmes that seek to isolate and characterize new activities from the aquatic ecosystem. One new antibiotic to emerge from these initiatives is the antibacterial compound tropodithietic acid (TDA). The aim of this study was to provide insight into the bioactivity of and the factors governing the production of TDA in marine Pseudovibrio isolates from a collection of marine sponges. The TDA produced by these Pseudovibrio isolates exhibited potent antimicrobial activity against a broad spectrum of clinical pathogens, while TDA tolerance was frequent in non-TDA producing marine isolates. Comparative genomics analysis suggested a high degree of conservation among the tda biosynthetic clusters while expression studies revealed coordinated regulation of TDA synthesis upon transition from log to stationary phase growth, which was not induced by TDA itself or by the presence of the C10-acyl homoserine lactone quorum sensing signal molecule.

Characterisation of Non-Autoinducing Tropodithietic Acid (TDA) Production from Marine Sponge Pseudovibrio Species

PubMed Central

Harrington, Catriona; Reen, F. Jerry; Mooij, Marlies J.; Stewart, Fiona A.; Chabot, Jean-Baptiste; Guerra, Antonio F.; Glöckner, Frank O.; Nielsen, Kristian F.; Gram, Lone; Dobson, Alan D. W.; Adams, Claire; O’Gara, Fergal

2014-01-01

The search for new antimicrobial compounds has gained added momentum in recent years, paralleled by the exponential rise in resistance to most known classes of current antibiotics. While modifications of existing drugs have brought some limited clinical success, there remains a critical need for new classes of antimicrobial compound to which key clinical pathogens will be naive. This has provided the context and impetus to marine biodiscovery programmes that seek to isolate and characterize new activities from the aquatic ecosystem. One new antibiotic to emerge from these initiatives is the antibacterial compound tropodithietic acid (TDA). The aim of this study was to provide insight into the bioactivity of and the factors governing the production of TDA in marine Pseudovibrio isolates from a collection of marine sponges. The TDA produced by these Pseudovibrio isolates exhibited potent antimicrobial activity against a broad spectrum of clinical pathogens, while TDA tolerance was frequent in non-TDA producing marine isolates. Comparative genomics analysis suggested a high degree of conservation among the tda biosynthetic clusters while expression studies revealed coordinated regulation of TDA synthesis upon transition from log to stationary phase growth, which was not induced by TDA itself or by the presence of the C10-acyl homoserine lactone quorum sensing signal molecule. PMID:25513851

SlpE is a calcium-dependent cytotoxic metalloprotease associated with clinical isolates of Serratia marcescens.

PubMed

Stella, Nicholas A; Callaghan, Jake D; Zhang, Liang; Brothers, Kimberly M; Kowalski, Regis P; Huang, Jean J; Thibodeau, Patrick H; Shanks, Robert M Q

Serralysin-like proteases are found in a wide variety of bacteria. These metalloproteases are frequently implicated in virulence and are members of the widely conserved RTX-toxin family. We identified a serralysin-like protease in the genome of a clinical isolate of Serratia marcescens that is highly similar to the canonical serralysin protein, PrtS. This gene was named serralysin-like protease E, SlpE, and was found in the majority (67%) of tested clinical isolates, but was absent from most tested non-clinical isolates including the insect pathogen and reference S. marcescens strain Db11. Purified recombinant SlpE exhibited calcium-dependent protease activity similar to metalloproteases PrtS and SlpB. Induction of slpE in the low-protease-producing S. marcescens strain PIC3611 highly elevated extracellular protease activity, and extracellular secretion required the lipD type 1 secretion system gene. Transcription of slpE was highly reduced in an eepR transcription factor mutant. Mutation of the slpE gene in a highly proteolytic clinical isolate reduced its protease activity, and evidence suggests that SlpE confers cytotoxicity of S. marcescens to the A549 airway carcinoma cell line. Together, these data reveal SlpE to be an EepR-regulated cytotoxic metalloprotease associated with clinical isolates of an important opportunistic pathogen. Copyright © 2017 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.

Prevalence of multi and pan drug resistant Pseudomonas aeruginosa with respect to ESBL and MBL in a tertiary care hospital.

PubMed

Jayakumar, S; Appalaraju, B

2007-10-01

Multi drug resistant Pseudomonas aeruginosa (MDRPA) and pan drug resistant Pseudomonas aeruginosa (PDRPA) isolates in critically ill patients are often difficult to treat. Prevalence of MDRPA and their antibiotic profile was investigated to select an appropriate empirical therapy. Moreover lack of sufficient data on prevalence of PDRPA in tertiary care hospitals indicated the need for this study. Pseudomonas aeruginosa was isolated in 245 patients over a period of one and half years from various clinical materials and their antibiotic profile was determined. Minimum inhibitory concentration (MIC) for Imipenem and Meropenam was determined by broth dilution method. Phenotypic confirmation test and EDTA double disk synergy test was used to detect Extended spectrum a-lactamase (ESBL) and Metallo-a-lactamase (MBL) producers respectively. Out of 245 isolates, 54 strains (22 %) and 11 strains (4%) were found to be MDRPA and PDRPA respectively. Carbapenem resistant isolates showed MICs ranging from 16 to > 64 microg/ml. Thirty eight strains (15.5%) were ESBL producers and six (54.5%) among 11 PDRPA were MBL producers. Prevalence of MDR and PDR isolates of Pseudomonas aeruginosa was found to be 22% and 4% respectively, which is less compared to other studies. Majority of the PDRPA isolates were MBL producers which have propensity to spread to other bacteria.

Frequency of enterotoxins, toxic shock syndrome toxin-1, and biofilm formation genes in Staphylococcus aureus isolates from cows with mastitis in the Northeast of Brazil.

PubMed

Costa, F N; Belo, N O; Costa, E A; Andrade, G I; Pereira, L S; Carvalho, I A; Santos, R L

2018-06-01

Staphylococcus aureus is among the microorganisms more frequently associated with subclinical bovine mastitis. S. aureus may produce several virulence factors. This study aimed at determining the frequency of virulence factors such as enterotoxins, toxic shock syndrome toxin 1, and ica adhesion genes. In addition, we assessed antimicrobial drug resistance in S. aureus isolated from clinical and subclinical cases of mastitis. A total of 88 cows with clinical or subclinical mastitis were sampled, resulting in 38 S. aureus isolates, from which 25 (65.78%) carried toxin genes, including seb, sec, sed, tst, and icaD adhesion gene. These S. aureus isolates belong to 21 ribotypes and three S. aureus strains belonged to the same ribotype producing ica adhesion gene. Approximately 90% of S. aureus strains obtained in our study demonstrated multiple resistance to different antimicrobial agents. The most efficacious antimicrobial agents against the isolates were gentamicin, amoxicillin, and norfloxacin. Gentamicin was the most efficacious agent inhibiting 78.95% of the S. aureus isolates. The least efficacious were penicillin, streptomycin, and ampicillin. Our results can help in understanding the relationship between virulence factors and subclinical mastitis caused by S. aureus. Further research about diversity of S. aureus isolates and genes responsible for the pathogenicity of subclinical mastitis is essential.

Evaluation of fosfomycin activity against ESBL producing Enterobacteriaceae isolated from Iran.

PubMed

Kazemian, Hossein

2018-05-16

Rising rates of antimicrobial resistance among Enterobacteriaceae limit the use of reliably active forms of available drugs. The aim of this study was to investigate the prevalence of fosfomycin (US6794490B2) resistance gene among ESBL producing isolates in Iran. We tested 355 isolates of Enterobacteriacea collected from various clinical samples including urine, wounds, blood and other sources during June 2016 to July 2017. Antibiotic sensitivity and extended spectrum beta lactamase (ESBL) production were tested using agar dilution method according to the Clinical and Laboratory Standards Institute (CLSI) guidelines. ESBL genes (blaTEM, bla SHV,bla CTX-M), plasmid-encoded fosfomycin resistance genes (fosA, fosB, fosA3 and fosC2) and chromosomal mutations (murA, glpT, uhpT) were detected by polymerase chain reaction (PCR). In this study, 151 of the 355 isolates were ESBL-positive. blaCTX-M (77%) was the most common gene followed by blaSHV (70%) and blaTEM (58%), either alone or in combination. Eighty nine percent (132/151) of the ESBL-positive isolates were MDR. Antimicrobial susceptibility rates were higher for fosfomycin (92.8%) and imipenem (35.5%) among ESBL-positive isolates. None of the ESBL- positive isolates harbored any mutations or plasmid-mediated fosfomycin resistance determinants. In conclusion, fosfomycin showed good antimicrobial activity against multidrug resistance ESBL- positive Enterobacteriaceae. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Whole genome sequencing as a tool to investigate a cluster of seven cases of listeriosis in Austria and Germany, 2011-2013.

PubMed

Schmid, D; Allerberger, F; Huhulescu, S; Pietzka, A; Amar, C; Kleta, S; Prager, R; Preußel, K; Aichinger, E; Mellmann, A

2014-05-01

A cluster of seven human cases of listeriosis occurred in Austria and in Germany between April 2011 and July 2013. The Listeria monocytogenes serovar (SV) 1/2b isolates shared pulsed-field gel electrophoresis (PFGE) and fluorescent amplified fragment length polymorphism (fAFLP) patterns indistinguishable from those from five food producers. The seven human isolates, a control strain with a different PFGE/fAFLP profile and ten food isolates were subjected to whole genome sequencing (WGS) in a blinded fashion. A gene-by-gene comparison (multilocus sequence typing (MLST)+) was performed, and the resulting whole genome allelic profiles were compared using SeqSphere(+) software version 1.0. On analysis of 2298 genes, the four human outbreak isolates from 2012 to 2013 had different alleles at ≤6 genes, i.e. differed by ≤6 genes from each other; the dendrogram placed these isolates in between five Austrian unaged soft cheese isolates from producer A (≤19-gene difference from the human cluster) and two Austrian ready-to-eat meat isolates from producer B (≤8-gene difference from the human cluster). Both food products appeared on grocery bills prospectively collected by these outbreak cases after hospital discharge. Epidemiological results on food consumption and MLST+ clearly separated the three cases in 2011 from the four 2012-2013 outbreak cases (≥48 different genes). We showed that WGS is capable of discriminating L. monocytogenes SV1/2b clones not distinguishable by PFGE and fAFLP. The listeriosis outbreak described clearly underlines the potential of sequence-based typing methods to offer enhanced resolution and comparability of typing systems for public health applications. © 2014 The Authors Clinical Microbiology and Infection © 2014 European Society of Clinical Microbiology and Infectious Diseases.

Pock forming ability of fowl pox virus isolated from layer chicken and its adaptation in chicken embryo fibroblast cell culture

PubMed Central

Gilhare, Varsha Rani; Hirpurkar, S. D.; Kumar, Ashish; Naik, Surendra Kumar; Sahu, Tarini

2015-01-01

Aim: The objective of the present study was to examine pock forming ability of field strain and vaccine strain of fowl pox virus (FPV) in chorioallantoic membrane (CAM) of embryonated chicken eggs and its adaptation in chicken embryo fibroblast (CEF) cell culture. Materials and Methods: Dry scabs were collected from 25 affected birds in glycerin-saline and preserved at 4°C until processed. Virus was isolated in 10-day-old embryonated chicken eggs by dropped CAM method. The identity of the virus is confirmed by clinical findings of affected birds, pock morphology and histopathology of infected CAM. In addition one field isolate and vaccine strain of FPV was adapted to CEF cell culture. CEF cell culture was prepared from 9-day-old embryonated chicken eggs. Result: Clinical symptoms observed in affected birds include pox lesion on comb, wattle, eyelids and legs, no internal lesions were observed. All field isolates produced similar findings in CAM. Pocks produced by field isolates ranged from 3 mm to 5 mm at the third passage while initial passages edematous thickening and necrosis of CAM was observed. Pocks formed by lyophilized strain were ranges from 0.5 mm to 2.5 mm in diameter scattered all over the membrane at the first passage. Intra-cytoplasmic inclusion bodies are found on histopathology of CAM. At third passage level, the CEF inoculated with FPV showed characteristic cytopathic effect (CPE) included aggregation of cells, syncytia and plaque formation. Conclusion: FPV field isolates and vaccine strain produced distinct pock lesions on CAMs. Infected CAM showed intracytoplasmic inclusion bodies. The CEF inoculated with FPV field isolate as well as a vaccine strain showed characteristic CPE at third passage level. PMID:27047081

Pock forming ability of fowl pox virus isolated from layer chicken and its adaptation in chicken embryo fibroblast cell culture.

PubMed

Gilhare, Varsha Rani; Hirpurkar, S D; Kumar, Ashish; Naik, Surendra Kumar; Sahu, Tarini

2015-03-01

The objective of the present study was to examine pock forming ability of field strain and vaccine strain of fowl pox virus (FPV) in chorioallantoic membrane (CAM) of embryonated chicken eggs and its adaptation in chicken embryo fibroblast (CEF) cell culture. Dry scabs were collected from 25 affected birds in glycerin-saline and preserved at 4°C until processed. Virus was isolated in 10-day-old embryonated chicken eggs by dropped CAM method. The identity of the virus is confirmed by clinical findings of affected birds, pock morphology and histopathology of infected CAM. In addition one field isolate and vaccine strain of FPV was adapted to CEF cell culture. CEF cell culture was prepared from 9-day-old embryonated chicken eggs. Clinical symptoms observed in affected birds include pox lesion on comb, wattle, eyelids and legs, no internal lesions were observed. All field isolates produced similar findings in CAM. Pocks produced by field isolates ranged from 3 mm to 5 mm at the third passage while initial passages edematous thickening and necrosis of CAM was observed. Pocks formed by lyophilized strain were ranges from 0.5 mm to 2.5 mm in diameter scattered all over the membrane at the first passage. Intra-cytoplasmic inclusion bodies are found on histopathology of CAM. At third passage level, the CEF inoculated with FPV showed characteristic cytopathic effect (CPE) included aggregation of cells, syncytia and plaque formation. FPV field isolates and vaccine strain produced distinct pock lesions on CAMs. Infected CAM showed intracytoplasmic inclusion bodies. The CEF inoculated with FPV field isolate as well as a vaccine strain showed characteristic CPE at third passage level.

Comparison of Molecular and Phenotypic Methods for the Detection and Characterization of Carbapenem Resistant Enterobacteriaceae.

PubMed

Somily, Ali M; Garaween, Ghada A; Abukhalid, Norah; Absar, Muhammad M; Senok, Abiola C

2016-03-01

In recent years, there has been a rapid dissemination of carbapenem resistant Enterobacteriaceae (CRE). This study aimed to compare phenotypic and molecular methods for detection and characterization of CRE isolates at a large tertiary care hospital in Saudi Arabia. This study was carried out between January 2011 and November 2013 at the King Khalid University Hospital (KKUH) in Saudi Arabia. Determination of presence of extended-spectrum beta-lactamases (ESBL) and carbapenem resistance was in accordance with Clinical and Laboratory Standards Institute (CLSI) guidelines. Phenotypic classification was done by the MASTDISCS(TM) ID inhibitor combination disk method. Genotypic characterization of ESBL and carbapenemase genes was performed by the Check-MDR CT102. Diversilab rep-PCR was used for the determination of clonal relationship. Of the 883 ESBL-positive Enterobacteriaceae detected during the study period, 14 (1.6%) isolates were carbapenem resistant. Both the molecular genotypic characterization and phenotypic testing were in agreement in the detection of all 8 metalo-beta-lactamases (MBL) producing isolates. Of these 8 MBL-producers, 5 were positive for blaNDM gene and 3 were positive for blaVIM gene. Molecular method identified additional blaOXA gene isolates while MASTDISCS(TM) ID detected one AmpC producer isolate. Both methods agreed in identifying 2 carbapenem resistant isolates which were negative for carbapenemase genes. Diversilab rep-PCR analysis of the 9 Klebsiella pneumoniae isolates revealed polyclonal distribution into eight clusters. MASTDISCS(TM) ID is a reliable simple cheap phenotypic method for detection of majority of carbapenemase genes with the exception of the blaOXA gene. We recommend to use such method in the clinical laboratory.

Phenotypic and genotypic characterization of biofilm formation among Staphylococcus aureus isolates from clinical specimens, an Atomic Force Microscopic (AFM) study.

PubMed

Bazari, Pelin Aslani Menareh; Honarmand Jahromy, Sahar; Zare Karizi, Shohreh

2017-09-01

Staphylococcus aureus is a major cause of nosocomial infections. Biofilm formation is an important factor for bacterial pathogenesis. Its mechanisms are complex and include of many genes depends on expression of icaADBC operon involved in the synthesis of a polysaccharide intercellular adhesion. The aim of study was to investigate biofilm forming ability of Staphylococcus aureus strains by phenotypic and genotypic methods. Also Atomic Force microscope (AFM) was used to visualize biofilm formation. 140 Isolates were collected from clinical specimens of patients in Milad Hospital, Tehran and diagnosed by biochemical tests. The ability of strains to produce slime was evaluated by CRA method. For diagnosing of bacterial EPS, Indian ink staining were used and finally biofilm surface of 3 isolates observed by AFM. The prevalence of icaA and icaD genes was determined by PCR. By CRA method 15% of samples considered as positive slime producers, 44.28% as intermediate and 40.71% indicative as negative slime producers. 118 staphylococcus aureus strains showed a distinct halo transparent zone but 22 strains showed no halo zone. AFM analysis of Slime positive isolates showed a distinct and complete biofilm formation. In slime negative strain, there was not observed biofilm. The prevalence of icaA, icaD genes was 44.2% and 10% of the isolates had both genes simultaneously. There is a relationship between exopolysaccharide layer and biofilm formation of Staphylococcus aureus isolates. The presence of icaAD genes among isolates is not associated with in vitro formation of biofilm. AFM is a useful tool for observation of bacterial biofilm formation. Copyright © 2017 Elsevier Ltd. All rights reserved.

Occurrence of efflux mechanism and cephalosporinase variant in a population of Enterobacter aerogenes and Klebsiella pneumoniae isolates producing extended-spectrum beta-lactamases.

PubMed

Tran, Que-Tien; Dupont, Myrielle; Lavigne, Jean-Philippe; Chevalier, Jacqueline; Pagès, Jean-Marie; Sotto, Albert; Davin-Regli, Anne

2009-04-01

We investigated the occurrence of multidrug resistance in 44 Enterobacter aerogenes and Klebsiella pneumoniae clinical isolates. Efflux was involved in resistance in E. aerogenes isolates more frequently than in K. pneumoniae isolates (100 versus 38% of isolates) and was associated with the expression of phenylalanine arginine beta-naphthylamide-susceptible active efflux. AcrA-TolC overproduction in E. aerogenes isolates was noted. An analysis of four E. aerogenes isolates for which cefepime MICs were high revealed no modification in porin expression but a new specific mutation in the AmpC beta-lactamase.

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±0.32), respectively. Also, MBL-positive strains produced robust biofilm compared to MBL-negative strains. Collectively, the production of site-dependent virulence factors can be highlighted as potential therapeutic targets for the treatment of infections caused by heterogeneous and resistant strains of P. aeruginosa. Copyright © 2014 Elsevier GmbH. All rights reserved.

Antimicrobial resistance pattern of Neisseria gonorrhoeae isolates from peripheral health centres and STD clinic attendees of a tertiary care centre in India.

PubMed

Bala, M; Ray, K; Gupta, S M

2008-06-01

The aim of the study was to compare the antimicrobial resistance pattern of Neisseria gonorrhoeae isolates from urban and rural peripheral health centres and from sexually transmitted disease (STD) clinic attendees. Antimicrobial susceptibility testing of 191 N. gonorrhoeae isolates (165 isolates from STD clinic attendees and 26 from peripheral health centres) was carried out in Delhi, India, using the calibrated dichotomous sensitivity technique for penicillin, tetracycline, ceftriaxone, ciprofloxacin, spectinomycin and nalidixic acid, and minimum inhibitory concentrations were determined using E-test. Penicillin-resistant, ciprofloxacin-resistant, penicillinase-producing N. gonorrhoeae and tetracycline-resistant N. gonorrhoeae strains were higher in STD clinic attendees than in peripheral health centres, probably because of less antibiotic pressure in the peripheral areas. High-level resistance to ciprofloxacin and multiresistant strains were also higher in STD clinic attendees. The present study emphasizes the importance of surveillance of antimicrobial resistance of N. gonorrhoeae in different population subgroups in order to monitor the spread of multiresistant strains and to update the national treatment recommendations.

An investigation of exudative epidermitis (greasy pig disease) and antimicrobial resistance patterns of Staphylococcus hyicus and Staphylococcus aureus isolated from clinical cases

PubMed Central

Park, Jeonghwa; Friendship, Robert M.; Poljak, Zvonimir; Weese, J. Scott; Dewey, Cate E.

2013-01-01

Exudative epidermitis (EE) is a common skin disease of young pigs, caused mainly by Staphylococcus hyicus. Increased prevalence of EE and poor response to treatment are reported. Common strategies used by Ontario pork producers to treat pigs with EE were determined using a survey. Injection of penicillin G was reported as the most common parenteral antibiotic choice. Antimicrobial resistance patterns of S. hyicus and Staphylococcus aureus isolated from clinical cases (30 herds with samples from approximately 6 pigs per farm) showed that 97% of S. hyicus isolates were resistant to penicillin G and ampicillin; 71% of these isolates were resistant to ceftiofur. Similar resistance was noted among S. aureus isolates. Antimicrobial resistance has become a problem in the treatment of EE in Ontario. PMID:23904636

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.

Microbiological characterisation of Burkholderia cepacia isolates from cystic fibrosis patients: investigation of the exopolysaccharides produced.

PubMed

Lagatolla, Cristina; Skerlavaj, Silvia; Dolzani, Lucilla; Tonin, Enrico A; Monti Bragadin, Carlo; Bosco, Marco; Rizzo, Roberto; Giglio, Luisella; Cescutti, Paola

2002-03-19

Eleven strains of Burkholderia cepacia were isolated directly from clinical specimens: 10 from sputum of cystic fibrosis patients, and one from a vaginal swab. They were biochemically identified using API20NE and confirmed by a PCR-based assay. The genomovar characterisation obtained by specific PCR amplification revealed seven strains belonging to genomovar I, three belonging to genomovar IIIA and one belonging to genomovar IV. All isolates were also typed by ribotyping and random amplification of polymorphic DNA analysis. Some of the characterised strains were examined for the ability to produce exopolysaccharides, with the aim of correlating the genomovar with the exopolysaccharide structure. The polysaccharides were analysed by means of methylation analysis and 1H-NMR spectroscopy in order to determine structural similarities. It was shown that different strains are capable of producing chemically different polysaccharides.

Mutation in the S-ribosylhomocysteinase (luxS) gene involved in quorum sensing affects biofilm formation and virulence in a clinical isolate of Aeromonas hydrophila

EPA Science Inventory

A diarrheal isolate SSU of Aeromonas hydrophila produces a cytotoxic enterotoxin (Act) with cytotoxic, enterotoxic, and hemolytic activities. Our laboratory has characterized from the above Aeromonas strain, in addition to Act, the type 3- and T6-secretion systems and their effec...

Antimicrobial activity of tigecycline against recent isolates of respiratory pathogens from Asian countries.

PubMed

Ko, Kwan Soo; Song, Jae-Hoon; Lee, Mi Young; Park, Sulhee; Kwon, Ki Tae; Heo, Sang Taek; Ryu, Seong Yeol; Oh, Won Sup; Peck, Kyong Ran; Lee, Nam Yong

2006-08-01

In vitro activities of tigecycline were compared with 15 other comparator agents against recent clinical isolates of respiratory pathogens (623 Streptococcus pneumoniae, 105 Staphylococcus aureus, 92 Klebsiella pneumoniae, and 84 Haemophilus influenzae isolates) collected from 11 Asian countries. All isolates of S. pneumoniae from Asian countries were susceptible to tigecycline regardless of penicillin susceptibility with MIC90 of

Genome-based analysis of Carbapenemase-producing Klebsiella pneumoniae isolates from German hospital patients, 2008-2014.

PubMed

Becker, Laura; Kaase, Martin; Pfeifer, Yvonne; Fuchs, Stephan; Reuss, Annicka; von Laer, Anja; Sin, Muna Abu; Korte-Berwanger, Miriam; Gatermann, Sören; Werner, Guido

2018-01-01

By using whole genome sequence data we aimed at describing a population snapshot of carbapenemase-producing K. pneumoniae isolated from hospitalized patients in Germany between 2008 and 2014. We selected a representative subset of 107 carbapenemase-producing K. pneumoniae clinical isolates possessing the four most prevalent carbapenemase types in Germany (KPC-2, KPC-3, OXA-48, NDM-1). Isolates were processed via illumina NGS. Data were analysed using different SNP-based mapping and de-novo assembly approaches. Relevant information was extracted from NGS data (antibiotic resistance determinants, wzi gene/ cps type, virulence genes). NGS data from the present study were also compared with 238 genome data from two previous international studies on K. pneumoniae. NGS-based analyses revealed a preferred prevalence of KPC-2-producing ST258 and KPC-3-producing ST512 isolates. OXA-48, being the most prevalent carbapenemase type in Germany, was associated with various K. pneumoniae strain types; most of them possessing IncL/M plasmid replicons suggesting a preferred dissemination of bla OXA-48 via this well-known plasmid type. Clusters ST15, ST147, ST258, and ST512 demonstrated an intermingled subset structure consisting of German and other European K. pneumoniae isolates. ST23 being the most frequent MLST type in Asia was found only once in Germany. This latter isolate contained an almost complete set of virulence genes and a K1 capsule suggesting occurrence of a hypervirulent ST23 strain producing OXA-48 in Germany. Our study results suggest prevalence of "classical" K. pneumonaie strain types associated with widely distributed carbapenemase genes such as ST258/KPC-2 or ST512/KPC-3 also in Germany. The finding of a supposed hypervirulent and OXA-48-producing ST23 K. pneumoniae isolates outside Asia is highly worrisome and requires intense molecular surveillance.

Prevalence of extended-spectrum beta-lactamases-producing microorganisms in nosocomial patients and molecular characterization of the shv type isolates

PubMed Central

de Oliveira, Caio Fernando; Salla, Adenilde; Lara, Valéria Maria; Rieger, Alexandre; Horta, Jorge André; Alves, Sydney Hartz

2010-01-01

The emergence of Extended-Spectrum Beta-Lactamase (ESBL)-producing microorganisms in Brazilian hospitals is a challenge that concerns scientists, clinicians and healthcare institutions due to the serious risk they pose to confined patients. The goal of this study was the detection of ESBL production by clinical strains of Escherichia coli and Klebsiella sp. isolated from pus, urine and blood of patients at Hospital Universitário Santa Maria, Rio Grande Sul, RS, Brazil and the genotyping of the isolates based on bla SHV genes. The ESBL study was carried out using the Combined Disc Method, while Polymerase Chain Reaction (PCR) was used to study the bla SHV genes. Of the 90 tested isolates, 55 (61.1%) were identified as ESBL-producing by the combined disk method. The bla SHV genes were found in 67.8% of these microorganisms. K. pneumoniae predominated in the samples, presenting the highest frequency of positive results from the combined disk and PCR. PMID:24031491

Clinical and veterinary isolates of Salmonella enterica serovar Enteritidis defective in lipopolysaccharide O-chain polymerization

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

Guard-Petter, J.; Parker, C.T.; Asokan, K.

1999-05-01

Twelve human and chicken isolates of Salmonella enterica serovar Enteritidis belonging to phage types 4, 8, 13a, and 23 were characterized for variability in lipopolysaccharide (LPS) composition. Isolates were differentiated into two groups, i.e., those that lacked immunoreactive O-chain, termed rough isolates, and those that had immunoreactive O-chain, termed smooth isolates. Isolates within these groups could be further differentiated by LPS compositional differences as detected by gel electrophoresis and gas liquid chromatography of samples extracted with water, which yielded significantly more LPS in comparison to phenol-chloroform extraction. The rough isolates were of two types, the O-antigen synthesis mutants and themore » O-antigen polymerization (wzy) mutants. Smooth isolates were also of two types, one producing low-molecular-weight (LMW) LPS and the other producing high-molecular-weight (HMW) LPS. To determine the genetic basis for the O-chain variability of the smooth isolates, the authors analyzed the effects of a null mutation in the O-chain length determinant gene, wzz (cld) of serovar Typhimurium. This mutation results in a loss of HMW LPS; however, the LMW LPS of this mutant was longer and more glucosylated than that from clinical isolates of serovar Enteritidis. Cluster analysis of these data and of those from two previously characterized isogenic strains of serovar Enteritidis that had different virulence attributes indicated that glucosylation of HMW LPS (via oafR function) is variable and results in two types of HMW structures, one that is highly glucosylated and one that is minimally glucosylated. These results strongly indicate that naturally occurring variability in wzy, wzz, and oafR function can be used to subtype isolates of serovar Enteritidis during epidemiological investigations.« less

Pseudomonas aeruginosa, an emerging pathogen among burn patients in Kurdistan Province, Iran.

PubMed

Kalantar, Enayat; Taherzadeh, Shadi; Ghadimi, Tayeb; Soheili, Fariborz; Salimizand, Heiman; Hedayatnejad, Alireza

2012-05-01

This study was conducted to determine the incidence of Pseudomonas aeruginosa infections among burn patients at Tohid Hospital, Iran. A total of 176 clinical specimens were obtained from 145 burn patients admitted to the burn unit of Tohid Hospital to detect the presence of P. aeruginosa. Antimicrobial susceptibility testing was conducted to detect extended spectrum beta-lactamase (ESBL) producing P. aeruginiosa using Clinical and Laboratory Standards Institute guidelines with the double disc synergy test (DDST). A polymerase chain reaction was used to detect PER-1 and OXA-10 among the isolates. The mean age, total body surface area and length of hospital stay among patients were 29 years, 37.7%, and 10 days, respectively. Kerosene was the commonest cause of burn (60%), followed by gas (30%). During the study, P. aeruginosa was detected in 100 isolates. The antibiotics they were most commonly resistant to were cefotaxime, ceftriaxone and ciprofloxacin. Of the 100 P. aeroginusa isolates, 28% were positive for ESBL production with the DDST, 48% and 52% were PER-1 and OXA-10 producers, respectively. The high frequency of PER-1 and OXA-10 producers at this hospital is of concern considering their potential spread among burn patients.

[Comparison of different methods for the identification of Candida species isolated from clinical specimens].

PubMed

Cetinkaya, Zafer; Altindiş, Mustafa; Aktepe, Orhan Cem; Karabiçak, Nilgün

2003-10-01

The aim of this study was to compare the different methods for the identification of Candida strains isolated from clinical specimens. The methods of germ tube examination, chlamydospore examination formed on the rice Tween-80 (RT-80) agar and evaluation of colony morphologies on the two chromogenic agars (CHROMagar Candida, Albicans ID), were compared with a reference API 20C AUX (bioMerieux, France) automated system based on the carbohydrate assimilation, for the identification of a total 255 Candida isolates. Of them, 173 (67.8%) were identified as C. albicans, 37 (14.5%) were C. glabrata, 23 (9%) were C. krusei, 9 (3.5%) were C. tropicalis, 9 (3.5%) were C. kefyr, 2 (0.8%) were C. guillermondii and 2 (0.8%) were C. parapsilosis, by API 20C AUX system. In the view of these results, 146 (84.4%) of C. albicans strains were identified by germ tube examination, 161 (93.1%) of C. albicans strains and 208 (81.5%) of total strains were identified by chlamydospore examination. 169 (97.7%) of C. albicans strains and 231 (90.6%) of total strains were identified by CHROMagar Candida method, and 168 (97.1%) of C. albicans strains were identified by Albicans ID method, correctly. In the CHROMagar Candida medium, 169 C. albicans isolates have produced bright green colored colonies, whereas 33 (89.2%) isolates which produced dark pink/purple colored colonies were identified as C. glabrata, 7 (77.8%) isolates which produced metalical blue colored colonies were identified as C. tropicalis and 22 (95.6%) isolates which produced pale pink colored colonies were identified as C. krusei. In the Albicans ID medium, four of the 172 isolates which were evaluated as C. albicans initially by producing blue colored colonies, have been identified as C. tropicalis by API 20C AUX system. The sensitivities and specificities of germ tube examination, RT-80, CHROMagar Candida and Albicans ID methods were found as follows, respectively; 84.4% and 100%, 93.1% and 100%, 97.7% and 100%, 99.4% and 95.3 percent. In conclusion, CHROMagar Candida medium seems the most favorable rapid and practical method with high sensitivity and specificity for the identification of Candida species, but its cost-effectiveness should be kept in view.

Clonality and Resistome analysis of KPC-producing Klebsiella pneumoniae strain isolated in Korea using whole genome sequencing.

PubMed

Lee, Yangsoon; Kim, Bong-Soo; Chun, Jongsik; Yong, Ji Hyun; Lee, Yeong Seon; Yoo, Jung Sik; Yong, Dongeun; Hong, Seong Geun; D'Souza, Roshan; Thomson, Kenneth S; Lee, Kyungwon; Chong, Yunsop

2014-01-01

We analyzed the whole genome sequence and resistome of the outbreak Klebsiella pneumoniae strain MP14 and compared it with those of K. pneumoniae carbapenemase- (KPC-) producing isolates that showed high similarity in the NCBI genome database. A KPC-2-producing multidrug-resistant (MDR) K. pneumoniae clinical isolate was obtained from a patient admitted to a Korean hospital in 2011. The strain MP14 was resistant to all tested β-lactams including monobactam, amikacin, levofloxacin, and cotrimoxazole, but susceptible to tigecycline and colistin. Resistome analysis showed the presence of β-lactamase genes including bla KPC-2, bla SHV-11, bla TEM-169, and bla OXA-9. MP14 also possessed aac(6'-)Ib, aadA2, and aph(3'-)Ia as aminoglycoside resistance-encoding genes, mph(A) for macrolides, oqxA and oqxB for quinolone, catA1 for phenicol, sul1 for sulfonamide, and dfrA12 for trimethoprim. Both SNP tree and cgMLST analysis showed the close relatedness with the KPC producers (KPNIH strains) isolated from an outbreak in the USA and colistin-resistant strains isolated in Italy. The plasmid-scaffold genes in plasmids pKpQil, pKpQil-IT, pKPN3, or pKPN-IT were identified in MP14, KPNIH, and Italian strains. The KPC-2-producing MDR K. pneumoniae ST258 stain isolated in Korea was highly clonally related with MDR K. pneumoniae strains from the USA and Italy. Global spread of KPC-producing K. pneumoniae is a worrying phenomenon.

Both epsilon-toxin and beta-toxin are important for the lethal properties of Clostridium perfringens type B isolates in the mouse intravenous injection model.

PubMed

Fernandez-Miyakawa, Mariano E; Fisher, Derek J; Poon, Rachael; Sayeed, Sameera; Adams, Vicki; Rood, Julian I; McClane, Bruce A; Uzal, Francisco A

2007-03-01

Clostridium perfringens is capable of producing up to 15 toxins, including alpha-toxin (CPA), beta-toxin (CPB), epsilon-toxin (ETX), enterotoxin, beta2-toxin (CPB2), and perfringolysin O. Type B isolates, which must produce CPA, CPB, and ETX, are associated with animal illnesses characterized by sudden death or acute neurological signs, with or without intestinal damage. Type B pathogenesis in ruminants is poorly understood, with some animals showing lesions and clinical signs similar to those caused by either type C or type D infections. It is unknown whether host or environmental conditions are dominant for determining the outcome of type B disease or if disease outcomes are determined by variable characteristics of type B isolates. To help clarify this issue, 19 type B isolates were evaluated for toxin production during late-log-phase growth via quantitative Western blotting and by biological activity assays. Most type B isolates produced CPB levels similar to those produced by type C isolates in vitro and have the potential to produce genotype C-like disease. The lethality of type B isolate supernatants administered intravenously to mice was evaluated with or without prior trypsin treatment, and monoclonal antibody neutralization studies also were performed. Correlation analyses comparing toxin levels in type B supernatants versus lethality and neutralization studies both found that the main contributor to lethality without pretreatment with trypsin was CPB, whereas neutralization studies indicated that CPB and ETX were both important after trypsin pretreatment. At least part of the CPB produced by type B isolates remained active after trypsin treatment. However, the overall lethalities of most supernatants were lower after trypsin pretreatment. Also, there was a significant association between ETX, CPB2, and CPA production in vitro among type B isolates. However, our results suggest that both CPB and ETX are likely the most important contributors to the pathogenesis of C. perfringens type B infections in domestic animals.

Prevalence of O25b-ST131 clone among Escherichia coli strains producing CTX-M-15, CTX-M-14 and CTX-M-92 β-lactamases.

PubMed

Giedraitienė, Agnė; Vitkauskienė, Astra; Pavilonis, Alvydas; Patamsytė, Vaiva; Genel, Nathalie; Decre, Dominique; Arlet, Guillaume

2017-02-01

Dissemination of multidrug-resistant Escherichia coli is closely associated with the worldwide spread of a single clone ST131, which is the main cause of urinary tract and bloodstream infections in patients from nursing homes and immunocompromised patients. The aim of our study was to determine the prevalence of ST131 clone and the replicons involved in the spread of bla CTX-M genes among O25b-ST131 CTX-M-producing E. coli isolates in Lithuania. The strains included in this study were screened for CTX-M β-lactamase-encoding genes, phylogenetic groups and ST131 clone by PCR. Bacterial conjugation was performed to identify plasmid replicon types responsible for bla CTX-M genes dissemination. A total of 158 E. coli clinical non-duplicate ESBL isolates were analyzed. Nearly half (n = 67, 42.4%) of the investigated E. coli isolates belonged to phylogenetic group B2. The isolates producing CTX-M-92 β-lactamases were identified to be the ST131 clone more frequently than the non-ST131 clone (11.5% vs. 3.1%, p = .035). The CTX-M-15 isolates were identified as ST131 isolates less frequently than non-ST131 isolates (50.8% vs. 71.1%; p = .015). The ST131 clone isolates contained type L/M and A/C replicons; a fused FII/FIB replicon was found in four isolates (23.5%). Type HI1 replicon was identified in ST131 E. coli isolates producing CTX-M-15 β-lactamases. This study demonstrates the predominance of the ST131 clone among CTX-M β-lactamase-producing E. coli isolates. Dissemination of bla CTX-M genes in ST131 strains can be linked not only to highly adapted IncF plasmids such as FII/FIB and FII, but also to plasmid replicon types A/C, L/M and HI1.

Previous Antibiotic Exposure Increases Risk of Infection with Extended-Spectrum-β-Lactamase- and AmpC-Producing Escherichia coli and Klebsiella pneumoniae in Pediatric Patients

PubMed Central

Miles-Jay, Arianna; Kronman, Matthew P.; Zhou, Chuan; Adler, Amanda L.; Haaland, Wren; Weissman, Scott J.; Elward, Alexis; Newland, Jason G.; Zaoutis, Theoklis; Qin, Xuan

2016-01-01

The objective of this study was to determine whether antibiotic exposure is associated with extended-spectrum-beta-lactamase- or AmpC-producing Escherichia coli or Klebsiella pneumoniae infections in children. We collected extended-spectrum-beta-lactamase- or AmpC-producing E. coli or K. pneumoniae isolates and same-species susceptible controls from normally sterile sites of patients aged ≤21 years, along with associated clinical data, at four free-standing pediatric centers. After controlling for potential confounders, the relative risk of having an extended-spectrum-beta-lactamase-producing isolate rather than a susceptible isolate was 2.2 times higher (95% confidence interval [CI], 1.49 to 3.35) among those with antibiotic exposure in the 30 days prior to infection than in those with no antibiotic exposure. The results were similar when analyses were limited to exposure to third-generation cephalosporins, other broad-spectrum beta-lactams, or trimethoprim-sulfamethoxazole. Conversely, the relative risk of having an AmpC-producing versus a susceptible isolate was not significantly elevated with any antibiotic exposure in the 30 days prior to infection (adjusted relative risk ratio, 1.12; 95% CI, 0.65 to 1.91). However, when examining subgroups of antibiotics, the relative risk of having an AmpC-producing isolate was higher for patients with exposure to third-generation cephalosporins (adjusted relative risk ratio, 4.48; 95% CI, 1.75 to 11.43). Dose-response relationships between antibiotic exposure and extended-spectrum-beta-lactamase-producing or AmpC-producing isolates were not demonstrated. These results reinforce the need to study and implement pediatric antimicrobial stewardship strategies, and they indicate that epidemiological studies of third-generation cephalosporin-resistant E. coli and K. pneumoniae isolates should include resistance mechanisms when possible. PMID:27139486

Antimicrobial potential of metabolites extracted from bacterial symbionts associated with marine sponges in coastal area of Gulf of Mannar Biosphere, India.

PubMed

Skariyachan, S; G Rao, A; Patil, M R; Saikia, B; Bharadwaj Kn, V; Rao Gs, J

2014-03-01

Marine coastal areas of India have vast diversity of sponges which harbours many endosymbiotic bacteria which are the source of many potential antimicrobial metabolites. This study focuses the screening and characterization of drug-producing bacteria symbiotically which are associated with marine sponges collected from Gulf of Mannar, South Coast India. Six different sponges were collected and they were identified on the basis of their morphology. The drug-producing isolates were screened by agar overlay method towards various clinical strains. The secondary metabolites were characterized and were found to be quinones, alkaloids, flavanoids and flavonyl glycosides. The metabolites showed significant inhibitory properties against clinical strains that were further identified as chromophoric and fluorophoric in nature. Ethyl acetate extracts of chromophore and floureophore substances showed significant inhibitory properties against Methicillin resistant Staphylococcus aureus (MRSA) and Salmonella typhi respectively. 16S rRNA gene sequencing of theses isolates revealed that chomophore-producing strain were closely related to Pseudomonas spp. RHLB12, isolated from Callyspongia spp. and floureophore-producing bacteria was related to Bacillus licheniformis T6-1 which was isolated from Haliclona spp. Hence, our study demonstrated that antimicrobial metabolites extracted from symbiotic bacteria associated with marine sponges have high therapeutic potential against many bacterial pathogens including multidrug-resistant strains. This is the first study demonstrating antimicrobial potential of flurophoric and chromophoric metabolites extracted from bacterial biosymbionts associated with marine sponges. Our study has significant scope as Indian coastal area especially harbours vast varieties of sponges with novel secondary metabolites-producing organisms. The natural metabolites extracted from sponge-derived bacteria pave novel therapeutic remedy against various pathogens when most of them are emerged as extreme drug resistant superbugs. Letters in Applied Microbiology © 2013 The Society for Applied Microbiology.

In Vitro Assessment of Combined Polymyxin B and Minocycline Therapy against Klebsiella pneumoniae Carbapenemase (KPC)-Producing K. pneumoniae.

PubMed

Huang, Dennis; Yu, Brenda; Diep, John K; Sharma, Rajnikant; Dudley, Michael; Monteiro, Jussimara; Kaye, Keith S; Pogue, Jason M; Abboud, Cely Saad; Rao, Gauri G

2017-07-01

The multidrug resistance profiles of Klebsiella pneumoniae carbapenemase (KPC) producers have led to increased clinical polymyxin use. Combination therapy with polymyxins may improve treatment outcomes, but it is uncertain which combinations are most effective. Clinical successes with intravenous minocycline-based combination treatments have been reported for infections caused by carbapenemase-producing bacteria. The objective of this study was to evaluate the in vitro activity of polymyxin B and minocycline combination therapy against six KPC-2-producing K. pneumoniae isolates (minocycline MIC range, 2 to 32 mg/liter). Polymyxin B monotherapy (0.5, 1, 2, 4, and 16 mg/liter) resulted in a rapid reduction of up to 6 log in bactericidal activity followed by regrowth by 24 h. Minocycline monotherapy (1, 2, 4, 8, and 16 mg/liter) showed no reduction of activity of >1.34 log against all isolates, although concentrations of 8 and 16 mg/liter prolonged the time to regrowth. When the therapies were used in combination, rapid bactericidal activity was followed by slower regrowth, with synergy (60 of 120 combinations at 24 h, 19 of 120 combinations at 48 h) and additivity (43 of 120 combinations at 24 h, 44 of 120 combinations at 48 h) against all isolates. The extent of killing was greatest against the more susceptible polymyxin B isolates (MICs of ≤0.5 mg/liter) regardless of the minocycline MIC. The pharmacodynamic activity of combined polymyxin B-minocycline therapy against KPC-producing K. pneumoniae is dependent on polymyxin B susceptibility. Further in vitro and animal studies must be performed to fully evaluate the efficacy of this drug combination. Copyright © 2017 American Society for Microbiology.

Comparison of growth inhibition and immunofluorescence tests in serotyping clinical isolates of Ureaplasma urealyticum.

PubMed Central

Piot, P

1977-01-01

Typestrains of the eight serotypes of Ureaplasma urealyticum and 39 genital isolates were typed by the growth inhibition (GI) and indirect immunofluorescence (IF) tests. The GI test proved to be very specific and simple, demanding large volumes of serum. It was less sensitive than the IF test, which produced more cross-reactions, was economical in serum, and able to detect mixed infections. Of the 39 isolates, 27 were serotyped by GI and 34 by IF. Mixed cultures occurred in as many as 36% of the isolates. PMID:326348

Characterisation of extended-spectrum β-lactamase and AmpC β-lactamase-producing Enterobacteriaceae isolated from companion animals in New Zealand.

PubMed

Karkaba, A; Grinberg, A; Benschop, J; Pleydell, E

2017-03-01

To assess the occurrence of, and characterise, extended-spectrum β-lactamase (ESBL) and AmpC β-lactamase (AmpC)-producing Enterobacteriaceae isolated by veterinary diagnostic laboratories from infection sites in companion animals in New Zealand. Selected Enterobacteriaceae isolates were submitted by seven New Zealand veterinary diagnostic laboratories. They were isolated from infection sites in companion animals between June 2012 and June 2013, and were resistant to amoxicillin-clavulanic acid, fluoroquinolones, or any combination of two or more antimicrobials. Based on disk diffusion test results, the isolates were phenotypically categorised according to production of ESBL and AmpC. Genes for ESBL and AmpC production were amplified by PCR and sequenced. Escherichia coli isolates were also typed by multilocus sequence typing. A total of 115 isolates matching the inclusion criteria were obtained from the participating laboratories, of which 74 (64%) originated from dogs and 29 (25%) from cats. Seven bacterial species were identified, of which E. coli was the most common (87/115, 76%). Of the 115 isolates, 10 (9%) expressed the ESBL phenotype, 43 (37%) the AmpC phenotype, and seven (6%) both ESBL and AmpC phenotypes. Of the 60 ESBL and AmpC-producing isolates, 36 (60%) were E. coli. Amongst these isolates, 27/60 (45%) were classified as multidrug resistant, compared with 15/55 (27%) non-ESBL or AmpC-producing isolates (p<0.01). Ninety five isolates were resistant to amoxicillin-clavulanic acid and 58 (61%) of these were ESBL or AmpC-producing. The predominant ESBL genes were bla CTX-M-14 and bla CTX-M-15 , and the dominant plasmid-encoded AmpC gene was bla CMY-2 . Thirty-eight E. coli multilocus sequence types (ST) were identified, and the most prevalent were ST12 (12/89, 13%), ST131 (6/89, 7%) and ST648 (6/89, 7%). ESBL and AmpC-producing isolates accounted for 35/1,082 (3.2%) of the Enterobacteriaceae isolated by one laboratory network over the study period. ESBL and AmpC-producing Enterobacteriaceae were associated with clinical infections in companion animals in New Zealand, and were often multidrug resistant. In this study, these organisms accounted for <5% of all Enterobacteriaceae isolated from infection sites by one laboratory network, but their prevalence among isolates resistant to amoxicillin-clavulanic acid was 61%. Therefore routine secondary testing for ESBL and AmpC production by Enterobacteriaceae that are resistant to amoxicillin-clavulanic acid in primary testing could improve the accuracy of definitive antimicrobial therapy in companion animals in New Zealand.

Rapid O serogroup identification of the ten most clinically relevant STECs by Luminex microbead-based suspension array

USDA-ARS?s Scientific Manuscript database

Identification and serotyping of Shiga toxin-producing Escherichia coli during foodborne outbreaks can aid in matching clinical, food, and environmental isolates when trying to identify the sources of illness and ultimately food contamination. Herein we describe a Luminex microbead-based suspension ...

Isolation Rearing Effects on Probabilistic Learning and Cognitive Flexibility in Rats

PubMed Central

AMITAI, Nurith; YOUNG, Jared W.; HIGA, Kerin; SHARP, Richard F.; GEYER, Mark A.; POWELL, Susan B.

2013-01-01

Isolation rearing is a neurodevelopmental manipulation that produces neurochemical, structural, and behavioral alterations in rodents that have consistencies with schizophrenia. Symptoms induced by isolation rearing that mirror clinically relevant aspects of schizophrenia, such as cognitive deficits, open up the possibility of testing putative therapeutics in isolation-reared animals prior to clinical development. We investigated what effect isolation rearing would have on cognitive flexibility, a cognitive function characteristically disrupted in schizophrenia. For this purpose, we assessed cognitive flexibility using between- and within-session probabilistic reversal-learning tasks based on clinical tests. Isolation-reared rats required more sessions, though not more task trials, to acquire criterion performance in the reversal phase of the task and were slower to adjust their task strategy after reward contingencies were switched. Isolation-reared rats also completed fewer trials and exhibited lower levels of overall activity in the probabilistic reversal-learning task compared to socially reared rats. This finding contrasted with the elevated levels of unconditioned investigatory activity and reduced levels of locomotor habituation that isolation-reared rats displayed in the behavioral pattern monitor. Finally, isolation-reared rats also exhibited sensorimotor gating deficits, reflected by decreased prepulse inhibition of the startle response, consistent with previous studies. We conclude that isolation rearing constitutes a valuable, noninvasive manipulation for modeling schizophrenia-like cognitive deficits and assessing putative therapeutics. PMID:23943516

Determination of Isavuconazole Susceptibility of Aspergillus and Candida Species by the EUCAST Method

PubMed Central

Howard, Susan J.; Lass-Flörl, Cornelia; Cuenca-Estrella, Manuel; Gomez-Lopez, Alicia

2013-01-01

Isavuconazole is a novel expanded-spectrum triazole, which has recently been approved by the FDA as an orphan drug to treat invasive aspergillosis and is currently being studied in phase III clinical trials for invasive candidiasis. The susceptibility of relatively few clinical isolates has been reported. In this study, the isavuconazole susceptibilities of 1,237 Aspergillus and 2,010 Candida geographically diverse clinical isolates were determined by EUCAST methodology at four European mycology laboratories, producing the largest multicenter data set thus far for this compound. In addition, a blinded collection of 30 cyp51A mutant Aspergillus fumigatus clinical isolates and 10 wild-type isolates was tested. From these two data sets, the following preliminary epidemiological cutoff (ECOFF) values were suggested: 2 mg/liter for Aspergillus fumigatus, Aspergillus terreus, and Aspergillus flavus; 4 mg/liter for Aspergillus niger; 0.25 mg/liter for Aspergillus nidulans; and 0.03 mg/liter for Candida albicans, Candida parapsilosis, and Candida tropicalis. Unfortunately, ECOFFs could not be determined for Candida glabrata or Candida krusei due to an unexplained interlaboratory MIC variation. For the blinded collection of A. fumigatus isolates, all MICs were ≤2 mg/liter for wild-type isolates. Differential isavuconazole MICs were observed for triazole-resistant A. fumigatus isolates with different cyp51A alterations: TR34/L98H mutants had elevated isavuconazole MICs, whereas isolates with G54 and M220 alterations had MICs in the wild-type range, suggesting that the efficacy of isavuconazole may not be affected by these alterations. This study will be an aid in interpreting isavuconazole MICs for clinical care and an important step in the future process of setting official clinical breakpoints. PMID:23959309

High prevalence of extended-spectrum ß-lactamase producing enterobacteriaceae among clinical isolates in Burkina Faso.

PubMed

Ouedraogo, Abdoul-Salam; Sanou, Mahamadou; Kissou, Aimée; Sanou, Soufiane; Solaré, Hermann; Kaboré, Firmin; Poda, Armel; Aberkane, Salim; Bouzinbi, Nicolas; Sano, Idrissa; Nacro, Boubacar; Sangaré, Lassana; Carrière, Christian; Decré, Dominique; Ouégraogo, Rasmata; Jean-Pierre, Hélène; Godreuil, Sylvain

2016-07-11

Nothing is known about the epidemiology and resistance mechanisms of extended-spectrum ß-lactamase-producing Enterobacteriaceae (ESBL-PE) in Burkina Faso. The objective of this study was to determine ESBL-PE prevalence and to characterize ESBL genes in Burkina Faso. During 2 months (June-July 2014), 1602 clinical samples were sent for bacteriologic investigations to the microbiology laboratories of the tree main hospitals of Burkina Faso. Isolates were identified by mass spectrometry using a matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) BioTyper. Antibiotic susceptibility was tested using the disk diffusion method on Müller-Hinton agar. The different ESBL genes in potential ESBL-producing isolates were detected by PCR and double stranded DNA sequencing. Escherichia coli phylogenetic groups were determined using a PCR-based method. ESBL-PE frequency was 58 % (179 strains among the 308 Enterobacteriaceae isolates identified in the collected samples; 45 % in outpatients and 70 % in hospitalized patients). The CTX-M-1 group was dominant (94 %, CTX-M-15 enzyme), followed by the CTX-M-9 group (4 %). ESBL producers were more often found in E. coli (67.5 %) and Klebsiella pneumoniae (26 %) isolates. E. coli isolates (n = 202; 60 % of all Enterobacteriaceae samples) were distributed in eight phylogenetic groups (A = 49, B1 = 15, B2 = 43, C = 22, Clade I = 7, D = 37, F = 13 and 16 unknown); 22 strains belonged to the sequence type ST131. No association between a specific strain and ESBL production was detected. This report shows the alarming spread of ESBL genes in Burkina Faso. Public health efforts should focus on education (population and healthcare professionals), surveillance and promotion of correct and restricted antibiotic use to limit their dissemination.

Genetic diversity of clinical and environmental isolates of Vibrio cholerae determined by amplified fragment length polymorphism fingerprinting.

PubMed

Jiang, S C; Matte, M; Matte, G; Huq, A; Colwell, R R

2000-01-01

Vibrio cholerae, the causative agent of major epidemics of diarrheal disease in Bangladesh, South America, Southeastern Asia, and Africa, was isolated from clinical samples and from aquatic environments during and between epidemics over the past 20 years. To determine the evolutionary relationships and molecular diversity of these strains, in order to understand sources, origin, and epidemiology, a novel DNA fingerprinting technique, amplified fragment length polymorphism (AFLP), was employed. Two sets of restriction enzyme-primer combinations were tested for fingerprinting of V. cholerae serogroup O1, O139, and non-O1, O139 isolates. Amplification of HindIII- and TaqI-digested genomic DNA produced 30 to 50 bands for each strain. However, this combination, although capable of separating environmental isolates of O1 and non-O1 strains, was unable to distinguish between O1 and O139 clinical strains. This result confirmed that clinical O1 and O139 strains are genetically closely related. On the other hand, AFLP analyses of restriction enzyme ApaI- and TaqI-digested genomic DNA yielded 20 to 30 bands for each strain, but were able to separate O1 from O139 strains. Of the 74 strains examined with the latter combination, 26 serogroup O1 strains showed identical banding patterns and were represented by the O1 El Tor strain of the seventh pandemic. A second group, represented by O139 Bengal, included 12 strains of O139 clinical isolates, with 7 from Thailand, 3 from Bangladesh, and 2 from India. Interestingly, an O1 clinical isolate from Africa also grouped with the O139 clinical isolates. Eight clinical O1 isolates from Mexico grouped separately from the O1 El Tor of the seventh pandemic, suggesting an independent origin of these isolates. Identical fingerprints were observed between an O1 environmental isolate from a river in Chile and an O1 clinical strain from Kenya, both isolated more than 10 years apart. Both strains were distinct from the O1 seventh pandemic strain. Two O139 clinical isolates from Africa clustered with environmental non-O1 isolates, independent of other O139 strains included in the study. These results suggest that although a single clone of pathogenic V. cholerae appears responsible for many cases of cholera in Asia, Africa, and Latin America during the seventh pandemic, other cases of clinical cholera were caused by toxigenic V. cholerae strains that appear to have been derived locally from environmental O1 or non-O1 strains.

Isolation and genetic characterization of metallo-β-lactamase and carbapenamase producing strains of Acinetobacter baumannii from patients at Tehran hospitals

PubMed Central

Shahcheraghi, F; Abbasalipour, M; Feizabadi, MM; Ebrahimipour, GH; Akbari, N

2011-01-01

Background and Objective Carbapenems are therapeutic choice against infections caused by gram-negative bacilli including strains of Acinetobacter baumannii. Resistance to these antibiotics is mediated by efflux pumps, porins, PBPs and ß-lactamases. The aim of this study was to determine the possibility of existence of MBLs, OXAs and GES-1 betalactamase genes among clinical isolates of Acinetobacter collected from Tehran hospitals. Material and Methods Two hundred and three Acinetobacter isolates were collected from patient at Tehran hospitals. The isolates were identified using biochemical tests. The susceptibility to different antibiotics was evaluated by disk diffusion method and MICs of imipenem were determined using Micro broth dilution method (CLSI). PCR was performed for detection of bla VIM-2, bla SPM-1, bla IMP-2, bla GES-1, bla OXA-51, bla OXA-23 betalactamase genes. Clonal relatedness was estimated by PFGE with the restriction enzyme SmaI. Results and Conclusion Of 100 isolates of imipenem resistant Acinetobacter spp. collected from Tehran hospitals in 2009 and 2010, 6 isolates produced metallo-beta-lactamases and 94 isolates produced OXA-type carbapenemase. The bla SPM-1, bla GES-1, bla OXA-51, bla OXA-23 genes were detected by PCR among 6, 2, 94 and 84 isolates of A. baumannii, respectively. The MICs of isolates to imipenem were 8–128 µg/mL. PFGE analysis of 29 bla OXA-51 and bla OXA-23-positive A. baumannii isolates gave 6 different patterns. This is the first report of SPM-1 and GES-1 beta-lactamase producing A. baumannii. Production of the OXA-23, OXA-51, GES-1 and SPM-1 enzyme presents an emerging threat of carbapenem resistance among A. baumannii in Iran. PMID:22347585

Fluoroquinolone-resistant and extended-spectrum β-lactamase-producing Escherichia coli from the milk of cows with clinical mastitis in Southern Taiwan.

PubMed

Su, Yaochi; Yu, Chang-You; Tsai, Yilin; Wang, Shao-Hung; Lee, Chihan; Chu, Chishih

2016-12-01

Escherichia coli is a common pathogen to cause clinical and subclinical mastitis in cows. A total of 57 E. coli isolates from raw milk from cows were characterized genetically and biochemically. Extended-spectrum β-lactamase (ESBL) genes, the mechanism for fluoroquinolone resistance, and variations in virulence genes and genomes of these E. coli isolates were investigated by the antimicrobial susceptibility test, simplex and multiplex polymerase chain reaction (PCR), and pulsed-field gel electrophoresis (PFGE). All E. coli isolates were resistant to cloxacillin (100%) and to a lesser extent (50%) to tetracycline, neomycin, gentamycin, ampicillin, ceftriaxone, cefotaxime (CTX), and ceftazidime (CAZ). Nearly 70% of the isolates were resistant to at least two antimicrobials and 28.1% carried AmpA and AmpC genes simultaneously. The predominant bla gene was bla TEM , followed by bla CMY , bla CTX , bla SHV , and bla DHA. Among the six (10.5%) ESBL-producing E. coli carrying bla CTX-M15 , bla CTX-M55 , or bla CTX-M14 , two isolates 31 of ST410 in the ST23 complex and 58 of ST167 in the ST10 complex were also resistant to ciprofloxacin, enrofloxacin, and levofloxacin, with mutations at codon 83 from serine to leucine and codon 87 from aspartic acid to asparagine in GyrA and at codon 80 from serine to isoleucine in ParC. These isolates were genetically diverse in pulsotype analysis, lacked toxin genes of human pathogenic E. coli and carried mostly the prevalent virulence genes fimH, papGII, and α-hemolysin. Lacking virulence genes examined, genetic diverse E. coli isolates are unrelated to human pathogenic E. coli. Enhancing sanitation in milk processing and transportation is needed to eliminate multidrug-resistant (MDR), fluoroquinolone-resistant, and ESBL-producing E. coli isolates. Copyright © 2014. Published by Elsevier B.V.

Identical plasmid AmpC beta-lactamase genes and plasmid types in E. coli isolates from patients and poultry meat in the Netherlands.

PubMed

Voets, Guido M; Fluit, Ad C; Scharringa, Jelle; Schapendonk, Claudia; van den Munckhof, Thijs; Leverstein-van Hall, Maurine A; Stuart, James Cohen

2013-11-01

The increasing prevalence of third-generation cephalosporin-resistant Enterobacteriaceae is a worldwide problem. Recent studies showed that poultry meat and humans share identical Extended-Spectrum Beta-Lactamase genes, plasmid types, and Escherichia coli strain types, suggesting that transmission from poultry meat to humans may occur. The aim of this study was to compare plasmid-encoded Ambler class C beta-lactamase (pAmpC) genes, their plasmids, and bacterial strain types between E. coli isolates from retail chicken meat and clinical isolates in the Netherlands. In total, 98 Dutch retail chicken meat samples and 479 third-generation cephalosporin non-susceptible human clinical E. coli isolates from the same period were screened for pAmpC production. Plasmid typing was performed using PCR-based replicon typing (PBRT). E coli strains were compared using Multi-Locus-Sequence-Typing (MLST). In 12 of 98 chicken meat samples (12%), pAmpC producing E. coli were detected (all blaCMY-2). Of the 479 human E. coli, 25 (5.2%) harboured pAmpC genes (blaCMY-2 n = 22, blaACT n = 2, blaMIR n = 1). PBRT showed that 91% of poultry meat isolates harboured blaCMY-2 on an IncK plasmid, and 9% on an IncI1 plasmid. Of the human blaCMY-2 producing isolates, 42% also harboured blaCMY-2 on an IncK plasmid, and 47% on an IncI1 plasmid. Thus, 68% of human pAmpC producing E. coli have the same AmpC gene (blaCMY-2) and plasmid type (IncI1 or IncK) as found in poultry meat. MLST showed one cluster containing one human isolate and three meat isolates, with an IncK plasmid. These findings imply that a foodborne transmission route of blaCMY-2 harbouring plasmids cannot be excluded and that further evaluation is required. © 2013.

Genetic characterization of Shiga toxin-producing Escherichia coli O26:H11 strains isolated from animal, food, and clinical samples

PubMed Central

Krüger, Alejandra; Lucchesi, Paula M. A.; Sanso, A. Mariel; Etcheverría, Analía I.; Bustamante, Ana V.; Burgán, Julia; Fernández, Luciana; Fernández, Daniel; Leotta, Gerardo; Friedrich, Alexander W.; Padola, Nora L.; Rossen, John W. A.

2015-01-01

The Shiga-toxin producing Escherichia coli (STEC) may cause serious illness in human. Here we analyze O26:H11 strains known to be among the most reported STEC strains causing human infections. Genetic characterization of strains isolated from animal, food, and clinical specimens in Argentina showed that most carried either stx1a or stx2a subtypes. Interestingly, stx2a-positive O26:H11 rarely isolated from cattle in other countries showed to be an important proportion of O26:H11 strains circulating in cattle and food in our region. Seventeen percent of the isolates harbored more than one gene associated with antimicrobial resistance. In addition to stx, all strains contained the virulence genes eae-β, tir, efa, iha, espB, cif, espA, espF, espJ, nleA, nleB, nleC, and iss; and all except one contained ehxA, espP, and cba genes. On the other hand, toxB and espI genes were exclusively observed in stx2-positive isolates, whereas katP was only found in stx1a-positive isolates. Our results show that O26:H11 STEC strains circulating in Argentina, including those isolated from humans, cattle, and meat products, present a high pathogenic potential, and evidence that cattle can be a reservoir of O26:H11 strains harboring stx2a. PMID:26539413

Genetic characterization of Shiga toxin-producing Escherichia coli O26:H11 strains isolated from animal, food, and clinical samples.

PubMed

Krüger, Alejandra; Lucchesi, Paula M A; Sanso, A Mariel; Etcheverría, Analía I; Bustamante, Ana V; Burgán, Julia; Fernández, Luciana; Fernández, Daniel; Leotta, Gerardo; Friedrich, Alexander W; Padola, Nora L; Rossen, John W A

2015-01-01

The Shiga-toxin producing Escherichia coli (STEC) may cause serious illness in human. Here we analyze O26:H11 strains known to be among the most reported STEC strains causing human infections. Genetic characterization of strains isolated from animal, food, and clinical specimens in Argentina showed that most carried either stx 1a or stx 2a subtypes. Interestingly, stx 2a-positive O26:H11 rarely isolated from cattle in other countries showed to be an important proportion of O26:H11 strains circulating in cattle and food in our region. Seventeen percent of the isolates harbored more than one gene associated with antimicrobial resistance. In addition to stx, all strains contained the virulence genes eae-β, tir, efa, iha, espB, cif, espA, espF, espJ, nleA, nleB, nleC, and iss; and all except one contained ehxA, espP, and cba genes. On the other hand, toxB and espI genes were exclusively observed in stx 2-positive isolates, whereas katP was only found in stx 1a-positive isolates. Our results show that O26:H11 STEC strains circulating in Argentina, including those isolated from humans, cattle, and meat products, present a high pathogenic potential, and evidence that cattle can be a reservoir of O26:H11 strains harboring stx 2a.

In vitro antibacterial activity of rifampicin in combination with imipenem, meropenem and doripenem against multidrug-resistant clinical isolates of Pseudomonas aeruginosa.

PubMed

Hu, Yi-Fan; Liu, Chang-Pan; Wang, Nai-Yu; Shih, Shou-Chuan

2016-08-24

Multidrug-resistant Pseudomonas aeruginosa has emerged as one of the most important healthcare-associated pathogens. Colistin is regarded as the last-resort antibiotic for multidrug-resistant Gram-negative bacteria, but is associated with high rates of acute kidney injury. The aim of this in vitro study is to search for an alternative treatment to colistin for multidrug-resistant P. aeruginosa infections. Multidrug and carbapenem-resistant P. aeruginosa isolates were collected between January 2009 and December 2012 at MacKay Memorial Hospital. Minimal inhibitory concentrations (MICs) were determined for various antibiotic combinations. Carbapenemase-producing genes including bla VIM, other β-lactamase genes and porin mutations were screened by PCR and sequencing. The efficacy of carbapenems (imipenem, meropenem, doripenem) with or without rifampicin was correlated with the type of porin mutation (frameshift mutation, premature stop codon mutation) in multidrug-resistant P. aeruginosa isolates without carbapenemase-producing genes. Of the 71 multidrug-resistant clinical P. aeruginosa isolates, only six harboured the bla VIM gene. Imipenem, meropenem and doripenem were significantly more effective (reduced fold-change of MICs) when combined with rifampicin in bla VIM-negative isolates, especially in isolates with porin frameshift mutation. Imipenem + rifampicin combination has a low MIC against multidrug-resistant P. aeruginosa, especially in isolates with porin frameshift mutation. The imipenem + rifampicin combination may provide an alternative treatment to colistin for multidrug -resistant P. aeruginosa infections, especially for patients with renal insufficiency.

Potentiating antibiotics in drug-resistant clinical isolates via stimuli-activated superoxide generation.

PubMed

Courtney, Colleen M; Goodman, Samuel M; Nagy, Toni A; Levy, Max; Bhusal, Pallavi; Madinger, Nancy E; Detweiler, Corrella S; Nagpal, Prashant; Chatterjee, Anushree

2017-10-01

The rise of multidrug-resistant (MDR) bacteria is a growing concern to global health and is exacerbated by the lack of new antibiotics. To treat already pervasive MDR infections, new classes of antibiotics or antibiotic adjuvants are needed. Reactive oxygen species (ROS) have been shown to play a role during antibacterial action; however, it is not yet understood whether ROS contribute directly to or are an outcome of bacterial lethality caused by antibiotics. We show that a light-activated nanoparticle, designed to produce tunable flux of specific ROS, superoxide, potentiates the activity of antibiotics in clinical MDR isolates of Escherichia coli , Salmonella enterica , and Klebsiella pneumoniae . Despite the high degree of antibiotic resistance in these isolates, we observed a synergistic interaction between both bactericidal and bacteriostatic antibiotics with varied mechanisms of action and our superoxide-producing nanoparticles in more than 75% of combinations. As a result of this potentiation, the effective antibiotic concentration of the clinical isolates was reduced up to 1000-fold below their respective sensitive/resistant breakpoint. Further, superoxide-generating nanoparticles in combination with ciprofloxacin reduced bacterial load in epithelial cells infected with S. enterica serovar Typhimurium and increased Caenorhabditis elegans survival upon infection with S. enterica serovar Enteriditis, compared to antibiotic alone. This demonstration highlights the ability to engineer superoxide generation to potentiate antibiotic activity and combat highly drug-resistant bacterial pathogens.

Prevalence of 16S rRNA methylase genes in Klebsiella pneumoniae isolates from a Chinese teaching hospital: coexistence of rmtB and armA genes in the same isolate.

PubMed

Yu, Fangyou; Wang, Liangxing; Pan, Jingye; Yao, Dan; Chen, Chan; Zhu, Tao; Lou, Qiang; Hu, Jian; Wu, Yang; Zhang, Xueqing; Chen, Zengqiang; Qu, Di

2009-05-01

16S rRNA methylase-mediated high-level resistance to aminoglycosides has been reported recently in clinical isolates of Gram-negative bacilli from several countries. Twenty-one (6.2%, 21/337) of 337 isolates of Klebsiella pneumoniae from a teaching hospital in Wenzhou, China, were positive for 16S rRNA methylase genes (3 for armA, 13 for rmtB, 5 for both armA and rmtB) and highly resistant to gentamicin, amikacin, and tobramycin (MICs, > or =256 microg/mL). Nineteen of 21 isolates harboring 16S rRNA methyalse genes were extended-spectrum beta-lactamase (ESBL) producers. The plasmids harboring 16S rRNA methylase genes from 14 of 21 donors were transferred into the recipients, Escherichia coli J53. The armA and the rmtB usually coexisted with ESBL genes in the same isolate in clinical isolates and cotransferred with ESBL genes on a self-transmissible conjugative plasmid to the recipients. Among 5 isolates harboring both armA and rmtB, the armA genes were located on the chromosomes, and the rmtB genes were located on the plasmids, as determined by Southern hybridization. The result of pulsed-field gel electrophoresis showed that horizontal gene transfer and clonal spread were responsible for the dissemination of the rmtB and the armA genes. 16S rRNA methylase-producing isolates of Klebsiella pneumoniae were commonly identified in the Chinese teaching hospital with coexistence of rmtB and armA genes in the same isolate.

Prevalence of Clinically Isolated Metallo-beta-lactamase-producing Pseudomonas aeruginosa, Coding Genes, and Possible Risk Factors in Iran

PubMed Central

Ghasemian, Abdolmajid; Salimian Rizi, Kobra; Rajabi Vardanjani, Hassan; Nojoomi, Farshad

2018-01-01

Background & Objective: The spread of carbapenem-resistant Pseudomonas aeruginosa is a global concern. Metallo-beta-lactamase (MBL) enzymes cause extensive drug resistance among Gram-negative bacteria. The current study aimed at determining the prevalence of MBL-producing P. aeruginosa in Iran. Data extraction: A total of 43 studies were found out of which 36 were adopted. Data were collected from Google, Google Scholar, Science Direct, PubMed, Scopus, Embase, and Sciverse. The terms “Pseudomonas aeruginosa”, “metallo-beta-lactamase”, “prevalence”, “carbapenems”, and “Iran” were searched. Data from the isolates not producing MBLs were excluded from the study. Data were analyzed with Graph Pad Prism 6, meta-analysis section. Results: According to the results of the current study, 36 surveys indicated that 55% of the clinically isolated P. aeruginosa in Iran were resistant to imipenem and meropenem, among which 37.72% were the MBL producers. Among genes encoding MBLs, blaVIM and blaIMP were predominant with the prevalence of 12.91%±11.01% and 12.50%±23.56%, respectively. No report of harboring blaNDM1 and blaSPM1 by P. aeruginosa was found, similar to most of the other countries in Asia. The prevalence of blaVIM and blaIMP from burn settings were 11.50%±3.5% and 24.65%±23%, respectively. Furthermore, the prevalence of these genes was not significantly different among burn and non-burn isolates (P=0.942 and P=0.597, respectively). Moreover, no relationship was observed between the MBL production and patients’ age range. Conclusion: Approximately half of P. aeruginosa isolates were carbapenem-resistant in Iran, and approximately half were the MBL producers. The blaVIM and blaIMP were the predominant MBLs among P. aeruginosa strains, while other genes were not found in P. aeruginosa. Moreover, there was no significant difference between blaVIM and blaIMP among burn and non-burn isolates. Due to the multiple drug resistance conferred by MBLs, detection and control of their spread alongside proper therapeutic regimens in hospitals and community settings are essential to prevent infection acquisition. PMID:29731790

Prevalence of Clinically Isolated Metallo-beta-lactamase-producing Pseudomonas aeruginosa, Coding Genes, and Possible Risk Factors in Iran.

PubMed

Ghasemian, Abdolmajid; Salimian Rizi, Kobra; Rajabi Vardanjani, Hassan; Nojoomi, Farshad

2018-01-01

The spread of carbapenem-resistant Pseudomonas aeruginosa is a global concern. Metallo-beta-lactamase (MBL) enzymes cause extensive drug resistance among Gram-negative bacteria. The current study aimed at determining the prevalence of MBL-producing P. aeruginosa in Iran. A total of 43 studies were found out of which 36 were adopted. Data were collected from Google, Google Scholar, Science Direct, PubMed, Scopus, Embase, and Sciverse. The terms " Pseudomonas aeruginosa ", "metallo-beta-lactamase", "prevalence", "carbapenems", and "Iran" were searched. Data from the isolates not producing MBLs were excluded from the study. Data were analyzed with Graph Pad Prism 6, meta-analysis section. According to the results of the current study, 36 surveys indicated that 55% of the clinically isolated P. aeruginosa in Iran were resistant to imipenem and meropenem, among which 37.72% were the MBL producers. Among genes encoding MBLs, bla VIM and bla IMP were predominant with the prevalence of 12.91%±11.01% and 12.50%±23.56%, respectively. No report of harboring bla NDM1 and bla SPM1 by P. aeruginosa was found, similar to most of the other countries in Asia. The prevalence of bla VIM and bla IMP from burn settings were 11.50%±3.5% and 24.65%±23%, respectively. Furthermore, the prevalence of these genes was not significantly different among burn and non-burn isolates (P=0.942 and P=0.597, respectively). Moreover, no relationship was observed between the MBL production and patients' age range. Approximately half of P. aeruginosa isolates were carbapenem-resistant in Iran, and approximately half were the MBL producers. The bla VIM and bla IMP were the predominant MBLs among P . aeruginosa strains, while other genes were not found in P. aeruginosa . Moreover, there was no significant difference between bla VIM and bla IMP among burn and non-burn isolates. Due to the multiple drug resistance conferred by MBLs, detection and control of their spread alongside proper therapeutic regimens in hospitals and community settings are essential to prevent infection acquisition.

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.

Exopolysaccharides produced by a clinical strain of Burkholderia cepacia isolated from a cystic fibrosis patient.

PubMed

Cescutti, Paola; Impallomeni, Giuseppe; Garozzo, Domenico; Sturiale, Luisa; Herasimenka, Yury; Lagatolla, Cristina; Rizzo, Roberto

2003-11-14

Burkholderia cepacia is an opportunistic pathogen involved in pulmonary infections related to cystic fibrosis. A clinical strain, BTS13, was isolated and the production of exopolysaccharides was tested growing the bacteria on two different media, one of which was rich in mannitol as carbon source. The primary structure of the polysaccharides was determined using mostly mass spectrometry and NMR spectroscopy. On both media an exopolysaccharide having the following repeating unit was produced: -->5)-beta-Kdop-(2-->3)-beta-D-Galp2Ac-(1-->4)-alpha-D-Galp-(1-->3)-beta-D-Galp-(1--> This polysaccharide has already been described as the biosynthetic product of another Burkholderia species, B. pseudomallei, the microbial agent causing melioidosis. In addition to this, when grown on the mannitol-rich medium, B. cepacia strain BTS13 produced another polysaccharide that was established to be levan: -->6)-beta-D-Fruf-(2-->. The content of levan was about 20% (w/w) of the total amount of polymers. The ability of B. cepacia to produce these two exopolysaccharides opens new perspectives in the investigation of the role of polysaccharides in lung infections.

Assessing the genome level diversity of Listeria monocytogenes from contaminated ice cream and environmental samples linked to a listeriosis outbreak in the United States.

PubMed

Chen, Yi; Luo, Yan; Curry, Phillip; Timme, Ruth; Melka, David; Doyle, Matthew; Parish, Mickey; Hammack, Thomas S; Allard, Marc W; Brown, Eric W; Strain, Errol A

2017-01-01

A listeriosis outbreak in the United States implicated contaminated ice cream produced by one company, which operated 3 facilities. We performed single nucleotide polymorphism (SNP)-based whole genome sequencing (WGS) analysis on Listeria monocytogenes from food, environmental and clinical sources, identifying two clusters and a single branch, belonging to PCR serogroup IIb and genetic lineage I. WGS Cluster I, representing one outbreak strain, contained 82 food and environmental isolates from Facility I and 4 clinical isolates. These isolates differed by up to 29 SNPs, exhibited 9 pulsed-field gel electrophoresis (PFGE) profiles and multilocus sequence typing (MLST) sequence type (ST) 5 of clonal complex 5 (CC5). WGS Cluster II contained 51 food and environmental isolates from Facility II, 4 food isolates from Facility I and 5 clinical isolates. Among them the isolates from Facility II and clinical isolates formed a clade and represented another outbreak strain. Isolates in this clade differed by up to 29 SNPs, exhibited 3 PFGE profiles and ST5. The only isolate collected from Facility III belonged to singleton ST489, which was in a single branch separate from Clusters I and II, and was not associated with the outbreak. WGS analyses clustered together outbreak-associated isolates exhibiting multiple PFGE profiles, while differentiating them from epidemiologically unrelated isolates that exhibited outbreak PFGE profiles. The complete genome of a Cluster I isolate allowed the identification and analyses of putative prophages, revealing that Cluster I isolates differed by the gain or loss of three putative prophages, causing the banding pattern differences among all 3 AscI-PFGE profiles observed in Cluster I isolates. WGS data suggested that certain ice cream varieties and/or production lines might have contamination sources unique to them. The SNP-based analysis was able to distinguish CC5 as a group from non-CC5 isolates and differentiate among CC5 isolates from different outbreaks/incidents.

Assessing the genome level diversity of Listeria monocytogenes from contaminated ice cream and environmental samples linked to a listeriosis outbreak in the United States

PubMed Central

Chen, Yi; Luo, Yan; Curry, Phillip; Timme, Ruth; Melka, David; Doyle, Matthew; Parish, Mickey; Hammack, Thomas S.; Allard, Marc W.; Brown, Eric W.; Strain, Errol A.

2017-01-01

A listeriosis outbreak in the United States implicated contaminated ice cream produced by one company, which operated 3 facilities. We performed single nucleotide polymorphism (SNP)-based whole genome sequencing (WGS) analysis on Listeria monocytogenes from food, environmental and clinical sources, identifying two clusters and a single branch, belonging to PCR serogroup IIb and genetic lineage I. WGS Cluster I, representing one outbreak strain, contained 82 food and environmental isolates from Facility I and 4 clinical isolates. These isolates differed by up to 29 SNPs, exhibited 9 pulsed-field gel electrophoresis (PFGE) profiles and multilocus sequence typing (MLST) sequence type (ST) 5 of clonal complex 5 (CC5). WGS Cluster II contained 51 food and environmental isolates from Facility II, 4 food isolates from Facility I and 5 clinical isolates. Among them the isolates from Facility II and clinical isolates formed a clade and represented another outbreak strain. Isolates in this clade differed by up to 29 SNPs, exhibited 3 PFGE profiles and ST5. The only isolate collected from Facility III belonged to singleton ST489, which was in a single branch separate from Clusters I and II, and was not associated with the outbreak. WGS analyses clustered together outbreak-associated isolates exhibiting multiple PFGE profiles, while differentiating them from epidemiologically unrelated isolates that exhibited outbreak PFGE profiles. The complete genome of a Cluster I isolate allowed the identification and analyses of putative prophages, revealing that Cluster I isolates differed by the gain or loss of three putative prophages, causing the banding pattern differences among all 3 AscI-PFGE profiles observed in Cluster I isolates. WGS data suggested that certain ice cream varieties and/or production lines might have contamination sources unique to them. The SNP-based analysis was able to distinguish CC5 as a group from non-CC5 isolates and differentiate among CC5 isolates from different outbreaks/incidents. PMID:28166293

Expanded spectrum β-lactamase producing Escherichia coli isolated from chickens with colibacillosis in Egypt.

PubMed

El-Shazly, D A; Nasef, S A; Mahmoud, F F; Jonas, Daniel

2017-07-01

Throughout the world, expanded spectrum β-lactamases (ESBL) are increasing among clinical isolates of Enterobacteriaceae, both in humans and animals. Unfortunately, there is a paucity of data on ESBL or Ampicillin class C β-lactamase (AmpC) in Egypt, although antimicrobial consumption is high in this developing country. This study aims to characterize the resistance mechanisms to expanded spectrum cephalosporins among resistant veterinary Escherichia coli isolates in Egypt. We investigated 50 clinical multi-resistant E. coli strains isolated from 20 chicken farms for production of ESBL or AmpC. Antibiotic susceptibility was tested by Clinical and Laboratory Standards Institute (CLSI) disk diffusion and ESBL confirmatory tests. PCR and sequencing were performed to screen for plasmid mediated ESBL genes and genes encoding AmpC β-lactamases. All the isolates were phylogentically classified, investigated for harboring class 1 integrons, and genotyped by amplified fragment length polymorphism (AFLP). Three strains showed ESBL and 6 strains AmpC phenotypic patterns, respectively, with confirmed ESBL genes of blaTEM-57, blaSHV-12, blaCTX-M-14, and blaCMY-2 for AmpC producing strains. All ESBL strains belonged to phylogroup D with different clones isolated from different flocks, while most of the AmpC strains belonged to phylogroup B1 (4/6) and were assigned to the same genotype distributed in 2 different farms. Class 1 integrons were disseminated in 60% of all tested strains and in 100% of ESBL and AmpC strains. These results highlight the antimicrobial resistance problem in Egypt, caused in all probability by unwise use of antimicrobials in animal husbandry. The results call for a nationwide surveillance program to monitor antimicrobial resistance. © 2017 Poultry Science Association Inc.

Bacteria isolated from companion animals in Japan (2014-2016) by blood culture.

PubMed

Tsuyuki, Yuzo; Kurita, Goro; Murata, Yoshiteru; Takahashi, Takashi

2018-02-24

We aimed to identify microorganisms isolated by blood culture (BC) from companion animals and to determine antimicrobial resistance of these isolates during 2014-2016 at veterinary laboratory, in comparison with those during 2010-2013, in Japan. Clinical data (animal species, visiting animals/hospitalized animals, and others except for disease type and clinical course including history of antimicrobial agent use) on ill animals at veterinary clinics or hospitals were obtained. We retrospectively analyzed animal-origin BC results extracted from the database in 2014-2016 and those obtained in 2010-2013. BC-positive samples were from most of dogs (n = 174 in 2014-2016 and n = 86 in 2010-2013). Escherichia coli (n = 50, 25.1%) and Staphylococcus intermedius group (SIG) bacteria (n = 23, 11.6%) were most prevalent in 2014-2016, while the percentages of E. coli (n = 22, 25.3%) and SIG (n = 9, 10.3%) in 2010-2013 were similar to those in 2014-2016. Percentages of extended-spectrum β-lactamase (ESBL)-producing E. coli and methicillin-resistant staphylococci (MRS) rate of SIG bacteria isolated in 2014-2016 were 28.0% and 69.6% (vs. 22.7% and 44.4% in 2010-2013), respectively. Fourteen ESBL-producing E. coli in 2014-2016 were isolated from 7 visiting animals and 7 hospitalized ones, whereas the sixteen MRS of SIG were from 7 visiting animals and 9 hospitalized ones. Our observations support the prevalent microorganisms isolated by BC and their antimicrobial resistance patterns for two study periods. Copyright © 2018 Japanese Society of Chemotherapy and The Japanese Association for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

Typing clinical and animal environment Aspergillus fumigatus gliotoxin producer strains isolated from Brazil by PCR-RFLP markers.

PubMed

Soleiro, C A; Pena, G A; Cavaglieri, L R; Coelho, I; Keller, L M; Dalcero, A M; Rosa, C A R

2013-12-01

Aspergillus fumigatus, a well-known human and animal pathogen causing aspergillosis, has been historically identified by morphological and microscopic features. However, recent studies have shown that species identification on the basis of morphology alone is problematic. The aim of this work was to confirm the taxonomic state at specie level of a set of clinical (human and animal) and animal environment A. fumigatus strains identified by morphological criteria applying a PCR-RFLP assay by an in silico and in situ analysis with three restriction enzymes. The A. fumigatus gliotoxin-producing ability was also determined. Previous to the in situ PCR-RFLP analysis, an in silico assay with BccI, MspI and Sau3AI restriction enzymes was carried out. After that, these enzymes were used for in situ assay. All A. fumigatus strains isolated from corn silage, human aspergillosis and bovine mastitis and high per cent of the strains isolated from cereals, animal feedstuff and sorghum silage were able to produce high gliotoxin levels. Also, all these strains identified by morphological criteria as A. fumigatus, regardless of its isolation source, had band patterns according to A. fumigatus sensu stricto by PCR-RFLP markers. Aspergillus fumigatus is a well-known human and animal pathogen causing aspergillosis. In this study, clinical (human and animal) and animal environment strains were able to produce high gliotoxin levels and had band profiles according to A. fumigatus sensu stricto by PCR-RFLP markers. The results obtained here suggest that strains involved in human and animal aspergillosis could come from the animal environment in which A. fumigatus is frequently found. Its presence in animal environments could affect animal health and productivity; in addition, there are risks of contamination for rural workers during handling and storage of animal feedstuffs. © 2013 The Society for Applied Microbiology.

Anti-mycobacterial screening of five Indian medicinal plants and partial purification of active extracts of Cassia sophera and Urtica dioica.

PubMed

Singh, Rambir; Hussain, Shariq; Verma, Rajesh; Sharma, Poonam

2013-05-13

To find out the anti-mycobacterial potential of Cassia sophera (C. sophera), Urtica dioica (U. dioica), Momordica dioica, Tribulus terrestris and Coccinia indica plants against multi-drug resistant (MDR) strain of Mycobacterium tuberculosis (M. tuberculosis). Plant materials were extracted successively with solvents of increasing polarity. Solvent extracts were screened for anti-mycobacterial activity against fast growing, non-pathogenic mycobacterium strain, Mycobacterium semegmatis, by disk diffusion method. The active extracts were tested against MDR and clinical isolates of M. tuberculosis by absolute concentration and proportion methods. The active extracts were subjected to bio-autoassay on TLC followed by silica column chromatography for isolation of potential drug leads. Hexane extract of U. dioica (HEUD) and methanol extract of C. sophera (MECS) produced inhibition zone of 20 mm in disc diffusion assay and MIC of 250 and 125 μ g/mL respectively in broth dilution assay against Mycobacterium semegmatis. Semipurified fraction F2 from MECS produced 86% inhibition against clinical isolate and 60% inhibition against MDR strain of M. tuberculosis. F18 from HEUD produced 81% inhibition against clinical isolate and 60% inhibition against MDR strain of M. tuberculosis. Phytochemical analysis indicated that anti-mycobacterial activity of MECS may be due to presence of alkaloids or flavonoids and that of HEUD due to terpenoids. C. sophera and U. dioica plant extracts exhibited promising anti-mycobacterial activity against MDR strain of M. tuberculosis. This is the first report of anti-mycobacterial activity form C. sophera. This study showed possibility of purifying novel anti-mycobacterial compound(s) from C. sophera and U. dioica. Copyright © 2013 Hainan Medical College. Published by Elsevier B.V. All rights reserved.

Tigecycline activity against metallo-β-lactamase-producing bacteria.

PubMed

Kumar, Simit; Bandyopadhyay, Maitreyi; Mondal, Soma; Pal, Nupur; Ghosh, Tapashi; Bandyopadhyay, Manas; Banerjee, Parthajit

2013-10-01

[corrected] Treatment of serious life-threatening multi-drug-resistant organisms poses a serious problem due to the limited therapeutic options. Tigecycline has been recently marketed as a broad-spectrum antibiotic with activity against both gram-positive and gram-negative bacteria. Even though many studies have demonstrated the activity of tigecycline against ESBL-producing Enterobacteriaceae, its activity is not well-defined against micro-organisms producing metallo-β-lactamases (MBLs), as there are only a few reports and the number of isolates tested is limited. The aim of the present study was to evaluate the activity of tigecycline against MBL-producing bacterial isolates. The isolates were tested for MBL production by (i) combined-disk test, (ii) double disc synergy test (DDST), (iii) susceptibility to aztreonam (30 μg) disk. Minimum inhibitory concentration to tigecycline was determined according to agar dilution method as per Clinical Laboratory Standards Institute (CLSI) guidelines. Disc diffusion susceptibility testing was also performed for all these isolates using tigecycline (15 μg) discs. Among the total 308 isolates included in the study, 99 were found to be MBL producers. MBL production was observed mostly in isolates from pus samples (40.47%) followed by urine (27.4%) and blood (13.09%). MBL production was observed in E. coli (41.48%), K. pneumoniae (26.67%), Proteus mirabilis (27.78%), Citrobacter spp. (41.67%), Enterobacter spp. (25.08%), and Acinetobacter spp. (27.27%). The result showed that tigecycline activity was unaffected by MBL production and it was showed almost 100% activity against all MBL-producing isolates, with most of the isolates exhibiting an MIC ranging from 0.25-8 μg/ml, except 2 MBL-producing E. coli isolates who had an MIC of 8 μg/ml. To conclude, tigecycline was found to be highly effective against MBL-producing Enterobacteriaceae and acinetobacter isolates, but the presence of resistance among organisms, even before the mass usage of the drug, warrants the need of its usage as a reserve drug. The study also found that the interpretative criteria for the disc diffusion method, recommended by the FDA, correlates well with the MIC detection methods. So, the microbiology laboratories might use the relatively easier method of disc diffusion, as compared to the comparatively tedious method of MIC determination.

Genotypic characterization of ESBL-producing E. coli from imported meat in South Korea.

PubMed

Kim, Young-Jo; Moon, Jin-San; Oh, Deog-Hwan; Chon, Jung-Whan; Song, Bo-Ra; Lim, Jong-Su; Heo, Eun-Jeong; Park, Hyun-Jung; Wee, Sung-Hwan; Sung, Kidon

2018-05-01

Twenty extended-spectrum β-lactamase (ESBL)-producing E. coli strains were isolated from imported meat in South Korea. ESBL strains of E. coli were detected in chicken (14/20) more often than in pork (6/20) and beef (0/20); the highest number (12/20) was detected in Brazilian meats. The bla CTX-M genes were predominant in meats from many countries. E. coli from pork imported from France produced the bla CTX-M-58 enzyme, which has never been documented previously in ESBL-producing bacteria from clinical or environmental sources. Additionally, the coexistence of the bla CTX-M-2 and bla OXA-1 enzymes in EC12-5 isolate was found for the first time in an ESBL E. coli isolate. A rare bla CTX-M type, bla CTX-M-25 , was found in 40% of ESBL E. coli isolates. Phenotypic susceptibility testing showed that E. coli isolates were resistant to up to eleven antibiotics, including ciprofloxacin. For the first time, a new combination in an integron gene cassette, aacA4-cmlA6-qacEΔ1, was found in an E. coli isolate from poultry imported from Brazil. Three E. coli ST117 isolates, from an avian pathogenic lineage producing CTX-M-94, harbored fimH, fyuA, iutA, papC, rfc, and traT virulence genes and were not susceptible to quinolones. For the first time, rfc and papG virulence factors were detected in ESBL E. coli strains isolated from meat products. Even though E. coli CC21 and CC22 were obtained from meats from the USA and Brazil, respectively, they had a similarity coefficient higher than 99% in rep-PCR and the same MLST type (ST117), phenotypic antibiotic resistance pattern, integron gene (qacEΔ1), and plasmid DNA profile. This study indicates that imported meat products may be a source of ESBL-producing E. coli strains in South Korea. Published by Elsevier Ltd.

Phenotypic and genotypic profile of clinical and animal multidrug-resistant Salmonella enterica isolates from Mexico.

PubMed

Aguilar-Montes de Oca, S; Talavera-Rojas, M; Soriano-Vargas, E; Barba-León, J; Vázquez-Navarrete, J; Acosta-Dibarrat, J; Salgado-Miranda, C

2018-01-01

The objective of this study was to obtain a phenotypic and genotypic profile of Salmonella enterica including multidrug-resistant (MDR) isolates from food-producing animals and clinical isolates, as well as their genetic relatedness in two different States of Mexico (Jalisco and State of Mexico). A total of 243 isolates were evaluated in terms of antimicrobial resistance (AMR) and related genes through a disk diffusion method and PCR respectively; we found 16 MDR isolates, all of them harbouring the bla CMY gene but not qnr genes, these isolates represent less than 10% of the collection. The pulsed-field gel electrophoresis revealed a higher genotypic similitude within isolates of State of Mexico than Jalisco. A low percentage of Salmonella isolates were resistant to relevant antibiotics in human health, nevertheless, the AMR and involved genes were similar despite the different serovars and origin of the isolates. This investigation provided an insight of the current status of AMR of Salmonella isolates in two States of Mexico and pinpoint the genes involved in AMR and their epidemiological relationship, the information could help to determine an adequate therapy in human and veterinary medicine. © 2017 The Society for Applied Microbiology.

Structural determination of the polysaccharide isolated from biofilms produced by a clinical strain of Klebsiella pneumoniae.

PubMed

Cescutti, Paola; De Benedetto, Gianluigi; Rizzo, Roberto

2016-07-22

Klebsiella pneumoniae are Gram negative opportunistic pathogens producing capsular (K) polysaccharides. Seventy-seven different K antigens have been described and they are the basis for K serotyping. Capsular polysaccharides are important virulence factors and have a relevant role for the structure of biofilm communities. Nevertheless, little information is available on the polysaccharides produced in biofilm matrices by Klebsiella spp. In the present study, a clinical isolate of Klebsiella pneumoniae was grown both on cellulose membranes deposited on agar plates, where it formed an adherent biofilm, and in liquid medium, where it formed floating biofilms (flocs). Extraction and purification of the polysaccharide fraction showed that only one main carbohydrate polymer was present in both adherent biofilms and flocs. Composition and linkage analysis, Smith degradation followed by ESI-MS, 1D and 2D NMR spectroscopy revealed that the polysaccharide belong to the type K24 and has the following structure. Copyright © 2016 Elsevier Ltd. All rights reserved.

Presence of extended-spectrum beta-lactamase-producing Enterobacteriaceae in waste waters, Kinshasa, the Democratic Republic of the Congo.

PubMed

De Boeck, H; Lunguya, O; Muyembe, J-J; Glupczynski, Y; Jacobs, J

2012-11-01

Extended-spectrum beta-lactamase (ESBL)-producing Enterobacteriaceae are a major public health concern. We previously demonstrated the presence of ESBL-producing Enterobacteriaceae in sachet-packaged water bags sold in Kinshasa, the Democratic Republic of the Congo. In complement to the previous study, we aimed to assess the presence of ESBL-producing Enterobacteriaceae in waste waters in Kinshasa.Enterobacteriaceae isolates recovered from environmental water samples were screened and phenotypically confirmed as ESBL-producers by disk diffusion according to Clinical and Laboratory Standards Institute (CLSI) guidelines (CLSI M100-S21). Final identification to the species level and further antimicrobial susceptibility testing were carried out with MicroScan® NBC42 panels and the identification of bla (ESBL) coding genes was performed by a commercial multiplex ligation polymerase chain reaction (PCR) microarray (Check-Points CT 101, Wageningen, the Netherlands). Overall, 194 non-duplicate Enterobacteriaceae were recovered from several sewer and river sites in nine out of 24 municipalities of Kinshasa. Fourteen isolates (7.4 %) were confirmed as ESBL-producers, the main species being Enterobacter cloacae (46.6 %) and Klebsiella pneumoniae (40.0 %). Associated resistance to both aminoglycoside and fluoroquinolone antibiotics was observed in ten isolates; the remaining isolates showed co-resistance to either fluoroquinolone (n = 3) or to aminoglycoside (n = 1) alone. All but one isolate carried bla (CTX-M) genes belonging to the CTX-M-1 group. ESBL-producing Enterobacteriaceae are increasingly being reported from various sources in the community. The present results suggest that ESBL-producing Enterobacteriaceae are widespread in the environment in the community of Kinshasa. Cities in Central Africa should be added to the map of potentially ESBL-contaminated environments and highlight the need to reinforce safe water supply and public sanitation.

Association of pellicle growth morphological characteristics and clinical presentation of Mycobacterium tuberculosis isolates.

PubMed

Arora, Ranjana; Armitige, Lisa; Wanger, Audrey; Hunter, Robert L; Hwang, Shen-An

2016-12-01

Trehalose 6,6'dimycolate (TDM) is a glycolipid found in nearly pure form on the surface of virulent Mycobacterium tuberculosis (MTB). This manuscript investigated the production of TDM, growth rate and colony morphology of multiple strains of MTB, each of which had been isolated from both pulmonary (sputum) and extrapulmonary sites of multiple patients. Since sputum contains MTB primarily from cavities and extrapulmonary biopsies are typically granulomas, this provided an opportunity to compare the behavior of single strains of MTB that had been isolated from cavities and granulomas. The results demonstrated that MTB isolated from pulmonary sites produced more TDM (3.23 ± 1.75 μg TDM/mg MTB), grew more rapidly as thin spreading pellicles, demonstrated early cording, and climbed culture well walls. In contrast, extrapulmonary isolates produced less TDM (1.42 ± 0.58 μg TDM/mg MTB) (p < 0.001) and grew as discrete patches with little tendency to spread or climb. Both Beijing pulmonary isolates and the non-Beijing pulmonary isolates produced significantly more TDM (1.64 ± 0.46 μg TDM/mg MTB) and grew faster than the Beijing and non-Beijing extrapulmonary isolates (1.14 ± 0.63 μg TDM/mg MTB) (p < 0.001 and p < 0.005 respectively). These results indicate that MTB from pulmonary sites (cavities) grows faster and produces more TDM than strains isolated from extrapulmonary sites (granulomas). This report suggests a critical role for TDM in cavitary TB. Copyright © 2016 Elsevier Ltd. All rights reserved.

Extended-Spectrum β-lactamase (ESBL) producing Enterobacter aerogenes phenotypically misidentified as Klebsiella pneumoniae or K. terrigena

PubMed Central

Claeys, Geert; De Baere, Thierry; Wauters, Georges; Vandecandelaere, Patricia; Verschraegen, Gerda; Muylaert, An; Vaneechoutte, Mario

2004-01-01

Background Enterobacter aerogenes and Klebsiella pneumoniae are common isolates in clinical microbiology and important as producers of extended spectrum β-lactamases (ESBL). The discrimination between both species, which is routinely based on biochemical characteristics, is generally accepted to be straightforward. Here we report that genotypically unrelated strains of E. aerogenes can be misidentified as K. pneumoniae by routine laboratories using standard biochemical identification and using identification automates. Results Ten clinical isolates, identified as K. pneumoniae or K. terrigena with the routinely used biochemical tests and with API-20E, were identified as E. aerogenes by tDNA-PCR – an identification that was confirmed by 16S rRNA gene sequencing for five of these isolates. Misidentification also occurred when using the automated identification systems Vitek 2 and Phoenix, and was due to delayed positivity for ornithine decarboxylase and motility. Subculture and prolonged incubation resulted in positive results for ornithine decarboxylase and for motility. It could be shown by RAPD-analysis that the E. aerogenes strains belonged to different genotypes. Conclusions Clinical E. aerogenes isolates can be easily misidentified as Klebsiella due to delayed positivity for ornithine decarboxylase and motility. The phenomenon may be widespread, since it was shown to occur among genotypically unrelated strains from different hospitals and different isolation dates. A useful clue for correct identification is the presence of an inducible β-lactamase, which is highly unusual for K. pneumoniae. In several instances, the use of genotypic techniques like tDNA-PCR may circumvent problems of phenotypic identification. PMID:15619329

Isolation of KPC 3-producing Enterobacter aerogenes in a patient colonized by MDR Klebsiella pneumoniae.

PubMed

Venditti, Carolina; Villa, Laura; Capone, Alessandro; Fortini, Daniela; D'Arezzo, Silvia; Nisii, Carla; Bordi, Eugenio; Puro, Vincenzo; Antonini, Mario; Carattoli, Alessandra; Cataldo, Maria Adriana; Petrosillo, Nicola; Di Caro, Antonino

2016-10-01

We describe the interspecies transmission of the plasmid-mediated blaKPC-3 gene, which confers carbapenem resistance, between clinically relevant gram-negative bacteria in a single patient. A KPC-3 producing Enterobacter aerogenes was isolated from a hospitalized patient previously colonized and then infected by a Klebsiella pneumoniae ST101 carrying the blaKPC-3 gene. The strains showed identical plasmids. Since intense horizontal exchanges among bacteria can occur in the gut, clinicians should be aware that patients colonized by carbapenem-resistant K. pneumoniae could become carriers of other carbapenem-resistant Enterobacteriaceae.

Detection of Ambler class A, B and D ß-lactamases among Pseudomonas aeruginosa and Acinetobacter baumannii clinical isolates from burn patients

PubMed Central

Hakemi Vala, M.; Hallajzadeh, M.; Hashemi, A.; Goudarzi, H.; Tarhani, M.; Sattarzadeh Tabrizi, M.; Bazmi, F.

2014-01-01

Summary In this study, we evaluated the existence of classes A, B and D ß-lactamases among Pseudomonas aeruginosa (P.aeruginosa) and Acinetobacter baumannii (A.baumannii) strains isolated from burn patients in Tehran during the years 2012 and 2013. From these strains, the frequency of MBL (metallo-beta-lactamase) and ESBL (extended-spectrum beta-lactamase) producers were evaluated using CDDT (Combined Disk Diffusion Tests). The prevalence of some related genes, including blaIMP, blaVIM, blaSPM, blaKPC, blaGIM, blaDIM, blaBIC, blaOXA-48, blaCTX-M-15 and blaNDM genes, was evaluated using PCR and sequencing methods. Of the 75 non-fermenter isolates, 47 P.aeruginosa and 28 A.baumannii were isolated and identified. A high rate of resistance to common antibiotics was detected among A.baumannii isolates in particular, showing 100% resistance to 9 tested antibiotics. CDDT showed that 21 (28%) and 25 (34.25%) of the non-fermenter isolates were ESBL and MBL producers respectively. The prevalence of blaCTX-M-15 and blaIMP genes among the 75 non-fermenter isolates was 7 (9.3%) and 1 (1.3%), respectively. Fortunately, no other genes were detected in either of the non-fermenters. The mortality rate due to MBL-producing isolates was 5 (20%). This study showed specific resistance genes exist among some MBL and ESBL gram-negative non-fermenters which were isolated from burn patients in Tehran. PMID:25249841

Detection of Ambler class A, B and D ß-lactamases among Pseudomonas aeruginosa and Acinetobacter baumannii clinical isolates from burn patients.

PubMed

Hakemi Vala, M; Hallajzadeh, M; Hashemi, A; Goudarzi, H; Tarhani, M; Sattarzadeh Tabrizi, M; Bazmi, F

2014-03-31

In this study, we evaluated the existence of classes A, B and D ß-lactamases among Pseudomonas aeruginosa (P.aeruginosa) and Acinetobacter baumannii (A.baumannii) strains isolated from burn patients in Tehran during the years 2012 and 2013. From these strains, the frequency of MBL (metallo-beta-lactamase) and ESBL (extended-spectrum beta-lactamase) producers were evaluated using CDDT (Combined Disk Diffusion Tests). The prevalence of some related genes, including blaIMP, blaVIM, blaSPM, blaKPC, blaGIM, blaDIM, blaBIC, blaOXA-48, blaCTX-M-15 and blaNDM genes, was evaluated using PCR and sequencing methods. Of the 75 non-fermenter isolates, 47 P.aeruginosa and 28 A.baumannii were isolated and identified. A high rate of resistance to common antibiotics was detected among A.baumannii isolates in particular, showing 100% resistance to 9 tested antibiotics. CDDT showed that 21 (28%) and 25 (34.25%) of the non-fermenter isolates were ESBL and MBL producers respectively. The prevalence of blaCTX-M-15 and blaIMP genes among the 75 non-fermenter isolates was 7 (9.3%) and 1 (1.3%), respectively. Fortunately, no other genes were detected in either of the non-fermenters. The mortality rate due to MBL-producing isolates was 5 (20%). This study showed specific resistance genes exist among some MBL and ESBL gram-negative non-fermenters which were isolated from burn patients in Tehran.

Bacteriological aspects implicated in abdominal surgical emergencies.

PubMed

Israil, A M; Delcaru, C; Palade, R S; Chifiriuc, C; Iordache, C; Vasile, D; Grigoriu, M; Voiculescu, D

2010-01-01

The purpose of the present study was to establish the microbial etiology of abdominal surgical emergencies as well as the relationship between the bacterial etiology and the virulence factors produced by the respective isolated strains. 110 bacterial strains were isolated from 100 randomized clinical cases, operated during 2009-2010 in the First Surgical Clinic of the University Hospital of Bucharest. The clinical cases (sex ratio 52 M/48F aged between 22-85 years old) were classified into three risk groups, as related to their severity. The isolated strains were characterized by cultural, microscopic and biochemical methods. After identification, the bacterial strains were investigated for their virulence potential (adherence to abiotic surface and production of soluble virulence factors). The specimens were collected from different clinical pathologies: diffuse acute peritonitis, biliary duct infections, severe acute pancreatitis followed by septic processes etc. The 110 bacterial (72 aerobic and 38 anaerobic) strains were isolated only in 70 out of 100 cases. Out of these 70 cases, in 45 already submitted to pre-operatory empiric broad spectrum antibiotic therapy, there were isolated 74 strains, whereas in 25 cases without any treatment, there were isolated 36 strains. The etiology was either mono-specific or multi-specific (aerobic-anaerobic associations, especially in old persons). Out of the 30 negative culture cases, 16 were already submitted to pre-operatory parenteral empiric antibiotic therapy at the moment of specimen collection. The aerobic etiology was dominated by Enterobacteriaceae. The most frequent anaerobic species belonged to Clostridium, Peptococcus and Bacteroides genera. It is to be mentioned that the isolation of Bifidobacterium and Veillonella spp. in 11 (10%) severe cases of the studied abdominal surgical emergencies is pleading for the fact that in certain conditions, bacteria belonging usually to commensal gut flora can turn to pathogenic becoming responsive for life-threatening cases. All aerobic and anaerobic strains exhibited some of the following virulence factors: mucinase, esculinase, pore-forming toxins (lecithinase), proteolytic enzymes, adherence ability (slime factor). The presence of these virulence factors (VF) could explain the severity of the clinical aspects. The bacterial etiology of the abdominal surgical emergencies exhibited a very large spectrum, the highest number of strains being of endogenous origin (Enterobacteriaceae and anaerobic strains). It was demonstrated that the isolated strains produced (cell associated and soluble) VF proving in this way their role as important virulence sources in the hospital environment and explaining the large diversity and severity of the clinical abdominal pathology. The results of the present study are also pleading for periodical readjustments of the pre-operatory empiric antibiotic therapy.

Comparison of the Modified-Hodge test, Carba NP test, and carbapenem inactivation method as screening methods for carbapenemase-producing Enterobacteriaceae.

PubMed

Yamada, Kageto; Kashiwa, Machiko; Arai, Katsumi; Nagano, Noriyuki; Saito, Ryoichi

2016-09-01

We compared three screening methods for carbapenemase-producing Enterobacteriaceae. While the Modified-Hodge test and Carba NP test produced false-negative results for OXA-48-like and mucoid NDM producers, the carbapenem inactivation method (CIM) showed positive results for these isolates. Although the CIM required cultivation time, it is well suited for general clinical laboratories. Copyright © 2016 Elsevier B.V. All rights reserved.

Flomoxef showed excellent in vitro activity against clinically important gram-positive and gram-negative pathogens causing community- and hospital-associated infections.

PubMed

Yang, Qiwen; Zhang, Hui; Cheng, Jingwei; Xu, Zhipeng; Hou, Xin; Xu, Yingchun

2015-04-01

The objective of this study was to better understand the in vitro activity of flomoxef against clinical pathogens. A total of 545 clinical isolates, including Escherichia coli, Klebsiella pneumoniae, Proteus mirabilis, methicillin-susceptible Staphylococcus aureus (MSSA), methicillin-resistant S. aureus, Streptococcus pneumoniae, and Streptococcus pyogenes, were isolated consecutively from clinical specimens from Peking Union Medical College Hospital in 2013. MICs were determined using broth microdilution method. esbl and ampC genes were detected by polymerase chain reaction and sequencing. Flomoxef showed excellent activity against E. coli, K. pneumoniae, and P. mirabilis isolates, with susceptibility rate of 88.8%, 88.3%, and 97.7%, separately. Moreover, flomoxef exhibited great activity against extended-spectrum beta-lactamase (ESBL) producers, with MIC50/MIC90 of 0.125/(0.5-1) μg/mL. Flomoxef showed MIC50/MIC90 of 0.5/0.5 μg/mL against MSSA, 0.125/0.25 μg/mL against S. pyogenes, and 2/16 μg/mL against S. pneumoniae. In conclusion, flomoxef is one of the cephamycins showing excellent activity against ESBL-producing or ESBL-nonproducing E. coli, K. pneumoniae, and P. mirabilis and was also potent against MSSA, S. pyogenes, and S. pneumoniae. Copyright © 2015 Elsevier Inc. All rights reserved.

Risk factors and impacts of clinical and subclinical mastitis in commercial meat-producing sheep flocks in Quebec, Canada.

PubMed

Arsenault, Julie; Dubreuil, Pascal; Higgins, Robert; Bélanger, Denise

2008-11-17

We conducted a prospective observational study on clinical and subclinical mastitis in 30 commercial meat-producing sheep flocks from 2 regions of the province of Quebec, Canada. A total of 2,792 ewes selected in late gestation were followed from lambing to weaning of lambs. The incidence of clinical mastitis for the total lactation period (average of 58 days) ranged among flocks from 0 to 6.6%, with a median of 1.2%. The most frequently isolated bacteria from the cases of clinical mastitis, in pure or mixed culture, were Mannheimia haemolytica (26%), Staphylococcus aureus (23%), and coagulase-negative staphylococci (17%). Incidence of clinical mastitis was higher in ewes that gave birth to 3 or more lambs and from the Estrie region, and was associated with an increase in ewe mortality, an increase in lamb mortality at the litter level, and a decrease in lamb's weaning weight for lambs born in multiple litter size or from ewes >or=4 years old. Among 354 selected ewes with clinically normal udder at the end of lactation, 28.8% had potentially pathogenic bacteria isolated from milk. The most prevalent bacteria were S. aureus (9.3%) and coagulase-negative staphylococci (9.3%). The risk of having a positive culture in at least one half was different between the two regions. Prevalence of ewes (n=261) with California Mastitis Test (CMT) positive result in at least one half was 24.1 and 14.9% using a cut-off of >or=1+ and >or=2+, respectively. Prevalence of culture-positive udder halves was 11.7% for CMT-negative compared with 53.6% for CMT 3+ halves. CMT status was positively associated with the isolation of coagulase-negative staphylococci, M. haemolytica, S. aureus, and various Streptococcus species, but not with other isolated bacteria. Additionally, prevalence of CMT-positive halves was higher in ewes from the Estrie region, aged of >or=4 years versus 1 year, having clinical mastitis previously detected in the lactation and/or with low body condition score. Lamb weaning weight was associated with CMT status of ewes, while weaning weight was not associated with milk culture results. More research is needed to understand the dynamic of milk SCC and IMI in ewes from meat-producing flocks, its economical impact and best ways to control it.

The spread of carbapenemase-producing bacteria in Africa: a systematic review

PubMed Central

Manenzhe, Rendani I.; Zar, Heather J.; Nicol, Mark P.; Kaba, Mamadou

2015-01-01

Background Carbapenems are the last line of defence against ever more prevalent MDR Gram-negative bacteria, but their efficacy is threatened worldwide by bacteria that produce carbapenemase enzymes. The epidemiology of bacteria producing carbapenemases has been described in considerable detail in Europe, North America and Asia; however, little is known about their spread and clinical relevance in Africa. Methods We systematically searched in PubMed, EBSCOhost, Web of Science, Scopus, Elsevier Masson Consulte and African Journals Online, international conference proceedings, published theses and dissertations for studies reporting on carbapenemase-producing bacteria in Africa. We included articles published in English or French up to 28 February 2014. We calculated the prevalence of carbapenemase producers only including studies where the total number of isolates tested was at least 30. Results Eighty-three studies were included and analysed. Most studies were conducted in North Africa (74%, 61/83), followed by Southern Africa (12%, 10/83), especially South Africa (90%, 9/10), West Africa (8%, 7/83) and East Africa (6%, 6/83). Carbapenemase-producing bacteria were isolated from humans, the hospital environment and community environmental water samples, but not from animals. The prevalence of carbapenemase-producing isolates in hospital settings ranged from 2.3% to 67.7% in North Africa and from 9% to 60% in sub-Saharan Africa. Conclusions Carbapenemase-producing bacteria have been described in many African countries; however, their prevalence is poorly defined and has not been systematically studied. Antibiotic stewardship and surveillance systems, including molecular detection and genotyping of resistant isolates, should be implemented to monitor and reduce the spread of carbapenemase-producing bacteria. PMID:25261423

[Four infants with upper urinary tract infection due to extended-spectrum bata lactamase (ESBL)-producing Escherichia coli].

PubMed

Hibino, Satoshi; Fukuchi, Kunihiko; Abe, Yoshifusa; Hoshino, Akihiro; Sakurai, Shunsuke; Mikawa, Takeshi; Fuke, Toshiya; Yoshida, Koichiro; Itabashi, Kazuo

2011-09-01

Bacteria producing extended-spectrum beta lactamase (ESBL) are detected mainly in adult urinary specimens, and are believed to cause hospital-acquired infection due to their resistance to many drugs. The incidence of community-acquired infection due to such bacteria is increasing, but few cases of infant upper urinary tract infection (UUTI) have been reported in Japan. We treated four infants with UUTI caused by ESBL-producing Escherichia coli, as determined by genotyping. Using medical records, we retrospectively evaluated the clinical course, antibiotic use and efficacy, antimicrobial susceptibility results, and the presence of underlying disease. One of the four had been previously hospitalized for occult bacteremia. Two developed UUTI after antibiotic treatment, indicating that previous antibiotic use may have been a risk factor in these cases. We could not identify the infection route in all cases. Two of the four had bilateral vesicoureteral reflux (VUR). Renal scintigraphy was done in three. Although an initial dimercaptosuccinic acid (DMSA) defect was detected in all four, only one had renal scarring. E. coli isolates from all four showed PCR signals for blaCTX-M-; one isolate positive for the blaCTX-M3 group and three positive for blaCTX-M14. Antimicrobial susceptibility test results showed all isolates to be resistant to cephalosporins, but discrepancies existed between antimicrobial susceptibility results and actual clinical efficacy. Clinically, cefazolin (CEZ) was effective in two subjects and ceftazidime (CAZ) effective in one. Panipenem/betamipron (PAPM/BP) was effective in one. None of the four developed sepsis or meningitis. Post hospitalization antibiotic prophylaxis showed that none of the four has had UUTI recur. Japan's ESBL-producing bacterial infection incidence is increasing, so medical professionals should watch for such UUTI even in first-case occurrence in infants.

[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.

Versatility of Biofilm Matrix Molecules in Staphylococcus epidermidis Clinical Isolates and Importance of Polysaccharide Intercellular Adhesin Expression during High Shear Stress

PubMed Central

Schaeffer, Carolyn R.; Hoang, Tra-My N.; Sudbeck, Craig M.; Alawi, Malik; Tolo, Isaiah E.; Robinson, D. Ashley; Horswill, Alexander R.; Rohde, Holger

2016-01-01

ABSTRACT Staphylococcus epidermidis is a leading cause of hospital-associated infections, including those of intravascular catheters, cerebrospinal fluid shunts, and orthopedic implants. Multiple biofilm matrix molecules with heterogeneous characteristics have been identified, including proteinaceous, polysaccharide, and nucleic acid factors. Two of the best-studied components in S. epidermidis include accumulation-associated protein (Aap) and polysaccharide intercellular adhesin (PIA), produced by the enzymatic products of the icaADBC operon. Biofilm composition varies by strain as well as environmental conditions, and strains producing PIA-mediated biofilms are more robust. Clinically, biofilm-mediated infections occur in a variety of anatomical sites with diverse physiological properties. To test the hypothesis that matrix composition exhibits niche specificity, biofilm-related genetic and physical properties were compared between S. epidermidis strains isolated from high-shear and low-shear environments. Among a collection of 105 clinical strains, significantly more isolates from high-shear environments carried the icaADBC operon than did those from low-shear settings (43.9% versus 22.9%, P < 0.05), while there was no significant difference in the presence of aap (77.2% versus 75.0%, P > 0.05). Additionally, a significantly greater number of high-shear isolates were capable of forming biofilm in vitro in a microtiter assay (82.5% versus 45.8%, P < 0.0001). However, even among high-shear clinical isolates, less than half contained the icaADBC locus; therefore, we selected for ica-negative variants with increased attachment to abiotic surfaces to examine PIA-independent biofilm mechanisms. Sequencing of selected variants identified substitutions capable of enhancing biofilm formation in multiple genes, further highlighting the heterogeneity of S. epidermidis biofilm molecules and mechanisms. IMPORTANCE Staphylococcus epidermidis is a leading cause of infections related to biomaterials, mostly due to their ability to form biofilm. Biofilm accumulation mechanisms vary, including those that are dependent on specific proteins, environmental DNA (eDNA), or polysaccharide intercellular adhesin (PIA). We found that those isolates obtained from high-shear environments, such as the lumen of a catheter, are more likely to produce PIA-mediated biofilms than those isolates obtained from a low-shear biomaterial-related infection. This suggests that PIA functions as a mechanism that is protective against shear flow. Finally, we performed selection experiments documenting the heterogeneity of biofilm accumulation molecules that function in the absence of PIA, further documenting the biofilm-forming potential of S. epidermidis. PMID:27747298

Stenotrophomonas maltophilia in Mexico: antimicrobial resistance, biofilm formation and clonal diversity.

PubMed

Flores-Treviño, Samantha; Gutiérrez-Ferman, Jessica Lizzeth; Morfín-Otero, Rayo; Rodríguez-Noriega, Eduardo; Estrada-Rivadeneyra, Diego; Rivas-Morales, Catalina; Llaca-Díaz, Jorge M; Camacho-Ortíz, Adrián; Mendoza-Olazarán, Soraya; Garza-González, Elvira

2014-11-01

Stenotrophomonas maltophilia is an important multidrug-resistant nosocomial pathogen associated with high mortality. Our aim was to examine antimicrobial susceptibility, biofilm production and clonal relatedness of clinical isolates of S. maltophilia. S. maltophilia isolates were collected between 2006 and 2013 from two tertiary care hospitals in Mexico. Antimicrobial susceptibility was evaluated by the broth microdilution method. PCR was used to determine the presence of β-lactamase genes L1 and L2. Biofilm formation was assessed with crystal violet staining. Clonal relatedness was determined by PFGE. Among the 119 collected S. maltophilia isolates, 73 (61.3%) were from the respiratory tract. Resistance levels exceeded 75% for imipenem, meropenem, ampicillin, aztreonam, gentamicin and tobramycin. Resistance to trimethoprim-sulfamethoxazole was 32.8%. L1 and L2 genes were detected in 77.1% (91/118) and 66.9% (79/118) of isolates, respectively. All S. maltophilia strains were able to produce biofilms. Strains were classified as weak (47.9%, 57/119), moderate (38.7%, 46/119), or strong (13.4%, 16/119) biofilm producers. A total of 89 distinct PFGE types were identified and 21.6% (22/102) of the isolates were distributed in nine clusters. This is the first study in Mexico to reveal characteristics of clinical isolates of S. maltophilia. Clonal diversity data indicate low cross-transmission of S. maltophilia in a hospital setting. The high antibiotic resistance underscores the need for continuous surveillance of S. maltophilia in hospital settings in Mexico. © 2014 The Authors.

Prevalence of Stx-producing Shigella species isolated from French Travelers Returning from the Caribbean: An Emerging Pathogen with International Implications

PubMed Central

Gray, Miranda D.; Lacher, David W.; Leonard, Susan R.; Abbott, Jason; Zhao, Shaohua; Lampel, Keith A.; Prothery, Estelle; Gouali, Malika; Weill, François-Xavier; Maurelli, Anthony T.

2015-01-01

Shiga toxins are potent cytotoxins that inhibit host cell protein synthesis, leading to cell death. Classically, these toxins are associated with intestinal infections due to Shiga toxin-producing Escherichia coli or Shigella dysenteriae serotype 1 and infections with these strains can lead to hemolytic uremic syndrome. Over the past decade there is increasing recognition that Shiga toxin is produced by additional Shigella species. We recently reported the presence and expression of stx genes in Shigella flexneri 2a clinical isolates. The toxin genes were carried by a new stx-encoding bacteriophage and infection with these strains correlated with recent travel to Haiti or the Dominican Republic. In this study we further explored the epidemiological link to this region by utilizing the French National Reference Center for Escherichia coli, Shigella and Salmonella collection to survey the frequency of Stx-producing Shigella species isolated from French travelers returning from the Caribbean. About 21% of the isolates tested were found to encode and produce Stx. These isolates included strains of S. flexneri 2a, S. flexneri Y, and S. dysenteriae 4. All of the travelers whom were infected with Stx-producing Shigella had recently traveled to Haiti, the Dominican Republic, or French Guiana. Furthermore, whole genome sequencing found that the toxin genes were encoded by a prophage that was highly identical to the phage we identified in our previous study. These findings demonstrate that this new stx-encoding prophage is circulating within that geographical area, has spread to other continents, and is capable of spreading to multiple Shigella serogroups. PMID:25980352

Association between isolation of Staphylococcus aureus one week after calving and milk yield, somatic cell count, clinical mastitis, and culling through the remaining lactation.

PubMed

Whist, Anne Cathrine; Osterås, Olav; Sølverød, Liv

2009-02-01

Cows with isolation of Staphylococcus aureus approximately 1 week after calving and milk yield, somatic cell count (SCC), clinical mastitis (CM), and culling risk through the remaining lactation were assessed in 178 Norwegian dairy herds. Mixed models with repeated measures were used to compare milk yield and SCC, and survival analyses were used to estimate the hazard ratio for CM and culling. On average, cows with an isolate of Staph. aureus had a significantly higher SCC than culture-negative cows. If no post-milking teat disinfection (PMTD) was used, the mean values of SCC were 42,000, 61,000, 68,000 and 77,000 cells/ml for cows with no Staph. aureus isolate, with Staph. aureus isolated in 1 quarter, in 2 quarters and more than 2 quarters respectively. If iodine PMTD was used, SCC means were 36,000; 63,000; 70,000 and 122,000, respectively. Primiparous cows testing positive for Staph. aureus had the same milk yield curve as culture-negative cows, except for those with Staph. aureus isolated in more than 2 quarters. They produced 229 kg less during a 305-d lactation. Multiparous cows with isolation of Staph. aureus in at least 1 quarter produced 94-161 kg less milk in 2nd and >3rd parity, respectively, and those with isolation in more than 2 quarters produced 303-390 kg less than multiparous culture-negative animals during a 305-d lactation. Compared with culture-negative cows, the hazard ratio for CM and culling in cows with isolation of Staph. aureus in at least 1 quarter was 2.0 (1.6-2.4) and 1.7 (1.5-1.9), respectively. There was a decrease in the SCC and in the CM risk in culture-negative cows where iodine PMTD had been used, indicating that iodine PMTD has a preventive effect on already healthy cows. For cows testing positive for Staph. aureus in more than 2 quarters at calving, iodine PMTD had a negative effect on the CM risk and on the SCC through the remaining lactation.

Clinical and molecular epidemiology of beta-hemolytic streptococcal infections in India.

PubMed

Mathur, Purva; Bhardwaj, Nidhi; Mathur, Kushal; Behera, Bijayini; Gupta, Gunjan; Kapil, Arti; Singh, Sarman; Misra, Mahesh Chandra

2014-03-13

Beta-hemolytic streptococci (βHS) cause a diverse array of human infections. Despite the high number of cases of streptococcal carriers and diseases, studies discerning the molecular epidemiology of βHS in India are limited. This study reports the molecular and clinical epidemiology of beta-hemolytic streptococcal infections from two geographically distinct regions of India. A total of 186 isolates of βHS from north and south India were included. The isolates were identified to species level and subjected to antimicrobial susceptibility testing. Polymerase chain reaction (PCR) was done to detect exotoxin genes, and emm types of group A streptococci (GAS) strains were ascertained by sequencing. GAS was the most common isolate (71.5%), followed by group G streptococci (GGS) (21%). A large proportion of GAS produced speB (97%), smeZ (89%), speF (91%), and speG (84%). SmeZ was produced by 21% and 50% of GGS and GGS, respectively. A total of 45 different emm types/subtypes were seen in GAS, with emm 11 being the most common. Resistance to tetracycline (73%) and erythromycin (34.5%) was commonly seen in GAS. A high diversity of emm types was seen in Indian GAS isolates with high macrolide and tetracycline resistance. SpeA was less commonly seen in Indian GAS isolates. There was no association between disease severity and exotoxin gene production.

Structural study and conformational behavior of the two different lipopolysaccharide O-antigens produced by the cystic fibrosis pathogen Burkholderia multivorans.

PubMed

Ieranò, Teresa; Silipo, Alba; Cescutti, Paola; Leone, Maria Rosaria; Rizzo, Roberto; Lanzetta, Rosa; Parrilli, Michelangelo; Molinaro, Antonio

2009-07-20

Lipopolysaccharides (LPSs) are virulence factors expressed by gram-negative bacteria; they are among those mainly responsible for bacterial virulence. In this work we define the primary structure and the conformational features of the O-chain from the LPS produced by the highly virulent clinical isolate Burkholderia multivorans strain C1576, an opportunistic human pathogen isolated in a cystic fibrosis center and causative of an outbreak with lethal outcome. We demonstrate that the LPS from this clinical isolate consists of two O-polysaccharide chains present in different amounts and made up of repeating units, both containing deoxy sugar. Additionally, conformational studies have been performed to establish and compare the spatial arrangements of the two polysaccharides and differences in their shape have been highlighted. The comprehension of the structural and conformational features of the two repeating units may help to explain their biological significance, the molecular shape of the bacterial external surface, and the comprehension at the molecular level of the recognition mechanisms of the antibodies.

High prevalence of sequence type 131 isolates producing CTX-M-15 among ESBL-producing Escherichia coli strains in north-east Iran.

PubMed

Moghanni, Marzie; Ghazvini, Kiarash; Farsiani, Hadi; Namaei, Mohammad Hasan; Derakhshan, Mohammad; Yousefi, Masoud; Maragheh, Alimohammad; Jamehdar, Saeid Amel

2018-05-25

The recent expansion of extended-spectrum-β-lactamase (ESBL)-producing Escherichia coli (E. coli) is a worldwide problem. The purpose of this study was to investigate the molecular characteristic of ESBL-producing E. coli strains in Mashhad, located in the northeast of Iran. A total of 455 clinical isolates of E. coli were collected at three hospitals in Mashhad between April and September 2015. Antibiotic susceptibility was determined with the Kirby Bauer disk diffusion test. The Combination Disc Test (CDT) was performed for the phenotypic detection of ESBLs. PCR was used for screening isolates for ESBLs typing. Phylogenetic groups and sequence type 131 were determined by multiplex-PCR. The prevalence of ESBL-producing E. coli in the collected E. coli strains was 52% (235/455). Among the 235 ESBL producer strains, 94.4% (222/235) tested positive for the CTX-M type, while 115 (48.9%), 92 (39%) and 21 (8.9%) strains were positive for TEM, OXA and SHV, respectively. Moreover, CTX-M-15 (94.1%, 209/222) was the most common ESBL-producing E. coli. Based on multiplex-PCR, the phylogenetic group B2 was the most predominant (169 isolates, 71.9%), followed by D (32, 13.6%), A (21, 8.9%), and B1 (13, 5.5%). Of the 169 groups of B2 isolates, ST131 (151, 89.3%) was the predominant clonal group. The obtained results revealed that an urgent investigation of the source and the transmission pathways of the CTX-M15-B2ST131 E. coli clone was needed to countervail this emergent public health problem. Copyright © 2018. Published by Elsevier Ltd.

Evaluation of Antioxidant Activity and Growth Control Properties of Nonoscale Structure Produced from Aloe vera var. littoralis Extract on Clinical Isolates of Salmonella.

PubMed

Ranjbar, Reza; Arjomandzadegan, Mohammad; Hosseiny, Hossein

2017-07-31

The aim of the study was to examine antibacterial properties of microemulsion structure produced from Aloe vera var. littoralis extract as a new tool of nanoscale drug-like materials. Aloe vera var. littoralis ( A. littoralis ) extract was prepared by distillation method. A nonocarrier structure in the microemulsion system was prepared from the extract. Serial concentrations were prepared from 8 mg/mL extract and the nonocarrier containing 0.1 mg/mL pure extract and were evaluated by a disk diffusion method for 35 Salmonella clinical isolates. Minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) were determined by microbroth dilution assay using MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) method by an enzyme-linked immunosorbent assay(ELISA) Microplate Reader apparatus. Antioxidant activity of the extract was determined by measuring the ferric reducing ability of plasma (FRAP) assay. From 35 clinical isolates of Salmonella , 17 isolates-including resistant isolates of S.E.1103 and S.E.49-had a zone of inhibition (ZI) of 7 to 32 mm in 0.007 mg/mL of the extract. S.E.76 isolate exposed to 30 µg/mL ceftazidime disk had a ZI of 12 mm but had 10 mm in 7µg/mL of A. littoralis extract. The inhibitory effect of a nanocarrier at a concentration of 25 µg/mL by 20 mm ZI was comparable by the ceftazidime (30 µg/mL) effect. MIC 50 was 0.25 mg/mL and MBC 50 was 0.5 mg/mL by MTT method for the extract. It was shown that A.littoralis extract had antioxidant activity of 31.67 µM/mg that could be increased based on concentration. It was concluded that the nanocarrier had a significant effect on the studied isolates in comparison with ordinary antibiotics and had potential for use as a natural antioxidant and antimicrobial material in complementary medicine.

Prevalence of plasmid-mediated quinolone resistance determinants in ESBL Enterobacteriaceae clinical isolates over a 1-year period in a French hospital.

PubMed

Crémet, L; Caroff, N; Dauvergne, S; Reynaud, A; Lepelletier, D; Corvec, S

2011-06-01

The prevalence of plasmid-mediated quinolone resistance (PMQR) determinants (qnrA, qnrB, qnrS, aac(6')-Ib-cr, and qepA) was investigated in a collection of 47 extended-spectrum β-lactamase (ESBL) producing enterobacterial isolates with reduced susceptibility to fluoroquinolones, recovered at Nantes University hospital, in 2006. qnr, aac(6')-Ib-cr, and qepA genes were screened by PCR, and positive results were subsequently confirmed by sequencing. The epidemiological relationship between positive isolates was studied by pulsed-field gel electrophoresis (PFGE). qnr-positive isolates were analyzed for antimicrobial susceptibility and presence of mutations in the quinolone resistance-determining region (QRDR) of gyrA and parC genes. ESBL genes were characterized by PCR and sequencing. Conjugation experiments were performed to determine whether the qnr-carrying plasmids were self-transferable. Two Klebsiella pneumoniae isolates (4.3%), not clonally related, harboured a qnrS1 gene, whereas no qnrA- or qnrB-positive isolate was detected. The aac(6')-Ib-cr gene was detected in 11 Escherichia coli and one K. pneumoniae isolates. None of the 47 isolates carried the qepA gene. ESBLs associated with QnrS1 were CTX-M-14 and CTX-M-15. The CTX-M-15 producing isolate was highly resistant to fluoroquinolones and harboured three mutations in the QRDR and two PMQR determinants (qnrS1 and aac(6')-Ib-cr). The CTX-M-14-producing isolate exhibited reduced susceptibility or resistance to fluoroquinolones without resistance to nalidixic acid. This strain harboured only a qnr gene on a single 170 kb transferable plasmid, without any mutation in the QRDR. In conclusion, our study showed that aac(6')-Ib-cr gene had occurred in multiclonal ESBL-producing enterobacterial isolates collected at Nantes University hospital in 2006, with a higher prevalence than qnr genes. Copyright © 2009 Elsevier Masson SAS. All rights reserved.

Herpes Simplex Virus Type 2 Glycoprotein G-Negative Clinical Isolates Are Generated by Single Frameshift Mutations

PubMed Central

Liljeqvist, Jan-Åke; Svennerholm, Bo; Bergström, Tomas

1999-01-01

Herpes simplex virus (HSV) codes for several envelope glycoproteins, including glycoprotein G-2 (gG-2) of HSV type 2 (HSV-2), which are dispensable for replication in cell culture. However, clinical isolates which are deficient in such proteins occur rarely. We describe here five clinical HSV-2 isolates which were found to be unreactive to a panel of anti-gG-2 monoclonal antibodies and therefore considered phenotypically gG-2 negative. These isolates were further examined for expression of the secreted amino-terminal and cell-associated carboxy-terminal portions of gG-2 by immunoblotting and radioimmunoprecipitation. The gG-2 gene was completely inactivated in four isolates, with no expression of the two protein products. For one isolate a normally produced secreted portion and a truncated carboxy-terminal portion of gG-2 were detected in virus-infected cell medium. Sequencing of the complete gG-2 gene identified a single insertion or deletion of guanine or cytosine nucleotides in all five strains, resulting in a premature termination codon. The frameshift mutations were localized within runs of five or more guanine or cytosine nucleotides and were dispersed throughout the gene. For the isolate for which a partially inactivated gG-2 gene was detected, the frameshift mutation was localized upstream of but adjacent to the nucleotides coding for the transmembranous region. Thus, this study demonstrates the existence of clinical HSV-2 isolates which do not express an envelope glycoprotein and identifies the underlying molecular mechanism to be a single frameshift mutation. PMID:10559290

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.

Purification and characterization of factors produced by Aspergillus fumigatus which affect human ciliated respiratory epithelium.

PubMed

Amitani, R; Taylor, G; Elezis, E N; Llewellyn-Jones, C; Mitchell, J; Kuze, F; Cole, P J; Wilson, R

1995-09-01

The mechanisms by which Aspergillus fumigatus colonizes the respiratory mucosa are unknown. Culture filtrates of eight of nine clinical isolates of A. fumigatus slowed ciliary beat frequency and damaged human respiratory epithelium in vitro. These changes appeared to occur concurrently. Culture filtrates of two clinical isolates of Candida albicans had no effect on ciliated epithelium. We have purified and characterized cilioinhibitory factors of a clinical isolate of A. fumigatus. The cilioinhibitory activity was heat labile, reduced by dialysis, and partially extractable into chloroform. The activity was associated with both high- and low-molecular-weight factors, as determined by gel filtration on Sephadex G-50. A low-molecular-weight cilioinhibitory factor was further purified by reverse-phase high-performance liquid chromatography and shown by mass spectrometry to be gliotoxin, a known metabolite of A. fumigatus. Gliotoxin significantly slowed ciliary beat frequency in association with epithelial damage at concentrations above 0.2 microgram/ml; other Aspergillus toxins, i.e., fumagillin and helvolic acid, were also cilioinhibitory but at much higher concentrations. High-molecular-weight (> or = 35,000 and 25,000) cilioinhibitory materials had neither elastolytic nor proteolytic activity and remain to be identified. Thus, A. fumigatus produces a number of biologically active substances which slow ciliary beating and damage epithelium and which may influence colonization of the airways.

Detection of Metallo-β-Lactamase Producing Pseudomonas aeruginosa Isolated from Public and Private Hospitals in Baghdad, Iraq.

PubMed

Al-Charrakh, Alaa H; Al-Awadi, Salwa J; Mohammed, Ahmed S

2016-02-01

Metallo-β-lactamase (MBL) producing Pseudomonas aeruginosa has been reported to be an important nosocomial infection. Its intrinsic and acquired resistance to various antimicrobial agents and its ability to develop multidrug resistance imposes a serious therapeutic problem. Different clinical samples were collected from public and private hospitals in Baghdad city, Iraq. Bacterial identification was done using conventional cultural, biochemical tests, and VITEk 2 system. Minimum inhibitory concentration (MIC) testing was performed using VITEK 2 automated system. Each P. aeruginosa isolates showed resistance to Carbapenems (Imipenem and Meropenem) were subjected to Imipenem-EDTA combined disc synergy test (CDST) to investigate the production of MBL (confirmative test). The presence of bla-genes encoded IMP, VIM, and SPM-1 was detected by conventional PCR technique. A total of 75 P. aeruginosa isolates were isolated, 16 (21.3%) were able to grow on MacConkey agar supplemented with Meropenem 4mg/L (MMAC). The MIC of different antibiotics showed that 6 (37.5 %) isolates were Carbapenem resistant, MIC ≥16 µg/ml while 4 (25%) isolates appear to be MBL producer using CDST test. PCR assay revealed that 3 (50%), 1 (16.6%) of the carbapenem resistant isolates harbored blaIMP, blaSPM-1 genes, respectively. blaVIM gene was not detected in this study. The prevalence of multi-drug resistant P. aeruginosa isolates especially Carbapenem resistant bacteria was increased in Baghdad province. The blaIMP was the predominant among the MBLs genes in P. aeruginosa in this study.

Exopolysaccharides produced by clinical strains belonging to the Burkholderia cepacia complex.

PubMed

Herasimenka, Yury; Cescutti, Paola; Impallomeni, Giuseppe; Campana, Silvia; Taccetti, Giovanni; Ravenni, Novella; Zanetti, Flavio; Rizzo, Roberto

2007-04-01

In the frame of a research line dedicated to better clarify the role of exopolysaccharides (EPS) in bacterial virulence, EPS produced by species of the Burkholderia cepacia complex (Bcc), namely Burkholderia multivorans, Burkholderia cenocepacia, and a Bcc member of undetermined genomovar, all isolated at the Cystic Fibrosis Regional Centre of Florence (Italy), were investigated for they structural properties. Three strains of B. multivorans, three of B. cenocepacia and one of a Bcc member of undetermined genomovar were isolated from CF patients. The reference strains C1576 and J2315, for genomovar II and III, respectively, were included in the study. The bacteria were grown on solid media, the exopolysaccharides produced were purified, and their structures were determined. In addition, sugar analysis of sputum samples was accomplished to search for EPS produced in vivo. Six strains out of seven produced the exopolysaccharide cepacian, while one strain of B. multivorans produced a completely different polymer, previously known in the literature as PS1. Two strains synthesised very small amounts of EPS. No definitive evidence for the presence of cepacian in sputum samples was found. Most strains examined produced abundant amounts of polysaccharides. Cepacian was the most common EPS isolated and its production was not associated to a particular genomovar.

Changing epidemiology of infections due to extended spectrum beta-lactamase producing bacteria

PubMed Central

2014-01-01

Background Community-associated infections caused by extended-spectrum beta-lactamase (ESBL) producing bacteria are a growing concern. Methods Retrospective cohort study of clinical infections due to ESBL-producing bacteria requiring admission from 2006-2011 at a tertiary care academic medical center in Providence, RI. Results A total of 321 infections due to ESBL-producing bacteria occurred during the study period. Fifty-eight cases (18%) were community-acquired, 170 (53%) were healthcare–associated, and 93 (29%) were hospital-acquired. The incidence of ESBL infections per 10,000 discharges increased during the study period for both healthcare-associated infections, 1.9 per year (95% CI 1-2.8), and for community-acquired infections, 0.85 per year (95% CI 0.3-1.4) but the rate remained unchanged for hospital-acquired infections. For ESBL-producing E. coli isolates, resistance to both ciprofloxacin and trimethoprim-sulfamethoxazole was 95% and 65%, respectively but 94% of isolates were susceptible to nitrofurantoin. Conclusions Community-acquired and healthcare-associated infections due to ESBL-producing bacteria are increasing in our community, particularly urinary tract infections due to ESBL-producing E. coli. Most isolates are resistant to oral antibiotics commonly used to treat urinary tract infections. Thus, our findings have important implications for outpatient management of such infections. PMID:24666610

Population structure of clinical Pseudomonas aeruginosa from West and Central African countries.

PubMed

Cholley, Pascal; Ka, Roughyatou; Guyeux, Christophe; Thouverez, Michelle; Guessennd, Nathalie; Ghebremedhin, Beniam; Frank, Thierry; Bertrand, Xavier; Hocquet, Didier

2014-01-01

Pseudomonas aeruginosa (PA) has a non-clonal, epidemic population with a few widely distributed and frequently encountered sequence types (STs) called 'high-risk clusters'. Clinical P. aeruginosa (clinPA) has been studied in all inhabited continents excepted in Africa, where a very few isolates have been analyzed. Here, we characterized a collection of clinPA isolates from four countries of West and Central Africa. 184 non-redundant isolates of clinPA from hospitals of Senegal, Ivory Coast, Nigeria, and Central African Republic were genotyped by MLST. We assessed their resistance level to antibiotics by agar diffusion and identified the extended-spectrum β-lactamases (ESBLs) and metallo-β-lactamases (MBLs) by sequencing. The population structure of the species was determined by a nucleotide-based analysis of the entire PA MLST database and further localized on the phylogenetic tree (i) the sequence types (STs) of the present collection, (ii) the STs by continents, (iii) ESBL- and MBL-producing STs from the MLST database. We found 80 distinct STs, of which 24 had no relationship with any known STs. 'High-risk' international clonal complexes (CC155, CC244, CC235) were frequently found in West and Central Africa. The five VIM-2-producing isolates belonged to CC233 and CC244. GES-1 and GES-9 enzymes were produced by one CC235 and one ST1469 isolate, respectively. We showed the spread of 'high-risk' international clonal complexes, often described as multidrug-resistant on other continents, with a fully susceptible phenotype. The MBL- and ESBL-producing STs were scattered throughout the phylogenetic tree and our data suggest a poor association between a continent and a specific phylogroup. ESBL- and MBL-encoding genes are borne by both successful international clonal complexes and distinct local STs in clinPA of West and Central Africa. Furthermore, our data suggest that the spread of a ST could be either due to its antibiotic resistance or to features independent from the resistance to antibiotics.

Outcome of urinary tract infections caused by extended spectrum β-lactamase-producing Enterobacteriaceae in children.

PubMed

Tratselas, Athanasios; Iosifidis, Elias; Ioannidou, Maria; Saoulidis, Stamatis; Kollios, Konstantinos; Antachopoulos, Charalampos; Sofianou, Danai; Roilides, Emmanuel J

2011-08-01

The outcome of patients with urinary tract infections caused by extended spectrum β-lactamases (ESBL)-producing bacteria (cases) was compared with that of matched controls with urinary tract infections caused by non-extended spectrum β-lactamases-producing isolates. Significantly, more case patients received inappropriate empiric therapy than controls. Nevertheless, clinical and microbiologic outcomes as well as formation of renal scars did not differ between the 2 groups.

The antimicrobial susceptibility, biofilm formation and genotypic profiles of Staphylococcus haemolyticus from bloodstream infections

PubMed Central

Silva, Patricia Vollú; Cruz, Raquel Souza; Keim, Luiz Sérgio; de Paula, Geraldo Renato; Carvalho, Bernadete Teixeira Ferreira; Coelho, Leonardo Rocchetto; Carvalho, Maria Cícera da Silva; da Rosa, Joel Mauricio Corrêa; Figueiredo, Agnes Marie Sá; Teixeira, Lenise Arneiro

2013-01-01

We analysed the antimicrobial susceptibility, biofilm formation and genotypic profiles of 27 isolates of Staphylococcus haemolyticus obtained from the blood of 19 patients admitted to a hospital in Rio de Janeiro, Brazil. Our analysis revealed a clinical significance of 36.8% and a multi-resistance rate of 92.6% among these isolates. All but one isolate carried the mecA gene. The staphylococcal cassette chromosome mec type I was the most prevalent mec element detected (67%). Nevertheless, the isolates showed clonal diversity based on pulsed-field gel electrophoresis analysis. The ability to form biofilms was detected in 66% of the isolates studied. Surprisingly, no icaAD genes were found among the biofilm-producing isolates. PMID:24037208

The antimicrobial susceptibility, biofilm formation and genotypic profiles of Staphylococcus haemolyticus from bloodstream infections.

PubMed

Silva, Patricia Vollú; Cruz, Raquel Souza; Keim, Luiz Sérgio; Paula, Geraldo Renato de; Carvalho, Bernadete Teixeira Ferreira; Coelho, Leonardo Rocchetto; Carvalho, Maria Cícera da Silva; Rosa, Joel Mauricio Corrêa da; Figueiredo, Agnes Marie Sá; Teixeira, Lenise Arneiro

2013-09-01

We analysed the antimicrobial susceptibility, biofilm formation and genotypic profiles of 27 isolates of Staphylococcus haemolyticus obtained from the blood of 19 patients admitted to a hospital in Rio de Janeiro, Brazil. Our analysis revealed a clinical significance of 36.8% and a multi-resistance rate of 92.6% among these isolates. All but one isolate carried the mecA gene. The staphylococcal cassette chromosome mec type I was the most prevalent mec element detected (67%). Nevertheless, the isolates showed clonal diversity based on pulsed-field gel electrophoresis analysis. The ability to form biofilms was detected in 66% of the isolates studied. Surprisingly, no icaAD genes were found among the biofilm-producing isolates.

A Bioreactor Method to Generate High-titer, Genetically Stable, Clinical-isolate Human Cytomegalovirus.

PubMed

Saykally, Victoria R; Rast, Luke I; Sasaki, Jeff; Jung, Seung-Yong; Bolovan-Fritts, Cynthia; Weinberger, Leor S

2017-11-05

Human cytomegalovirus (HCMV) infection is a major cause of morbidity and mortality in transplant patients and a leading cause of congenital birth defects (Saint Louis, 2016). Vaccination and therapeutic studies often require scalable cell culture production of wild type virus, represented by clinical isolates. Obtaining sufficient stocks of wild-type clinical HCMV is often labor intensive and inefficient due to low yield and genetic loss, presenting a barrier to studies of clinical isolates. Here we report a bioreactor method based on continuous infection, where retinal pigment epithelial (ARPE-19) cells adhered to microcarrier beads are infected in a bioreactor and used to produce high-titers of clinical isolate HCMV that maintain genetic integrity of key viral tropism factors and the viral genome. In this bioreactor, an end-stage infection can be maintained by regular addition of uninfected ARPE-19 cells, providing convenient preparation of 10 7 -10 8 pfu/ml of concentrated TB40/E IE2-EYFP stocks without daily cell passaging or trypsinization. Overall, this represents a 100-fold increase in gain of virus production of 100-times compared to conventional static-culture plates, while requiring 90% less handling time. Moreover, this continuous infection environment has the potential to monitor infection dynamics with applications for real-time tracking of viral evolution.

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.

Performance of the CLSI Carba NP and the Rosco Carb Screen Assays Using North American Carbapenemase-Producing Enterobacteriaceae and Pseudomonas aeruginosa Isolates

PubMed Central

Gallagher, Lauren C.; Roundtree, Sylvester S.; Lancaster, Diana P.; Rudin, Susan D.; Bard, Jennifer Dien; Roberts, Amity L.; Marshall, Steven H.; Bonomo, Robert A.

2015-01-01

This study compared the performance of the Carba NP assay, published by the Clinical and Laboratory Standards Institute, and the Rosco Rapid Carb Screen kit. Carba NP had superior sensitivity, but both assays required an increased inoculum to detect carbapenemase production in isolates with blaNDM, blaIMP, and blaOXA-48. PMID:26269624

Transcriptomics in human blood incubation reveals the importance of oxidative stress response in Saccharomyces cerevisiae clinical strains.

PubMed

Llopis, Silvia; Querol, Amparo; Heyken, Antje; Hube, Bernhard; Jespersen, Lene; Fernández-Espinar, M Teresa; Pérez-Torrado, Roberto

2012-08-23

In recent years an increasing number of yeast infections in humans have been related to certain clinical isolates of Saccharomyces cerevisiae. Some clinical strains showed in vivo and in vitro virulence traits and were able to cause death in mice whereas other clinical strains were avirulent. In this work, we studied the transcriptional profiles of two S. cerevisiae clinical strains showing virulent traits and two control non-virulent strains during a blood incubation model and detected a specific transcriptional response of clinical strains. This response involves an mRNA levels increase of amino acid biosynthesis genes and especially oxidative stress related genes. We observed that the clinical strains were more resistant to reactive oxygen species in vitro. In addition, blood survival of clinical isolates was high, reaching similar levels to pathogenic Candida albicans strain. Furthermore, a virulent strain mutant in the transcription factor Yap1p, unable to grow in oxidative stress conditions, presented decreased survival levels in human blood compared with the wild type or YAP1 reconstituted strain. Our data suggest that this enhanced oxidative stress response in virulent clinical isolates, presumably induced in response to oxidative burst from host defense cells, is important to increase survival in human blood and can help to infect and even produce death in mice models.

Survey for collagenase gene prtC in Porphyromonas gingivalis and Porphyromonas endodontalis isolated from endodontic infections.

PubMed

Odell, L J; Baumgartner, J C; Xia, T; David, L L

1999-08-01

Collagenase is a potential virulence factor shown to be expressed by Porphyromonas gingivalis associated with periodontal disease. The purpose of this study was to use the polymerase chain reaction (PCR) to detect the presence of the collagenase gene (prtC) in 21 strains of Porphyromonas species isolated from endodontic infections. Type strains for P. gingivalis (ATCC 33277), P. endodontalis (ATCC 35406), Prevotella intermedia (ATCC 25611), and Prevotella nigrescens (ATCC 33563) were used as controls. When PCR primers specific for the 16S ribosomal RNA gene of P. gingivalis or P. endodontalis were used, 16 of the strains were identified as P. gingivalis, and five strains were identified as P. endodontalis. The presence of the prtC gene for collagenase was detected using PCR. Amplicons were analyzed by agarose gel electrophoresis, with an 815 bp amplicon representing the presence of the collagenase gene. Type strain ATCC 33277 and all 16 clinical isolates of P. gingivalis produced the collagenase gene amplicon. Neither type strain ATCC 35406 nor the five strains from clinical isolates of P. endodontalis produced the collagenase gene amplicon. These results indicate that P. gingivalis from endodontic infections possesses the prtC gene. P. endodontalis does not seem to exhibit prtC. The virulence of P. gingivalis may be related to its production of collagenase.

Epidemiology of extended-spectrum β-lactamase, AmpC, and carbapenemase production in Proteus mirabilis.

PubMed

Datta, Priya; Gupta, Varsha; Arora, Shilpa; Garg, Shivani; Chander, Jagdish

2014-01-01

Proteus mirabilis strains that produce extended-spectrum β-lactamase (ESBL), AmpC β-lactamase, and carbapenemase pose potential threats to patient care because most clinical diagnostic laboratories may not attempt to detect these three major groups of enzymes. Therefore, the objective of this study was to ascertain if P. mirabilis isolates collected from our heathcare facility possess various mechanisms of resistance to β-lactams (i.e., ESBL, AmpC, and carbapenemases) and to additionally arrive at conclusions regarding concurrent testing for these three mechanism of drug resistance in order to reduce cost and time in routine diagnostic testing. Between January 2011 and June 2011, 60 consecutive non-repeated strains of P. mirabilis were evaluated for production of ESBLs, AmpC β-lactamases, and carbapenemases. Of these, 36 isolates were found to be ESBL producers, and 7 (12%) were positive for production of AmpC β-lactamases and ESBLs. Therefore, 19.4% of ESBL-producing Proteus strains coproduced AmpC enzymes. The modified Hodge test confirmed carbapenemase production in only 1 isolate (1.7%), which was also ESBL- and AmpC-positive. The clinical impact of additional AmpC expression in ESBL-producing P. mirabilis results in a newly acquired resistance to β-lactamase inhibitors. In addition, to save time and costs, we recommend the use of cefepime/cefepime-clavulanate or boronic acid for the ESBL detection but in only those strains that were positive for ESBL by screening and negative by confirmatory tests.

Analysis of the cellulose synthase operon genes, bcsA, bcsB, and bcsC in Cronobacter species: Prevalence among species and their roles in biofilm formation and cell-cell aggregation.

PubMed

Hu, Lan; Grim, Christopher J; Franco, Augusto A; Jarvis, Karen G; Sathyamoorthy, Vengopal; Kothary, Mahendra H; McCardell, Barbara A; Tall, Ben D

2015-12-01

Cronobacter species are emerging food-borne pathogens that cause severe sepsis, meningitis, and necrotizing entercolitis in neonates and infants. Bacterial pathogens such as Escherichia coli and Salmonella species produce extracellular cellulose which has been shown to be involved in rugosity, biofilm formation, and host colonization. In this study the distribution and prevalence of cellulose synthase operon genes (bcsABZC) were determined by polymerase chain reaction (PCR) analysis in 231 Cronobacter strains isolated from clinical, food, environmental, and unknown sources. Furthermore, bcsA and bcsB isogenic mutants were constructed in Cronobacter sakazakii BAA894 to determine their roles. In calcofluor binding assays bcsA and bcsB mutants did not produce cellulose, and their colonial morphotypes were different to that of the parent strain. Biofilm formation and bacterial cell-cell aggregation were significantly reduced in bcsA and bcsB mutants compared to the parental strain. bcsA or bcsAB PCR-negative strains of C. sakazakii did not bind calcofluor, and produced less biofilm and cell-cell aggregation compared to strains possessing bcsAB genes. These data indicated that Cronobacter bcsABZC were present in all clinical isolates and most of food and environmental isolates. bcsA and bcsB genes of Cronobacter were necessary to produce cellulose, and were involved in biofilm formation and cell-cell aggregation. Published by Elsevier Ltd.

Emerge of blaNDM-1 and blaOXA-48-like harboring carbapenem-resistant Klebsiella pneumoniae isolates from hospitalized patients in southwestern Iran.

PubMed

Hosseinzadeh, Zahra; Sedigh Ebrahim-Saraie, Hadi; Sarvari, Jamal; Mardaneh, Jalal; Dehghani, Behzad; Rokni-Hosseini, Seyed Mohammad Hossein; Motamedifar, Mohammad

2018-06-01

One of the most important emerging carbapenem-resistant bacteria is Klebsiella pneumoniae (K. pneumoniae). The present study aimed to investigate the antibiotic susceptibility pattern of K. pneumoniae isolates and detection of carbapenemase producing K. pneumoniae obtained from Iranian hospitalized patients. This cross-sectional study was performed on 211 K. pneumoniae isolates which were recovered from different clinical specimens from 2014 to 2015. Modified Hodge test (MHT) and double disk synergy test (DDST) were done for detection of carbapenemase and metallo-beta-lactamase (MBL) producing K. pneumoniae. The presence of antibiotic resistance determinants was investigated by polymerase chain reaction (PCR) method. The results of antibiotic susceptibility showed that all isolates were resistant to ampicillin, and then mostly resistant to piperacillin and ceftazidime with 76.3% and 66.8%, respectively. On the other hand, the highest sensitivity was toward polymyxin B, followed by carbapenems. Of 29 carbapenem-resistant isolates, all were high-level imipenem-resistant isolates (Minimum inhibitory concentration ≥4), except 4 isolates. The results of MHT and DDST showed that 93.1% (27/29) of carbapenem-resistant isolates were carbapenemase and MBL producing isolates, respectively. The presence of bla NDM-1 and bla OXA-48-like genes was detected in 27 (10.9%) and 2 (0.9%) isolates, respectively. This is the first identification of bla NDM-1 and bla OXA-48-like in K. pneumoniae in Southwestern Iran and the highest reported prevalence of bla NDM in this bacterium from Iran. Since carbapenem-resistant isolates containing New Delhi metallo-beta-lactamase 1 (NDM-1) were almost resistant to all the tested antibiotics, the resistance due to this gene may be increased in the near future as a potential health threat. Copyright © 2017. Published by Elsevier Taiwan LLC.

Candida species biofilm and Candida albicans ALS3 polymorphisms in clinical isolates

PubMed Central

Bruder-Nascimento, Ariane; Camargo, Carlos Henrique; Mondelli, Alessandro Lia; Sugizaki, Maria Fátima; Sadatsune, Terue; Bagagli, Eduardo

2014-01-01

Over the last decades, there have been important changes in the epidemiology of Candida infections. In recent years, Candida species have emerged as important causes of invasive infections mainly among immunocompromised patients. This study analyzed Candida spp. isolates and compared the frequency and biofilm production of different species among the different sources of isolation: blood, urine, vulvovaginal secretions and peritoneal dialysis fluid. Biofilm production was quantified in 327 Candida isolates obtained from patients attended at a Brazilian tertiary public hospital (Botucatu, Sao Paulo). C. albicans ALS3 gene polymorphism was also evaluated by determining the number of repeated motifs in the central domain. Of the 198 total biofilm-positive isolates, 72 and 126 were considered as low and high biofilm producers, respectively. Biofilm production by C. albicans was significantly lower than that by non-albicans isolates and was most frequently observed in C. tropicalis. Biofilm production was more frequent among bloodstream isolates than other clinical sources, in urine, the isolates displayed a peculiar distribution by presenting two distinct peaks, one containing biofilm-negative isolates and the other containing isolates with intense biofilm production. The numbers of tandem-repeat copies per allele were not associated with biofilm production, suggesting the evolvement of other genetic determinants. PMID:25763043

First report of extended-spectrum β-lactamases among clinical isolates of Escherichia coli in Gaza Strip, Palestine.

PubMed

Tayh, Ghassan; Ben Sallem, Rym; Ben Yahia, Houssem; Gharsa, Haythem; Klibi, Naouel; Boudabous, Abdellatif; Ben Slama, Karim

2016-09-01

This study aimed to assess the occurrence of extended-spectrum β-lactamases (ESBLs) in clinical Escherichia coli isolates from Palestine and to characterise their type, genetic environments and associated resistance genes. Twenty-seven broad-spectrum-cephalosporin-resistant E. coli isolates were recovered between April and June 2013 in Gaza Strip hospitals. Characterisation of ESBL genes and their genetic environments, detection of associated resistance genes, and the presence and characterisation of integrons were performed by PCR and sequencing. The clonal relationship among ESBL-positive E. coli strains was determined by pulsed-field gel electrophoresis (PFGE) using the restriction enzyme XbaI. Phylogroup typing and virulence factors were studied by PCR. The following ESBL genes were identified: blaCTX-M-15 (21 isolates); blaCTX-M-14a (2 isolates); blaCTX-M-1 (2 isolates); blaCTX-M-3 (1 isolate); and blaCTX-M-27 (1 isolate). The blaTEM-1 gene was also detected in eight CTX-M-producing strains. The ISEcp1 sequence was found upstream of blaCTX-M in 23 isolates, and orf477 was found downstream of this gene in 24 isolates. IS903 was also detected downstream in three isolates. Six isolates carried class 1 integrons with the gene cassette arrangement dfrA17-aadA5. High clonal diversity was observed among the studied isolates by PFGE (24 unrelated profiles). The virulence gene fimA was detected in 23 isolates, the aer gene in 8 isolates and the papC gene in 7 isolates. The studied isolates belonged to phylogroups B2 (12 isolates), D (12 isolates) and A (3 isolates). This is the first report of the detection of CTX-M class β-lactamases in E. coli of clinical origin in Gaza Strip hospitals. Copyright © 2016 International Society for Chemotherapy of Infection and Cancer. Published by Elsevier Ltd. All rights reserved.

Phylogeny of Shiga toxin-producing Escherichia coli O157 isolated from cattle and clinically ill humans

USDA-ARS?s Scientific Manuscript database

Cattle are a major reservoir for Shiga toxin-producing Escherichia coli O157 (STEC O157) and harbor multiple genetic subtypes that do not all associate with human disease. STEC O157 evolved from an E. coli O55:H7 progenitor, however, a lack of genome sequence has hindered investigations on the dive...

Exopolysaccharides produced by Burkholderia cenocepacia recA lineages IIIA and IIIB.

PubMed

Chiarini, Luigi; Cescutti, Paola; Drigo, Laura; Impallomeni, Giuseppe; Herasimenka, Yury; Bevivino, Annamaria; Dalmastri, Claudia; Tabacchioni, Silvia; Manno, Graziana; Zanetti, Flavio; Rizzo, Roberto

2004-08-01

Clinical and environmental strains of Burkholderia cenocepacia belonging to the recA lineages IIIA and IIIB were examined for exopolysaccharide (EPS) production. The exopolysaccharides structure was determined using mainly gas chromatography coupled to mass spectrometry and NMR spectroscopy. All the strains produced Cepacian, a highly branched polysaccharide constituted of a heptasaccharide repeating unit, composed of one rhamnose, one glucose, one glucuronic acid, one mannose and three galactose residues. This polymer is the most common exopolysaccharide produced by strains of the Burkholderia cepacia (Bcc) complex. One clinical strain produced also another polysaccharide constituted of three galactose units and one 3-deoxy-D-manno-2-octulosonic acid residues, a polymer that was previously isolated from two strains of B. cepacia genomovar I and B. cenocepacia IIIA.

Rapid optical determination of β-lactamase and antibiotic activity

PubMed Central

2014-01-01

Background The absence of rapid tests evaluating antibiotic susceptibility results in the empirical prescription of antibiotics. This can lead to treatment failures due to escalating antibiotic resistance, and also furthers the emergence of drug-resistant bacteria. This study reports a rapid optical method to detect β-lactamase and thereby assess activity of β-lactam antibiotics, which could provide an approach for targeted prescription of antibiotics. The methodology is centred on a fluorescence quenching based probe (β-LEAF – β-Lactamase Enzyme Activated Fluorophore) that mimics the structure of β-lactam antibiotics. Results The β-LEAF assay was performed for rapid determination of β-lactamase production and activity of β-lactam antibiotic (cefazolin) on a panel of Staphylococcus aureus ATCC strains and clinical isolates. Four of the clinical isolates were determined to be lactamase producers, with the capacity to inactivate cefazolin, out of the twenty-five isolates tested. These results were compared against gold standard methods, nitrocefin disk test for β-lactamase detection and disk diffusion for antibiotic susceptibility, showing results to be largely consistent. Furthermore, in the sub-set of β-lactamase producers, it was demonstrated and validated that multiple antibiotics (cefazolin, cefoxitin, cefepime) could be assessed simultaneously to predict the antibiotic that would be most active for a given bacterial isolate. Conclusions The study establishes the rapid β-LEAF assay for β-lactamase detection and prediction of antibiotic activity using S. aureus clinical isolates. Although the focus in the current study is β-lactamase-based resistance, the overall approach represents a broad diagnostic platform. In the long-term, these studies form the basis for the development of assays utilizing a broader variety of targets, pathogens and drugs. PMID:24708478

Identification and antimicrobial susceptibility of obligate anaerobic bacteria from clinical samples of animal origin.

PubMed

Mayorga, Melissa; Rodríguez-Cavallini, Evelyn; López-Ureña, Diana; Barquero-Calvo, Elías; Quesada-Gómez, Carlos

2015-12-01

The etiology of veterinary infectious diseases has been the focus of considerable research, yet relatively little is known about the causative agents of anaerobic infections. Susceptibility studies have documented the emergence of antimicrobial resistance and indicate distinct differences in resistance patterns related to veterinary hospitals, geographic regions, and antibiotic-prescribing regimens. The aim of the present study was to identify the obligate anaerobic bacteria from veterinary clinical samples and to determinate the in vitro susceptibility to eight antimicrobials and their resistance-associated genes. 81 clinical specimens obtained from food-producing animals, pets and wild animals were examined to determine the relative prevalence of obligate anaerobic bacteria, and the species represented. Bacteroides spp, Prevotella spp and Clostridium spp represented approximately 80% of all anaerobic isolates. Resistance to metronidazole, clindamycin, tetracycline and fluoroquinolones was found in strains isolated from food-producing animals. Ciprofloxacin, enrofloxacin and cephalotin showed the highest resistance in all isolates. In 17%, 4% and 14% of tetracycline-resistant isolates, the resistance genes tetL, tetM and tetW were respectively amplified by PCR whereas in 4% of clindamycin-resistant strains the ermG gene was detected. 26% of the isolates were positive for cepA, while only 6% harbored the cfxA (resistance-conferring genes to beta-lactams). In this study, the obligate anaerobic bacteria from Costa Rica showed a high degree of resistance to most antimicrobials tested. Nevertheless, in the majority of cases this resistance was not related to the resistance acquired genes usually described in anaerobes. It is important to address and regulate the use of antimicrobials in the agricultural industry and the empirical therapy in anaerobic bacterial infections in veterinary medicine, especially since antibiotics and resistant bacteria can persist in the environment. Copyright © 2015 Elsevier Ltd. All rights reserved.

Genetic & virulence profiling of ESBL-positive E. coli from nosocomial & veterinary sources.

PubMed

Tyrrell, J M; Wootton, M; Toleman, M A; Howe, R A; Woodward, M; Walsh, T R

2016-04-15

CTX-M genes are the most prevalent ESBL globally, infiltrating nosocomial, community and environmental settings. Wild and domesticated animals may act as effective vectors for the dissemination of CTX-producing Enterobacteriaceae. This study aimed to contextualise blaCTX-M-14-positive, cephalosporin-resistant Enterobacteriaceae human infections and compared resistance and pathogenicity markers with veterinary isolates. Epidemiologically related human (n=18) and veterinary (n=4) blaCTX-M-14-positive E. coli were fully characterised. All were typed by XbaI pulsed field gel electrophoresis and ST. Chromosomal/plasmidic locations of blaCTX-M-14 were deduced by S1-nuclease digestion, and association with ISEcp1 was investigated by sequencing. Conjugation experiments assessed transmissibility of plasmids carrying blaCTX-M-14. Presence of virulence determinants was screened by PCR assay and pathogenicity potential was determined by in vitro Galleria mellonella infection models. 84% of clinical E. coli originated from community patients. blaCTX-M-14 was found ubiquitously downstream of ISEcp1 upon conjugative plasmids (25-150 kb). blaCTX-M-14 was also found upon the chromosome of eight E. coli isolates. CTX-M-14-producing E. coli were found at multiple hospital sites. Clonal commonality between patient, hospitals and livestock microbial populations was found. In vivo model survival rates from clinical isolates (30%) and veterinary isolates (0%) were significantly different (p<0.05). Co-transfer of blaCTX-M-14 and virulence determinants was demonstrated. There is evidence of clonal spread of blaCTX-M-14-positive E. coli involving community patients and farm livestock. blaCTX-M-14 positive human clinical isolates carry a lower intrinsic pathogenic potential than veterinary E. coli highlighting the need for greater veterinary practices in preventing dissemination of MDR E. coli among livestock. Copyright © 2016. Published by Elsevier B.V.

Analysis of ESBL- and AmpC-positive Enterobacteriaceae at the Department of Neonatology, University Hospital Olomouc.

PubMed

Husičková, Vendula; Chromá, Magdaléna; Kolář, Milan; Hricová, Kristýna; Stosová, Taťána; Kantor, Lumír; Dubrava, Lubomír

2011-06-01

Bacterial infections are an important issue in current clinical medicine. The severity of infectious diseases has increased dramatically in recent years, which is also due to increasing numbers of resistant bacteria, including strains producing broad-spectrum beta-lactamases. The study aimed at determining the prevalence of ESBL- and AmpC-positive Enterobacteriaceae at the Department of Neonatology, University Hospital Olomouc. Enterobacteriaceae were isolated from clinical samples from infants hospitalized at the Department of Neonatology, University Hospital Olomouc over a period of 2 years. ESBL- and AmpC-positive isolates were subjected to basic genetic analysis. In the study period, a total of 1,526 isolates of the Enterobacteriaceae family were identified, including 55 (3.6%) cases of the ESBL phenotype and 17 (1.1%) AmpC-positive isolates. Genetic analysis of ESBL-positive isolates revealed a majority of CTX-M enzymes. Among AmpC beta-lactamases, the EBC, CIT, DHA, and MOX types were detected. An Escherichia coli strain was isolated with mutations in the promoter region of the ampC chromosomal gene that are associated with overproduction of the relevant enzyme.

Characterization of multidrug-resistant diabetic foot ulcer enterococci.

PubMed

Semedo-Lemsaddek, Teresa; Mottola, Carla; Alves-Barroco, Cynthia; Cavaco-Silva, Patrícia; Tavares, Luís; Oliveira, Manuela

2016-02-01

Diabetes mellitus is a highly prevalent chronic progressive disease with complications that include diabetic-foot ulcers. Enterococci isolated from diabetic-foot infections were identified, evaluated by macro-restriction analysis, and screened for virulence traits and antimicrobial resistance. All isolates were considered multidrug-resistant, cytolysin and gelatinase producers, and the majority also demonstrated the ability to produce biofilms. These results indicate the importance of enterococci in diabetic-foot infection development and persistence, especially regarding their biofilm-forming ability and resistance to clinically relevant antibiotics. Copyright © 2015 Elsevier España, S.L.U. y Sociedad Española de Enfermedades Infecciosas y Microbiología Clínica. All rights reserved.

Potential of Ocimum basilicum L. and Salvia officinalis L. essential oils against biofilms of P. aeruginosa clinical isolates.

PubMed

Stojanović-Radić, Z; Pejcić, M; Stojanović, N; Sharifi-Rad, J; Stanković, N

2016-08-29

Biofilms are complex communities of microorganisms, responsible for more than 60% of the chronic human infections and they represent one of the leading concerns in medicine. Pseudomonas aeruginosa is human pathogenic bacteria which causes numerous diseases and is known for its ability to produce biofilm. Ocimum basilicum L. (basil) and Salvia officinalis L. (sage) are widely used plants in traditional medicine for the treatment of different conditions. Therefore, the aim of this study was to investigate the potential of basil and sage essential oils against P. aeruginosa biofilm producing strains. The efficacy of two essential oils on P. aeruginosa biofilm forming ability was determined using crystal violet method. Out of 15 strains isolated from different clinical biological samples, two were strong, 11 moderate and one weak biofilm producer. Good efficacy of sage essential oil towards strong and weak biofilm producers, but not of basil essential oil, was observed. In the case of moderate biofilm producers, 81.8% showed lower biofilm production after incubation with the sage oil, while 63.6% showed the reduction of biofilm production after basil essential oil treatment. The obtained results showed high potential of both oils for the treatment of persistent infections caused by Pseudomonas aeruginosa biofilms.

Bacteriocins of Non-aureus Staphylococci Isolated from Bovine Milk.

PubMed

Carson, Domonique A; Barkema, Herman W; Naushad, Sohail; De Buck, Jeroen

2017-09-01

Non- aureus staphylococci (NAS), the bacteria most commonly isolated from the bovine udder, potentially protect the udder against infection by major mastitis pathogens due to bacteriocin production. In this study, we determined the inhibitory capability of 441 bovine NAS isolates (comprising 26 species) against bovine Staphylococcus aureus Furthermore, inhibiting isolates were tested against a human methicillin-resistant S. aureus (MRSA) isolate using a cross-streaking method. We determined the presence of bacteriocin clusters in NAS whole genomes using genome mining tools, BLAST, and comparison of genomes of closely related inhibiting and noninhibiting isolates and determined the genetic organization of any identified bacteriocin biosynthetic gene clusters. Forty isolates from 9 species ( S. capitis , S. chromogenes , S. epidermidis , S. pasteuri , S. saprophyticus , S. sciuri , S. simulans , S. warneri , and S. xylosus ) inhibited growth of S. aureus in vitro , 23 isolates of which, from S. capitis , S. chromogenes , S. epidermidis , S. pasteuri , S. simulans , and S. xylosus , also inhibited MRSA. One hundred five putative bacteriocin gene clusters encompassing 6 different classes (lanthipeptides, sactipeptides, lasso peptides, class IIa, class IIc, and class IId) in 95 whole genomes from 16 species were identified. A total of 25 novel bacteriocin precursors were described. In conclusion, NAS from bovine mammary glands are a source of potential bacteriocins, with >21% being possible producers, representing potential for future characterization and prospective clinical applications. IMPORTANCE Mastitis (particularly infections caused by Staphylococcus aureus ) costs Canadian dairy producers $400 million/year and is the leading cause of antibiotic use on dairy farms. With increasing antibiotic resistance and regulations regarding use, there is impetus to explore bacteriocins (bacterially produced antimicrobial peptides) for treatment and prevention of bacterial infections. We examined the ability of 441 NAS bacteria from Canadian bovine milk samples to inhibit growth of S. aureus in the laboratory. Overall, 9% inhibited growth of S. aureus and 58% of those also inhibited MRSA. In NAS whole-genome sequences, we identified >21% of NAS as having bacteriocin genes. Our study represents a foundation to further explore NAS bacteriocins for clinical use. Copyright © 2017 American Society for Microbiology.

Bacteriocins of Non-aureus Staphylococci Isolated from Bovine Milk

PubMed Central

Carson, Domonique A.; Barkema, Herman W.; Naushad, Sohail

2017-01-01

ABSTRACT Non-aureus staphylococci (NAS), the bacteria most commonly isolated from the bovine udder, potentially protect the udder against infection by major mastitis pathogens due to bacteriocin production. In this study, we determined the inhibitory capability of 441 bovine NAS isolates (comprising 26 species) against bovine Staphylococcus aureus. Furthermore, inhibiting isolates were tested against a human methicillin-resistant S. aureus (MRSA) isolate using a cross-streaking method. We determined the presence of bacteriocin clusters in NAS whole genomes using genome mining tools, BLAST, and comparison of genomes of closely related inhibiting and noninhibiting isolates and determined the genetic organization of any identified bacteriocin biosynthetic gene clusters. Forty isolates from 9 species (S. capitis, S. chromogenes, S. epidermidis, S. pasteuri, S. saprophyticus, S. sciuri, S. simulans, S. warneri, and S. xylosus) inhibited growth of S. aureus in vitro, 23 isolates of which, from S. capitis, S. chromogenes, S. epidermidis, S. pasteuri, S. simulans, and S. xylosus, also inhibited MRSA. One hundred five putative bacteriocin gene clusters encompassing 6 different classes (lanthipeptides, sactipeptides, lasso peptides, class IIa, class IIc, and class IId) in 95 whole genomes from 16 species were identified. A total of 25 novel bacteriocin precursors were described. In conclusion, NAS from bovine mammary glands are a source of potential bacteriocins, with >21% being possible producers, representing potential for future characterization and prospective clinical applications. IMPORTANCE Mastitis (particularly infections caused by Staphylococcus aureus) costs Canadian dairy producers $400 million/year and is the leading cause of antibiotic use on dairy farms. With increasing antibiotic resistance and regulations regarding use, there is impetus to explore bacteriocins (bacterially produced antimicrobial peptides) for treatment and prevention of bacterial infections. We examined the ability of 441 NAS bacteria from Canadian bovine milk samples to inhibit growth of S. aureus in the laboratory. Overall, 9% inhibited growth of S. aureus and 58% of those also inhibited MRSA. In NAS whole-genome sequences, we identified >21% of NAS as having bacteriocin genes. Our study represents a foundation to further explore NAS bacteriocins for clinical use. PMID:28667105

Shiga Toxin 1–Producing Shigella sonnei Infections, California, United States, 2014–2015

PubMed Central

Nelson, Jennifer A.; Kimura, Akiko C.; Poe, Alyssa; Collins, Joan; Kao, Annie S.; Cruz, Laura; Inami, Gregory; Vaishampayan, Julie; Garza, Alvaro; Chaturvedi, Vishnu; Vugia, Duc J.

2016-01-01

Shiga toxins (Stx) are primarily associated with Shiga toxin–producing Escherichia coli and Shigella dysenteriae serotype 1. Stx production by other shigellae is uncommon, but in 2014, Stx1-producing S. sonnei infections were detected in California. Surveillance was enhanced to test S. sonnei isolates for the presence and expression of stx genes, perform DNA subtyping, describe clinical and epidemiologic characteristics of case-patients, and investigate for sources of infection. During June 2014–April 2015, we identified 56 cases of Stx1-producing S. sonnei, in 2 clusters. All isolates encoded stx1 and produced active Stx1. Multiple pulsed-field gel electrophoresis patterns were identified. Bloody diarrhea was reported by 71% of case-patients; none had hemolytic uremic syndrome. Some initial cases were epidemiologically linked to travel to Mexico, but subsequent infections were transmitted domestically. Continued surveillance of Stx1-producing S. sonnei in California is necessary to characterize its features and plan for reduction of its spread in the United States. PMID:26982255

Comparative Genomic Analysis of a Clinical Isolate of Klebsiella quasipneumoniae subsp. similipneumoniae, a KPC-2 and OKP-B-6 Beta-Lactamases Producer Harboring Two Drug-Resistance Plasmids from Southeast Brazil

PubMed Central

Nicolás, Marisa F.; Ramos, Pablo Ivan Pereira; Marques de Carvalho, Fabíola; Camargo, Dhian R. A.; de Fátima Morais Alves, Carlene; Loss de Morais, Guilherme; Almeida, Luiz G. P.; Souza, Rangel C.; Ciapina, Luciane P.; Vicente, Ana C. P.; Coimbra, Roney S.; Ribeiro de Vasconcelos, Ana T.

2018-01-01

The aim of this study was to unravel the genetic determinants responsible for multidrug (including carbapenems) resistance and virulence in a clinical isolate of Klebsiella quasipneumoniae subsp. similipneumoniae by whole-genome sequencing and comparative analyses. Eighty-three clinical isolates initially identified as carbapenem-resistant K. pneumoniae were collected from nosocomial infections in southeast Brazil. After RAPD screening, the KPC-142 isolate, showing the most divergent DNA pattern, was selected for complete genome sequencing in an Illumina HiSeq 2500 instrument. Reads were assembled into scaffolds, gaps between scaffolds were resolved by in silico gap filling and extensive bioinformatics analyses were performed, using multiple comparative analysis tools and databases. Genome sequencing allowed to correct the classification of the KPC-142 isolate as K. quasipneumoniae subsp. similipneumoniae. To the best of our knowledge this is the first complete genome reported to date of a clinical isolate of this subspecies harboring both class A beta-lactamases KPC-2 and OKP-B-6 from South America. KPC-142 has one 5.2 Mbp chromosome (57.8% G+C) and two plasmids: 190 Kbp pKQPS142a (50.7% G+C) and 11 Kbp pKQPS142b (57.3% G+C). The 3 Kbp region in pKQPS142b containing the blaKPC−2 was found highly similar to that of pKp13d of K. pneumoniae Kp13 isolated in Southern Brazil in 2009, suggesting the horizontal transfer of this resistance gene between different species of Klebsiella. KPC-142 additionally harbors an integrative conjugative element ICEPm1 that could be involved in the mobilization of pKQPS142b and determinants of resistance to other classes of antimicrobials, including aminoglycoside and silver. We present the completely assembled genome sequence of a clinical isolate of K. quasipneumoniae subsp. similipneumoniae, a KPC-2 and OKP-B-6 beta-lactamases producer and discuss the most relevant genomic features of this important resistant pathogen in comparison to several strains belonging to K. quasipneumoniae subsp. similipneumoniae (phylogroup II-B), K. quasipneumoniae subsp. quasipneumoniae (phylogroup II-A), K. pneumoniae (phylogroup I), and K. variicola (phylogroup III). Our study contributes to the description of the characteristics of a novel K. quasipneumoniae subsp. similipneumoniae strain circulating in South America that currently represent a serious potential risk for nosocomial settings. PMID:29503635

Genetic Diversity, Antimicrobial Susceptibility, and Biofilm Formation of Cronobacter spp. Recovered from Spices and Cereals

PubMed Central

Li, Yuanhong; Yu, Huan; Jiang, Hua; Jiao, Yang; Zhang, Yaodong; Shao, Jihong

2017-01-01

Cronobacter species are important food-borne opportunistic pathogens which have been implicated in the cause of necrotizing enterocolitis, sepsis, and meningitis in neonates and infants. However, these bacteria are routinely found in foodstuffs, clinical specimens, and environmental samples. This study investigated the genetic diversity, antimicrobial susceptibility, and biofilm formation of Cronobacter isolates (n = 40) recovered from spices and cereals in China during 2014–2015. Based on the fusA sequencing analysis, we found that the majority (23/40, 57.5%) of Cronobacter isolates in spices and cereals were C. sakazakii, while the remaining strains were C. dublinensis (6/40, 15.0%), C. malonaticus (5/40, 12.5%), C. turicensis (4/40, 10.0%), and C. universalis (2/40, 5.0%). Multilocus sequence typing (MLST) analysis produced 30 sequence types (STs) among the 40 Cronobacter isolates, with 5 STs (ST4, ST13, ST50, ST129, and ST158) related to neonatal meningitis. The pattern of the overall ST distribution was diverse; in particular, it was revealed that ST148 was the predominant ST, presenting 12.5% within the whole population. MLST assigned 12 isolates to 7 different clonal complexes (CCs), 4, 13, 16, 17, 72, 129, and 143, respectively. The results of O-antigen serotyping indicated that C. sakazakii serotype O1 and O2 were the most two prevalent serotypes. The antimicrobial susceptibility testing showed that the 40 Cronobacter isolates were susceptible to most of the antibiotics tested except for ceftriaxone, meropenem, and aztreona. Of the 40 Cronobacter strains tested, 13 (32.5%) were assessed as weak bioflim producers, one (2.5%) was a moderate biofilm producer, one (2.5%) was strong biofilm producer, and the others (62.5%) were non-biofilm producers. MLST and O-antigen serotyping have indicated that Cronobacter strains recovered from spices and cereals were genetically diverse. Isolates of clinical origin, particularly the C. sakazakii ST4 neonatal meningitic pathovar, have been identified from spices and cereals. Moreover, antimicrobial resistance of Cronobacter strains was observed, which may imply a potential public health risk. Therefore, the surveillance of Cronobacter spp. in spices and cereals should be strengthened to improve epidemiological understandings of Cronobacter infections. PMID:29312246

The spread of carbapenemase-producing bacteria in Africa: a systematic review.

PubMed

Manenzhe, Rendani I; Zar, Heather J; Nicol, Mark P; Kaba, Mamadou

2015-01-01

Carbapenems are the last line of defence against ever more prevalent MDR Gram-negative bacteria, but their efficacy is threatened worldwide by bacteria that produce carbapenemase enzymes. The epidemiology of bacteria producing carbapenemases has been described in considerable detail in Europe, North America and Asia; however, little is known about their spread and clinical relevance in Africa. We systematically searched in PubMed, EBSCOhost, Web of Science, Scopus, Elsevier Masson Consulte and African Journals Online, international conference proceedings, published theses and dissertations for studies reporting on carbapenemase-producing bacteria in Africa. We included articles published in English or French up to 28 February 2014. We calculated the prevalence of carbapenemase producers only including studies where the total number of isolates tested was at least 30. Eighty-three studies were included and analysed. Most studies were conducted in North Africa (74%, 61/83), followed by Southern Africa (12%, 10/83), especially South Africa (90%, 9/10), West Africa (8%, 7/83) and East Africa (6%, 6/83). Carbapenemase-producing bacteria were isolated from humans, the hospital environment and community environmental water samples, but not from animals. The prevalence of carbapenemase-producing isolates in hospital settings ranged from 2.3% to 67.7% in North Africa and from 9% to 60% in sub-Saharan Africa. Carbapenemase-producing bacteria have been described in many African countries; however, their prevalence is poorly defined and has not been systematically studied. Antibiotic stewardship and surveillance systems, including molecular detection and genotyping of resistant isolates, should be implemented to monitor and reduce the spread of carbapenemase-producing bacteria. © 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.

Dissemination of a Multidrug-Resistant VIM-1- and CMY-99-Producing Proteus mirabilis Clone in Bulgaria.

PubMed

Markovska, Rumyana; Schneider, Ines; Keuleyan, Emma; Ivanova, Dobrinka; Lesseva, Magdalena; Stoeva, Temenuga; Sredkova, Mariya; Bauernfeind, Adolf; Mitov, Ivan

2017-04-01

The aim of this study was to analyze the beta-lactamases and the molecular epidemiology of 19 clinically significant isolates of Proteus mirabilis with decreased susceptibility to imipenem, which have been collected from seven hospitals, located in different Bulgarian towns (Sofia, Varna, and Pleven). The isolates were obtained from blood, urine, tracheal and wound specimens. One additional isolate from hospital environment was included. Susceptibility testing, conjugation experiments, and plasmid replicon typing were carried out. Beta-lactamases were characterized by isoelectric focusing, PCR, and sequencing. Clonal relatedness was investigated by RAPD and PFGE. Integron mapping was performed by PCR and sequencing. All isolates showed a multidrug-resistance profile, but remained susceptible to piperacillin/tazobactam, cefepime, meropenem, and fosfomycin. They produced identical beta-lactamases, namely: TEM-1, VIM-1, and CMY-99. PCR mapping revealed that the bla VIM-1 gene was part of a class 1 integron that additionally included the aac(6')-I, dhfrA1, and ant(3″)-Ia genes. In addition, 17 of the isolates carried the armA gene. Conjugation experiments and plasmid replicon typing were unsuccessful. The isolates were clonally related according to RAPD and PFGE typing. This study reveals the nationwide distribution of a multidrug-resistant P. mirabilis clone producing VIM-1 and CMY-99 along with the presence of different aminoglycoside resistance mechanisms.

Long-Term, Low-Frequency Cluster of a German-Imipenemase-1-Producing Enterobacter hormaechei ssp. steigerwaltii ST89 in a Tertiary Care Hospital in Germany.

PubMed

Wendel, Andreas F; Meyer, Sebastian; Deenen, René; Köhrer, Karl; Kolbe-Busch, Susanne; Pfeffer, Klaus; Willmann, Matthias; Kaasch, Achim J; MacKenzie, Colin R

2018-05-11

Enterobacter cloacae complex is a common cause of hospital outbreaks. A retrospective and prospective molecular analysis of carbapenem-resistant clinical isolates in a tertiary care center demonstrated an outbreak of a German-imipenemase-1 (GIM-1) metallo-beta-lactamase-producing Enterobacter hormaechei ssp. steigerwaltii affecting 23 patients between 2009 and 2016. Thirty-three isolates were sequence type 89 by conventional multilocus sequence typing (MLST) and displayed a maximum difference of 49 out of 3,643 targets in the ad-hoc core-genome MLST (cgMLST) scheme (SeqSphere+ software; Ridom, Münster, Germany). The relatedness of all isolates was confirmed by further maximum-likelihood phylogeny. One clonal complex of highly related isolates (≤15 allele difference in cgMLST) contained 17 patients, but epidemiological data only suggested five transmission events. The bla GIM-1 -gene was embedded in a class-1-integron (In770) and the Tn21-subgroup transposon Tn6216 (KC511628) on a 25-kb plasmid. Environmental screening detected one colonized sink trap in a service room. The outbreak was self-limited as no further bla GIM-1 -positive E. hormaechei has been isolated since 2016. Routine molecular screening of carbapenem-nonsusceptible gram-negative isolates detected a long-term, low-frequency outbreak of a GIM-1-producing E. hormaechei ssp. steigerwaltii clone. This highlights the necessity of molecular surveillance.

Dissemination and genetic support of broad-spectrum beta-lactam-resistant Escherichia coli strain isolated from two Tunisian hospitals during 2004-2012.

PubMed

Ayari, Khaoula; Bourouis, Amel; Chihi, Hela; Mahrouki, Sihem; Naas, Thierry; Belhadj, Omrane

2017-06-01

The dissemination of extended-spectrum β-lactamase (ESBL)-producing bacteria presented a great concern worldwide. Gram-negative organisms such as Escherichia coli and Klebsiella pneumoniae are the most frequently isolated pathogens responsible for nosocomial infections. The aim of this study was to investigate and to follow the emergence of resistance and the characterization of Extended-Spectrum Beta-Lactamases (ESBL) among broad-spectrum beta-lactam- Escherichia coli clinical isolates recovered from the military hospital and Habib Thameur hospital in Tunisia. A total of 113 E.coli isolates obtained during the period 2004 through 2012 showed a significant degree of multi-resistance. Among these strains, the double-disk synergy test confirmed the ESBL phenotype in 46 isolates. These included 32(70%) strains from Hospital A and 14(30%) from Hospital B. The ESBL was identified as CTX-M-15. The ESBL resistance was transferred by a 60 kb plasmid CTXM-15-producing isolates were unrelated according to the PFGE analysis and characterization of the regions surrounding the blaCTX-M-15 showed the ISEcp1 elements located in the upstream region of the bla gene and 20 of them truncated by IS26. ESBL producing E. coli strains are a serious threat in the community in Tunisia and we should take into consideration any possible spread of such epidemiological resistance.

Unacceptably High Error Rates in Vitek 2 Testing of Cefepime Susceptibility in Extended-Spectrum-β-Lactamase-Producing Escherichia coli

PubMed Central

Rhodes, Nathaniel J.; Richardson, Chad L.; Heraty, Ryan; Liu, Jiajun; Malczynski, Michael; Qi, Chao

2014-01-01

While a lack of concordance is known between gold standard MIC determinations and Vitek 2, the magnitude of the discrepancy and its impact on treatment decisions for extended-spectrum-β-lactamase (ESBL)-producing Escherichia coli are not. Clinical isolates of ESBL-producing E. coli were collected from blood, tissue, and body fluid samples from January 2003 to July 2009. Resistance genotypes were identified by PCR. Primary analyses evaluated the discordance between Vitek 2 and gold standard methods using cefepime susceptibility breakpoint cutoff values of 8, 4, and 2 μg/ml. The discrepancies in MICs between the methods were classified per convention as very major, major, and minor errors. Sensitivity, specificity, and positive and negative predictive values for susceptibility classifications were calculated. A total of 304 isolates were identified; 59% (179) of the isolates carried blaCTX-M, 47% (143) carried blaTEM, and 4% (12) carried blaSHV. At a breakpoint MIC of 8 μg/ml, Vitek 2 produced a categorical agreement of 66.8% and exhibited very major, major, and minor error rates of 23% (20/87 isolates), 5.1% (8/157 isolates), and 24% (73/304), respectively. The sensitivity, specificity, and positive and negative predictive values for a susceptibility breakpoint of 8 μg/ml were 94.9%, 61.2%, 72.3%, and 91.8%, respectively. The sensitivity, specificity, and positive and negative predictive values for a susceptibility breakpoint of 2 μg/ml were 83.8%, 65.3%, 41%, and 93.3%, respectively. Vitek 2 results in unacceptably high error rates for cefepime compared to those of agar dilution for ESBL-producing E. coli. Clinicians should be wary of making treatment decisions on the basis of Vitek 2 susceptibility results for ESBL-producing E. coli. PMID:24752253

Comparison of the Carba NP, Modified Carba NP, and Updated Rosco Neo-Rapid Carb Kit Tests for Carbapenemase Detection

PubMed Central

AbdelGhani, Sameh; Thomson, Gina K.; Snyder, James W.

2015-01-01

The accurate detection of carbapenemase-producing organisms is a major challenge for clinical laboratories. The Carba NP test is highly accurate but inconvenient, as it requires frequent preparation of fresh imipenem solution. The current study was designed to compare the Carba NP test to two alternative tests for accuracy and convenience. These were a modified Carba NP test that utilized intravenous (i.v.) imipenem-cilastatin, which is less expensive than reference standard imipenem powder, and an updated version of the Rosco Neo-Rapid Carb kit, which does not require the preparation of imipenem solution and has a shelf life of 2 years. The comparison included 87 isolates that produced class A carbapenemases (including KPC-2, -3, -4, -5, -6, and -8, NMC-A, and SME type), 40 isolates that produced metallo-β-lactamases (including NDM-1, GIM-1, SPM-1, IMP-1, -2, -7, -8, -18, and -27, and VIM-1, -2, and -7), 11 isolates that produced OXA-48, and one isolate that produced OXA-181. Negative controls consisted of 50 isolates that produced extended-spectrum β-lactamases (ESBLs), AmpCs (including hyperproducers), K1, other limited-spectrum β-lactamases, and porin and efflux mutants. Each test exhibited 100% specificity and high sensitivity (Carba NP, 100%; Rosco, 99% using modified interpretation guidelines; and modified Carba NP, 96%). A modified approach to interpretation of the Rosco test was necessary to achieve the sensitivity of 99%. If the accuracy of the modified interpretation is confirmed, the Rosco test is an accurate and more convenient alternative to the Carba NP test. PMID:26311862

Prevalence of Shiga toxin-producing Shigella species isolated from French travellers returning from the Caribbean: an emerging pathogen with international implications.

PubMed

Gray, M D; Lacher, D W; Leonard, S R; Abbott, J; Zhao, S; Lampel, K A; Prothery, E; Gouali, M; Weill, F-X; Maurelli, A T

2015-08-01

Shiga toxins (Stxs) are potent cytotoxins that inhibit host cell protein synthesis, leading to cell death. Classically, these toxins are associated with intestinal infections due to Stx-producing Escherichia coli or Shigella dysenteriae serotype 1, and infections with these strains can lead to haemolytic-uraemic syndrome. Over the past decade, there has been increasing recognition that Stx is produced by additional Shigella species. We recently reported the presence and expression of stx genes in Shigella flexneri 2a clinical isolates. The toxin genes were carried by a new stx-encoding bacteriophage, and infection with these strains correlated with recent travel to Haiti or the Dominican Republic. In this study, we further explored the epidemiological link to this region by utilizing the French National Reference Centre for Escherichia coli, Shigella and Salmonella collection to survey the frequency of Stx-producing Shigella species isolated from French travellers returning from the Caribbean. Approximately 21% of the isolates tested were found to encode and produce Stx. These isolates included strains of S. flexneri 2a, S. flexneri Y, and S. dysenteriae 4. All of the travellers who were infected with Stx-producing Shigella had recently travelled to Haiti, the Dominican Republic, or French Guiana. Furthermore, whole genome sequencing showed that the toxin genes were encoded by a prophage that was highly identical to the phage that we identified in our previous study. These findings demonstrate that this new stx-encoding prophage is circulating within that geographical area, has spread to other continents, and is capable of spreading to multiple Shigella serogroups. Published by Elsevier Ltd.

Genetic diversity and virulence potential of shiga toxin-producing Escherichia coli O113:H21 strains isolated from clinical, environmental, and food sources.

PubMed

Feng, Peter C H; Delannoy, Sabine; Lacher, David W; Dos Santos, Luis Fernando; Beutin, Lothar; Fach, Patrick; Rivas, Marta; Hartland, Elizabeth L; Paton, Adrienne W; Guth, Beatriz E C

2014-08-01

Shiga toxin-producing Escherichia coli strains of serotype O113:H21 have caused severe human diseases, but they are unusual in that they do not produce adherence factors coded by the locus of enterocyte effacement. Here, a PCR microarray was used to characterize 65 O113:H21 strains isolated from the environment, food, and clinical infections from various countries. In comparison to the pathogenic strains that were implicated in hemolytic-uremic syndrome in Australia, there were no clear differences between the pathogens and the environmental strains with respect to the 41 genetic markers tested. Furthermore, all of the strains carried only Shiga toxin subtypes associated with human infections, suggesting that the environmental strains have the potential to cause disease. Most of the O113:H21 strains were closely related and belonged in the same clonal group (ST-223), but CRISPR analysis showed a great degree of genetic diversity among the O113:H21 strains. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

Low Prevalence of carbapenem-resistant Enterobacteriaceae among wounded military personnel.

PubMed

Mende, Katrin; Beckius, Miriam L; Zera, Wendy C; Onmus-Leone, Fatma; Murray, Clinton K; Tribble, David R

2017-01-01

Multidrug-resistant organisms (MDROs) are a global health problem that affect both civilian and military populations. Among wounded warriors, MDROs further complicate the care of trauma-related infections, resulting in extended duration of hospitalization, as well as increased morbidity and mortality. During the wars in Iraq and Afghanistan, extended spectrum β-lactamase-producing Enterobacteriaceae were frequently isolated from wounded warriors. The potential emergence of difficult-to-treat carbapenem-resistant Enterobacteriaceae represented a serious challenge for clinicians. We examined carbapenem-resistant Enterobacteriaceae prevalence among wounded military personnel over a 6-year period (2009-2015). Among 4090 Enterobacteriaceae isolates collected, 16 (0.4%) were carbapenem-resistant, of which the majority was Enterobacter aerogenes (44%) followed by Klebsiella pneumoniae (37%), and Escherichia coli (19%). Five isolates (31%) collected from 2 patients were carbapenemase-producers with one associated with an infection. All 5 carbapenemase-producing isolates were resistant to all tested carbapenems and each carried one carbapenemase gene (4 with blaKPC-3 and 1 with blaNDM-1). Overall, although a large number of Enterobacteriaceae isolates were collected, only a small proportion was carbapenem-resistant and data indicate a lack of a cluster. Due to these limited numbers, it is difficult to make any conclusions regarding the association between carbapenem resistance, antibiotic exposure, and clinical outcomes.

Extended-spectrum beta-lactamase-producing bacteria isolated from hematologic patients in Manaus, State of Amazonas, Brazil

PubMed Central

Ferreira, Cristina Motta; Ferreira, William Antunes; Almeida, Nayanne Cristina Oliveira da Silva; Naveca, Felipe Gomes; Barbosa, Maria das Graças Vale

2011-01-01

Antibiotic therapy in hematologic patients, often weak and susceptible to a wide range of infections, particularly nosocomial infections derived from long hospitalization periods, is a challenging issue. This paper presents ESBL-producing strains isolated from such hematologic patients treated at the Amazon Hematology and Hemotherapy Foundation (HEMOAM) in the Brazilian Amazon Region to identify the ESBL genes carried by them as well as the susceptibility to 11 antimicrobial agents using the E-test method. A total of 146 clinical samples were obtained from July 2007 to August 2008, when 17 gram-negative strains were isolated in our institution. The most frequent isolates confirmed by biochemical tests and 16S rRNA sequencing were E. coli (8/17), Serratia spp. (3/17) and B.cepacia (2/17). All gram-negative strains were tested for extended-spectrum-beta-lactamases (ESBLs), where: (12/17) strains carried ESBL; among these, (8/12) isolates carried blaTEM, blaCTX-M, blaOXA , blaSHV genes, (1/12) blaTEM gene and (3/12) blaTEM, blaCTX-M, blaOXA genes. Antibiotic resistance was found in (15/17) of the isolates for tetracycline, (12/17) for ciprofloxacin, (1/17) resistance for cefoxitin and chloramphenicol, (1/17) for amikacin and (3/17) cefepime. This research showed the presence of gram-negative ESBL-producing bacteria infecting hematologic patients in HEMOAM. These strains carried the blaTEM, blaSHV, blaCTX-M and blaOXA genes and were resistant to different antibiotics used in the treatment. This finding was based on a period of 13 months, during which clinical samples from specific populations were obtained. Therefore, caution is required when generalizing the results that must be based on posological orientations and new breakpoints for disk diffusion and microdilution published by CLSI 2010. PMID:24031725

Molecular epidemiology of an outbreak of clinical mastitis in sheep caused by Mannheimia haemolytica.

PubMed

Omaleki, Lida; Browning, Glenn F; Allen, Joanne L; Markham, Philip F; Barber, Stuart R

2016-08-15

The aetiology and epidemiology of outbreaks of clinical mastitis in sheep under extensive pastoral conditions are incompletely understood. The objective of this study was to conduct a detailed investigation of a clinical mastitis outbreak that affected more than 10% of 230 at-risk ewes on a sheep and grain producing property in south east Australia during drought conditions in 2009. Milk samples were collected aseptically from all affected ewes and plated on sheep blood agar for bacterial identification. M. haemolytica was isolated from 80% of the samples that yielded cultivable microorganisms and thus was the main microorganism responsible for the outbreak. Analysis of the restriction endonuclease cleavage patterns of the isolates using pulsed field gel electrophoresis revealed some evidence of clonality, suggesting the possibility of horizontal transmission, but there was also considerable diversity between the clusters of closely related isolates. Multilocus sequence typing of the M. haemolytica isolates revealed most of the isolates belonged to ST1 with no association between the PFGE and MLST fingerprints of the isolates. Resistance to neomycin, streptomycin and sulphafurazole was detected in some of the isolates, but they were all susceptible to penicillin, ampicillin, ceftiofur, amoxycillin/clavulanic acid, ciprofloxacin, tetracycline, erythromycin and trimethoprim. This is the first published record of a comparison of the strains of M. haemolytica involved in a clinical mastitis outbreak in sheep and demonstrates the importance of this pathogen in sheep production systems, particularly during adverse climatic conditions and increased stocking rate. Copyright © 2016 Elsevier B.V. All rights reserved.

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.

Effect of multipurpose solutions against Acinetobacter carrying QAC genes.

PubMed

Boost, Maureen V; Chan, Jessica; Shi, Guang-sen; Cho, Pauline

2014-03-01

Acinetobacter has low virulence but causes infections in subjects with reduced immunity. It has been reported in ocular infections including those of patients using contact lenses. Treatment is difficult because Acinetobacter is frequently multidrug resistant. Antibiotic-resistant strains frequently also harbor genes for antiseptic resistance (quaternary ammonium compound [QAC]) genes. Because Acinetobacter is part of the normal flora, it may contaminate contact lens and accessories. This study aims to investigate carriage rates of QAC genes in household and clinical isolates of Acinetobacter and to determine the effectiveness of two multipurpose solutions (MPSs) for soft lenses against organisms carrying QAC genes. DNA was extracted from 11 bathroom isolates and 15 clinical isolates and amplified by polymerase chain reaction to determine the presence of qacEΔ1. Gene-positive and gene-negative control strains were used to challenge the two MPSs, and minimum inhibitory concentrations (MICs) of these organisms to benzalkonium chloride and chlorhexidine gluconate were determined. More than 90% of isolates carried qacEΔ1. The MICs of clinical isolates were higher than those of isolates of bathrooms. Both MPSs were able to produce a 3-log reduction in the numbers of all isolates. Although most isolates carried qacEΔ1 and elevated MICs to benzalkonium chloride and chlorhexidine gluconate were observed, all were susceptible to both MPSs tested. However, if there were to be poor compliance with care procedures, it is probable that such organisms could survive in the presence of diluted or expired solutions.

Spread of ISCR1 elements containing blaDHA-₁ and multiple antimicrobial resistance genes leading to increase of flomoxef resistance in extended-spectrum-beta-lactamase-producing Klebsiella pneumoniae.

PubMed

Lee, Chen-Hsiang; Liu, Jien-Wei; Li, Chia-Chin; Chien, Chun-Chih; Tang, Ya-Fen; Su, Lin-Hui

2011-09-01

Increasing resistance to quinolones, aminoglycosides, and/or cephamycins in extended-spectrum-β-lactamase (ESBL)-producing Enterobacteriaceae exacerbates the already limited antibiotic treatment options for infections due to these microbes. In this study, the presence of resistance determinants for these antimicrobial agents was examined by PCR among ESBL-producing Klebsiella pneumoniae (ESBL-KP) isolates that caused bacteremia. Pulsed-field gel electrophoresis was used to differentiate the clonal relationship among the isolates studied. Transferability and the location of the resistance genes were analyzed by conjugation experiments, followed by DNA-DNA hybridization. Among the 94 ESBL-KP isolates studied, 20 isolates of flomoxef-resistant ESBL-KP were identified. They all carried a DHA-1 gene and were genetically diverse. CTX-M genes were found in 18 of the isolates. Among these DHA-1/CTX-M-producing K. pneumoniae isolates, ISCR1 was detected in 13 (72%) isolates, qnr genes (1 qnrA and 17 qnrB genes) were detected in 18 (100%), aac(6')-Ib-cr was detected in 11 (61%), and 16S rRNA methylase (all armA genes) was detected in 14 (78%). Four transconjugants were available for further analysis, and qnrB4, aac(6')-Ib-cr, armA, and bla(DHA-1) were all identified on these self-transferable bla(CTX-M)-carrying plasmids. The genetic environments of ISCR1 associated with armA, bla(DHA-1), and qnrB4 genes in the four transconjugants were identical. Replicon-type analysis revealed a FIIA plasmid among the four self-transferable plasmids, although the other three were nontypeable. The cotransfer of multiple resistance genes with the ISCR1 element-carrying plasmids has a clinical impact and warrants close monitoring and further study.

Diversity of β-lactamases produced by imipenem resistant, Pseudomonas aeruginosa isolates from the bloodstream.

PubMed

Najar Peerayeh, Shahin; Pirhajati Mahabadi, Rahim; Pakbaten Toupkanlou, Sanaz; Siadat, Seyed Davar

2014-11-01

The emergence of imipenem non-susceptible Pseudomonas aeruginosa isolates is a matter of great concern because these isolates can become resistant to all available antibiotics. This study conducted to characterize β-lactamase genes in imipenem resistant P. aeruginosa isolates from bloodstream. 56 non-duplicate clinical isolates of P. aeruginosa were collected in Tehran hospitals. Antibacterial susceptibility was determined by disk diffusion and MIC methods. ESBL and MBL production was confirmed by combined disk. β-Lactamase classes A, B and D genes were identified by PCR. Seventeen (30.3%) isolates were imipenem resistant for which 16 isolates simultaneously were resistant to all tested antibiotics. While among 39 imipenem susceptible isolates, only two isolates were resistant to all tested antibiotics. In imipenem resistant isolates, blaTEM, blaSHV and blaOXA-10 were found in 41.1% of isolates and blaVIM, blaIMP and blaPER were identified in 47%, 11.7% and 5.8% of isolates respectively, while in imipenem susceptible isolates, blaTEM, blaSHV and blaOXA-10 were determined in 2.5%, 7.6% and 33.3% of isolates, respectively. The imipenem resistant isolates had been recovered mostly (67.7%) from patients in the Burn hospital. The result of this study indicated the emergence of multidrug resistant MBL and non-MBL producing P. aeruginosa, particularly in the Burn hospital and blaVIM was dominant β-lactamase genes in imipenem resistant isolates. The isolation of carrier patients may lead to prevent a further dissemination. Copyright © 2014 Elsevier Ltd and ISBI. All rights reserved.

In Vitro Activity of Aztreonam-Avibactam against Enterobacteriaceae and Pseudomonas aeruginosa Isolated by Clinical Laboratories in 40 Countries from 2012 to 2015.

PubMed

Karlowsky, James A; Kazmierczak, Krystyna M; de Jonge, Boudewijn L M; Hackel, Meredith A; Sahm, Daniel F; Bradford, Patricia A

2017-09-01

The combination of the monobactam aztreonam and the non-β-lactam β-lactamase inhibitor avibactam is currently in clinical development for the treatment of serious infections caused by metallo-β-lactamase (MBL)-producing Enterobacteriaceae , a difficult-to-treat subtype of carbapenem-resistant Enterobacteriaceae for which therapeutic options are currently very limited. The present study tested clinically significant isolates of Enterobacteriaceae ( n = 51,352) and Pseudomonas aeruginosa ( n = 11,842) collected from hospitalized patients in 208 medical center laboratories from 40 countries from 2012 to 2015 for in vitro susceptibility to aztreonam-avibactam, aztreonam, and comparator antimicrobial agents using a standard broth microdilution methodology. Avibactam was tested at a fixed concentration of 4 μg/ml in combination with 2-fold dilutions of aztreonam. The MIC 90 s of aztreonam-avibactam and aztreonam were 0.12 and 64 μg/ml, respectively, for all Enterobacteriaceae isolates; >99.9% of all isolates and 99.8% of meropenem-nonsusceptible isolates ( n = 1,498) were inhibited by aztreonam-avibactam at a concentration of ≤8 μg/ml. PCR and DNA sequencing identified 267 Enterobacteriaceae isolates positive for MBL genes (NDM, VIM, IMP); all Enterobacteriaceae carrying MBLs demonstrated aztreonam-avibactam MICs of ≤8 μg/ml and a MIC 90 of 1 μg/ml. Against all P. aeruginosa isolates tested, the MIC 90 of both aztreonam-avibactam and aztreonam was 32 μg/ml; against MBL-positive P. aeruginosa isolates ( n = 452), MIC 90 values for aztreonam-avibactam and aztreonam were 32 and 64 μg/ml, respectively. The current study demonstrated that aztreonam-avibactam possesses potent in vitro activity against a recent, sizeable global collection of Enterobacteriaceae clinical isolates, including isolates that were meropenem nonsusceptible, and against MBL-positive isolates of Enterobacteriaceae , for which there are few treatment options. Copyright © 2017 American Society for Microbiology.

In Vitro Activity of Aztreonam-Avibactam against Enterobacteriaceae and Pseudomonas aeruginosa Isolated by Clinical Laboratories in 40 Countries from 2012 to 2015

PubMed Central

Karlowsky, James A.; de Jonge, Boudewijn L. M.; Hackel, Meredith A.; Sahm, Daniel F.

2017-01-01

ABSTRACT The combination of the monobactam aztreonam and the non-β-lactam β-lactamase inhibitor avibactam is currently in clinical development for the treatment of serious infections caused by metallo-β-lactamase (MBL)-producing Enterobacteriaceae, a difficult-to-treat subtype of carbapenem-resistant Enterobacteriaceae for which therapeutic options are currently very limited. The present study tested clinically significant isolates of Enterobacteriaceae (n = 51,352) and Pseudomonas aeruginosa (n = 11,842) collected from hospitalized patients in 208 medical center laboratories from 40 countries from 2012 to 2015 for in vitro susceptibility to aztreonam-avibactam, aztreonam, and comparator antimicrobial agents using a standard broth microdilution methodology. Avibactam was tested at a fixed concentration of 4 μg/ml in combination with 2-fold dilutions of aztreonam. The MIC90s of aztreonam-avibactam and aztreonam were 0.12 and 64 μg/ml, respectively, for all Enterobacteriaceae isolates; >99.9% of all isolates and 99.8% of meropenem-nonsusceptible isolates (n = 1,498) were inhibited by aztreonam-avibactam at a concentration of ≤8 μg/ml. PCR and DNA sequencing identified 267 Enterobacteriaceae isolates positive for MBL genes (NDM, VIM, IMP); all Enterobacteriaceae carrying MBLs demonstrated aztreonam-avibactam MICs of ≤8 μg/ml and a MIC90 of 1 μg/ml. Against all P. aeruginosa isolates tested, the MIC90 of both aztreonam-avibactam and aztreonam was 32 μg/ml; against MBL-positive P. aeruginosa isolates (n = 452), MIC90 values for aztreonam-avibactam and aztreonam were 32 and 64 μg/ml, respectively. The current study demonstrated that aztreonam-avibactam possesses potent in vitro activity against a recent, sizeable global collection of Enterobacteriaceae clinical isolates, including isolates that were meropenem nonsusceptible, and against MBL-positive isolates of Enterobacteriaceae, for which there are few treatment options. PMID:28630192

NDM-5 Carbapenemase-Encoding Gene in Multidrug-Resistant Clinical Isolates of Escherichia coli from Algeria

PubMed Central

Sassi, Asma; Loucif, Lotfi; Gupta, Sushim Kumar; Dekhil, Mazouz; Chettibi, Houria

2014-01-01

Here, we report the first autochthonous cases of infections caused by blaNDM-5 New Delhi metallo-β-lactamase-producing Escherichia coli strains recovered from urine and blood specimens of three patients from Algeria between January 2012 and February 2013. The three isolates belong to sequence type 2659 and they coexpress blaCTX-M-15 with the blaTEM-1 and blaaadA2 genes. PMID:24982080

Mastitis Pathogens with High Virulence in a Mouse Model Produce a Distinct Cytokine Profile In Vivo

PubMed Central

Johnzon, Carl-Fredrik; Artursson, Karin; Söderlund, Robert; Guss, Bengt; Rönnberg, Elin; Pejler, Gunnar

2016-01-01

Mastitis is a serious medical condition of dairy cattle. Here, we evaluated whether the degree of virulence of mastitis pathogens in a mouse model can be linked to the inflammatory response that they provoke. Clinical isolates of Staphylococcus aureus (S. aureus) (strain 556 and 392) and Escherichia coli (E. coli) (676 and 127), and laboratory control strains [8325-4 (S. aureus) and MG1655 (E. coli)], were injected i.p. into mice, followed by the assessment of clinical scores and inflammatory parameters. As judged by clinical scoring, E. coli 127 exhibited the largest degree of virulence among the strains. All bacterial strains induced neutrophil recruitment. However, whereas E. coli 127 induced high peritoneal levels of CXCL1, G-CSF, and CCL2, strikingly lower levels of these were induced by the less virulent bacterial strains. High concentrations of these compounds were also seen in blood samples taken from animals infected with E. coli 127, suggesting systemic inflammation. Moreover, the levels of CXCL1 and G-CSF, both in the peritoneal fluid and in plasma, correlated with clinical score. Together, these findings suggest that highly virulent clinical mastitis isolates produce a distinct cytokine profile that shows a close correlation with the severity of the bacterial infection. PMID:27713743

Sub-typing of extended-spectrum-β-lactamase-producing isolates from a nosocomial outbreak: application of a 10-loci generic Escherichia coli multi-locus variable number tandem repeat analysis.

PubMed

Karami, Nahid; Helldal, Lisa; Welinder-Olsson, Christina; Ahrén, Christina; Moore, Edward R B

2013-01-01

Extended-spectrum β-lactamase producing Escherichia coli (ESBL-E. coli) were isolated from infants hospitalized in a neonatal, post-surgery ward during a four-month-long nosocomial outbreak and six-month follow-up period. A multi-locus variable number tandem repeat analysis (MLVA), using 10 loci (GECM-10), for 'generic' (i.e., non-STEC) E. coli was applied for sub-species-level (i.e., sub-typing) delineation and characterization of the bacterial isolates. Ten distinct GECM-10 types were detected among 50 isolates, correlating with the types defined by pulsed-field gel electrophoresis (PFGE), which is recognized to be the 'gold-standard' method for clinical epidemiological analyses. Multi-locus sequence typing (MLST), multiplex PCR genotyping of bla CTX-M, bla TEM, bla OXA and bla SHV genes and antibiotic resistance profiling, as well as a PCR assay specific for detecting isolates of the pandemic O25b-ST131 strain, further characterized the outbreak isolates. Two clusters of isolates with distinct GECM-10 types (G06-04 and G07-02), corresponding to two major PFGE types and the MLST-based sequence types (STs) 131 and 1444, respectively, were confirmed to be responsible for the outbreak. The application of GECM-10 sub-typing provided reliable, rapid and cost-effective epidemiological characterizations of the ESBL-producing isolates from a nosocomial outbreak that correlated with and may be used to replace the laborious PFGE protocol for analyzing generic E. coli.

[Prevalence of ESBL-positive enterobacteriaceae in large moravian hospitals (Czech Republic)].

PubMed

Kolář, Milan; Chromá, Magdaléna; Hricová, Kristýna; Husičková, Vendula; Lovečková, Yvona; Chmelařová, Eva; Bartoníková, Nataša; Rybníkářová, Petra

2010-10-01

bacterial infections have become an important issue in current medicine. Recently, their frequency and severity have significantly increased as a result of the rising number of resistant bacteria. One of important mechanisms of resistance is production of broad-spectrum beta-lactamases, namely the ESBL type. The study aimed at determining the frequency of ESBL-positive Enterobacteriaceae in three large hospitals in Moravia, the eastern part of the Czech Republic. enterobacteriaceae were isolated from clinical material obtained from patients hospitalized in the University Hospital Olomouc, Teaching Hospital Ostrava and Bata Regional Hospital Zlín throughout 2009. Standard microbiology techniques were used for identification. The production of ESBLs was determined by the modified Double-Disk Synergy Test. ESBL-positive isolates of Escherichia coli from ICU patients were subjected to basic genetic analysis. during the study period, a total of 12,922 strains from the Enterobacteriaceae family were detected. The ESBL phenotype was found in 907 cases, i.e. 7 % of all isolates. The most prevalent species of ESBL-producing Enterobacteriaceae were Klebsiella pneumoniae, Klebsiella oxytoca and Escherichia coli. A comparison of general wards and ICUs revealed a higher percentage of ESBL-positive strains of Klebsiella pneumoniae and a lower proportion of ESBL-positive Escherichia coli isolates in intensive care patients. When assessing the patients' clinical material, ESBL-producing strains were most frequently detected in urine. Genetic analysis of ESBL-positive Escherichia coli strains from ICU patients revealed the CTX-M type of ESBL production in most isolates.

Phenotypic and genotypic characterization of vancomycin-resistant Enterococcus faecium clinical isolates from two hospitals in Mexico: First detection of VanB phenotype-vanA genotype.

PubMed

Bocanegra-Ibarias, Paola; Flores-Treviño, Samantha; Camacho-Ortiz, Adrián; Morfin-Otero, Rayo; Villarreal-Treviño, Licet; Llaca-Díaz, Jorge; Martínez-Landeros, Erik Alan; Rodríguez-Noriega, Eduardo; Calzada-Güereca, Andrés; Maldonado-Garza, Héctor Jesús; Garza-González, Elvira

2016-01-01

Enterococcus faecium has emerged as a multidrug-resistant nosocomial pathogen involved in outbreaks worldwide. Our aim was to determine the antimicrobial susceptibility, biofilm production, and clonal relatedness of vancomycin-resistant E. faecium (VREF) clinical isolates from two hospitals in Mexico. Consecutive clinical isolates (n=56) were collected in two tertiary care hospitals in Mexico from 2011 to 2014. VREF isolates were characterized by phenotypic and molecular methods including pulsed-field gel electrophoresis (PFGE). VREF isolates were highly resistant to vancomycin, erythromycin, norfloxacin, high-level streptomycin, and teicoplanin, and showed lower resistance to tetracycline, nitrofurantoin and quinupristin-dalfopristin. None of the isolates were resistant to linezolid. The vanA gene was detected in all isolates. Two VanB phenotype-vanA genotype isolates, highly resistant to vancomycin and susceptible to teicoplanin, were detected. Furthermore, 17.9% of the isolates were classified as biofilm producers, and the espfm gene was found in 98.2% of the isolates. A total of 37 distinct PFGE patterns and 6 clones (25% of the isolates as clone A, 5.4% as clone B, and 3.6% each as clone C, D, E, and F) were detected. Clone A was detected in 5 different wards of the same hospital during 14 months of surveillance. The high resistance to most antimicrobial agents and the moderate cross-transmission of VREF detected accentuates the need for continuous surveillance of E. faecium in the hospital setting. This is also the first reported incidence of the E. faecium VanB phenotype-vanA genotype in the Americas. Copyright © 2015 Elsevier España, S.L.U. and Sociedad Española de Enfermedades Infecciosas y Microbiología Clínica. All rights reserved.

Klebsiella pneumoniae Strains Producing Extended-Spectrum β-Lactamases in Spain: Microbiological and Clinical Features▿

PubMed Central

de Alegría, C. Ruiz; Rodríguez-Baño, J.; Cano, M. E.; Hernández-Bello, J. R.; Calvo, J.; Román, E.; Díaz, M. A.; Pascual, A.; Martínez-Martínez, L.

2011-01-01

Extended-spectrum β-lactamases (ESBL) of the CTX-M, SHV, and TEM families were recognized in 76 (67%), 31 (27%), and 6 (5%) isolates, respectively, among 162 ESBL-producing Klebsiella pneumoniae (ESBL-Kp) strains obtained in a multicenter study in Spain. Predisposing factors for ESBL-Kp acquisition included invasive procedures, mechanical ventilation, and previous antimicrobial use. PMID:21191059

Imipenem resistance in clinical Escherichia coli from Qom, Iran.

PubMed

Shams, Saeed; Hashemi, Ali; Esmkhani, Mohammad; Kermani, Somaye; Shams, Elham; Piccirillo, Alessandra

2018-05-18

The emergence of metallo-β-lactamase-producing Enterobacteriaceae is a worldwide health concern. In this study, the first evaluation of MBL genes, bla IMP and bla VIM , in Escherichia coli resistant to imipenem isolated from urine and blood specimens in Qom, Iran is described. Three hundred urine and blood specimens were analysed to detect the presence of E. coli. Resistance to imipenem and other antimicrobials was determined by disk diffusion and MIC. MBL production was screened using CDDT. PCR was also carried out to determine the presence of bla IMP and bla VIM genes in imipenem-resistant isolates. In total, 160 E. coli isolates were collected from March to May 2016. According to disk diffusion, high-level of resistance (20%) to cefotaxime was observed, whereas the lowest (1%) was detected for tetracycline. In addition, five isolates showed resistance to imipenem with a MIC ≥ 4 µg/mL. CDDT test confirmed that five isolates were MBL-producing strains, but no bla IMP and bla VIM genes were detected. Results of this study show a very low level of resistance to imipenem in our geographical area.

Prevalence of the mcr-1 colistin resistance gene in extended-spectrum β-lactamase-producing Escherichia coli from human faecal samples collected in 2012 in rural villages in Shandong Province, China.

PubMed

Bi, Zhenwang; Berglund, Björn; Sun, Qiang; Nilsson, Maud; Chen, Baoli; Tärnberg, Maria; Ding, Lilu; Stålsby Lundborg, Cecilia; Bi, Zhenqiang; Tomson, Göran; Yao, Jingjing; Gu, Zhanying; Yin, Xiao; Kou, Zengqiang; Nilsson, Lennart E

2017-04-01

Since its initial discovery in China in 2015, the plasmid-mediated colistin resistance gene mcr-1 has been reported in Escherichia coli isolated from clinical samples, animals and meat worldwide. In this study, 706 extended-spectrum β-lactamase (ESBL)-producing E. coli from 411 persons were detected in a collection of faecal samples from 1000 rural residents in three counties in Shandong Province, China. These isolates were screened for mcr-1 and phenotypic colistin resistance. The gene was found in 3.5% of the isolates (from 4.9% of persons) from all three counties. All isolates with phenotypic colistin resistance carried mcr-1. These data indicate that commensal carriage of ESBL-producing E. coli with mcr-1 among persons in rural China was already present in 2012 and that mcr-1 was the most important colistin resistance mechanism. Interventions are necessary to minimise further dissemination of mcr-1, which would limit the future usefulness of colistin as a last-resort antibiotic. Copyright © 2017 Elsevier B.V. and International Society of Chemotherapy. All rights reserved.

Escherichia coli Sequence Type 131 (ST131) Subclone H30 as an Emergent Multidrug-Resistant Pathogen Among US Veterans

PubMed Central

Colpan, Aylin; Johnston, Brian; Porter, Stephen; Clabots, Connie; Anway, Ruth; Thao, Lao; Kuskowski, Michael A.; Tchesnokova, Veronika; Sokurenko, Evgeni V.; Johnson, James R.; Allen, Bradley L.; Baracco, Gio J.; Bedimo, Roger; Bessesen, Mary; Bonomo, Robert A.; Brecher, Stephen M.; Brown, Sheldon T.; Castellino, Laila; Desai, Arundhati S.; Fernau, Fletcher; Fisher, Mark A.; Fleckenstein, James; Fleming, Carol S.; Fries, Narla J.; Kan, Virginia L.; Kauffman, Carol A.; Klutts, Stacey; Ohl, Michael; Russo, Thomas; Swiatlo, Andrea; Swiatlo, Edwin

2013-01-01

Background. Escherichia coli sequence type 131 (ST131), typically fluoroquinolone-resistant (FQ-R) and/or extended-spectrum β-lactamase (ESBL)–producing, has emerged globally. We assessed its prevalence and characteristics among US veterans. Methods. In 2011, 595 de-identified E. coli clinical isolates were collected systematically within 3 resistance groups (FQ-susceptible [FQ-S], FQ-R, and ESBL-producing) from 24 nationally distributed Veterans Affairs Medical Centers (VAMCs). ST131 and its H30 subclone were detected by polymerase chain reaction and compared with other E. coli for molecular traits, source, and resistance profiles. Results. ST131 accounted for 78% (184/236) of FQ-R and 64.2% (79/123) of ESBL-producing isolates, but only 7.2% (17/236) of FQ-S isolates (P < .001). The H30 subclone accounted for ≥95% of FQ-R and ESBL-producing, but only 12.5% of FQ-S, ST131 isolates (P < .001). By back-calculation, 28% of VAMC E. coli isolates nationally represented ST131. Overall, ST131 varied minimally in prevalence by specimen type, inpatient/outpatient source, or locale; was the most prevalent ST, followed distantly by ST95 and ST12 (13% each); and accounted for ≥40% (β-lactams), >50% (trimethoprim-sulfamethoxazole , multidrug), or >70% (ciprofloxacin, gentamicin) of total antimicrobial resistance. FQ-R and ESBL-producing ST131 isolates had higher virulence scores than corresponding non-ST131 isolates. ST131 pulsotypes overlapped extensively among VAMCs. Conclusions. Among US veterans, ST131, primarily its H30 subclone, accounts for most antimicrobial-resistant E. coli and is the dominant E. coli strain overall. Possible contributors include multidrug resistance, extensive virulence gene content, and ongoing transmission. Focused attention to ST131, especially its H30 subclone, could reduce infection-related morbidity, mortality, and costs among veterans. PMID:23926176

Psl Produced by Mucoid Pseudomonas aeruginosa Contributes to the Establishment of Biofilms and Immune Evasion.

PubMed

Jones, Christopher J; Wozniak, Daniel J

2017-06-20

Despite years of research and clinical advances, chronic pulmonary infections with mucoid Pseudomonas aeruginosa remain the primary concern for cystic fibrosis patients. Much of the research on these strains has focused on the contributions of the polysaccharide alginate; however, it is becoming evident that the neutral polysaccharide Psl also contributes to biofilm formation and the maintenance of chronic infections. Here, we demonstrate that Psl produced by mucoid strains has significant roles in biofilm structure and evasion of immune effectors. Though mucoid strains produce less Psl than nonmucoid strains, the Psl that is produced is functional, since it mediates adhesion to human airway cells and epithelial cell death. Additionally, Psl protects mucoid bacteria from opsonization and killing by complement components in human serum. Psl production by mucoid strains stimulates a proinflammatory response in the murine lung, leading to reduced colonization. To determine the relevance of these data to clinical infections, we tested Psl production and biofilm formation of a panel of mucoid clinical isolates. We demonstrated three classes of mucoid isolates, those that produce Psl and form robust biofilms, those that did not produce Psl and have a poor biofilm phenotype, and exopolysaccharide (EPS) redundant strains. Collectively, these experimental results demonstrate that Psl contributes to the biofilm formation and immune evasion of many mucoid strains. This is a novel role for Psl in the establishment and maintenance of chronic pulmonary infections by mucoid strains. IMPORTANCE Cystic fibrosis patients are engaged in an ongoing battle against chronic lung infections by the bacterium Pseudomonas aeruginosa One key factor contributing to the maintenance of chronic infections is the conversion to a mucoid phenotype, where the bacteria produce copious amounts of the polysaccharide alginate. Once the bacteria become mucoid, existing treatments are poorly effective. We proposed that mucoid bacteria produce an additional polysaccharide, Psl, which is important for their establishment and maintenance of chronic infections. This work demonstrates that Psl enhances attachment of mucoid bacteria to lung surfaces and leads to inflammation and damage in the lung. Additionally, we find that 50% of mucoid bacteria isolated from patients with chronic infections rely on Psl for the structure of their biofilm communities, suggesting that treatments against Psl should be investigated to enhance the success of current therapies. Copyright © 2017 Jones and Wozniak.

Purification and characterization of factors produced by Aspergillus fumigatus which affect human ciliated respiratory epithelium.

PubMed Central

Amitani, R; Taylor, G; Elezis, E N; Llewellyn-Jones, C; Mitchell, J; Kuze, F; Cole, P J; Wilson, R

1995-01-01

The mechanisms by which Aspergillus fumigatus colonizes the respiratory mucosa are unknown. Culture filtrates of eight of nine clinical isolates of A. fumigatus slowed ciliary beat frequency and damaged human respiratory epithelium in vitro. These changes appeared to occur concurrently. Culture filtrates of two clinical isolates of Candida albicans had no effect on ciliated epithelium. We have purified and characterized cilioinhibitory factors of a clinical isolate of A. fumigatus. The cilioinhibitory activity was heat labile, reduced by dialysis, and partially extractable into chloroform. The activity was associated with both high- and low-molecular-weight factors, as determined by gel filtration on Sephadex G-50. A low-molecular-weight cilioinhibitory factor was further purified by reverse-phase high-performance liquid chromatography and shown by mass spectrometry to be gliotoxin, a known metabolite of A. fumigatus. Gliotoxin significantly slowed ciliary beat frequency in association with epithelial damage at concentrations above 0.2 microgram/ml; other Aspergillus toxins, i.e., fumagillin and helvolic acid, were also cilioinhibitory but at much higher concentrations. High-molecular-weight (> or = 35,000 and 25,000) cilioinhibitory materials had neither elastolytic nor proteolytic activity and remain to be identified. Thus, A. fumigatus produces a number of biologically active substances which slow ciliary beating and damage epithelium and which may influence colonization of the airways. PMID:7543879

Plasmid analyses in clinical isolates of Bacteroides fragilis and other Bacteroides species.

PubMed Central

Wallace, B L; Bradley, J E; Rogolsky, M

1981-01-01

Plasmid analyses were performed on Bacteroides strains isolated from clinical specimens. Of 32 Bacteroides strains, 8 were found to contain plasmids. Seven of these eight strains were B. fragilis, and the other one was B. distasonis. Three of these eight strains harbored only a 3.0-megadalton plasmid. Two strains had only a 2.0-megadalton plasmid, and one had 2.0-, 3.0-megadalton plasmid. Of the remaining two strains, one had 2.0-, 3.0-, and 5.0-megadalton plasmids, and the other had 3.0- and 5.0-megadalton plasmids. Beta-Lactamase was produced by 93% of the clinical isolates. Seven of the eight plasmid-carrying strains were cadmium resistant, five were zinc resistant, four were mercury resistant, and two expressed a brick-red fluorescence under ultraviolet light. None of these traits could be associated with a plasmid after performing either curing experiments or genetic transfer experiments by cell-to-cell contact. Images PMID:6974737

Antimicrobial resistance and beta-lactamase production of clinical isolates of prevotella and porphyromonas species.

PubMed

Bahar, Hrisi; Torun, Muzeyyen Mamal; Demirci, Mehmet; Kocazeybek, Bekir

2005-03-01

This study determined the beta-lactamase production and the antimicrobial resistance of 72 Prevotella species and 48 Porphyromonas species isolated from different clinical specimens. All strains were identified using API 32 ID. The beta-lactamase production was determined by nitrocefin disks. E test strips of benzylpenicillin, ampicillin + sulbactam, cefoxitin, clindamycin, metronidazole and imipenem were tested for each strain. Nineteen Prevotella melaninogenica, 18 Prevotella intermedia, 16 Prevotella denticola, 11 Prevotella loescheii and 8 Prevotella bivia strains were identified. Four were clindamycin resistant. The highest beta-lactamase production was found at a rate of 68.4% in P. melaninogenica species. Additionally, 33 Porphyromonas asaccharolytica and 15 Porphyromonas gingivalis strains were identified. None of them produced beta-lactamase. In view of the emerging antibiotic resistance among anaerobes, the current local susceptibility profile of our Prevotella and Porphyromonas species will establish the basis for additional surveys tracing significant changes in the antimicrobial resistance of our clinical isolates. Copyright 2005 S. Karger AG, Basel.

Epidemiology and clinical features of community-onset bacteremia caused by extended-spectrum β-lactamase-producing Klebsiella pneumoniae.

PubMed

Lee, Jeong-a; Kang, Cheol-In; Joo, Eun-Jeong; Ha, Young Eun; Kang, Seung-Ji; Park, So Yeon; Chung, Doo Ryeon; Peck, Kyong Ran; Ko, Kwan Soo; Lee, Nam Yong; Song, Jae-Hoon

2011-06-01

There is limited clinical information regarding community-onset bacteremia caused by extended-spectrum β-lactamase (ESBL)-producing Klebsiella pneumoniae. This study was performed to evaluate risk factors and clinical outcomes of community-onset bacteremia caused by ESBL-producing K. pneumoniae. A total of 435 patients with community-onset K. pneumoniae bacteremia were included and data from patients with ESBL-producing K. pneumoniae bacteremia were compared to those with non-ESBL-producing bacteremia. Isolates with ESBLs were microbiologically characterized. Of 435 patients with community-onset K. pneumoniae bacteremia, 33 (7.6%) were infected with ESBL producers, of which 25 were further classified as healthcare-associated infections. The most common underlying diseases were solid tumors (n = 20, 60.6%) and diabetes mellitus (n = 10, 30.3%), and the most common infection was intra-abdominal infection (n = 20, 60.6%). Multivariate analysis showed that corticosteroid use (odds ratio [OR] = 13.73, 95% confidence interval [CI] = 1.93-97.6, p = 0.009), percutaneous tubes (OR = 7.30, 95% CI = 2.41-22.12, p < 0.001), and prior receipt of antibiotics (OR = 5.65, 95% CI = 2.43-14.16, p < 0.001) were significant factors associated with ESBL producers. When the 30-day mortality rate was evaluated, no significant difference was found between ESBL group and non-ESBL group (12.1% [4/32] vs. 16.0% [35/192]; p = 0.429). Among 16 isolates, for which the ESBL characterization was performed by PCR, the most common types of ESBLs were SHV (n = 16) and cefotaxime-M-2 (n = 5). Pulsed-field gel electrophoresis analysis of the ESBL-producing organisms showed extensive clonal diversity. ESBL-producing K. pneumoniae is a significant cause of bacteremia, even in patients with community-onset infections, particularly in patients with corticosteroid use, percutaneous tube, prior receipt of antibiotics, or healthcare-associated infections.

Characteristics of environmental isolates of Legionella pneumophila. [Legionella

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

Orrison, L.H.; Cherry, W.B.; Fliermans, C.B.

1981-07-01

Thirty-eight cultures of Legionella pneumophila isolated from survace waters were characterized by their morphological, tinctorial, biochemical, and serological properties and by their ability to produce disease in guinea pigs. Their susceptibility to antimicrobial agents also was tested. When they were compared with clinical isolates, no important differences were found between cultures from the two sources. Sodium hippurate hydrolysis, gelatin liquefaction, pigment formation, and ..beta..-lactamase and alkaline phosphatase activity were useful in differentiating the four described species of Legionella. Hydrolysis of diacetylfluorescein and the inability to reduce nitrate help to distinguish Legionella species from other gram-negative bacterial rods.

Prevalence of bla TEM-220 gene in Penicillinase-producing Neisseria gonorrhoeae strains carrying Toronto/Rio plasmid in Argentina, 2002 - 2011.

PubMed

Gianecini, Ricardo; Oviedo, Claudia; Guantay, Cristina; Piccoli, Laura; Stafforini, Graciela; Galarza, Patricia

2015-12-16

Penicillinase-producing Neisseria gonorroheae (PPNG) was first isolated in 1976. PPNG strains carrying bla TEM-1 and bla TEM-135 gene have been described in different countries. Recently, a novel bla TEM-220 allele was detected in PPNG isolates carrying Toronto/Rio plasmid. The prevalence and characteristics of TEM-220 strains worldwide are unknown, and therefore, it needs to be studied. The purpose of this study was to detect bla TEM-220 gene in PPNG strains possessing Toronto/Rio plasmid over a period of ten years in Argentina, and to evaluate the proportion of isolates producing non-TEM-220 containing the T539C substitution in the bla TEM allele. One hundred and fifty one PPNG isolates carrying Toronto/Rio plasmid were studied between 2002 and 2011. A mismatch amplification mutation assay (MAMA) PCR was used to identify the T539C substitution in the bla TEM allele and a MAMA-PCR protocol was developed to detect the G547A substitution in the bla TEM-220. The reference agar dilution method of the Clinical and Laboratory Standard Institute (CLSI) was used for susceptibility testing to five β-lactams antibiotics, ciprofloxacin, tetracycline and azithromycin. In all TEM-220-producing isolates, the whole bla TEM gene was sequenced and the isolates were typed using N. gonorroheae multiantigen sequence typing (NG-MAST). MAMA PCR successfully identified the G547A substitution in the bla TEM-220 allele. The proportion of isolates that possessed the bla TEM-220 allele was 2.6 %, and 93.2 % MAMA TEM-220 PCR-negative isolates showed the T539C substitution in the bla TEM gene. No differences in the susceptibility to five beta-lactam antibiotics tested were observed in PPNG isolates TEM-220-producing and PPNG isolates carrying the T539C substitution in the bla TEM gene. All TEM-220 isolates were indistinguishable by NG-MAST. This is the first study which shows the prevalence of bla TEM-220 in N. gonorrhoeae isolates carrying Toronto/Rio plasmid in Argentina. Although the bla TEM-220 allele does not appear to be associated with an extended spectrum beta-lactamase (ESBL) phenotype of resistance, a single nucleotide polymorphism added to the bla TEM-220 or bla TEM containing the T539C substitution could lead to the emergence of ESBL. Thus, it is imperative to investigate in surveillance programs, not only the plasmid type in PPNG isolates and the bla TEM allele associated, but phenotypical characteristics and geographical distribution of isolates.

Transcriptomics in human blood incubation reveals the importance of oxidative stress response in Saccharomyces cerevisiae clinical strains

PubMed Central

2012-01-01

Background In recent years an increasing number of yeast infections in humans have been related to certain clinical isolates of Saccharomyces cerevisiae. Some clinical strains showed in vivo and in vitro virulence traits and were able to cause death in mice whereas other clinical strains were avirulent. Results In this work, we studied the transcriptional profiles of two S. cerevisiae clinical strains showing virulent traits and two control non-virulent strains during a blood incubation model and detected a specific transcriptional response of clinical strains. This response involves an mRNA levels increase of amino acid biosynthesis genes and especially oxidative stress related genes. We observed that the clinical strains were more resistant to reactive oxygen species in vitro. In addition, blood survival of clinical isolates was high, reaching similar levels to pathogenic Candida albicans strain. Furthermore, a virulent strain mutant in the transcription factor Yap1p, unable to grow in oxidative stress conditions, presented decreased survival levels in human blood compared with the wild type or YAP1 reconstituted strain. Conclusions Our data suggest that this enhanced oxidative stress response in virulent clinical isolates, presumably induced in response to oxidative burst from host defense cells, is important to increase survival in human blood and can help to infect and even produce death in mice models. PMID:22916735

Dissemination of blaNDM-5 gene via an IncX3-type plasmid among non-clonal Escherichia coli in China.

PubMed

Li, Xi; Fu, Ying; Shen, Mengyuan; Huang, Danyan; Du, Xiaoxing; Hu, Qingfeng; Zhou, Yonglie; Wang, Dairong; Yu, Yunsong

2018-01-01

The emergence and spread of New Delhi metallo-β-lactamase-producing Enterobacteriaceae has been a serious challenge to manage in the clinic due to its rapid dissemination of multi-drug resistance worldwide. As one main type of carbapenemases, New Delhi metallo-β-lactamase (NDM)is able to confer resistance to almost all β-lactams, including carbapenems, in Enterobacteriaceae . Recently, New Delhi metallo-β-lactamase-5 attracted extensive attention because of increased resistance to carbapenems and widespread dissemination. However, the dissemination mechanism of bla NDM-5 gene remains unclear. A total of 224 carbapenem-resistant Enterobacteriaceae isolates (CRE) were collected from different hospitals in Zhejiang province. NDM-5-positive isolates were identified and subjected to genotyping, susceptibility testing, and clinical data analysis. We established the genetic location of bla NDM-5 with southern blot hybridisation, and analysed plasmids containing bla NDM-5 with filter mating and DNA sequencing. Eleven New Delhi metallo-β-lactamase-5 (NDM-5)-producing strains were identified, including 9 Escherichia coli strains, 1 Klebsiella pneumoniae strain, and 1 Citrobacter freundii strain. No epidemiological links for E. coli isolates were identified by multilocus sequence typing (MLST) and pulsed-field gel electrophoresis (PFGE). S1-PFGE and southern blot suggested that the bla NDM-5 gene was located on a 46-kb IncX3-type plasmid in all isolates. Nine of the 11 isolates (81.8%) tested could successfully transfer their carbapenem-resistant phenotype to E. coli strain C600. Moreover, sequence analysis further showed that this plasmid possessed high sequence similarity to most of previously reported bla NDM-5 -habouring plasmids in China. The present data in this study showed the IncX3 type plasmid played an important role in the dissemination of bla NDM-5 in Enterobacteriaceae . In addition, to the best of our knowledge, this report is the first to isolate both E. coli and C. freundii strains carrying bla NDM-5 from one single patient, which further indicated the possibility of bla NDM-5 transmission among diverse species. Close surveillance is urgently needed to monitor the further dissemination of NDM-5-producing isolates.

Detection of the frequency of PER-1 type extended-spectrum β-lactamase-producing Acinetobacter baumannii clinical isolates in Turkey: a multicenter study.

PubMed

Aşik, Gülşah; Özdemir, Mehmet; Kurtoğlu, Muhammet Güzel; Yağci, Server; Öksüz, Lütfiye; Gül, Mustafa; Koçoğlu, Mücahide Esra; Sesli Çetin, Emel; Seyrek, Adnan; Berktaş, Mustafa; Ayyildiz, Ahmet; Çiftci, İhsan Hakkı

2014-01-01

β-Lactamases are an important resistance mechanism in Acinetobacter baumannii. Pseudomonas extended-resistance (PER-1) type β-lactamase-producing strains have been reported from various geographic locations; however, PER-1 type β-lactamases from Turkish hospitals have not been investigated extensively. The aim of this study was to determine the prevalence of PER-1 type β-lactamases in A. baumannii isolates in various regions of Turkey. A total of 763 clinical A. baumannii isolates were collected from 9 university hospitals and 2 state hospitals between 2008 and 2011. Molecular amplification of the OXA-51 gene from the A. baumannii genome was performed in order to verify identification of the species. Real-time polymerase chain reaction was used to detect blaPER-1 genes. PER-1 was detected in 24.6% of the isolates. The annual frequencies of the PER-1 enzyme were detected as 52.2%, 35.9%, and 8.3% in 2008, 2009, and 2010, respectively. PER-1 prevalence decreased gradually over time. The differences observed in PER-1 prevalence among the regions of Turkey were statistically significant (chi-square test; P < 0.001). These data demonstrate that the frequency of detection of PER-1 type β-lactamases in A. baumannii species has decreased in Turkey. However, the increased carbapenem resistance, together with multidrug resistance, has created a worrisome situation regarding this pathogen.

Antimicrobial activity of ceftaroline and other anti-infective agents against microbial pathogens recovered from the surgical intensive care patient population: a prevalence analysis.

PubMed

Edmiston, Charles E; Krepel, Candace J; Leaper, David; Ledeboer, Nathan A; Mackey, Tami-Lea; Graham, Mary Beth; Lee, Cheong; Rossi, Peter J; Brown, Kellie R; Lewis, Brian D; Seabrook, Gary R

2014-12-01

Ceftaroline is a new parenteral cephalosporin agent with excellent activity against methicillin-sensitive (MSSA) and resistant strains of Staphylococcus aureus (MRSA). Critically ill surgical patients are susceptible to infection, often by multi-drug-resistant pathogens. The activity of ceftaroline against such pathogens has not been described. Three hundred thirty-five consecutive microbial isolates were collected from surgical wounds or abscesses, respiratory, urine, and blood cultures from patients in the surgical intensive care unit (SICU) of a major tertiary medical center. Using Clinical and Laboratory Standards Institute (CLSI) standard methodology and published breakpoints, all aerobic, facultative anaerobic isolates were tested against ceftaroline and selected comparative antimicrobial agents. All staphylococcal isolates were susceptible to ceftaroline at a breakpoint of ≤1.0 mcg/mL. In addition, ceftaroline exhibited excellent activity against all streptococcal clinical isolates and non-ESBL-producing strains of Enterobacteriaceae (93.5%) recovered from SICU patients. Ceftaroline was inactive against ESBL-producing Enterobacteriaceae, Pseudomonas aeruginosa, vancomycin-resistant enterococci, and selective gram-negative anaerobic bacteria. At present, ceftaroline is the only cephalosporin agent that is active against community and healthcare-associated MRSA. Further studies are needed to validate the benefit of this novel broad-spectrum anti-infective agent for the treatment of susceptible serious infections in the SICU patient population.

Molecular characteristics, biofilm-forming abilities, and quorum sensing molecules in Vibrio parahaemolyticus strains isolated from marine and clinical environments in Korea.

PubMed

Mizan, Md Furkanur Rahaman; Bang, Hyeon-Jo; Sadekuzzaman, Mohammad; Lee, Nari; Kim, Tae-Jo; Ha, Sang-Do

2017-05-01

Vibrio parahaemolyticus is an inhabitant of marine and estuarine environments and causes seafood-borne gastroenteritis in humans. In this study, an UltraFast LabChip Real-Time PCR assay was evaluated for rapid detection and quantification of pathogenic V. parahaemolyticus isolates. Escherichia coli and Vibrio harveyi were used as negative controls. Twenty-six tdh-positive, biofilm-producing V. parahaemolyticus isolates were analyzed by repetitive extragenic palindromic-polymerase chain reaction (REP-PCR). REP-PCR analysis showed that the majority of the V. parahaemolyticus isolates originated from seafood and that clinical specimens formed two major clusters at 92.8% and 32% similarity levels. The presence and quantification of Autoinducer-2 was carried out using high-performance liquid chromatography with fluorescence detection (HPLC-FLD) after derivatization of Autoinducer-2 with 2, 3-diaminonaphthalene. The presence of tdh-positive V. parahaemolyticus in marine samples highlights the need for constant environmental monitoring to protect public health.

Molecular typing and antimicrobial susceptibility of Clostridium perfringens from broiler chickens.

PubMed

Gharaibeh, Saad; Al Rifai, Rami; Al-Majali, Ahmad

2010-12-01

Clostridium perfringens (Cp) causes necrotic enteritis disease in commercial poultry. Antimicrobials are used to control and treat this disease and sometimes clinical outbreaks do not respond well to certain treatments. This study was designed to isolate Cp from clinical cases, type these isolates by multiplex PCR, and determine their antimicrobial susceptibility by micro-dilution method. A total of 67 Cp isolates were obtained from 155 broiler chicken flocks. All isolates were classified as type A and non-enterotoxin producers. Lincomycin, erythromycins, and tilmicosin showed very high minimal inhibitory concentration (MIC) 50 of ≥256 μg/ml. However, tylosin, amoxicillin, ampicillin, penicillin, florfenicol, danofloxacin, enrofloxacin, chlortetracycline, doxycycline, and oxytetracycline had variable MIC₅₀ of 64, 0.5, 1, 1, 8, 4, 8, 4, 8, 0.5 μg/ml, respectively. It is recommended that Cp infections in Jordan be treated with either penicillins or tetracyclines especially amoxicillin and oxytetracycline. Copyright © 2010 Elsevier Ltd. All rights reserved.

Enterococcus faecium isolated from honey synthesized bacteriocin-like substances active against different Listeria monocytogenes strains.

PubMed

Ibarguren, Carolina; Raya, Raúl R; Apella, María C; Audisio, M Carina

2010-02-01

Four Enterococcus faecium strains, isolated from honeycombs (C1 and M2d strains) and feral combs (Mori1 and M1b strains) secreted antimicrobial substances active against fourteen different Listeria spp. strains. The antimicrobial compound(s) present in the cell free supernatant were highly thermostable (121 degrees C for 15 min) and inactivated by proteolytic enzymes, but not by alpha-amylase and lipase, thus suggesting a peptidic nature. Since the structural bacteriocin gene determinants of enterocins A and B were PCR amplified from the four E. faecium isolates, only the bacteriocin produced by strain C1 was further characterized: it showed a broad band of approximately 4.0-7.0 kDa in SDS-PAGE and was bactericidal (4 log decrease) against L. monocytogenes 99/287. L. monocytogenes 99/287R, a clone spontaneously resistant to the enterocin produced by E. avium DSMZ17511 (ex PA1), was not inhibited by the enterocin-like compounds produced by strain C1. However, it was inhibited in mixed culture fermentations by E. faecium C1 and a bacteriostatic effect was observed. The bacteriocin-producer Enterococcus strains were not haemolytic; gelatinase negative and sensitive to vancomycin and other clinically relevant antibiotics.

[Antibacterial activity of rare Streptomyces species against clinical resistant bacteria].

PubMed

Boughachiche, Faiza; Reghioua, Sihem; Zerizer, Habiba; Boulahrouf, Abderrahmane

2012-01-01

In the search for new antibiotics from Steptomyces, investigating extremes habitats enhances the probability of isolating novel producers. In this context, the antibacterial activity of four Streptomyces strains isolated from Ezzmoul saltpans was studied. Two of them showed antibacterial activity against antibiotic's resistant bacteria (Bacillus cereus: β-lactamines and sulfamides resistant, Streptococcus faecalis: penicillin, tetracycline and cotrimoxazole resistant, and Staphylococcus aureus Mu 50: vancomycine resistant). The most active Streptomyces strain produces one type of polar bioactive molecules that resists to temperature variation and light exposition. Its activity appears in the first culture day and reaches its maximal value in the fourth day. The second strain presents themoresistant activity that reaches its maximal value in the first culture day. It produces two types of bioactive molecules, one is polar and the second is non polar (according to thin layer chromatography technique results).

Macromolecular and solution properties of Cepacian: the exopolysaccharide produced by a strain of Burkholderia cepacia isolated from a cystic fibrosis patient.

PubMed

Sist, Paola; Cescutti, Paola; Skerlavaj, Silvia; Urbani, Ranieri; Leitão, Jorge H; Sá-Correia, Isabel; Rizzo, Roberto

2003-09-01

Light scattering and viscosity measurements were carried out on the previously chemically characterised exopolysaccharide produced by a strain of Burkholderia cepacia isolated from a cystic fibrosis patient. The same exopolysaccharide was also produced by other clinical strains in different laboratories. Therefore, the name Cepacian is now proposed for this exopolysaccharide. Experiments performed as a function of the ionic strength on the native polymer revealed a change in the overall shape of the polymer at low ionic strength. This behaviour was absent in the de-acetylated sample. Potentiometric titrations and light scattering experiments carried out on the acidic form of the native polymer revealed the formation of macromolecular aggregates with a stoichiometry n and 2n stabilised by interactions involving the uronic acid residues.

Insights from the genome of a high alkaline cellulase producing Aspergillus fumigatus strain obtained from Peruvian Amazon rainforest.

PubMed

Paul, Sujay; Zhang, Angel; Ludeña, Yvette; Villena, Gretty K; Yu, Fengan; Sherman, David H; Gutiérrez-Correa, Marcel

2017-06-10

Here, we report the complete genome sequence of a high alkaline cellulase producing Aspergillus fumigatus strain LMB-35Aa isolated from soil of Peruvian Amazon rainforest. The genome is ∼27.5mb in size, comprises of 228 scaffolds with an average GC content of 50%, and is predicted to contain a total of 8660 protein-coding genes. Of which, 6156 are with known function; it codes for 607 putative CAZymes families potentially involved in carbohydrate metabolism. Several important cellulose degrading genes, such as endoglucanase A, endoglucanase B, endoglucanase D and beta-glucosidase, are also identified. The genome of A. fumigatus strain LMB-35Aa represents the first whole sequenced genome of non-clinical, high cellulase producing A. fumigatus strain isolated from forest soil. Copyright © 2017 Elsevier B.V. All rights reserved.

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.

Isolation, Analysis and Antimicrobial Activity of the Acidic Fractions of Mastic, Kurdica, Mutica and Cabolica Gums from Genus Pistacia

PubMed Central

Sharifi, Mohammad Sharif; Hazell, Stuart Loyd

2012-01-01

The chemical entities of Mastic, Kurdica, Mutica and Cabolica gums from genus Pistacia have been isolated and characterised by GC-Mass Spectrometry, High Performance Liquid Chromatography and Column Chromatography. These chemical entities were screened for anti-microbial activities against nine strains of Helicobacter pylori and some other Gram-negative and Gram-positive bacteria. The most bioactive components were structurally analysed. These components mimic steroid compounds, in particular, the known antibiotic Fusidic acid. Some of these chemical entities have produced promising data that could lead to the development of a novel class of antimicrobial agents that may have application in the treatment of infectious disease. Kill kinetics have been also performed, and the produced data were evaluated by Generalized Multiplicative Analysis Of Variance (GEMANOVA) for the bactericidal and bacteriostatic activities which can be clinically significant. The isolated components were all bactericidal. PMID:22980113

Inter-laboratory comparison of multi-locus variable-number tandem repeat analysis (MLVA) for verocytotoxin-producing Escherichia coli O157 to facilitate data sharing.

PubMed

Holmes, A; Perry, N; Willshaw, G; Hanson, M; Allison, L

2015-01-01

Multi-locus variable number tandem repeat analysis (MLVA) is used in clinical and reference laboratories for subtyping verocytotoxin-producing Escherichia coli O157 (VTEC O157). However, as yet there is no common allelic or profile nomenclature to enable laboratories to easily compare data. In this study, we carried out an inter-laboratory comparison of an eight-loci MLVA scheme using a set of 67 isolates of VTEC O157. We found all but two isolates were identical in profile in the two laboratories, and repeat units were homogeneous in size but some were incomplete. A subset of the isolates (n = 17) were sequenced to determine the actual copy number of representative alleles, thereby enabling alleles to be named according to international consensus guidelines. This work has enabled us to realize the potential of MLVA as a portable, highly discriminatory and convenient subtyping method.

ESBL, MBL and Ampc β Lactamases Producing Superbugs – Havoc in the Intensive Care Units of Punjab India

PubMed Central

Oberoi, Loveena; Singh, Nachhatarjit; Sharma, Poonam; Aggarwal, Aruna

2013-01-01

Background: An alarming rise in the rates of the antibiotic resistance has now become a serious and an increasingly common public health concern, with severe implications, especially in the intensive care units. A variety of β-lactamases which include ESBLs, AmpC β-lactamases and metallo-βlactamases, have emerged as the most worrisome mechanism of resistance among the gram negative bacteria, which pose a therapeutic challenge to the health care settings. Materials and Methods: The present study was aimed at knowing the prevalence of various β-lactamases in the gram negative isolates which were obtained from ICU patients. A total 273 gram negative isolates from 913 clinical samples which were received over a period of one year were processed for their identification and their antimicrobial susceptibility pattern was determined. They were then screened for the β-lactamase production. Results: Among the 273 isolates, the β-lactamase production was observed in 193 strains. 96 (35.16%) strains were ESBL producers, followed by 30 (10.98%) metallo β- lactamase (MBL) producers and 15(5.4%) AmpC producers. The major ESBL and AmpC producer was Escherichia coli, while Klebsiella pneumonia was the predominant MBL producer. The co production of the ESBL/MBL/ AmpC β- lactamases was observed in 52 (19.04%) strains and it was more common in Escherichia coli. A multidrug resistance to the fluoroquinolones and the aminoglycosides was also observed in the β- lactamase producing organisms. Conclusion: The high prevalence of the β- lactamases in the ICU isolates emphasizes the need for a continuous surveillance in the ICUs to detect the resistant strains, strict guidelines for the antibiotic therapy and the implementation of infection control measures to reduce the increasing burden of antibiotic resistance. PMID:23450498

Detection of ESBL among ampc producing enterobacteriaceae using inhibitor-based method

PubMed Central

Bakthavatchalu, Sasirekha; Shakthivel, Uma; Mishra, Tannu

2013-01-01

Introduction The occurrence of multiple β-lactamases among bacteria only limits the therapeutic options but also poses a challenge. A study using boronic acid (BA), an AmpC enzyme inhibitor, was designed to detect the combined expression of AmpC β-lactamases and extended-spectrum β-lactamases (ESBLs) in bacterial isolates further different phenotypic methods are compared to detect ESBL and AmpC. Methods A total of 259 clinical isolates of Enterobacteriaceae were isolated and screened for ESBL production by (i) CLSI double-disk diffusion method (ii) cefepime- clavulanic acid method (iii) boronic disk potentiation method. AmpC production was detected using cefoxitin alone and in combination with boronic acid and confirmation was done by three dimensional disk methods. Isolates were also subjected to detailed antibiotic susceptibility test. Results Among 259 isolates, 20.46% were coproducers of ESBL and AmpC, 26.45% were ESBL and 5.40% were AmpC. All of the 53 AmpC and ESBL coproducers were accurately detected by boronic acid disk potentiation method. Conclusion The BA disk test using Clinical and Laboratory Standards Institute methodology is simple and very efficient method that accurately detects the isolates that harbor both AmpCs and ESBLs. PMID:23504148

Emergence of antibiotic-resistant bacteria in patients with Fournier gangrene.

PubMed

Lin, Wei-Ting; Chao, Chien-Ming; Lin, Hsin-Lan; Hung, Ming-Chran; Lai, Chih-Cheng

2015-04-01

This study was conducted to investigate the bacteriology and associated patterns of antibiotic resistance Fournier gangrene. Patients with Fournier's gangrene from 2008 to 2012 were identified from the computerized database in a medical center in southern Taiwan. The medical records of all patients with Fournier's gangrene were reviewed retrospectively. There were 61 microorganisms, including 60 bacteria and one Candida spp, isolated from clinical wound specimens from 32 patients. The most common isolates obtained were Streptococcus spp. (n=12), Peptoniphilus spp. (n=8), Staphylococcus aureus (n=7), Escherichia coli (n=7), and Klebsiella pneumoniae (n=7). Among 21 strains of gram-negative bacilli, five (23.8%) were resistant to fluoroquinolones, and three isolates were resistant to ceftriaxone. Two E. coli strains produced extended-spectrum beta-lactamase. Four of the seven S. aureus isolates were methicillin-resistant. Among 15 anaerobic isolates, nine (60%) were resistant to penicillin, and eight (53.3%) were resistant to clindamycin. Four (26.7%) isolates were resistant to metronidazole. The only independent risk factor associated with mortality was inappropriate initial antibiotic treatment (p=0.021). Antibiotic-resistant bacteria are emerging in the clinical setting of Fournier gangrene. Clinicians should use broad-spectrum antibiotics initially to cover possible antibiotic-resistant bacteria.

Development of an algorithm for phenotypic screening of carbapenemase-producing Enterobacteriaceae in the routine laboratory.

PubMed

Robert, Jérôme; Pantel, Alix; Merens, Audrey; Meiller, Elodie; Lavigne, Jean-Philippe; Nicolas-Chanoine, Marie-Hélène

2017-01-17

Carbapenemase-producing Enterobacteriaceae (CPE) are difficult to identify among carbapenem non-susceptible Enterobacteriaceae (NSE). We designed phenotypic strategies giving priority to high sensitivity for screening putative CPE before further testing. Presence of carbapenemase-encoding genes in ertapenem NSE (MIC > 0.5 mg/l) consecutively isolated in 80 French laboratories between November 2011 and April 2012 was determined by the Check-MDR-CT103 array method. Using the Mueller-Hinton (MH) disk diffusion method, clinical diameter breakpoints of carbapenems other than ertapenem, piperazicillin+tazobactam, ticarcillin+clavulanate and cefepime as well as diameter cut-offs for these antibiotics and temocillin were evaluated alone or combined to determine their performances (sensitivity, specificity, positive and negative likelihood ratios) for identifying putative CPE among these ertapenem-NSE isolates. To increase the screening specificity, these antibiotics were also tested on cloxacillin-containing MH when carbapenem NSE isolates belonged to species producing chromosomal cephalosporinase (AmpC) but Escherichia coli. Out of the 349 ertapenem NSE, 52 (14.9%) were CPE, including 39 producing OXA-48 group carbapenemase, eight KPC and five MBL. A screening strategy based on the following diameter cut offs, ticarcillin+clavulanate <15 mm, temocillin <15 mm, meropenem or imipenem <22 mm, and cefepime <26 mm, showed 100% sensitivity and 68.1% specificity with the better likelihood ratios combination. The specificity increased when a diameter cut-off <32 mm for imipenem (76.1%) or meropenem (78.8%) further tested on cloxacillin-containing MH was added to the previous strategy for AmpC-producing isolates. The proposed strategies that allowed for increasing the likelihood of CPE among ertapenem-NSE isolates should be considered as a surrogate for carbapenemase production before further CPE confirmatory testing.

Role of AcrR and RamA in Fluoroquinolone Resistance in Clinical Klebsiella pneumoniae Isolates from Singapore

PubMed Central

Schneiders, T.; Amyes, S. G. B.; Levy, S. B.

2003-01-01

The MICs of ciprofloxacin for 33 clinical isolates of K. pneumoniae resistant to extended-spectrum cephalosporins from three hospitals in Singapore ranged from 0.25 to >128 μg/ml. Nineteen of the isolates were fluoroquinolone resistant according to the NCCLS guidelines. Strains for which the ciprofloxacin MIC was ≥0.5 μg/ml harbored a mutation in DNA gyrase A (Ser83→Tyr, Leu, or IIe), and some had a secondary Asp87→Asn mutation. Isolates for which the MIC was 16 μg/ml possessed an additional alteration in ParC (Ser80→IIe, Trp, or Arg). Tolerance of the organic solvent cyclohexane was observed in 10 of the 19 fluoroquinolone-resistant strains; 3 of these were also pentane tolerant. Five of the 10 organic solvent-tolerant isolates overexpressed AcrA and also showed deletions within the acrR gene. Complementation of the mutated acrR gene with the wild-type gene decreased AcrA levels and produced a two- to fourfold reduction in the fluoroquinolone MICs. None of the organic solvent-tolerant clinical isolates overexpressed another efflux-related gene, acrE. While marA and soxS were not overexpressed, another marA homologue, ramA, was overexpressed in 3 of 10 organic solvent-tolerant isolates. These findings indicate that multiple target and nontarget gene changes contribute to fluoroquinolone resistance in K. pneumoniae. Besides AcrR mutations, ramA overexpression (but not marA or soxS overexpression) was related to increased AcrAB efflux pump expression in this collection of isolates. PMID:12936981

Identification and Pathogenic Potential of Clinical Bacillus and Paenibacillus Isolates

PubMed Central

Celandroni, Francesco; Salvetti, Sara; Gueye, Sokhna Aissatou; Mazzantini, Diletta; Lupetti, Antonella; Senesi, Sonia; Ghelardi, Emilia

2016-01-01

The soil-related Bacillus and Paenibacillus species have increasingly been implicated in various human diseases. Nevertheless, their identification still poses problems in the clinical microbiology laboratory and, with the exception of Bacillus anthracis and Bacillus cereus, little is known on their pathogenicity for humans. In this study, we evaluated the use of matrix-assisted laser desorption—ionization time of flight mass spectrometry (MALDI-TOF MS) in the identification of clinical isolates of these genera and conducted genotypic and phenotypic analyses to highlight specific virulence properties. Seventy-five clinical isolates were subjected to biochemical and MALDI-TOF MS identification. 16S rDNA sequencing and supplemental tests were used to solve any discrepancies or failures in the identification results. MALDI-TOF MS significantly outperformed classical biochemical testing for correct species identification and no misidentification was obtained. One third of the collected strains belonged to the B. cereus species, but also Bacillus pumilus and Bacillus subtilis were isolated at high rate. Antimicrobial susceptibility testing showed that all the B. cereus, B. licheniformis, B. simplex, B. mycoides, Paenibacillus glucanolyticus and Paenibacillus lautus isolates are resistant to penicillin. The evaluation of toxin/enzyme secretion, toxin-encoding genes, motility, and biofilm formation revealed that B. cereus displays the highest virulence potential. However, although generally considered nonpathogenic, most of the other species were shown to swim, swarm, produce biofilms, and secrete proteases that can have a role in bacterial virulence. In conclusion, MALDI-TOF MS appears useful for fast and accurate identification of Bacillus and Paenibacillus strains whose virulence properties make them of increasing clinical relevance. PMID:27031639

Isolates of Cryptococcus neoformans from Infected Animals Reveal Genetic Exchange in Unisexual, α Mating Type Populations▿ †

PubMed Central

Bui, Tien; Lin, Xiaorong; Malik, Richard; Heitman, Joseph; Carter, Dee

2008-01-01

Sexual reproduction and genetic exchange are important for the evolution of fungal pathogens and for producing potentially infective spores. Studies to determine whether sex occurs in the pathogenic yeast Cryptococcus neoformans var. grubii have produced enigmatic results, however: basidiospores are the most likely infective propagules, and clinical isolates are fertile and genetically diverse, consistent with a sexual species, but almost all populations examined consist of a single mating type and have little evidence for genetic recombination. The choice of population is critical when looking for recombination, particularly when significant asexual propagation is likely and when latency may complicate assessing the origin of an isolate. We therefore selected isolates from infected animals living in the region of Sydney, Australia, with the assumption that the relatively short life spans and limited travels of the animal hosts would provide a very defined population. All isolates were mating type α and were of molecular genotype VNI or VNII. A lack of linkage disequilibrium among loci suggested that genetic exchange occurred within both genotype groups. Four diploid VNII isolates that produced filaments and basidium-like structures when cultured in proximity to an a mating type strain were found. Recent studies suggest that compatible α-α unions can occur in C. neoformans var. neoformans populations and in populations of the sibling species Cryptococcus gattii. As a mating type strains of C. neoformans var. grubii have never been found in Australia, or in the VNII molecular type globally, the potential for α-α unions is evidence that α-α unisexual mating maintains sexual recombination and diversity in this pathogen and may produce infectious propagules. PMID:18552280

Respiratory disease due to current egg drop syndrome virus in Pekin ducks.

PubMed

Cha, Se-Yeoun; Kang, Min; Moon, Oun-Kyoung; Park, Choi-Kyu; Jang, Hyung-Kwan

2013-08-30

Severe acute respiratory symptoms with coughing, dyspnea, and gasping were reported in two flocks of 9-day-old Pekin ducklings from different provinces. Gross lesions, white exudate and mucous membrane congestion in the trachea as well as blue to purple color changes and sclerosis in lungs were observed. Histological lesions revealed that the trachea and bronchial epithelium were hyperplastic and infiltrated by neutrophil granulocytes. Egg drop syndrome virus (EDSV) was differentially diagnosed using polymerase chain reaction, and the strains were isolated from tracheas and lungs by inoculation of 10-day-old embryonated duck eggs. The virus isolates were designated strain D11-JW-012 and D11-JW-017. The clinical and pathological signs were reproduced by intra-tracheal inoculation of the isolates in 3-day-old ducklings. Although the two isolates produced similar clinical signs, pathological lesions and ciliostasis, the D11-JW-017 strain resulted in more severe clinical signs with progressive symptoms compared to those of D11-JW-012 strain-infected ducklings. We suggest that different EDSV strains with mild or severe to moderate pathogenicity coexist and have potential risks in poultry. Hereby, we report an EDSV infection in ducklings. Copyright © 2013 Elsevier B.V. All rights reserved.

Molecular typing of Epidermophyton floccosum isolated from patients with dermatophytosis by RAPD-PCR.

PubMed

Khosravi, Alireza; Behzad, Forough; Sabokbar, Azar; Shokri, Hojjatollah; Haddadi, Siamak; Masoudi-Nejad, Ali

2010-12-01

We evaluated the ability of randomly amplified polymorphic DNA (RAPD) to type Epidermophyton floccosum isolates recovered from patients with dermatophytosis originating from different regions of Iran. A total of 13 clinical isolates of E. floccosum obtained from Iranian patients were analyzed by RAPD with 7 arbitrary primers (OPN16, OPD18' OPU15, OPX19, R28, OPA04 and OPAA17). Among the applied primers, OPN16 produced banding patterns from all the isolates. In addition, some of the isolates had very close relation. The phenon line which represented the mean similarities was at the value of 73%. At this level, 4 groups were characterized. Two isolates of a patient had different molecular patterns, suggesting infection transmission from different sources in the case of a single patient. RAPD-PCR provided a rapid and practical tool for identification of E. floccosum isolates, which was independent of morphological characteristics, and enhanced laboratory diagnosis of dermatophytosis.

Biofilm formation by Staphylococcus haemolyticus.

PubMed

Fredheim, Elizabeth Gladys Aarag; Klingenberg, Claus; Rohde, Holger; Frankenberger, Stephanie; Gaustad, Peter; Flaegstad, Trond; Sollid, Johanna Ericson

2009-04-01

Infections due to coagulase-negative staphylococci (CoNS) most frequently occur after the implantation of medical devices and are attributed to the biofilm-forming potential of CoNS. Staphylococcus haemolyticus is the second most frequently isolated CoNS from patients with hospital-acquired infections. There is only limited knowledge of the nature of S. haemolyticus biofilms. The aim of this study was to characterize S. haemolyticus biofilm formation. We analyzed the biofilm-forming capacities of 72 clinical S. haemolyticus isolates. A detachment assay with NaIO(4), proteinase K, or DNase was used to determine the main biofilm components. Biofilm-associated genes, including the ica operon, were analyzed by PCR, and the gene products were sequenced. Confocal laser scanning microscopy (CLSM) was used to elucidate the biofilm structure. Fifty-three isolates (74%) produced biofilms after growth in Trypticase soy broth (TSB) with glucose, but only 22 (31%) produced biofilms after growth in TSB with NaCl. It was necessary to dissolve the biofilm in ethanol-acetone to measure the optical density of the full biofilm mass. DNase, proteinase K, and NaIO(4) caused biofilm detachment for 100%, 98%, and 38% of the isolates, respectively. icaRADBC and polysaccharide intercellular adhesin (PIA) production were found in only two isolates. CLSM indicated that the biofilm structure of S. haemolyticus clearly differs from that of S. epidermidis. We conclude that biofilm formation is a common phenotype in clinical S. haemolyticus isolates. In contrast to S. epidermidis, proteins and extracellular DNA are of functional relevance for biofilm accumulation, whereas PIA plays only a minor role. The induction of biofilm formation and determination of the biofilm mass also needed to be optimized for S. haemolyticus.

Detection of Antibiotic Resistance Determinants and their Transmissibility among Clinically-isolated Carbapenem-resistant Escherichia coli from South India.

PubMed

Mahalingam, Niranjana; Manivannan, Bhavani; Khamari, Balaram; Siddaramappa, Shivakumara; Adak, Sudeshna; Bulagonda, Eswarappa Pradeep

2018-05-08

The aim of this study was to analyze the prevalence of CTX-M, TEM, SHV, VIM, NDM and OXA genes in carbapenemase-producing E. coli and their transmissibility at a tertiary care hospital in south India. Twenty-one carbapenem-resistant E. coli (CRE) were collected from Sri Sathya Sai Institute of Higher Medical Sciences, Prasanthigram, India. Resistance to antibiotics was analyzed by Vitek-2, and identity of the isolates was confirmed by 16S rDNA sequencing. RAPD and ERIC-PCR was performed for molecular typing. Metallo beta-lactamase production was confirmed by double disc synergy test. Presence of the extended spectrum beta lactamases CTX-M, TEM and SHV, and the carbapenemases NDM, VIM and OXA were determined by PCR. Carbapenemase variants were further confirmed by sequencing. Transmissibility of the genes was tested by conjugation. Twelve of the twenty-one (57%) carbapenem-resistant E. coli isolates were community-acquired, indicating the spread of CRE in environmental samples. TEM and NDM-5 were found to be the major beta-lactamases produced by the pathogens. OXA-181 was found in five of the isolates. All the 21 isolates were found to harbor more than one of the tested beta-lactamases, and all the isolates were found to have the capacity to participate in conjugation; fifteen of the transconjugants were found to have acquired the tested beta-lactamases, substantiating their ability to get transferred to other strains of bacteria. Monitoring of community-acquired carbapenem-resistant bacteria is very important as the association of resistance determinants with mobile genetic elements would present a serious clinical challenge. ©2018The Author(s). Published by S. Karger AG, Basel.

Comparison of CHROMagar Salmonella Medium and Xylose-Lysine-Desoxycholate and Salmonella-Shigella Agars for Isolation of Salmonella Strains from Stool Samples

PubMed Central

Maddocks, Susan; Olma, Tom; Chen, Sharon

2002-01-01

The growth and appearance of 115 stock Salmonella isolates on a new formulation of CHROMagar Salmonella (CAS) medium were compared to those on xylose-lysine-desoxycholate agar (XLD), Salmonella-Shigella agar (SS), and Hektoen enteric agar (HEA) media. CAS medium was then compared prospectively to XLD and SS for the detection and presumptive identification of Salmonella strains in 500 consecutive clinical stool samples. All stock Salmonella isolates produced typical mauve colonies on CAS medium. Nine Salmonella strains were isolated from clinical specimens. The sensitivities for the detection of salmonellae after primary plating on CAS medium and the combination of XLD and SS after enrichment were 100%. The specificity for the detection of salmonellae after primary plating on CAS medium (83%) was significantly (P < 0.0001) higher than that after primary plating on the combination of SS and XLD media (55%) (a 28% difference in rates; 95% confidence interval, 23.0 to 34%). Twenty-nine non-Salmonella organisms produced mauve colonies on CAS medium, including 17 Candida spp. (59%) and 8 Pseudomonas spp. (28%). These were easily excluded as salmonellae by colony morphology, microscopic examination of a wet preparation, or oxidase testing. One biochemically inert Escherichia coli isolate required further identification to differentiate it from Salmonella spp. The use of plating on CAS medium demonstrated high levels of sensitivity and specificity and reduced the time to final identification of Salmonella spp., resulting in substantial cost savings. It can be recommended for use for the primary isolation of Salmonella spp. from stool specimens. Other media (e.g., XLD) are required to detect Shigella spp. concurrently. PMID:12149365

Population Structure of Clinical Pseudomonas aeruginosa from West and Central African Countries

PubMed Central

Cholley, Pascal; Ka, Roughyatou; Guyeux, Christophe; Thouverez, Michelle; Guessennd, Nathalie; Ghebremedhin, Beniam; Frank, Thierry; Bertrand, Xavier; Hocquet, Didier

2014-01-01

Background Pseudomonas aeruginosa (PA) has a non-clonal, epidemic population with a few widely distributed and frequently encountered sequence types (STs) called ‘high-risk clusters’. Clinical P. aeruginosa (clinPA) has been studied in all inhabited continents excepted in Africa, where a very few isolates have been analyzed. Here, we characterized a collection of clinPA isolates from four countries of West and Central Africa. Methodology 184 non-redundant isolates of clinPA from hospitals of Senegal, Ivory Coast, Nigeria, and Central African Republic were genotyped by MLST. We assessed their resistance level to antibiotics by agar diffusion and identified the extended-spectrum β-lactamases (ESBLs) and metallo-β-lactamases (MBLs) by sequencing. The population structure of the species was determined by a nucleotide-based analysis of the entire PA MLST database and further localized on the phylogenetic tree (i) the sequence types (STs) of the present collection, (ii) the STs by continents, (iii) ESBL- and MBL-producing STs from the MLST database. Principal Findings We found 80 distinct STs, of which 24 had no relationship with any known STs. ‘High-risk’ international clonal complexes (CC155, CC244, CC235) were frequently found in West and Central Africa. The five VIM-2-producing isolates belonged to CC233 and CC244. GES-1 and GES-9 enzymes were produced by one CC235 and one ST1469 isolate, respectively. We showed the spread of ‘high-risk’ international clonal complexes, often described as multidrug-resistant on other continents, with a fully susceptible phenotype. The MBL- and ESBL-producing STs were scattered throughout the phylogenetic tree and our data suggest a poor association between a continent and a specific phylogroup. Conclusions ESBL- and MBL-encoding genes are borne by both successful international clonal complexes and distinct local STs in clinPA of West and Central Africa. Furthermore, our data suggest that the spread of a ST could be either due to its antibiotic resistance or to features independent from the resistance to antibiotics. PMID:25187957

Mesenchymal Stem/Stromal Cells: A New "Cells as Drugs" Paradigm. Efficacy and Critical Aspects in Cell Therapy

PubMed Central

de Girolamo, Laura; Lucarelli, Enrico; Alessandri, Giulio; Avanzini, Maria Antonietta; Bernardo, Maria Ester; Biagi, Ettore; Brini, Anna Teresa; D’Amico, Giovanna; Fagioli, Franca; Ferrero, Ivana; Locatelli, Franco; Maccario, Rita; Marazzi, Mario; Parolini, Ornella; Pessina, Augusto; Torre, Maria Luisa

2013-01-01

Mesenchymal stem cells (MSCs) were first isolated more than 50 years ago from the bone marrow. Currently MSCs may also be isolated from several alternative sources and they have been used in more than a hundred clinical trials worldwide to treat a wide variety of diseases. The MSCs mechanism of action is undefined and currently under investigation. For in vivo purposes MSCs must be produced in compliance with good manufacturing practices and this has stimulated research on MSCs characterization and safety. The objective of this review is to describe recent developments regarding MSCs properties, physiological effects, delivery, clinical applications and possible side effects. PMID:23278600

Comparison of Meropenem MICs and Susceptibilities for Carbapenemase-Producing Klebsiella pneumoniae Isolates by Various Testing Methods▿

PubMed Central

Bulik, Catharine C.; Fauntleroy, Kathy A.; Jenkins, Stephen G.; Abuali, Mayssa; LaBombardi, Vincent J.; Nicolau, David P.; Kuti, Joseph L.

2010-01-01

We describe the levels of agreement between broth microdilution, Etest, Vitek 2, Sensititre, and MicroScan methods to accurately define the meropenem MIC and categorical interpretation of susceptibility against carbapenemase-producing Klebsiella pneumoniae (KPC). A total of 46 clinical K. pneumoniae isolates with KPC genotypes, all modified Hodge test and blaKPC positive, collected from two hospitals in NY were included. Results obtained by each method were compared with those from broth microdilution (the reference method), and agreement was assessed based on MICs and Clinical Laboratory Standards Institute (CLSI) interpretative criteria using 2010 susceptibility breakpoints. Based on broth microdilution, 0%, 2.2%, and 97.8% of the KPC isolates were classified as susceptible, intermediate, and resistant to meropenem, respectively. Results from MicroScan demonstrated the most agreement with those from broth microdilution, with 95.6% agreement based on the MIC and 2.2% classified as minor errors, and no major or very major errors. Etest demonstrated 82.6% agreement with broth microdilution MICs, a very major error rate of 2.2%, and a minor error rate of 2.2%. Vitek 2 MIC agreement was 30.4%, with a 23.9% very major error rate and a 39.1% minor error rate. Sensititre demonstrated MIC agreement for 26.1% of isolates, with a 3% very major error rate and a 26.1% minor error rate. Application of FDA breakpoints had little effect on minor error rates but increased very major error rates to 58.7% for Vitek 2 and Sensititre. Meropenem MIC results and categorical interpretations for carbapenemase-producing K. pneumoniae differ by methodology. Confirmation of testing results is encouraged when an accurate MIC is required for antibiotic dosing optimization. PMID:20484603

Environmental monitoring and assessment of antibacterial metabolite producing actinobacteria screened from marine sediments in south coastal regions of Karnataka, India.

PubMed

Skariyachan, Sinosh; Garka, Shruthi; Puttaswamy, Sushmitha; Shanbhogue, Shobitha; Devaraju, Raksha; Narayanappa, Rajeswari

2017-06-01

Assessment of the therapeutic potential of secondary metabolite producing microorganisms from the marine coastal areas imparts scope and application in the field of environmental monitoring. The present study aims to screen metabolites with antibacterial potential from actionbacteria associated with marine sediments collected from south coastal regions of Karnataka, India. The actinobacteria were isolated and characterized from marine sediments by standard protocol. The metabolites were extracted, and antibacterial potential was analyzed against eight hospital associated bacteria. The selected metabolites were partially characterized by proximate analysis, SDS-PAGE, and FTIR-spectroscopy. The antibiogram of the test clinical isolates revealed that they were emerged as multidrug-resistant strains (P ≤ 0.05). Among six actinobacteria (IS1-1S6) screened, 100 μl -1 metabolite from IS1 showed significant antibacterial activities against all the clinical isolates except Pseudomonas aeruginosa. IS2 demonstrated antimicrobial potential towards Proteus mirabilis, Streptococcus pyogenes, and Escherichia coli. The metabolite from IS3 showed activity against Strep. pyogenes and E. coli. The metabolites from IS4, IS5, and IS6 exhibited antimicrobial activities against Ps. aeruginosa (P ≤ 0.05). The two metabolites that depicted highest antibacterial activities against the test strains were suggested to be antimicrobial peptides with low molecular weight. These isolates were characterized and designated as Streptomyces sp. strain mangaluru01 and Streptomyces sp. mangaloreK01 by 16S ribosomal DNA (rDNA) sequencing. This study suggests that south coastal regions of Karnataka, India, are one of the richest sources of antibacterial metabolites producing actinobacteria and monitoring of these regions for therapeutic intervention plays profound role in healthcare management.

[Identification of Candida dubliniensis strains using heat tolerance tests, morphological characteristics and molecular methods].

PubMed

Arikan, Sevtap; Darka, Ozge; Hasçelik, Gülşen; Günalp, Ayfer

2003-01-01

Described in 1995, Candida dubliniensis is a novel Candida species closely related to Candida albicans due primarily to its ability to produce germ tube and chlamydospores. Given these phenotypic similarities between the two species, C. dubliniensis cannot be readily distinguished from Candida albicans by routine laboratory work-up. We explored the frequency of isolation of C. dubliniensis among 213 strains previously defined as C. albicans based on their ability to produce germ tube. The test isolates were initially examined for their morphological features on cornmeal tween 80 agar, inability to grow at 45 degrees C, and the biochemical assimilation profile (ID 32C system, bioMerieux, France). Among all, 2 (0.9%) of the isolates were identified as C. dubliniensis based on the production of numerous chlamydospores in chains on cornmeal tween 80 agar and the lack of growth at 45 degrees C. The assimilation profile of these isolates was found to be in accordance with this identification. In an effort to confirm the identification, polymerase chain reaction (PCR) studies were carried out by using the C. dubliniensis specific primer set, DUBF and DUBR. Both of the isolates yielded C. dubliniensis-specific 288 base pair amplification products, confirming the previous identification obtained with the initial screening tests. The isolates were found to be susceptible to fluconazole and itraconazole, and generated amphotericin B minimal inhibitory concentrations of 0.5-1 microgram/ml by NCCLS M27-A2 microdilution method. These data suggest that the isolation rate of C. dubliniensis among our clinical isolates is low. The morphological features on cornmeal tween 80 agar and the lack of ability to grow at 45 degrees C appear as reliable, cheap, and practical screening tests in initial identification of C. dubliniensis among germ tube-producing Candida strains.

Clinical evaluation of the Vitek ANI card for identification of anaerobic bacteria.

PubMed Central

Schreckenberger, P C; Celig, D M; Janda, W M

1988-01-01

An evaluation of the Vitek Anaerobe Identification (ANI) card was performed with 341 bacterial isolates, including 313 clinical isolates and 28 stock strains of anaerobic microorganisms. Identifications obtained with the ANI card were compared with those determined by conventional methods. The card identified 73.2% of 149 anaerobic gram-negative bacilli, 63.6% of 44 Clostridium spp., 65.8% of 38 anaerobic nonsporeforming gram-positive bacilli, and 69.1% of 110 anaerobic cocci, with no further testing required. When genus-level identifications were included, 83.9% of the anaerobic gram-negative bacilli, 70.5% of Clostridium spp., 73.7% of the anaerobic nonsporeforming gram-positive bacilli, and 73.6% of the anaerobic cocci were identified. Nineteen isolates (5.6%) produced identifications of good confidence but marginal separation or questionable biotype, in which the correct identification was listed with one or two other possible choices and extra tests were required and suggested. A total of 28 (8.2%) were not identified and 29 isolates (8.5%) were misidentified by the ANI card. Among the commonly isolated clinically significant anaerobes, the ANI card identified 100% of 55 Bacteroides fragilis and 100% of 8 Clostridium perfringens. Use of supplemental tests and expansion of the data base to include additional strains of organisms that are difficult to separate even with conventional methods may improve the accuracy of the ANI card as a method for identification of anaerobic bacteria in the clinical laboratory. PMID:3343321

Limits of patient isolation measures to control extended-spectrum beta-lactamase–producing Enterobacteriaceae: model-based analysis of clinical data in a pediatric ward

PubMed Central

2013-01-01

Background Extended-spectrum beta-lactamase–producing Enterobacteriaceae (ESBL-E) are a growing concern in hospitals and the community. How to control the nosocomial ESBL-E transmission is a matter of debate. Contact isolation of patients has been recommended but evidence supporting it in non-outbreak settings has been inconclusive. Methods We used stochastic transmission models to analyze retrospective observational data from a two-phase intervention in a pediatric ward, successively implementing single-room isolation and patient cohorting in an isolation ward, combined with active ESBL-E screening. Results For both periods, model estimates suggested reduced transmission from isolated/cohorted patients. However, most of the incidence originated from sporadic sources (i.e. independent of cross-transmission), unaffected by the isolation measures. When sporadic sources are high, our model predicted that even substantial efforts to prevent transmission from carriers would have limited impact on ESBL-E rates. Conclusions Our results provide evidence that, considering the importance of sporadic acquisition, e.g. endogenous selection of resistant strains following antibiotic treatment, contact-isolation measures alone might not suffice to control ESBL-E. They also support the view that estimating cross-transmission extent is key to predicting the relative success of contact-isolation measures. Mathematical models could prove useful for those estimations and guide decisions concerning the most effective control strategy. PMID:23618041

Potential for biocontrol of melanized fungi by actinobacteria isolated from intertidal region of Ilha Do Mel, Paraná, Brazil.

PubMed

Dalitz, Camila de Araújo; Porsani, Mariana Vieira; Figel, Izabel Cristina; Pimentel, Ida C; Dalzoto, Patrícia R

Actinobacteria occur in many environments and have the capacity to produce secondary metabolites with antibiotic potential. Identification and taxonomy of actinobacteria that produce antimicrobial substances is essential for the screening of new compounds, and sequencing of the 16S region of ribosomal DNA (rDNA), which is conserved and present in all bacteria, is an important method of identification. Melanized fungi are free-living organisms, which can also be pathogens of clinical importance. This work aimed to evaluate growth inhibition of melanized fungi by actinobacteria and to identify the latter to the species level. In this study, antimicrobial activity of 13 actinobacterial isolates from the genus Streptomyces was evaluated against seven melanized fungi of the genera Exophiala, Cladosporium, and Rhinocladiella. In all tests, all actinobacterial isolates showed inhibitory activity against all isolates of melanized fungi, and only one actinobacterial isolate had less efficient inhibitory activity. The 16S rDNA region of five previously unidentified actinobacterial isolates from Ilha do Mel, Paraná, Brazil, was sequenced; four of the isolates were identified as Streptomyces globisporus subsp. globisporus, and one isolate was identified as Streptomyces aureus. This work highlights the potential of actinobacteria with antifungal activity and their role in the pursuit of novel antimicrobial substances. Copyright © 2016 Sociedade Brasileira de Microbiologia. Published by Elsevier Editora Ltda. All rights reserved.

Species identification of Aspergillus, Fusarium and Mucorales with direct surface analysis by matrix-assisted laser desorption ionization time-of-flight mass spectrometry.

PubMed

De Carolis, E; Posteraro, B; Lass-Flörl, C; Vella, A; Florio, A R; Torelli, R; Girmenia, C; Colozza, C; Tortorano, A M; Sanguinetti, M; Fadda, G

2012-05-01

Accurate species discrimination of filamentous fungi is essential, because some species have specific antifungal susceptibility patterns, and misidentification may result in inappropriate therapy. We evaluated matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) for species identification through direct surface analysis of the fungal culture. By use of culture collection strains representing 55 species of Aspergillus, Fusarium and Mucorales, a reference database was established for MALDI-TOF MS-based species identification according to the manufacturer's recommendations for microflex measurements and MALDI BioTyper 2.0 software. The profiles of young and mature colonies were analysed for each of the reference strains, and species-specific spectral fingerprints were obtained. To evaluate the database, 103 blind-coded fungal isolates collected in the routine clinical microbiology laboratory were tested. As a reference method for species designation, multilocus sequencing was used. Eighty-five isolates were unequivocally identified to the species level (≥99% sequence similarity); 18 isolates producing ambiguous results at this threshold were initially rated as identified to the genus level only. Further molecular analysis definitively assigned these isolates to the species Aspergillus oryzae (17 isolates) and Aspergillus flavus (one isolate), concordant with the MALDI-TOF MS results. Excluding nine isolates that belong to the fungal species not included in our reference database, 91 (96.8%) of 94 isolates were identified by MALDI-TOF MS to the species level, in agreement with the results of the reference method; three isolates were identified to the genus level. In conclusion, MALDI-TOF MS is suitable for the routine identification of filamentous fungi in a medical microbiology laboratory. © 2011 The Authors. Clinical Microbiology and Infection © 2011 European Society of Clinical Microbiology and Infectious Diseases.

Characterization of the RpoS Status of Clinical Isolates of Salmonella enterica

PubMed Central

Robbe-Saule, Véronique; Algorta, Gabriela; Rouilhac, Isabelle; Norel, Françoise

2003-01-01

The stationary-phase-inducible sigma factor, σS (RpoS), is the master regulator of the general stress response in Salmonella and is required for virulence in mice. rpoS mutants can frequently be isolated from highly passaged laboratory strains of Salmonella. We examined the rpoS status of 116 human clinical isolates of Salmonella, including 41 Salmonella enterica serotype Typhi strains isolated from blood, 38 S. enterica serotype Typhimurium strains isolated from blood, and 37 Salmonella serotype Typhimurium strains isolated from feces. We examined the abilities of these strains to produce the σS protein, to express RpoS-dependent catalase activity, and to resist to oxidative stress in the stationary phase of growth. We also carried out complementation experiments with a cloned wild-type rpoS gene. Our results showed that 15 of the 41 Salmonella serotype Typhi isolates were defective in RpoS. We sequenced the rpoS allele of 12 strains. This led to identification of small insertions, deletions, and point mutations resulting in premature stop codons or affecting regions 1 and 2 of σS, showing that the rpoS mutations are not clonal. Thus, mutant rpoS alleles can be found in freshly isolated clinical strains of Salmonella serotype Typhi, and they may affect virulence properties. Interestingly however, no rpoS mutants were found among the 75 Salmonella serotype Typhimurium isolates. Strains that differed in catalase activity and resistance to hydrogen peroxide were found, but the differences were not linked to the rpoS status. This suggests that Salmonella serotype Typhimurium rpoS mutants are counterselected because rpoS plays a role in the pathogenesis of Salmonella serotype Typhimurium in humans or in the transmission cycle of the disease. PMID:12902215

Synergy between baicalein and penicillins against penicillinase-producing Staphylococcus aureus.

PubMed

Qian, Minyi; Tang, Shusheng; Wu, Congming; Wang, Yang; He, Tao; Chen, Tingting; Xiao, Xilong

2015-09-01

The combination of baicalein (the active constituent of Scutellaria baicalensis) with penicillin G/amoxicillin showed potent synergy against 20 clinical penicillinase-producing Staphylococcus aureus strains including 10 isolates that were additionally methicillin-resistant (MRSA). The fractional inhibitory concentration (FIC) indices of penicillins+baiclein ranged from 0.14 to 0.38. Baicalein protected penicillins (penicillin G and amoxicillin) from penicillinase and increased the susceptibility of penicillinase-supplemented S. aureus ATCC 29213 in a dose-dependent manner. The inhibition of penicillinase activity by baicalein should be responsible for the synergism and protective effect. These findings offer us good evidence that the penicillins combined with baicalein showed potent synergistic activity against penicillinase-producing S. aureus and penicillinase-producing MRSA in vitro and might provide promising implications for clinical treatment of these bacterial infections. Copyright © 2015 Elsevier GmbH. All rights reserved.

Comparison of two commercial carbapenemase gene confirmatory assays in multiresistant Enterobacteriaceae and Acinetobacter baumannii-complex.

PubMed

Rösner, Stephan; Gehlweiler, Kevin; Küsters, Uta; Kolbert, Mathias; Hübner, Kirsten; Pfennigwerth, Niels; Mack, Dietrich

2018-01-01

Multidrug-resistant Gram-negative bacilli (MDR-GNB) producing carbapenemases are increasing at an alarming speed. Rapid confirmation of carbapenemase type will be an important diagnostic step in clinical microbiology laboratories not only to reduce the risk of transmissions but also for optimising antibiotic therapy in the future. We compared diagnostic reliability of two commercially available molecular assays (Check-Direct CPE vs. AID line probe assay) for detection and typing of carbapenemase genes in 80 well-characterized isolates of MDR-GNB. Respective strains were isolated in various clinical specimens at our clinical microbiology laboratory. The reference standard included confirmation of carbapenemase-production at the molecular level at the German National Reference Laboratory for Multidrug-resistant Gram-negative bacteria (Ruhr-University Bochum, Germany). 53 Enterobacteriaceae and 27 members of the A. baumannii-complex were used in this study. The tested assays appeared highly reliable to confirm carbapenemase-producing Enterobacteriaceae (CPE) with respective sensitivities of 97.7%, but are currently unsuitable for analysis of members of the A. baumannii-complex. Both assays are easy to perform and rapid tools for confirmation and typing of the most common carbapenemase genes in Enterobacteriaceae. Implementation should be possible for any clinical microbiology laboratory with Check-Direct CPE being easier to handle and having less technological requirements.

Molecular epidemiology and drug resistant mechanism in carbapenem-resistant Klebsiella pneumoniae isolated from pediatric patients in Shanghai, China.

PubMed

Zhang, Xingyu; Chen, Di; Xu, Guifeng; Huang, Weichun; Wang, Xing

2018-01-01

Infection by carbapenem-resistant Klebsiella pneumoniae (CR-KP) is a public health challenge worldwide, in particular among children, which was associated with high morbidity and mortality rates. There was limited data in pediatric populations, thus this study aimed to investigate molecular epidemiology and drug resistant mechanism of CR-KP strains from pediatric patients in Shanghai, China. A total of 41 clinical CR-KP isolates from sputum, urine, blood or drainage fluid were collected between July 2014 and May 2015 in Shanghai Children's Medical Center. Multilocus sequence typing (MLST), antibiotic susceptibility testing, PCR amplification and sequencing of the drug resistance associated genes were applied to all these isolates. MLST analysis revealed 16 distinct STs identified within the 41 isolates, among which the most frequently represented were ST11(19.5%),ST25(14.6%),ST76(14.6%),ST37(9.8%).One new ST was first identified. All CR-KP isolates showed MDR phenotypes and were resistance to ceftazidime, imipenem, piperacillin / tazobactam, ceftriaxone, ampicillin /sulbactam, aztreonam. They were confirmed as carbapenemase producer, NDM-1 (56.1%, 23/41), IMP (26.8%, 11/41), KPC-2 (22.0%, 9/41) were detected. Of note, two isolates carried simultaneously both NDM-1 and IMP-4. All CR-KP strains contained at least one of extended spectrum β-lactamase genes tested(TEM, SHV, OXA-1, CTX-M group) and six isolates carried both ESBL and AmpC genes(DHA-1). Among the penicllinase and β-lactamase genes, the most frequently one is SHV(92.7%,38/41), followed by TEM-1(68.3%,28/41), CTX-M-14(43.9%,18/41), CTX-M-15(43.9%,14/41), OXA-1(14.6%,6/41). In the present study, NDM-1-producing isolates was the predominant CR-KP strains in children, follow by IMP and KPC-producing strains. NDM-1and IMP-4 were more frequent than KPC-2 and showed a multiclonal background. Those suggested carbapenem-resistant in children is diverse, and certain resistance mechanisms differ from prevalent genotypes in adults in the same region. Knowledge of the molecular epidemiology and drug resistant mechanism of CR-KP can have a profound effect on clinical treatment, infection control measures and public health policies for children.

Genetic makeup of Shiga toxin-producing Escherichia coli in relation to clinical symptoms and duration of shedding: a microarray analysis of isolates from Swedish children.

PubMed

Matussek, A; Jernberg, C; Einemo, I-M; Monecke, S; Ehricht, R; Engelmann, I; Löfgren, S; Mernelius, S

2017-08-01

Shiga toxin (Stx)-producing Escherichia coli (STECs) cause non-bloody diarrhea, hemorrhagic colitis, and hemolytic uremic syndrome, and are the primary cause of acute renal failure in children worldwide. This study investigated the correlation of genetic makeup of STEC strains as revealed by DNA microarray to clinical symptoms and the duration of STEC shedding. All STEC isolated (n = 96) from patients <10 years of age in Jönköping County, Sweden from 2003 to 2015 were included. Isolates were characterized by DNA microarray, including almost 280 genes. Clinical data were collected through a questionnaire and by reviewing medical records. Of the 96 virulence genes (including stx) in the microarray, 62 genes were present in at least one isolate. Statistically significant differences in prevalence were observed for 21 genes when comparing patients with bloody diarrhea (BD) and with non-bloody stool (18 of 21 associated with BD). Most genes encode toxins (e.g., stx2 alleles, astA, toxB), adhesion factors (i.e. espB_O157, tir, eae), or secretion factors (e.g., espA, espF, espJ, etpD, nleA, nleB, nleC, tccP). Seven genes were associated with prolonged stx shedding; the presence of three genes (lpfA, senB, and stx1) and the absence of four genes (espB_O157, espF, astA, and intI1). We found STEC genes that might predict severe disease outcome already at diagnosis. This can be used to develop diagnostic tools for risk assessment of disease outcome. Furthermore, genes associated with the duration of stx shedding were detected, enabling a possible better prediction of length of STEC carriage after infection.

The first human report of mobile colistin resistance gene, mcr-1, in Finland.

PubMed

Gröndahl-Yli-Hannuksela, Kirsi; Lönnqvist, Emilia; Kallonen, Teemu; Lindholm, Laura; Jalava, Jari; Rantakokko-Jalava, Kaisu; Vuopio, Jaana

2018-05-01

Colistin resistance mediated by mobile mcr-1 gene has raised concern during the last years. After steep increase in mcr-1 reports, other mcr-gene variants (mcr-2 to mcr-5) have been revealed as well. In 2016, a clinical study was conducted on asymptomatic stool carriage of extended spectrum beta-lactamase (ESBL) producing Escherichia coli and Klebsiella pneumoniae among Finnish adults. All suspected ESBL producing bacterial isolates were first tested by phenotypic ESBL-confirmation methods, and then further analyzed with whole genome sequencing to identify the resistance genes. We found one study subject carrying a colistin resistant E. coli with a transferrable mcr-1 gene. This multi-drug resistant isolate, although initially suspected to be an ESBL producer, did not carry any ESBL genes, but was proven to carry several other resistance genes by using whole genome sequencing. Sequence type was ST93. The mcr-1 gene was connected to IncX4 plasmid which suggests that the colistin resistance gene locates in the respective plasmid. Here, we report the finding of a mcr-1 harboring human E. coli isolate from Finland. Clinical antimicrobial resistance (AMR) rates are low in Finland, and mobile colistin resistance has not been reported previously. This highlights the importance of AMR surveillance also in populations with low levels of resistance. © 2018 The Authors. APMIS published by John Wiley & Sons Ltd on behalf of Scandinavian Societies for Medical Microbiology and Pathology.

Resistance Pattern and Detection of Metallo-beta-lactamase Genes in Clinical Isolates of Pseudomonas aeruginosa in a Central Nigeria Tertiary Hospital.

PubMed

Zubair, K O; Iregbu, K C

2018-02-01

Acquired metallo-β-lactamases (MBLs) pose serious problem both in terms of treatment and infection control in the hospitals and report across the world showed an increase in their prevalence. However, there is a paucity of data from Africa, and their report is rare in Nigeria. This study aimed to determine the prevalence of acquired MBL-resistant genes in carbapenem-resistant Pseudomonas aeruginosa in Abuja, North Central Nigeria. Two hundred nonduplicate, consecutive isolates of P. aeruginosa from clinical samples submitted to the Medical Microbiology Laboratory of National Hospital, Abuja were screened for carbapenem resistance using imipenem and meropenem. Phenotypic detection of MBL-producing strains was determined using Total MBL confirm kits and E-test strips on isolates that were resistant to both Imipenem and meropenem. The MBL genes were detected using multiplex polymerase chain reaction, while the gene variant was determined by sequencing. Twenty-two MBL-producing strains were detected phenotypically, but only 5 harbored the blaVIM-1 gene, giving a prevalence of 2.5%. These 5 strains were resistant to all the antipseudomonal antibiotics tested except Aztreonam and Colistin. Other common MBL-genes were not detected. The prevalence of MBL-producing strains of P. aeruginosa which poses serious challenge for therapeutics and infection control is currently low in Abuja, North Central, Nigeria. Therefore, rational use of the carbapenems and other antipseudomonal antibiotics, regular surveillance and adequate infection control measures should be instituted to limit further spread.

Alleviation of hyperglycemia in diabetic rats by intraportal injection of insulin-producing cells generated from surgically resected human pancreatic tissue.

PubMed

Shyu, Jia-Fwu; Wang, Hwai-Shi; Shyr, Yi-Ming; Wang, Shin-E; Chen, Chia-Hsiang; Tan, Joo-Shin; Lin, Meng-Feng; Hsieh, Po-Shiuan; Sytwu, Huey-Kang; Chen, Tien-Hua

2011-03-01

Although islet transplantation holds promise for the treatment of diabetes, the scarcity of donor tissue remains a major drawback. The aim of this study is to generate insulin-producing cells from adult human pancreatic cells isolated from surgically resected pancreatic tissue. To isolate pancreatic endocrine precursor cells from 57 surgically resected pancreases, the cells were cultured and propagated in conditioned medium after which they were differentiated in Matrigel. The resultant cells were characterized using morphology, immunofluorescent studies, expression of differentiated pancreatic islet-specific genes using quantitative reverse transcription-PCR, and glucose-induced insulin secretion through analysis of C-peptide secretion. The relationships between propagation of insulin-producing cells and clinical variables of the donor were also analyzed. Finally, insulin-producing cell function was examined in streptozotocin-induced diabetic rats. Pancreatic endocrine precursor cells were successfully cultured; insulin-producing cells cultured from soft pancreas parenchyma had a significantly higher success rate. Morphological examination revealed islet-like cluster formation upon transfer to Matrigel. The presence of the neural stem cell marker nestin, duct cell marker cytokeratin 19, and endocrine cell markers C-peptide and pancreatic and duodenal homeobox 1, was also observed. In addition, glucose-stimulated C-peptide release was significantly increased in the insulin-producing cells. Furthermore, in diabetic rats, transplantation of insulin-producing cells reduced hyperglycemia. Isolated pancreatic endocrine precursor cells from surgically resected pancreatic tissue differentiated into insulin-producing cells and showed characteristics of functional endocrine cells. Thus, surgically resected pancreatic tissue may represent an alternative source of functional insulin-producing cells.

Current perspectives on the epidemiology and pathogenesis of clinically significant Vibrio spp.

PubMed Central

Janda, J M; Powers, C; Bryant, R G; Abbott, S L

1988-01-01

Recent taxonomic advances have now implicated several different Vibrio species as human pathogens. While the most common clinical presentation of Vibrio infection continues to be gastroenteritis, an increasing number of extraintestinal infections are being reported, particularly in immunocompromised individuals. Detection of Vibrio infections requires a good clinical history and the use of appropriate isolation and identification procedures by the laboratory to confirm illnesses attributed to Vibrio species. Except for Vibrio cholerae O1 and Vibrio parahaemolyticus, there is little direct evidence linking the production of a myriad of cell-associated or extracellular factors produced by each species with human disease and pathogenesis. Many questions regarding pathogenic Vibrio species remain unanswered, including their frequency and distribution in environmental specimens (water, shellfish), infective doses, virulence potential of individual isolates, and markers associated with such strains. Images PMID:3058295

Occurrence of Multidrug Resistant Extended Spectrum Beta-Lactamase-Producing Bacteria on Iceberg Lettuce Retailed for Human Consumption.

PubMed

Bhutani, Natasha; Muraleedharan, Chithra; Talreja, Deepa; Rana, Sonia Walia; Walia, Sandeep; Kumar, Ashok; Walia, Satish K

2015-01-01

Antibiotic resistance in bacteria is a global problem exacerbated by the dissemination of resistant bacteria via uncooked food, such as green leafy vegetables. New strains of bacteria are emerging on a daily basis with novel expanded antibiotic resistance profiles. In this pilot study, we examined the occurrence of antibiotic resistant bacteria against five classes of antibiotics on iceberg lettuce retailed in local convenience stores in Rochester, Michigan. In this study, 138 morphologically distinct bacterial colonies from 9 iceberg lettuce samples were randomly picked and tested for antibiotic resistance. Among these isolates, the vast majority (86%) demonstrated resistance to cefotaxime, and among the resistant bacteria, the majority showed multiple drug resistance, particularly against cefotaxime, chloramphenicol, and tetracycline. Three bacterial isolates (2.17%) out of 138 were extended spectrum beta-lactamase (ESBL) producers. Two ESBL producers (T1 and T5) were identified as Klebsiella pneumoniae, an opportunistic pathogen with transferable sulfhydryl variable- (SHV-) and TEM-type ESBLs, respectively. The DNA sequence analysis of the bla SHV detected in K. pneumoniae isolate T1 revealed 99% relatedness to bla SHV genes found in clinical isolates. This implies that iceberg lettuce is a potential reservoir of newly emerging and evolving antibiotic resistant bacteria and its consumption poses serious threat to human health.

Occurrence of Multidrug Resistant Extended Spectrum Beta-Lactamase-Producing Bacteria on Iceberg Lettuce Retailed for Human Consumption

PubMed Central

Talreja, Deepa; Rana, Sonia Walia; Walia, Sandeep; Walia, Satish K.

2015-01-01

Antibiotic resistance in bacteria is a global problem exacerbated by the dissemination of resistant bacteria via uncooked food, such as green leafy vegetables. New strains of bacteria are emerging on a daily basis with novel expanded antibiotic resistance profiles. In this pilot study, we examined the occurrence of antibiotic resistant bacteria against five classes of antibiotics on iceberg lettuce retailed in local convenience stores in Rochester, Michigan. In this study, 138 morphologically distinct bacterial colonies from 9 iceberg lettuce samples were randomly picked and tested for antibiotic resistance. Among these isolates, the vast majority (86%) demonstrated resistance to cefotaxime, and among the resistant bacteria, the majority showed multiple drug resistance, particularly against cefotaxime, chloramphenicol, and tetracycline. Three bacterial isolates (2.17%) out of 138 were extended spectrum beta-lactamase (ESBL) producers. Two ESBL producers (T1 and T5) were identified as Klebsiella pneumoniae, an opportunistic pathogen with transferable sulfhydryl variable- (SHV-) and TEM-type ESBLs, respectively. The DNA sequence analysis of the bla SHV detected in K. pneumoniae isolate T1 revealed 99% relatedness to bla SHV genes found in clinical isolates. This implies that iceberg lettuce is a potential reservoir of newly emerging and evolving antibiotic resistant bacteria and its consumption poses serious threat to human health. PMID:26064922

Isolation, Phylogenetic Analysis and Anti-infective Activity Screening of Marine Sponge-Associated Actinomycetes

PubMed Central

Abdelmohsen, Usama Ramadan; Pimentel-Elardo, Sheila M.; Hanora, Amro; Radwan, Mona; Abou-El-Ela, Soad H.; Ahmed, Safwat; Hentschel, Ute

2010-01-01

Terrestrial actinomycetes are noteworthy producers of a multitude of antibiotics, however the marine representatives are much less studied in this regard. In this study, 90 actinomycetes were isolated from 11 different species of marine sponges that had been collected from offshore Ras Mohamed (Egypt) and from Rovinj (Croatia). Phylogenetic characterization of the isolates based on 16S rRNA gene sequencing supported their assignment to 18 different actinomycete genera representing seven different suborders. Fourteen putatively novel species were identified based on sequence similarity values below 98.2% to other strains in the NCBI database. A putative new genus related to Rubrobacter was isolated on M1 agar that had been amended with sponge extract, thus highlighting the need for innovative cultivation protocols. Testing for anti-infective activities was performed against clinically relevant, Gram-positive (Enterococcus faecalis, Staphylococcus aureus) and Gram-negative (Escherichia coli, Pseudomonas aeruginosa) bacteria, fungi (Candida albicans) and human parasites (Leishmania major, Trypanosoma brucei). Bioactivities against these pathogens were documented for 10 actinomycete isolates. These results show a high diversity of actinomycetes associated with marine sponges as well as highlight their potential to produce anti-infective agents. PMID:20411105

Antimicrobial susceptibility and biochemical patterns of Neisseria gonorrhoeae strains in Vejle area, Denmark.

PubMed

Lis-Tønder, J; Cybulski, Z

2009-10-01

Strains of Neisseria gonorrhoeae resistant to penicillin and ciprofloxacin have been isolated worldwide. Increasing number of N. gonorrhoeae that lack the enzyme proline aminopeptidase (proA-negative N. gonorrhoeae) have been detected in many countries all over the world. This study aims to assess the resistance profiles of N. gonorrhoeae isolates sent to the Department of Clinical Microbiology in Vejle, Denmark, between 2003 and 2007, and to analyse their biochemical patterns. Sixty-two strains of N. gonorrhoeae were retrospectively analysed for their susceptibility to penicillin, ciprofloxacin and ceftriaxone. The identification of isolated strains was confirmed using both biochemical and immunological tests. Twenty-one (33.9%) N. gonorrhoeae isolates were resistant to penicillin and 30 (48.4%) were resistant to ciprofloxacin. All strains were susceptible to ceftriaxone. Fifty-six (90.3%) N. gonorrhoeae strains showed API NH biochemical code 10,010 (produced acid from glucose and proline aminopeptidase). Six strains showed code 10,000 that lack the enzyme proline aminopeptidase (proA-negative N. gonorrhoeae). Ceftriaxone should be used as the first-line treatment of gonorrhoea in Vejle community area, Denmark, both for infections with proA-producing and proA-negative N. gonorrhoeae isolates, which circulate in the region.

Factors responsible for subclinical mastitis in cows caused by Staphylococcus chromogenes and its susceptibility to antibiotics based on bap, fnbA, eno, mecA, tetK, and ermA genes.

PubMed

Bochniarz, M; Adaszek, Ł; Dzięgiel, B; Nowaczek, A; Wawron, W; Dąbrowski, R; Szczubiał, M; Winiarczyk, S

2016-12-01

The aim of this study was to recognize selected factors of virulence determining the adhesion of Staphylococcus chromogenes to cows' udder tissues in subclinical mastitis and to evaluate the susceptibility of this pathogen to antibiotics. The subjects of the study were 38 isolates of Staph. chromogenes from 335 samples of milk from cows with subclinical coagulase-negative staphylococci mastitis. Somatic cell count ranged between 216,000 and 568,000/mL of milk (average 356,000/mL of milk). We confirmed the ability to produce slime in 24 isolates (63.2%), and the ability to produce protease in 29 isolates (76.3%). In each slime-producing isolate, the bap gene was not found, and the fnbA and eno genes were not detected. In vitro tests showed that ceftiofur had the highest effectiveness against Staph. chromogenes (89.5% of susceptible isolates). Minimum inhibitory concentrations ranged from 0.06 to 2µg/mL for susceptible isolates. The minimum concentrations required to inhibit growth of 90 and 50% of the isolates for ceftiofur were at or below the cutoffs recommended by the Clinical and Laboratory Standards Institute (2 and 0.06µg/mL, respectively). A significant percentage of the isolates were susceptible to other β-lactam antibiotics: amoxicillin with clavulanic acid (84.2%) and ampicillin (81.6%). The lowest effectiveness among β-lactams was for penicillin (73.7% of susceptible isolates), and the minimum inhibitory concentration for penicillin ranged from <0.06 to 8µg/mL. None of the examined isolates had the mecA gene, but β-lactamase was detected in 4 isolates (10.5%). Erythromycin and oxytetracycline exhibited the lowest activity against Staph. chromogenes (71.1 and 63.2% of susceptible isolates, respectively). The genes tetK (6 isolates) and ermA (1 isolate) were also detected. Copyright © 2016 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Dissemination of Multidrug-Resistant, Class I and II Integrons and Molecular Typing of CTX-M-producing Klebsiella pneumoniae.

PubMed

Akya, Alisha; Elahi, Azam; Chegenelorestani, Roya; Rezaee, Mahya

2018-01-01

Klebsiella pneumoniae ( K. pneumoniae ) is an important opportunistic pathogen causes serious community and hospital-acquired infections, which is highly resistant to antibiotics. We aimed to determine the frequency of multidrug resistant (MDR) and molecular typing of clinical isolates of K. pneumoniae . One hundred isolates of K. pneumoniae were collected from clinical samples in three general hospitals in Kermanshah. The antimicrobial susceptibility and extended-spectrum beta-lactamases (ESBL) production of isolates were determined using disk diffusion and combined disk methods, respectively. The bla CTX-M gene, class I and II integrons were detected using polymerase chain reaction. The bla CTX-M positive isolates were selected for genotyping using pulsed-field gel electrophoresis (PFGE). MDR phenotype was observed in 56% of isolates. The 40% of isolates were ESBL positive and 35 isolates contained bla CTX-M . Class I and II of integrons were detected in 50 (89.2%) and 39 (69.6%) of MDR isolates, respectively. PFGE patterns of K. pneumoniae bla CTX-M positive isolates indicated 19 clusters (X 1-19 ) with different genotype patterns. The study findings highlight the concern of circulating MDR strains of K. pneumoniae with bla CTX-M and class I and II integrons in Kermanshah hospitals. The presence of integrons among isolates may facilitate the spread of new resistance genes in this bacterium. Therefore, surveillance for the spread of MDR strains of this bacterium is recommended in hospitals.

KPC and VIM producing Enterobacter cloacae strain from a hospital in northeastern Venezuela.

PubMed

Martínez, Dianny; Marcano, Daniel; Rodulfo, Hectorina; Salgado, Nurys; Cuaical, Nirvia; Rodriguez, Lucy; Caña, Luisa; Medina, Belkis; Guzman, Militza; De Donato, Marcos

2015-06-01

An 83-year-old male patient is admitted to the central hospital in Cumana, Venezuela with severe urinary infection, history of hospitalizaions and prolonged antimicrobial treatments. A strain of Enterobacter cloacae was isolated showing resistance to multiple types of antibiotics (only sensitive to gentamicin), with phenotype of serine- and metallo-carbapenemases. Both, bla(VIM-2) and bla(KPC) genes were detected in the isolate. This is the first report of an Enterobacteriaceae species producing both KPC carbapenemase and VIM metallo carbapenemase in Venezuela. This finding has a great clinical and epidemiological impact in the region, because of the feasibility of transferring these genes, through mobile elements to other strains of Enterobacter and to other infection-causing species of bacteria.

Slime production by clinical isolates of Blastoschizomyces capitatus from patients with hematological malignancies and catheter-related fungemia.

PubMed

D'Antonio, D; Parruti, G; Pontieri, E; Di Bonaventura, G; Manzoli, L; Sferra, R; Vetuschi, A; Piccolomini, R; Romano, F; Staniscia, T

2004-10-01

In order to expand the present knowledge of the pathogenic potential of Blastoschizomyces capitatus in central venous catheter (CVC)-related bloodstream infections, six strains of the organism recovered from three leukemic patients with CVC-related fungemia in different years were investigated. Isolates and control strains were tested for their genetic relatedness and for their ability to produce slime in glucose-containing solutions. DNA restriction enzyme analysis revealed that all strains of B. capitatus were identical, whereas slime production assays and examination of ex vivo material showed that they were able to produce large amounts of slime. Slime production may therefore play a relevant pathogenic role in cases of CVC-related fungemia caused by B. capitatus.

Comparison of CTX-M-14- and CTX-M-15-producing Escherichia coli and Klebsiella pneumoniae isolates from patients with bacteremia.

PubMed

Shin, Juyoun; Kim, Dae Hun; Ko, Kwan Soo

2011-07-01

Recently, CTX-M-15-producing Enterobacteriaceae has disseminated worldwide. To better understand the success of CTX-M-15-type extended-spectrum β-lactamase, we compared the CTX-M-15-producing Escherichia coli and Klebsiella pneumoniae isolates with CTX-M-14-producing E. coli and K. pneumoniae isolates that had been more prevalent before the recent increase of CTX-M-15 in Korea. Eighty-nine CTX-M-producing E. coli bloodstream infection isolates and 33 K. pneumoniae bloodstream infection isolates were collected in 2008 from nine hospitals in Korea. In vitro susceptibility testing and multilocus sequence typing were performed for all isolates. Phylogenetic groupings and distribution of virulence determinants and addiction systems were examined for only E. coli isolates. Among the 89 CTX-M-producing E. coli isolates, 54 isolates (60.7%) contained bla(CTx-M-15) and bla(CTx-M-14) was identified in 31 isolates (34.8%). Among 33 CTX-M-producing K. pneumoniae isolates, bla(CTx-M-14) and bla(CTx-M-15) were identified in 18 (54.5%) and 15 (45.5%) isolates, respectively. While CTX-M-14- and CTX-M-15-producing E. coli isolates displayed similar antimicrobial resistance rates, CTX-M-15-producing K. pneumoniae isolates showed significantly higher resistance rates of ciprofloxacin and piperacillin-tazobactam than CTX-M-14-producing isolates. ST131 and ST405 were the main clones in both CTX-M-14- and CTX-M-15-producing E. coli isolates. Although the frequency of virulence determinants was similar between two E. coli groups, ST131 and ST405 isolates producing CTX-M-15 showed higher frequency of determinants. In addition, CTX-M-15-producing E. coli isolates showed higher prevalence of addiction systems, particularly vagCD. ST405 showed the highest prevalence rates among main E. coli clones. In K. pneumoniae, ST15 and ST11, with high resistance rates, were the main clones of CTX-M-15-producing isolates, but no main clones was found among CTX-M-14-producing isolates because of extreme diversity. Rapid increase of CTX-M-15-producing E. coli isolates was due to certain clone with high frequency of virulence determinants and addiction systems. High antimicrobial resistance rates of CTX-M-15-producing K. pneumoniae isolates may contribute to their increase. Copyright © 2011 The British Infection Association. Published by Elsevier Ltd. All rights reserved.

A simple electroelution method for rapid protein purification: isolation and antibody production of alpha toxin from Clostridium septicum.

PubMed

Vázquez-Iglesias, Lorena; Estefanell-Ucha, Borja; Barcia-Castro, Leticia; Páez de la Cadena, María; Álvarez-Chaver, Paula; Ayude-Vázquez, Daniel; Rodríguez-Berrocal, Francisco Javier

2017-01-01

Clostridium septicum produces a number of diseases in human and farm animals which, in most of the cases, are fatal without clinical intervention. Alpha toxin is an important agent and the unique lethal virulent factor produced by Clostridium septicum. This toxin is haemolytic, highly lethal and necrotizing activities but is being used as an antigen to develop animal vaccines. The aim of this study was to isolate the alpha toxin of Clostridium septicum and produce highly specific antibodies against it. In this work, we have developed a simple and efficient method for alpha toxin purification, based on electroelution that can be used as a time-saving method for purifying proteins. This technique avoids contamination by other proteins that could appear during other protein purification techniques such chromatography. The highly purified toxin was used to produce polyclonal antibodies. The specificity of the antibodies was tested by western blot and these antibodies can be applied to the quantitative determination of alpha toxin by slot blot.

A simple electroelution method for rapid protein purification: isolation and antibody production of alpha toxin from Clostridium septicum

PubMed Central

Estefanell-Ucha, Borja; Barcia-Castro, Leticia; Páez de la Cadena, María; Álvarez-Chaver, Paula; Ayude-Vázquez, Daniel; Rodríguez-Berrocal, Francisco Javier

2017-01-01

Clostridium septicum produces a number of diseases in human and farm animals which, in most of the cases, are fatal without clinical intervention. Alpha toxin is an important agent and the unique lethal virulent factor produced by Clostridium septicum. This toxin is haemolytic, highly lethal and necrotizing activities but is being used as an antigen to develop animal vaccines. The aim of this study was to isolate the alpha toxin of Clostridium septicum and produce highly specific antibodies against it. In this work, we have developed a simple and efficient method for alpha toxin purification, based on electroelution that can be used as a time-saving method for purifying proteins. This technique avoids contamination by other proteins that could appear during other protein purification techniques such chromatography. The highly purified toxin was used to produce polyclonal antibodies. The specificity of the antibodies was tested by western blot and these antibodies can be applied to the quantitative determination of alpha toxin by slot blot. PMID:28652930

SME-type carbapenem-hydrolyzing class A beta-lactamases from geographically diverse Serratia marcescens strains.

PubMed

Queenan, A M; Torres-Viera, C; Gold, H S; Carmeli, Y; Eliopoulos, G M; Moellering, R C; Quinn, J P; Hindler, J; Medeiros, A A; Bush, K

2000-11-01

Three sets of carbapenem-resistant Serratia marcescens isolates have been identified in the United States: 1 isolate in Minnesota in 1985 (before approval of carbapenems for clinical use), 5 isolates in Los Angeles (University of California at Los Angeles [UCLA]) in 1992, and 19 isolates in Boston from 1994 to 1999. All isolates tested produced two beta-lactamases, an AmpC-type enzyme with pI values of 8.6 to 9.0 and one with a pI value of approximately 9.5. The enzyme with the higher pI in each strain hydrolyzed carbapenems and was not inhibited by EDTA, similar to the chromosomal class A SME-1 beta-lactamase isolated from the 1982 London strain S. marcescens S6. The genes encoding the carbapenem-hydrolyzing enzymes were cloned in Escherichia coli and sequenced. The enzyme from the Minnesota isolate had an amino acid sequence identical to that of SME-1. The isolates from Boston and UCLA produced SME-2, an enzyme with a single amino acid change relative to SME-1, a substitution from valine to glutamine at position 207. Purified SME enzymes from the U. S. isolates had beta-lactam hydrolysis profiles similar to that of the London SME-1 enzyme. Pulsed-field gel electrophoresis analysis revealed that the isolates showed some similarity but differed by at least three genetic events. In conclusion, a family of rare class A carbapenem-hydrolyzing beta-lactamases first described in London has now been identified in S. marcescens isolates across the United States.

SME-Type Carbapenem-Hydrolyzing Class A β-Lactamases from Geographically Diverse Serratia marcescens Strains

PubMed Central

Queenan, Anne Marie; Torres-Viera, Carlos; Gold, Howard S.; Carmeli, Yehuda; Eliopoulos, George M.; Moellering, Robert C.; Quinn, John P.; Hindler, Janet; Medeiros, Antone A.; Bush, Karen

2000-01-01

Three sets of carbapenem-resistant Serratia marcescens isolates have been identified in the United States: 1 isolate in Minnesota in 1985 (before approval of carbapenems for clinical use), 5 isolates in Los Angeles (University of California at Los Angeles [UCLA]) in 1992, and 19 isolates in Boston from 1994 to 1999. All isolates tested produced two β-lactamases, an AmpC-type enzyme with pI values of 8.6 to 9.0 and one with a pI value of approximately 9.5. The enzyme with the higher pI in each strain hydrolyzed carbapenems and was not inhibited by EDTA, similar to the chromosomal class A SME-1 β-lactamase isolated from the 1982 London strain S. marcescens S6. The genes encoding the carbapenem-hydrolyzing enzymes were cloned in Escherichia coli and sequenced. The enzyme from the Minnesota isolate had an amino acid sequence identical to that of SME-1. The isolates from Boston and UCLA produced SME-2, an enzyme with a single amino acid change relative to SME-1, a substitution from valine to glutamine at position 207. Purified SME enzymes from the U.S. isolates had β-lactam hydrolysis profiles similar to that of the London SME-1 enzyme. Pulsed-field gel electrophoresis analysis revealed that the isolates showed some similarity but differed by at least three genetic events. In conclusion, a family of rare class A carbapenem-hydrolyzing β-lactamases first described in London has now been identified in S. marcescens isolates across the United States. PMID:11036019

Degenerate primer MOB typing of multiresistant clinical isolates of E. coli uncovers new plasmid backbones.

PubMed

Garcillán-Barcia, M Pilar; Ruiz del Castillo, Belén; Alvarado, Andrés; de la Cruz, Fernando; Martínez-Martínez, Luis

2015-01-01

Degenerate Primer MOB Typing is a PCR-based protocol for the classification of γ-proteobacterial transmissible plasmids in five phylogenetic relaxase MOB families. It was applied to a multiresistant E. coli collection, previously characterized by PCR-based replicon-typing, in order to compare both methods. Plasmids from 32 clinical isolates of multiresistant E. coli (19 extended spectrum beta-lactamase producers and 13 non producers) and their transconjugants were analyzed. A total of 95 relaxases were detected, at least one per isolate, underscoring the high potential of these strains for antibiotic-resistance transmission. MOBP12 and MOBF12 plasmids were the most abundant. Most MOB subfamilies detected were present in both subsets of the collection, indicating a shared mobilome among multiresistant E. coli. The plasmid profile obtained by both methods was compared, which provided useful data upon which decisions related to the implementation of detection methods in the clinic could be based. The phylogenetic depth at which replicon and MOB-typing classify plasmids is different. While replicon-typing aims at plasmid replication regions with non-degenerate primers, MOB-typing classifies plasmids into relaxase subfamilies using degenerate primers. As a result, MOB-typing provides a deeper phylogenetic depth than replicon-typing and new plasmid groups are uncovered. Significantly, MOB typing identified 17 plasmids and an integrative and conjugative element, which were not detected by replicon-typing. Four of these backbones were different from previously reported elements. Copyright © 2014 Elsevier Inc. All rights reserved.

Synergetic Antimicrobial Effects of Mixtures of Ethiopian Honeys and Ginger Powder Extracts on Standard and Resistant Clinical Bacteria Isolates

PubMed Central

Ewnetu, Yalemwork; Lemma, Wossenseged; Birhane, Nega

2014-01-01

Purpose. To evaluate antimicrobial effects of mixtures of Ethiopian honeys and ginger rhizome powder extracts on Staphylococcus aureus (ATCC 25923), Escherichia coli (ATCC 25922), Staphylococcus aureus (MRSA), Escherichia coli (R), and Klebsiella pneumoniae (R). Methods. Agar diffusion and broth assays were performed to determine susceptibility of these standard and resistant clinical bacteria isolates using honey-ginger powder extract mixtures. Results. Honey-ginger powder extract mixtures produced the highest mean inhibition (25.62 mm ± 2.55) compared to the use of honeys (21.63 mm ± 3.30) or ginger extracts (19.23 mm ± 3.42) individually. The ranges of inhibitions produced by honey-ginger extract mixtures on susceptible test organisms (26–30 mm) and resistant strains (range: 19–27 mm) were higher compared to 7–22 mm and 0–14 mm by standard antibiotic discs. Minimum inhibitory concentrations (MIC) of mixture of honeys-ginger extracts were 6.25% (0.625 v/mL) on the susceptible bacteria compared to 75% for resistant clinical isolates. Minimum bactericidal concentration (MBC) of honey-ginger extracts was 12.5% (0.125 g/mL) for all the test organisms. Conclusion. The result of this study showed that honey-ginger powder extract mixtures have the potential to serve as cheap source of antibacterial agents especially for the drug resistant bacteria strains. PMID:24772182

Lithium ameliorates autistic-like behaviors induced by neonatal isolation in rats

PubMed Central

Wu, Xiaoyan; Bai, Yanrui; Tan, Tao; Li, Hongjie; Xia, Shuting; Chang, Xinxia; Zhou, Zikai; Zhou, Weihui; Li, Tingyu; Wang, Yu Tian; Dong, Zhifang

2014-01-01

Neonatal isolation is a widely accepted model to study the long-term behavioral changes produced by the early life events. However, it remains unknown whether neonatal isolation can induce autistic-like behaviors, and if so, whether pharmacological treatment can overcome it. Here, we reported that newborn rats subjected to individual isolations from their mother and nest for 1 h per day from postnatal days 1–9 displayed apparent autistic-like symptoms including social deficits, excessive repetitive self-grooming behavior, and increased anxiety- and depressive-like behaviors tested in young adult (postnatal days 42–56) compared to normal reared controls. Furthermore, these behavioral changes were accompanied by impaired adult hippocampal neurogenesis and reduced the ratio of excitatory/inhibitory synaptic transmissions, as reflected by an increase in spontaneous inhibitory postsynaptic current (sIPSC) and normal spontaneous excitatory postsynaptic current (sEPSC) in the hippocampal CA1 pyramidal neuron. More importantly, chronic administration of lithium, a clinically used mood stabilizer, completely overcame neonatal isolation-induced autistic-like behaviors, and restored adult hippocampal neurogenesis as well as the balance between excitatory and inhibitory activities to physiological levels. These findings indicate that neonatal isolation may produce autistic-like behaviors, and lithium may be a potential therapeutic agent against autism spectrum disorders (ASD) during development. PMID:25018711

[Selective-differential nutrient medium "Shewanella IRHLS agar" for isolation of Shewanella genus bacteria].

PubMed

Sivolodsky, E P

2015-01-01

Development of a selective-differential nutrient medium for isolation of Shewanella genus bacteria. 73 strains of Shewanella bacteria (S. algae--3, S. baltica--26, S. putrefaciens--44) and 80 strains of 22 other bacteria genera were used. Shewanella species were identified by methods and criteria proposed by Nozue H. et al., 1992; Khashe S. et al., 1998. Nutrient media "Shewanella IRHLS Agar" for shewanella isolation was developed. Medium selective factors: irgazan DP-300 (I). 0.14-0.2 g/l and rifampicin (R) 0.0005-0.001 g/l. Shevanella colonies were detected by the production of hydrogen sulfide (H), lipase presence (L), lack of sorbitol fermentation (S). The medium suppressed the growth of hydrogen sulfide producers (Salmonella, Proteus) and blocked hydrogen sulfide production by Citrobacter. Growth of Escherichia, Enterobacter, Klebsiella, Shigella, Staphylococcus, Bacillus was also suppressed, Analytical sensitivity of the medium was 1-2 CFU/ml for Shewanella and Stenotrophomonas, Aerombnas, Serratia genera bacteria. 72 strains of Shewanella were isolated from water of Neva river in this medium, 91.7 ± 3.2% of those produced H2S. 1 strain of S. algae was isolated from clinical material. The developed media allows to use it in a complex for Stenotrophomo- nas sp., Aeromonas sp., Serratia sp., Citrobactersp. and Shewanella bacteria isolation.

Phylogenetic study of metallo-β-lactamase producing multidrug resistant Pseudomonas aeruginosa isolates from burn patients.

PubMed

Jena, Jayanti; Debata, Nagen Kumar; Sahoo, Rajesh Kumar; Subudhi, Enketeswara

2015-12-01

The present study was carried out to understand the clonal relationship using enterobacteriaceae repetitive intergenic consensus polymerase chain reaction (ERIC-PCR) among metallo-β-lactamase (MBL) producing multidrug resistant Pseudomonas aeruginosa isolates from burn victims and their susceptibility to commonly used anti-pseudomonal agents. In the present study 94 non-duplicate P. aeruginosa strains from the wound samples of burn patients were included. Identification of the isolates was done by biochemical methods and antibiotic sensitivity was done by disc diffusion method following CLSI (Clinical Laboratory Standard Institute) guidelines. By using imipenem (IPM)-EDTA disk diffusion/double disc synergy method carbapenem resistant organisms were tested for MBL. To define the clonal relationship ERIC-PCR was used. Of the 94 isolates, 18 (19.14%) were found resistant to IPM and MBL production was shown 11 (11.70%) by the IPM-EDTA disc diffusion method. From dendrogram of the ERIC-PCR profile four major clusters were obtained (A, B, C and D). Cluster B contained the majority of the isolates (6 strains 1, 4, 8, 9, 10 and 11). This study using ERIC-PCR of randomly collected isolates exhibits high genetic diversity which rules out cross contamination frequency. Copyright © 2015 Elsevier Ltd and ISBI. All rights reserved.

Fermentation products in the cystic fibrosis airways induce aggregation and dormancy-associated expression profiles in a CF clinical isolate of Pseudomonas aeruginosa.

PubMed

Phan, Joann; Gallagher, Tara; Oliver, Andrew; England, Whitney E; Whiteson, Katrine

2018-05-01

Pseudomonas aeruginosa is a well-known dominant opportunistic pathogen in cystic fibrosis (CF) with a wide range of metabolic capacities. However, P. aeruginosa does not colonize the airways alone, and benefits from the metabolic products of neighboring cells-especially volatile molecules that can travel between different parts of the airways easily. Here, we present a study that investigates the metabolic, gene expression profiles and phenotypic responses of a P. aeruginosa clinical isolate to fermentation products lactic acid and 2,3-butanediol, metabolites that are produced by facultative anaerobic members of the CF polymicrobial community and potential biomarkers of disease progression. Although previous studies have successfully investigated the metabolic and transcriptional profiles of P. aeruginosa, most have used common lab reference strains that may differ in important ways from clinical isolates. Using transcriptomics and metabolomics with gas chromatography time of flight mass spectrometry, we observe that fermentation products induce pyocyanin production along with the expression of genes involved in P. aeruginosa amino acid utilization, dormancy and aggregative or biofilm modes of growth. These findings have important implications for how interactions within the diverse CF microbial community influence microbial physiology, with potential clinical consequences.

Plasmid-mediated AmpC beta-lactamase-producing Escherichia coli causing urinary tract infection in the Auckland community likely to be resistant to commonly prescribed antimicrobials.

PubMed

Drinkovic, Dragana; Morris, Arthur J; Dyet, Kristin; Bakker, Sarah; Heffernan, Helen

2015-03-13

To estimate the prevalence and characterise plasmid-mediated AmpC beta-lactamase (PMACBL)- producing Escherichia coli in the Auckland community. All cefoxitin non-susceptible (NS) E. coli identified at the two Auckland community laboratories between 1 January and 31 August 2011 were referred to ESR for boronic acid double-disc synergy testing, to detect the production of AmpC beta-lactamase, and polymerase chain reaction (PCR) to identify the presence of PMACBL genes. PMACBL-producing isolates were typed using pulsed-field gel electrophoresis (PFGE), and PCR was used to determine their phylogenetic group and to identify multilocus sequence type (ST)131. Antimicrobial susceptibility testing and detection of extended-spectrum beta-lactamases (ESBLs) were performed according to the Clinical and Laboratory Standards Institute recommendations. 101 (51%) and 74 (37%) of 200 non-duplicate cefoxitin-NS E. coli were PMACBL producers or assumed hyper-producers of chromosomal AmpC beta-lactamase, respectively. The prevalence of PMACBL-producing E. coli was 0.4%. PMACBL-producing E. coli were significantly less susceptible to norfloxacin, trimethoprim and nitrofurantoin than E. coli that produced neither a PMACBL nor an ESBL. Very few (4%) PMACBL-producing E. coli co-produced an ESBL. Most (88%) of the PMACBL-producing isolates had a CMY-2-like PMACBL. The PMACBL-producing E. coli isolates were diverse based on their PFGE profiles, 44% belonged to phylogenetic group D, and only four were ST131. 100 of the 101 PMACBL-producing E. coli were cultured from urine, and were causing urinary tract infection (UTI) in the majority of patients. The median patient age was 56 years and most (94%) of the patients were women. A greater proportion of patients with community-acquired UTI caused by PMACBL-producing E. coli received a beta-lactam antimicrobial than patients with community-acquired UTI caused by other non-AmpC, non-ESBL-producing E. coli. Thirty-six (43%) patients with community-acquired UTI due to PMACBL-producing E. coli were neither hospitalised nor had any antimicrobial treatment in the previous 6 months. The prevalence of PMACBL-producing E. coli was relatively low in the Auckland community, but has increased in recent years. Typing revealed that the majority of the PMACBL-producing E. coli in the Auckland region were genetically unrelated meaning that a point source or direct person to person transmission are not drivers of local community spread currently. The isolates were more resistant to non-beta-lactam antimicrobials than other non-AmpC, non-ESBL-producing E. coli, leaving few treatment options. The majority of the PMACBL-producing E. coli isolates seemed to be acquired in the community and were most frequently isolated from women with UTI. A large proportion of patients with community-acquired UTI had not been hospitalised nor had any antimicrobial treatment in the previous 6 months.

Molecular epidemiology over an 11-year period (2000 to 2010) of extended-spectrum β-lactamase-producing Escherichia coli causing bacteremia in a centralized Canadian region.

PubMed

Peirano, Gisele; van der Bij, Akke K; Gregson, Daniel B; Pitout, Johann D D

2012-02-01

A study was designed to assess the importance of sequence types among extended-spectrum β-lactamase (ESBL)-producing Escherichia coli isolates causing bacteremia over an 11-year period (2000 to 2010) in a centralized Canadian region. A total of 197 patients with incident infections were identified; the majority presented with community-onset urosepsis, with a significant increase in the prevalence of ESBL-producing E. coli during the later part of the study. The majority of E. coli isolates produced either CTX-M-15 or CTX-M-14. We identified 7 different major sequence types among 91% of isolates (i.e., the ST10 clonal complex, ST38, ST131, ST315, ST393, ST405, and ST648) and provided insight into their clinical and molecular characteristics. ST38 was the most antimicrobial-susceptible sequence type and predominated during 2000 to 2004 but disappeared after 2008. ST131 was the most antimicrobial-resistant sequence type, and the influx of a single pulsotype of this sequence type was responsible for the significant increase of ESBL-producing E. coli strains since 2007. During 2010, 49/63 (78%) of the ESBL-producing E. coli isolates belonged to ST131, and this sequence type had established itself as a major drug-resistant pathogen in Calgary, Alberta, Canada, posing an important new public health threat within our region. We urgently need well-designed epidemiological and molecular studies to understand the dynamics of transmission, risk factors, and reservoirs for E. coli ST131. This will provide insight into the emergence and spread of this multiresistant sequence type.

In vitro evaluation of antibiotic synergy for polymyxin B-resistant carbapenemase-producing Klebsiella pneumoniae.

PubMed

Elemam, Azza; Rahimian, Joseph; Doymaz, Mehmet

2010-10-01

Since carbapenemase-producing Klebsiella pneumoniae strains were first reported in North Carolina, these highly resistant organisms have been isolated with increasing frequency, especially in the New York City area. Polymyxin B is one of the few antimicrobials that retain reliable activity against these organisms. However, polymyxin B MICs are elevated against K. pneumoniae isolates with increasing frequency, leaving clinicians with few therapeutic options. We investigated several antimicrobial agents for potential synergy with polymyxin B against 12 clinical strains of carbapenemase-producing K. pneumoniae. A broth microdilution assay using a 96-well plate was developed in which graded dilutions of polymyxin B and the study drug were incubated with resistant isolates in a checkerboard pattern. Polymyxin B was studied in combination with cefazolin, ceftriaxone, cefepime, imipenem, gentamicin, tigecycline, doxycycline, and rifampin. All K. pneumoniae strains tested positive for K. pneumoniae carbapenemase (KPC) genes by real-time PCR and had elevated polymyxin B MIC values ranging from 16 to 128 μg/ml. Synergy was observed with the combination of polymyxin B and rifampin as well as with polymyxin B and doxycycline, resulting in at least a 4-fold decrease in the polymyxin B MIC. For both combinations, this effect occurred at physiologically achievable concentrations. Less pronounced synergy was noted with tigecycline and polymyxin B. No synergy was observed at physiologic concentrations with the other antimicrobials studied. These results suggest that rifampin, doxycycline, and tigecycline may be useful additions to polymyxin B in the treatment of infections caused by highly resistant carbapenemase-producing K. pneumoniae. Further studies are warranted to determine if these in vitro findings translate into clinical efficacy.

Spread of ISCR1 Elements Containing blaDHA-1 and Multiple Antimicrobial Resistance Genes Leading to Increase of Flomoxef Resistance in Extended-Spectrum-β-Lactamase-Producing Klebsiella pneumoniae▿

PubMed Central

Lee, Chen-Hsiang; Liu, Jien-Wei; Li, Chia-Chin; Chien, Chun-Chih; Tang, Ya-Fen; Su, Lin-Hui

2011-01-01

Increasing resistance to quinolones, aminoglycosides, and/or cephamycins in extended-spectrum-β-lactamase (ESBL)-producing Enterobacteriaceae exacerbates the already limited antibiotic treatment options for infections due to these microbes. In this study, the presence of resistance determinants for these antimicrobial agents was examined by PCR among ESBL-producing Klebsiella pneumoniae (ESBL-KP) isolates that caused bacteremia. Pulsed-field gel electrophoresis was used to differentiate the clonal relationship among the isolates studied. Transferability and the location of the resistance genes were analyzed by conjugation experiments, followed by DNA-DNA hybridization. Among the 94 ESBL-KP isolates studied, 20 isolates of flomoxef-resistant ESBL-KP were identified. They all carried a DHA-1 gene and were genetically diverse. CTX-M genes were found in 18 of the isolates. Among these DHA-1/CTX-M-producing K. pneumoniae isolates, ISCR1 was detected in 13 (72%) isolates, qnr genes (1 qnrA and 17 qnrB genes) were detected in 18 (100%), aac(6′)-Ib-cr was detected in 11 (61%), and 16S rRNA methylase (all armA genes) was detected in 14 (78%). Four transconjugants were available for further analysis, and qnrB4, aac(6′)-Ib-cr, armA, and blaDHA-1 were all identified on these self-transferable blaCTX-M-carrying plasmids. The genetic environments of ISCR1 associated with armA, blaDHA-1, and qnrB4 genes in the four transconjugants were identical. Replicon-type analysis revealed a FIIA plasmid among the four self-transferable plasmids, although the other three were nontypeable. The cotransfer of multiple resistance genes with the ISCR1 element-carrying plasmids has a clinical impact and warrants close monitoring and further study. PMID:21746945

Clinical and molecular epidemiology of veterinary blastomycosis in Wisconsin.

PubMed

Anderson, Jennifer L; Sloss, Brian L; Meece, Jennifer K

2013-04-22

Several studies have shown that Blastomyces dermatitidis, the etiologic agent of blastomycosis, is a genetically diverse pathogen. Blastomycosis is a significant health issue in humans and other mammals. Veterinary and human isolates matched with epidemiological case data from the same geographic area and time period were used to determine: (i) if differences in genetic diversity and structure exist between clinical veterinary and human isolates of B. dermatitidis and (ii) if comparable epidemiologic features differ among veterinary and human blastomycosis cases. Genetic typing of 301 clinical B. dermatitidis isolates produced 196 haplotypes (59 unique to veterinary isolates, 134 unique to human isolates, and 3 shared between canine and human isolates). Private allelic richness was higher in veterinary (median 2.27) compared to human isolates (median 1.14) (p = 0.005). Concordant with previous studies, two distinct genetic groups were identified among all isolates. Genetic group assignment was different between human and veterinary isolates (p < 0.001), with more veterinary isolates assigned to Group 2. The mean age of dogs diagnosed with blastomycosis was 6 years. Thirty cases were in male dogs (52%) and 24 were females (41%). The breed of dog was able to be retrieved in 38 of 58 cases with 19 (50%) being sporting breeds. Three of four felines infected with blastomycosis were domestic shorthair males between ages 6-12, and presented with disseminated disease. The other was a lynx with pulmonary disease. The equine isolate was from an 11-year-old male Halflinger with disseminated disease. Disseminated disease was reported more often in veterinary (62%) than human cases (19%) (p < 0.001). Isolates from all hosts clustered largely into previously identified genetic groups, with 3 haplotypes being shared between human and canine isolates confirming that B. dermatitidis isolates capable of infecting both species occur in nature. Allelic diversity measures trended higher in veterinary samples, with a higher number of total alleles and private alleles. Veterinary isolates of B. dermatitidis contributed a substantial amount of diversity to the overall population genetic structure demonstrating the importance of including veterinary isolates in genetic studies of evolution and virulence in this organism.

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

Characterization of Clostridium perfringens in the feces of adult horses and foals with acute enterocolitis

PubMed Central

Gohari, Iman Mehdizadeh; Arroyo, Luis; MacInnes, Janet I.; Timoney, John F.; Parreira, Valeria R.; Prescott, John F.

2014-01-01

Up to 60% of cases of equine colitis have no known cause. To improve understanding of the causes of acute colitis in horses, we hypothesized that Clostridium perfringens producing enterotoxin (CPE) and/or beta2 toxin (CPB2) are common and important causes of severe colitis in horses and/or that C. perfringens producing an as-yet-undescribed cytotoxin may also cause colitis in horses. Fecal samples from 55 horses (43 adults, 12 foals) with clinical evidence of colitis were evaluated by culture for the presence of Clostridium difficile, C. perfringens, and Salmonella. Feces were also examined by enzyme-linked immunosorbent assay (ELISA) for C. difficile A/B toxins and C. perfringens alpha toxin (CPA), beta2 toxin (CPB2), and enterotoxin (CPE). Five C. perfringens isolates per sample were genotyped for the following genes: cpa, cpb, cpb2 consensus, cpb2 atypical, cpe (enterotoxin), etx (epsilon toxin), itx (iota toxin), netB (necrotic enteritis toxin B), and tpeL (large C. perfringens cytotoxin). The supernatants of these isolates were also evaluated for toxicity for an equine cell line. All fecal samples were negative for Salmonella. Clostridium perfringens and C. difficile were isolated from 40% and 5.4% of samples, respectively. All fecal samples were negative for CPE. Clostridium perfringens CPA and CPB2 toxins were detected in 14.5% and 7.2% of fecal samples, respectively, all of which were culture-positive for C. perfringens. No isolates were cpe, etx, netB, or tpeL gene-positive. Atypical cpb2 and consensus cpb2 genes were identified in 15 (13.6%) and 4 (3.6%) of 110 isolates, respectively. All equine C. perfringens isolates showed far milder cytotoxicity effects than a CPB-producing positive control, although cpb2-positive isolates were slightly but significantly more cytotoxic than negative isolates. Based on this studied population, we were unable to confirm our hypothesis that CPE and CPB2-producing C. perfringens are common in horses with colitis in Ontario and we failed to identify cytotoxic activity in vitro in the type A isolates recovered. PMID:24396174

Characterization of Clostridium perfringens in the feces of adult horses and foals with acute enterocolitis.

PubMed

Gohari, Iman Mehdizadeh; Arroyo, Luis; Macinnes, Janet I; Timoney, John F; Parreira, Valeria R; Prescott, John F

2014-01-01

Up to 60% of cases of equine colitis have no known cause. To improve understanding of the causes of acute colitis in horses, we hypothesized that Clostridium perfringens producing enterotoxin (CPE) and/or beta2 toxin (CPB2) are common and important causes of severe colitis in horses and/or that C. perfringens producing an as-yet-undescribed cytotoxin may also cause colitis in horses. Fecal samples from 55 horses (43 adults, 12 foals) with clinical evidence of colitis were evaluated by culture for the presence of Clostridium difficile, C. perfringens, and Salmonella. Feces were also examined by enzyme-linked immunosorbent assay (ELISA) for C. difficile A/B toxins and C. perfringens alpha toxin (CPA), beta2 toxin (CPB2), and enterotoxin (CPE). Five C. perfringens isolates per sample were genotyped for the following genes: cpa, cpb, cpb2 consensus, cpb2 atypical, cpe (enterotoxin), etx (epsilon toxin), itx (iota toxin), netB (necrotic enteritis toxin B), and tpeL (large C. perfringens cytotoxin). The supernatants of these isolates were also evaluated for toxicity for an equine cell line. All fecal samples were negative for Salmonella. Clostridium perfringens and C. difficile were isolated from 40% and 5.4% of samples, respectively. All fecal samples were negative for CPE. Clostridium perfringens CPA and CPB2 toxins were detected in 14.5% and 7.2% of fecal samples, respectively, all of which were culture-positive for C. perfringens. No isolates were cpe, etx, netB, or tpeL gene-positive. Atypical cpb2 and consensus cpb2 genes were identified in 15 (13.6%) and 4 (3.6%) of 110 isolates, respectively. All equine C. perfringens isolates showed far milder cytotoxicity effects than a CPB-producing positive control, although cpb2-positive isolates were slightly but significantly more cytotoxic than negative isolates. Based on this studied population, we were unable to confirm our hypothesis that CPE and CPB2-producing C. perfringens are common in horses with colitis in Ontario and we failed to identify cytotoxic activity in vitro in the type A isolates recovered.

Carbapenem-resistant Serratia marcescens isolates producing Bush group 2f beta-lactamase (SME-1) in the United States: results from the MYSTIC Programme.

PubMed

Gales, A C; Biedenbach, D J; Winokur, P; Hacek, D M; Pfaller, M A; Jones, R N

2001-02-01

Two carbapenem (imipenem, meropenem)-resistant Serratia marcescens strains were isolated in the United States (Chicago, IL) through the 1999 MYSTIC (Meropenem Yearly Susceptibility Test Information Collection) Programme. The S. marcescens antimicrobial susceptible patterns were: susceptible to ceftriaxone, ceftazidime, and cefepime (MICs, < or = 0.25 microg/ml), and resistance to the carbapenems (imipenem and meropenem; MIC, > 32 microg/ml) and aztreonam (MIC, > = 16 microg/ml). Each S. marcescens isolate shared an identical epidemiologic type (ribotype and PFGE) and the outer membrane protein profile was also identical to those of the wild type susceptible strains from the same medical center. The PCR utilizing bla(sme-1) primers amplified a gene product that was identified as consistent with SME-1 after DNA sequencing. Imipenem and meropenem resistance due to production of carbapenem-hydrolyzing enzymes among clinical isolates is still very rare, but microbiology laboratories should be aware of these chromosomally encoded enzymes among class C beta-lactamases producing enteric bacilli such as S. marcescens and Enterobacter cloacae.

High prevalence of Escherichia coli sequence type 131 among antimicrobial-resistant E. coli isolates from geriatric patients.

PubMed

Ho, Pak-Leung; Chu, Yuki Pui-Shan; Lo, Wai-U; Chow, Kin-Hung; Law, Pierra Y; Tse, Cindy Wing-Sze; Ng, Tak-Keung; Cheng, Vincent Chi-Chung; Que, Tak-Lun

2015-03-01

Previous work on the subclones within Escherichia coli ST131 predominantly involved isolates from Western countries. This study assessed the prevalence and antimicrobial resistance attributed to this clonal group. A total of 340 consecutive, non-duplicated urinary E. coli isolates originating from four clinical laboratories in Hong Kong in 2013 were tested. ST131 prevalence among the total isolates was 18.5 % (63/340) and was higher among inpatient isolates (23.0 %) than outpatient isolates (11.8 %, P<0.001), and higher among isolates from patients aged ≥65 years than from patients aged 18-50 years and 51-64 years (25.4 vs 3.4 and 4.0 %, respectively, P<0.001). Of the 63 ST131 isolates, 43 (68.3 %) isolates belonged to the H30 subclone, whereas the remaining isolates belonged to H41 (n = 17), H54 (n = 2) and H22 (n = 1). All H30 isolates were ciprofloxacin-resistant, of which 18.6 % (8/43) belonged to the H30-Rx subclone. Twenty-six (41.3 %) ST131 isolates were ESBL-producers, of which 19 had blaCTX-M-14 (12 non-H30-Rx, two H30-Rx and five H41), six had blaCTX-M-15 (five non-H30-Rx and one H30-Rx) and one was blaCTX-M-negative (H30). In conclusion, ST131 accounts for a large share of the antimicrobial-resistant E. coli isolates from geriatric patients. Unlike previous reports, ESBL-producing ST131 strains mainly belonged to non-H30-Rx rather than the H30-Rx subclone, with blaCTX-M-14 as the dominant enzyme type. © 2015 The Authors.

Antimicrobial susceptibility and emerging resistance determinants (blaCTX-M, rmtB, fosA3) in clinical isolates from urinary tract infections in the Bolivian Chaco.

PubMed

Bartoloni, Alessandro; Sennati, Samanta; Di Maggio, Tiziana; Mantella, Antonia; Riccobono, Eleonora; Strohmeyer, Marianne; Revollo, Carmen; Villagran, Ana Liz; Pallecchi, Lucia; Rossolini, Gian Maria

2016-02-01

Bolivia is among the lowest-resourced South American countries, with very few data available on antibiotic resistance in bacterial pathogens. The phenotypic and molecular characterization of bacterial isolates responsible for urinary tract infections (UTIs) in the Bolivian Chaco are reported here. All clinical isolates from UTIs collected in the Hospital Basico Villa Montes between June 2010 and January 2014 were analyzed (N=213). Characterization included susceptibility testing, extended-spectrum beta-lactamase (ESBL) detection, identification of relevant resistance determinants (e.g., CTX-M-type ESBLs, 16S rRNA methyltransferases, glutathione S-transferases), and genotyping of CTX-M producers. Very high resistance rates were observed. Overall, the lowest susceptibility was observed for trimethoprim-sulphamethoxazole, tetracycline, nalidixic acid, amoxicillin-clavulanic acid, ciprofloxacin, and gentamicin. Of E. coli and K. pneumoniae, 11.6% were ESBL producers. Resistance to nitrofurantoin, amikacin, and fosfomycin remained low, and susceptibility to carbapenems was fully preserved. CTX-M-15 was the dominant CTX-M variant. Four E. coli ST131 (two being H30-Rx) were identified. Of note, isolates harbouring rmtB and fosA3 were detected. Bolivia is not an exception to the very high resistance burden affecting many South American countries. Optimization of alternative approaches to monitor local antibiotic resistance trends in resource-limited settings is strongly encouraged to support the implementation of effective empiric treatment guidelines. Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.

Identification of Clinical Coryneform Bacterial Isolates: Comparison of Biochemical Methods and Sequence Analysis of 16S rRNA and rpoB Genes▿

PubMed Central

Adderson, Elisabeth E.; Boudreaux, Jan W.; Cummings, Jessica R.; Pounds, Stanley; Wilson, Deborah A.; Procop, Gary W.; Hayden, Randall T.

2008-01-01

We compared the relative levels of effectiveness of three commercial identification kits and three nucleic acid amplification tests for the identification of coryneform bacteria by testing 50 diverse isolates, including 12 well-characterized control strains and 38 organisms obtained from pediatric oncology patients at our institution. Between 33.3 and 75.0% of control strains were correctly identified to the species level by phenotypic systems or nucleic acid amplification assays. The most sensitive tests were the API Coryne system and amplification and sequencing of the 16S rRNA gene using primers optimized for coryneform bacteria, which correctly identified 9 of 12 control isolates to the species level, and all strains with a high-confidence call were correctly identified. Organisms not correctly identified were species not included in the test kit databases or not producing a pattern of reactions included in kit databases or which could not be differentiated among several genospecies based on reaction patterns. Nucleic acid amplification assays had limited abilities to identify some bacteria to the species level, and comparison of sequence homologies was complicated by the inclusion of allele sequences obtained from uncultivated and uncharacterized strains in databases. The utility of rpoB genotyping was limited by the small number of representative gene sequences that are currently available for comparison. The correlation between identifications produced by different classification systems was poor, particularly for clinical isolates. PMID:18160450

Molecular and epidemiological characterization of carbapenemase-producing Enterobacteriaceae in Norway, 2007 to 2014.

PubMed

Samuelsen, Ørjan; Overballe-Petersen, Søren; Bjørnholt, Jørgen Vildershøj; Brisse, Sylvain; Doumith, Michel; Woodford, Neil; Hopkins, Katie L; Aasnæs, Bettina; Haldorsen, Bjørg; Sundsfjord, Arnfinn

2017-01-01

The prevalence of carbapenemase-producing Enterobacteriaceae (CPE) is increasing worldwide. Here we present associated patient data and molecular, epidemiological and phenotypic characteristics of all CPE isolates in Norway from 2007 to 2014 confirmed at the Norwegian National Advisory Unit on Detection of Antimicrobial Resistance. All confirmed CPE isolates were characterized pheno- and genotypically, including by whole genome sequencing (WGS). Patient data were reviewed retrospectively. In total 59 CPE isolates were identified from 53 patients. Urine was the dominant clinical sample source (37%) and only 15% of the isolates were obtained from faecal screening. The majority of cases (62%) were directly associated with travel or hospitalization abroad, but both intra-hospital transmission and one inter-hospital outbreak were observed. The number of CPE cases/year was low (2-14 cases/year), but an increasing trend was observed. Klebsiella spp. (n = 38) and E. coli (n = 14) were the dominant species and blaKPC (n = 20), blaNDM (n = 19), blaOXA-48-like (n = 12) and blaVIM (n = 7) were the dominant carbapenemase gene families. The CPE isolates were genetically diverse except for K. pneumoniae where clonal group 258 associated with blaKPC dominated. All isolates were multidrug-resistant and a significant proportion (21%) were resistant to colistin. Interestingly, all blaOXA-48-like, and a large proportion of blaNDM-positive Klebsiella spp. (89%) and E. coli (83%) isolates were susceptible in vitro to mecillinam. Thus, mecillinam could have a role in the treatment of uncomplicated urinary tract infections caused by OXA-48- or NDM-producing E. coli or K. pneumoniae. In conclusion, the impact of CPE in Norway is still limited and mainly associated with travel abroad, reflected in the diversity of clones and carbapenemase genes.

Strain-specific transmission in an outbreak of ESBL-producing Enterobacteriaceae in the hemato-oncology care unit: a cohort study.

PubMed

Uemura, Makiko; Imataki, Osamu; Uchida, Shumpei; Nakayama-Imaohji, Haruyuki; Ohue, Yukiko; Matsuka, Harumi; Mori, Hatsune; Dobashi, Hiroaki; Kuwahara, Tomomi; Kadowaki, Norimitsu

2017-01-05

Extended-spectrum β-lactamase (ESBL)-producing bacteria are resistant to several types of antibiotics excluding carbapenems. A transmissibility of ESBL-producing Enterobacteriaceae would be depending on each bacterial property, however, that has not been elucidated in clinical setting. In this study, we attempted to identify the source of an outbreak of ESBL-producing bacteria in a medical oncology and immunology care unit. An ESBL-producing Enterobacteriaceae (ESBL-E) outbreak observed between July 2012 and August 2012 in Kagawa University Hospital was surveyed using various molecular microbiology techniques. We used Pulsed-field gel electrophoresis (PFGE), PCR-based ESBL gene typing, and direct sequence of ESBL gene as molecular microbiology typing method to distinguish each strain. The typical prevalence of ESBL-E isolation in the unit was 7.0 per month (1.7 per week). The prevalence of ESBL-E isolation during the target research period was 20.0 per month (5.0 per week). In total, 19 isolates (11 K. pneumoniae and 8 E. coli) were obtained from clinical samples, including four control strains (two each of both bacteria), that were physically different from those obtained from other inpatient units in our hospital. Pulsed-field gel electrophoresis (PFGE) for K. pneumoniae (digested by XbaI) produced similar patterns excluding one control strain. PCR classification of the ESBL gene for K. pneumoniae revealed that all strains other than the control strain carried SHV and CTX-M-9. This result was reconfirmed by direct DNA sequencing. Although the outbreak of K. pneumoniae was considered to be "clonal," PFGE and PCR classification of the ESBL genes for E. coli uncovered at least six different "non-clonal" strains possessing individual ESBL gene patterns. According to the result of an antibiogram, the pattern of antimicrobial susceptibility was more variable for K. pneumoniae than for E. coli. Typing by PFGE and ESBL gene PCR analysis is practical for discriminating various organisms. In our cohort, two outbreaks were concomitantly spread with different transmission strategies, namely clonal and non-clonal, in the same unit. This might represent clinical evidence that transmissibility differs according to the type of strain. We speculated that patient-to-patient transmission of ESBL-E occurred according to the properties of each individual strain.

Evaluation of Antioxidant Activity and Growth Control Properties of Nonoscale Structure Produced from Aloe vera var. littoralis Extract on Clinical Isolates of Salmonella

PubMed Central

Hosseiny, Hossein

2017-01-01

The aim of the study was to examine antibacterial properties of microemulsion structure produced from Aloe vera var. littoralis extract as a new tool of nanoscale drug-like materials. Aloe vera var. littoralis (A. littoralis) extract was prepared by distillation method. A nonocarrier structure in the microemulsion system was prepared from the extract. Serial concentrations were prepared from 8 mg/mL extract and the nonocarrier containing 0.1 mg/mL pure extract and were evaluated by a disk diffusion method for 35 Salmonella clinical isolates. Minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) were determined by microbroth dilution assay using MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) method by an enzyme-linked immunosorbent assay(ELISA) Microplate Reader apparatus. Antioxidant activity of the extract was determined by measuring the ferric reducing ability of plasma (FRAP) assay. From 35 clinical isolates of Salmonella, 17 isolates—including resistant isolates of S.E.1103 and S.E.49—had a zone of inhibition (ZI) of 7 to 32 mm in 0.007 mg/mL of the extract. S.E.76 isolate exposed to 30 µg/mL ceftazidime disk had a ZI of 12 mm but had 10 mm in 7µg/mL of A. littoralis extract. The inhibitory effect of a nanocarrier at a concentration of 25 µg/mL by 20 mm ZI was comparable by the ceftazidime (30 µg/mL) effect. MIC50 was 0.25 mg/mL and MBC50 was 0.5 mg/mL by MTT method for the extract. It was shown that A.littoralis extract had antioxidant activity of 31.67 µM/mg that could be increased based on concentration. It was concluded that the nanocarrier had a significant effect on the studied isolates in comparison with ordinary antibiotics and had potential for use as a natural antioxidant and antimicrobial material in complementary medicine. PMID:28758958

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.

Characterization of Mycobacterium bohemicum Isolated from Human, Veterinary, and Environmental Sources

PubMed Central

Torkko, Pirjo; Suomalainen, Sini; Iivanainen, Eila; Suutari, Merja; Paulin, Lars; Rudbäck, Eeva; Tortoli, Enrico; Vincent, Véronique; Mattila, Rauni; Katila, Marja-Leena

2001-01-01

Chemotaxonomic and genetic properties were determined for 14 mycobacterial isolates identified as members of a newly described species Mycobacterium bohemicum. The isolates recovered from clinical, veterinary, and environmental sources were compared for lipid composition, biochemical test results, and sequencing of the 16S ribosomal DNA (rDNA) and the 16S-23S rDNA internal transcribed spacer (ITS) regions. The isolates had a lipid composition that was different from those of other known species. Though the isolates formed a distinct entity, some variations were detected in the features analyzed. Combined results of the phenotypic and genotypic analyses were used to group the isolates into three clusters. The major cluster (cluster A), very homogenous in all respects, comprised the M. bohemicum type strain, nine clinical and veterinary isolates, and two of the five environmental isolates. Three other environmental isolates displayed an insertion of 14 nucleotides in the ITS region; they also differed from cluster A in fatty alcohol composition and produced a positive result in the Tween 80 hydrolysis test. Among these three, two isolates were identical (cluster B), but one isolate (cluster C) had a unique high-performance liquid chromatography profile, and its gas liquid chromatography profile lacked 2-octadecanol, which was present in all other isolates analyzed. Thus, sequence variation in the 16S-23S ITS region was associated with interesting variations in lipid composition. Two of the isolates analyzed were regarded as potential inducers of human or veterinary infections. Each of the environmental isolates, all of which were unrelated to the cases presented, was cultured from the water of a different stream. Hence, natural waters are potential reservoirs of M. bohemicum. PMID:11136772

Presence of quinolone resistance to qnrB1 genes and blaOXA-48 carbapenemase in clinical isolates of Klebsiella pneumoniae in Spain.

PubMed

Rodríguez Martínez, J M; Díaz-de Alba, P; Lopez-Cerero; Ruiz-Carrascoso, G; Gomez-Gil, R; Pascual, A

2014-01-01

A study is presented on the presence of quinolone resistance qnrB1 genes in clinical isolates belonging to the largest series of infections caused by OXA-48-producing Klebsiella pneumoniae in a single-centre outbreak in Spain. Evidence is also provided, according to in vitro results, that there is a possibility of co-transfer of plasmid harbouring blaOXA-48 with an other plasmid harbouring qnrB1 in presence of low antibiotic concentrations of fluoroquinolones, showing the risk of multi-resistance screening. Copyright © 2013 Elsevier España, S.L.U. y Sociedad Española de Enfermedades Infecciosas y Microbiología Clínica. All rights reserved.

A matched-pair comparison of inlay and onlay trochlear designs for patellofemoral arthroplasty: no differences in clinical outcome but less progression of osteoarthritis with inlay designs.

PubMed

Feucht, Matthias J; Cotic, Matthias; Beitzel, Knut; Baldini, Julia F; Meidinger, Gebhart; Schöttle, Philip B; Imhoff, Andreas B

2017-09-01

To compare clinical and radiographic results after isolated patellofemoral arthroplasty (PFA) using either a second-generation inlay or onlay trochlear design. The hypothesis was that an inlay design will produce better clinical results and less progression of tibiofemoral osteoarthritis (OA) compared to an onlay design. Fifteen consecutive patients undergoing isolated PFA with an onlay design trochlear component (Journey™ PFJ, Smith & Nephew) were matched with 15 patients after isolated PFA with an inlay design trochlear component (HemiCAP ® Wave, Arthrosurface). Matching criteria were age, gender, body mass index, and follow-up period. An independent observer evaluated patients prospectively, whereas data were compared retrospectively. Clinical outcome was assessed using WOMAC, Lysholm score, and pain VAS. Kellgren-Lawrence grading was used to assess progression of tibiofemoral OA. Conversion to total knee arthroplasty was necessary in one patient within each group, leaving 14 patients per group for final evaluation. The mean follow-up was 26 months in the inlay group and 25 months in the onlay group (n.s.). Both groups displayed significant improvements of all clinical scores (p < 0.05). No significant differences were found between the two groups with regard to the clinical outcome and reoperation rate. No significant progression of tibiofemoral OA was observed in the inlay group, whereas 53 % of the onlay group showed progression of medial and/or lateral tibiofemoral OA (p = 0.009). Isolated PFA using either a second-generation inlay or onlay trochlear component significantly improves functional outcome scores and pain. The theoretical advantages of an inlay design did not result in better clinical outcome scores; however, progression of tibiofemoral OA was significantly less common in patients with an inlay trochlear component. This implant design may therefore improve long-term results and survival rates after isolated PFA. III.

Production of Pseudomonas aeruginosa Intercellular Small Signaling Molecules in Human Burn Wounds

PubMed Central

Que, Yok-Ai; Hazan, Ronen; Ryan, Colleen M.; Milot, Sylvain; Lépine, François; Lydon, Martha; Rahme, Laurence G.

2011-01-01

Pseudomonas aeruginosa has developed a complex cell-to-cell communication system that relies on low-molecular weight excreted molecules to control the production of its virulence factors. We previously characterized the transcriptional regulator MvfR, that controls a major network of acute virulence functions in P. aeruginosa through the control of its ligands, the 4-hydroxy-2-alkylquinolines (HAQs)—4-hydroxy-2-heptylquinoline (HHQ) and 3,4-dihydroxy-2-heptylquinoline (PQS). Though HHQ and PQS are produced in infected animals, their ratios differ from those in bacterial cultures. Because these molecules are critical for the potency of activation of acute virulence functions, here we investigated whether they are also produced during human P. aeruginosa acute wound infection and whether their ratio is similar to that observed in P. aeruginosa-infected mice. We found that a clinically relevant P. aeruginosa isolate produced detectable levels of HAQs with ratios of HHQ and PQS that were similar to those produced in burned and infected animals, and not resembling ratios in bacterial cultures. These molecules could be isolated from wound tissue as well as from drainage liquid. These results demonstrate for the first time that HAQs can be isolated and quantified from acute human wound infection sites and validate the relevance of previous studies conducted in mammalian models of infection. PMID:23533774

First Report of Group CTX-M-9 Extended Spectrum Beta-Lactamases in Escherichia coli Isolates from Pediatric Patients in Mexico

PubMed Central

Merida-Vieyra, Jocelin; De Colsa, Agustin; Calderon Castañeda, Yair; Arzate Barbosa, Patricia; Aquino Andrade, Alejandra

2016-01-01

The aim of this study was to identify the presence of group CTX-M-9 extended spectrum beta-lactamases (ESBL) in clinical Escherichia coli isolates from pediatric patients. A total of 404 non-repeated positive ESBL E. coli isolates were collected from documented clinical infections in pediatric patients over a 2-year period. The identification and susceptibility profiles were determined using an automated system. Isolates that suggested ESBL production based on their resistance profiles to third and fourth generation cephalosporin and monobactam were selected. ESBL production was phenotypically confirmed using a diffusion method with cefotaxime and ceftazidime discs alone and in combination with clavulanic acid. blaESBL gene identification was performed through PCR amplification and sequencing. Pulsed Field Gel Electrophoresis (PFGE) and Multilocus Sequence Typing (MLST) were performed to establish the clonal relationships of the E. coli isolates. CTX-M-9-type ESBLs were detected in 2.5% of the isolates. The subtypes corresponded to blaCTX-M-14 (n = 4) and blaCTX-M-27 (n = 6). Additionally, coexistence with other beta-lactamases was observed. A clonal relationship was established in three isolates; the rest were classified as non-related. We found seven different sequence type (ST) in CTX-M-9- producing E. coli isolates. ST38 was the most frequent. This study is the first report in Mexico to document the presence of group CTX-M-9 ESBLs in E. coli isolates from pediatric patients. PMID:27992527

Direct Isolation of Candida spp. from Blood Cultures on the Chromogenic Medium CHROMagar Candida

PubMed Central

Horvath, Lynn L.; Hospenthal, Duane R.; Murray, Clinton K.; Dooley, David P.

2003-01-01

CHROMagar Candida is a selective and differential chromogenic medium that has been shown to be useful for identification of Candida albicans, Candida krusei, Candida tropicalis, and perhaps Candida glabrata. Colony morphology and color have been well defined when CHROMagar Candida has been used to isolate yeast directly from clinical specimens, including stool, urine, respiratory, vaginal, oropharyngeal, and esophageal sources. Direct isolation of yeast on CHROMagar Candida from blood cultures has not been evaluated. We evaluated whether the color and colony characteristics produced by Candida spp. on CHROMagar Candida were altered when yeasts were isolated directly from blood cultures. Fifty clinical isolates of Candida were inoculated into aerobic and anaerobic blood culture bottles and incubated at 35°C in an automated blood culture system. When growth was detected, an aliquot was removed and plated onto CHROMagar Candida. As a control, CHROMagar Candida plates were inoculated with the same isolate of yeast grown on Sabouraud dextrose agar simultaneously. No significant difference was detected in color or colony morphology between the blood and control isolates in any of the tested organisms. All C. albicans (n = 12), C. tropicalis (n = 12), C. glabrata (n = 9), and C. krusei (n = 5) isolates exhibited the expected species-specific colony characteristics and color, whether isolated directly from blood or from control cultures. CHROMagar Candida can be reliably used for direct isolation of yeast from blood cultures. Direct isolation could allow mycology laboratories to more rapidly identify Candida spp., enable clinicians to more quickly make antifungal agent selections, and potentially decrease patient morbidity and mortality. PMID:12791890

Emergence of carbapenem-resistant Enterobacteriaceae isolated from patients in a university hospital in Saudi Arabia. Epidemiology, clinical profiles and outcomes.

PubMed

Alotaibi, Fawzia E; Bukhari, Elham E; Al-Mohizea, Maha M; Hafiz, Taghreed; Essa, Eman B; AlTokhais, Yasmeen I

Carbapenemase-producing Enterobacteriaceae have been steadily spreading worldwide during the last decade. Nine patients were identified prospectively and were followed during their hospitalization course to identify the epidemiology, clinical profiles and outcomes. These patients had one or more cultures positive for a CRE isolate, contributing to a total of eleven positive cultures from various sites without including duplicates of isolates obtained from the same site. Isolates from these patients included five Klebseilla pneumoniae, three Escherichia coli, and one Enterobacter aerogenes. Five isolates were grown from blood cultures, three from wound cultures, one from urine cultures, one from respiratory cultures and one from an abscess collection. Five survived the hospital course. The other five patients died due to severe sepsis, septic shock or multi-organ failure. Of the nine isolates of CRE identified for which molecular analysis were available, four K. pneumonia were confirmed as blaNDM and one as OXA-48. For the purpose of controlling the spread of CRE in our institution, we recommend considering active surveillance cultures and screening patients transferred from other hospitals or coming from highly endemic settings at admission for these organisms. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

Development of a Phenotypic Method for Detection of Fecal Carriage of OXA-48-Producing Enterobacteriaceae after Incidental Detection from Clinical Specimen▿

PubMed Central

Ruppé, Etienne; Armand-Lefèvre, Laurence; Lolom, Isabelle; El Mniai, Assiya; Muller-Serieys, Claudette; Ruimy, Raymond; Woerther, Paul-Louis; Bilariki, Kalliopi; Marre, Michel; Massin, Philippe; Andremont, Antoine; Lucet, Jean-Christophe

2011-01-01

We report incidental isolation of an OXA-48-producing Escherichia coli strain in urine of a 62-year-old woman recently returning from a 2-month vacation in Morocco. Commercially available extended-spectrum beta-lactamase (ESBL)-targeting medium failed to detect it in the patient's stools, although a locally developed and easy-to-implement method using ertapenem-supplemented brain heart infusion (BHI) broths could. PMID:21562103

Evaluation of Carbapenem Resistance Mechanisms and Its Association with Pseudomonas aeruginosa Infections in the Northwest of Iran.

PubMed

Akhi, Mohammad Taghi; Khalili, Younes; Ghotaslou, Reza; Yousefi, Saber; Kafil, Hossein Samadi; Naghili, Behrooz; Sheikhalizadeh, Vajihe

2018-03-01

The aims of this study were to determine carbapenem resistance mechanisms, molecular epidemiological relationship, clinical impact, and patient outcome of carbapenem-resistant Pseudomonas aeruginosa (CRPA) infections. A total of 42 nonduplicated CRPA were recovered from Urmia, Iran. Antimicrobial susceptibility tests were carried out using phenotypic methods. The carbapenem resistance mechanisms such as carbapenemase genes, efflux pump hyperexpression, AmpC overproduction, and OprD gene downregulation were determined by phenotypic and molecular methods. Eighteen metallo-β-lactamase (MBL) producer isolates were found to be sensitive to amikacin. Among the CRPA, 52.3%, 26.1%, 26.1%, and 59.5% were identified as carbapenemase, efflux pump hyperexpression, AmpC overproduction, and reduced expression OprD gene, respectively. Random Amplified Polymorphic DNA analysis yielded 25 distinct profiles. Most MBL-positive isolates were recovered from patients hospitalized in urology and internal wards with urinary tract infections. Most of the strains showed downregulation of porin. The clonal distribution of the strains was related to carbapenem resistance mechanisms (most of MBL producers belong to the same clones) and the same hospital wards where the isolates were collected. The study demonstrates that the main risk factor of MBL-related infections was hospitalization in non-intensive wards. Amikacin was considered a very efficient antibiotic to treatment of MBL-producing CRPA isolates. Our results showed that OprD downregulation and IMP-type MBL are the main carbapenem resistance mechanisms in CRPA isolates from northwest of Iran.

Characterization of OXA-48-like-producing Enterobacteriaceae isolated from river water in Algeria.

PubMed

Tafoukt, Rima; Touati, Abdelaziz; Leangapichart, Thongpan; Bakour, Sofiane; Rolain, Jean-Marc

2017-09-01

The spread of carbapenemase-producing Enterobacteriaceae (CPE) is a significant problem for healthcare worldwide. The prevalence of carbapenem-resistant Enterobacteriaceae (CPE) in water environments in Algeria are unknown. The aim of this study was to screen for the presence of CPE isolates in the Soummam River in Bejaia, Algeria. Isolates of Enterobacteriaceae recovered from twelve samples of river water and showing reduced susceptibility to carbapenems were included in this study. The isolates were identified by matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS). Isolates were subjected to antimicrobial susceptibility testing and the modified Carba NP test. Carbapenemase and extended-spectrum β-lactamase (ESBL) determinants were studied by PCR amplification and sequencing. The clonal relatedness between isolates was studied by Multilocus Sequence Typing (MLST) method. A total of 20 carbapenem-resistant Enterobacteriaceae strains were included in this study, identified as Escherichia coli (n = 12), Klebsiella pneumoniae (n = 3), Raoultella ornithinolytica (n = 3), Citrobacter freundii (n = 1) and Citrobacter braakii (n = 1). Carbapenemase genes identified in this study included bla OXA-48 , observed in 17 isolates (9 E. coli, 3 K. pneumoniae, 3 R. ornithinolytica, 1 C. freundii and 1 C. braakii), and bla OXA-244 , a variant of bla OXA-48 , was found in three E. coli isolates. MLST showed that 12 E. coli strains belonged to six different sequence types (ST559, ST38, ST212, ST3541, 1972 and ST2142), and we identified three different STs in K. pneumoniae isolates, including ST133, ST2055, and a new sequence type: ST2192. This study showed the presence of OXA-48-like-producing Enterobacteriaceae in water environments and highlighted the potential role of aquatic environments as reservoirs of clinically relevant antimicrobial-resistant bacteria, with the potential to spread throughout the community. Copyright © 2017 Elsevier Ltd. All rights reserved.

Molecular characteristic of mcr-1 producing Escherichia coli in a Chinese university hospital.

PubMed

He, Qing-Wen; Xu, Xiao-Hong; Lan, Fang-Jun; Zhao, Zhi-Chang; Wu, Zhi-Yun; Cao, Ying-Ping; Li, Bin

2017-04-19

Colistin has been considered as a last-line treatment option in severe infections caused by multidrug-resistant (MDR) gram-negative pathogens. However, the emergence of the mobile colistin resistance gene (mcr-1) has challenged this viewpoint. The aim of this study is to explore the prevalence of mcr-1 in Escherichia coli (E. coli) in a Chinese teaching hospital, and investigate their molecular characteristics. A total of 700 E. coli isolates were used to screen mcr-1 by PCR and sequencing in a Chinese university hospital from August 2014 to August 2015. Susceptibility test of mcr-1-producing isolates was determined by Vitek -2 Compact system. 26 virulence factors (VFs), phylogenetic groups, Multi-locus sequence typing (MLST), and DNA Fingerprinting (ERIC-PCR) of strains were investigated by PCR. Four (0.6%) mcr-1 producing E. coli isolates were found in this study. The results of antibiotic susceptibility test showed that all four isolates were resistant to colistin, ciprofloxacin, levofloxacin, cefazolin, and trimethoprim/sulfamethoxazole, and were susceptible to amikacin, ertapenem and imipenem. In addition, all 4 isolates exhibited high-level resistance to aztreonam, cefotaxime and gentamicin. The numbers of VFs contained in mcr-1 positive isolates were no more than 4 in our study. MLST result demonstrated that these isolates were assigned to two sequence types: ST156 and ST167. The result of phylogenetic analysis showed that four mcr-1-positive isolates belong to two phylogenetic groups: A and B1 group. ERIC-PCR showed that four mcr-1 positive strains were categorized into three different genotypes. Our study demonstrated a low prevalence of mcr-1 in E. coli clinical isolates in a Chinese teaching hospital, and we have gained insights into the molecular characteristics of these mcr-1-positive strains. Increasing the surveillance of these infections, as well as taking effective infection control measures are urgently needed to take to control the transmission of mcr-1 gene.

Wild Birds, Frequent Carriers of Extended-Spectrum β-Lactamase (ESBL) Producing Escherichia coli of CTX-M and SHV-12 Types.

PubMed

Alcalá, Leticia; Alonso, Carla Andrea; Simón, Carmen; González-Esteban, Chabier; Orós, Jesús; Rezusta, Antonio; Ortega, Carmelo; Torres, Carmen

2016-11-01

To get a better insight into the role of birds as reservoirs of extended-spectrum β-lactamase (ESBL) and plasmidic AmpC β-lactamase (pAmpC) Escherichia coli producers, 100 fecal samples belonging to 15 different wild avian species from Northern Spain were analyzed. Cefotaxime-resistant (CTX R ) E. coli isolates were identified in 16 of the 100 tested birds, which corresponded to 9 animal species (Gyps fulvus-griffon vulture, Larus michahellis-yellow-legged gull, Milvus migrans-black kite, Milvus milvus-red kite, Ciconia ciconia-white stork, Sturnus unicolor-spotless starling, Aquila chrysaetos-golden eagle, Cuculus canorus-common cuckoo, Tyto alba-barn owl). Fifteen isolates harbored ESBL or pAmpC-encoding genes (number of isolates): bla SHV-12 (9), bla CTX-M-1 (3), bla CTX-M-14 (2), and bla CMY-2 (1). The last CTX R isolate presented a -42-point-mutation in the chromosomal ampC promoter. Eleven out of 15 ESBL/pAmpC E. coli isolates were multiresistant (most common resistance phenotype: β-lactams-quinolones-tetracycline-sulfamethoxazole/trimethoprim). A plasmid-mediated quinolone resistance determinant (qnrS1) was identified in one E. coli from a barn owl. High genetic diversity was observed among ESBL/pAmpC E. coli isolates, with 12 different sequence types (STs), including several strains of STs frequently detected among human clinical isolates (ST38/D, ST131/B2, ST155/B1, ST10/A). The ST131 isolate belonged to the emergent ciprofloxacin-resistant H30R subclone. This study reveals a high percentage of bird as carriers of ESBL/pAmpC E. coli isolates in Spain, highlighting the elevated rate among storks, kites, and vultures. Wild birds can contribute to the global spread of ESBL/pAmpC-producing E. coli in natural ecosystems.

Communication between hospitals and isolated aboriginal community health clinics.

PubMed

Mackenzie, G; Currie, B J

1999-04-01

This study described the communication dynamics, identified problems and recommended changes to improve patient follow-up and communication between Royal Darwin Hospital (RDH) and isolated Aboriginal community health clinics (CHC) in the Northern Territory (NT). In 1995, staff interviews were conducted and an audit of isolated Aboriginal patients' RDH discharge summaries (DS). Eighteen per cent of RDH DSs never arrived in CHCs. DSs were often prepared late and more likely to be in CHC records if written on time and if the referral source was specified. Interviews revealed discontent between CHCs and RDH regarding: communication, DS documentation, the supply of discharge medication, as well as different hospital and community perceptions of Aboriginies' reliability to carry a DS and CHC desire for patients to be given DSs at discharge. Aboriginal patients should be given a DS at discharge and resident medical officers should be educated as to the function and importance of the DS. In 18 months following this study, RDH appointed unit-based Aboriginal health workers and a policy was produced for written communication between hospital and CHCs, as well as a discharge planning manual for Aboriginal communities. Projects investigating communication between hospitals and isolated Aboriginal clinics and patient follow-up may result in significant policy changes concerning these processes.

Mycological survey of Korean cereals and production of mycotoxins by Fusarium isolates.

PubMed Central

Lee, U S; Jang, H S; Tanaka, T; Toyasaki, N; Sugiura, Y; Oh, Y J; Cho, C M; Ueno, Y

1986-01-01

The fungal species isolated from Korean cereals (barley, polished barley, wheat, rye, and malt) were Alternaria spp., Aspergillus spp., Chaetomium spp., Drechslera spp., Epicoccum sp., Fusarium spp., and Penicillium spp., etc. The number of Fusarium strains isolated was 36, and their ability to produce Fusarium mycotoxins on rice was tested. Nivalenol (NIV) was produced by Fusarium graminearum (7 of 9 isolates), Fusarium oxysporum (3 of 10 isolates), and Fusarium spp. (7 of 15 isolates). Of 15 isolates of Fusarium spp., 6 formed deoxynivalenol (DON). Fusarenon-X and 3-acetyl-DON were produced by most NIV- and DON-forming isolates, respectively. Zearalenone was produced by 3 isolates of F. graminearum, 1 isolate of Fusarium equiseti, and 11 isolates of Fusarium spp. T-2 toxin was not produced by any Fusarium isolates. The highest concentrations of mycotoxins produced by Fusarium isolates were 77.4 (NIV), 5.3 (DON), 138.3 (fusarenon-X), 40.6 (3-acetyl-DON), and 23.2 (zearalenone) micrograms/g. PMID:2947538

Mycological survey of Korean cereals and production of mycotoxins by Fusarium isolates.

PubMed

Lee, U S; Jang, H S; Tanaka, T; Toyasaki, N; Sugiura, Y; Oh, Y J; Cho, C M; Ueno, Y

1986-12-01

The fungal species isolated from Korean cereals (barley, polished barley, wheat, rye, and malt) were Alternaria spp., Aspergillus spp., Chaetomium spp., Drechslera spp., Epicoccum sp., Fusarium spp., and Penicillium spp., etc. The number of Fusarium strains isolated was 36, and their ability to produce Fusarium mycotoxins on rice was tested. Nivalenol (NIV) was produced by Fusarium graminearum (7 of 9 isolates), Fusarium oxysporum (3 of 10 isolates), and Fusarium spp. (7 of 15 isolates). Of 15 isolates of Fusarium spp., 6 formed deoxynivalenol (DON). Fusarenon-X and 3-acetyl-DON were produced by most NIV- and DON-forming isolates, respectively. Zearalenone was produced by 3 isolates of F. graminearum, 1 isolate of Fusarium equiseti, and 11 isolates of Fusarium spp. T-2 toxin was not produced by any Fusarium isolates. The highest concentrations of mycotoxins produced by Fusarium isolates were 77.4 (NIV), 5.3 (DON), 138.3 (fusarenon-X), 40.6 (3-acetyl-DON), and 23.2 (zearalenone) micrograms/g.

E. coli encoding blaNDM-5 associated with community-acquired UTI cases with unusual MIC creep like phenomenon against Imipenem.

PubMed

Gajamer, Varsha Rani; Bhattacharjee, Amitabha; Paul, Deepjyoti; Deshamukhya, Chandrayee; Singh, Ashish Kr; Pradhan, Nilu; Tiwari, Hare Krishna

2018-05-15

Carbapenemase-producing Escherichia coli are of major clinical concern. The present study aimed to identify NDM-5 producing E.coli associated with community-acquired urinary tract infection, co-harboring ESBL genes and a pattern of imipenem MIC creep. A total of 973 urine samples were collected from females of age between 18-49 diagnosed with UTI. Isolates were identified by standard microbiological procedures. Presence of bla NDM and ESBL genes was determined by PCR assay and sequencing.PCR based replicon typing was performed. Plasmid stability of all bla producers and their transformants study were analyzed by serial passages and MIC creep phenomenon was analysed by studying revertants. Among 34 bla NDM-5 positive E.coli isolates, 21 isolates co harbored bla CTX-M-15 , followed by multiple combinations of genes. The study revealed diverse types of plasmids type viz; HI1, I1, FIA+FIB, FIA and Y. The strains showed progressive plasmid loss after 31 passages.Twenty-eight isolates mostly had MIC value of 0.5μg/ml and 1μg/ml against imipenem, ertapenem and meropenem. However, on studying the MIC creep activity; the MIC was found elevated from 0.5ug/ml to 64μg/ml and from 1μg/ml to 128μg/ml. While analyzing the revertants, MIC of most of the NDM positive isolates was reduced to 16μg/ml after the 30th serial passages. This study observed a unique phenotype of NDM producers which is not been reported earlier. In the current study, the observed phenomenon poses a global threat as these pathogens may evade phenotypic screening by routine laboratories. Copyright © 2018. Published by Elsevier Ltd.

An Improved Extended-Spectrum-β-Lactamase Detection Test Utilizing Aztreonam plus Clavulanate.

PubMed

Thomson, Gina K; Ayaz, Maaz; Lutes, Kelli; Thomson, Kenneth S

2018-01-01

Clinical laboratories test for extended-spectrum β-lactamases (ESBLs) for epidemiological and infection control purposes and also for the potential of cephalosporins to cause therapeutic failures. Testing can be problematic, because the CLSI does not recommend the testing of all producers of ESBLs and also falsely negative results may occur with isolates that coproduce AmpC. Boronic acid-supplemented tests can enhance ESBL detection in AmpC producers. Because aztreonam inhibits AmpCs, a study was designed to compare ESBL detection by the CLSI disk test (CLSI), a boronic acid-supplemented CLSI disk test (CLSI plus BA), and an aztreonam plus clavulanate disk test (ATM plus CA). The study tested 100 well-characterized Enterobacteriaceae , Acinetobacter baumannii , and Pseudomonas aeruginosa isolates. Seventy produced TEM, SHV, or CTX-M ESBLs, with 15 coproducing an AmpC and 11 coproducing a metallo-β-lactamase. Thirty ESBL-negative isolates were also tested. Tests were inoculated by CLSI methodology and interpreted as positive if an inhibitor caused a zone diameter increase of ≥5 mm. The percentages of ESBL producers detected were as follows: ATM plus CA, 95.7%; CLSI plus BA, 88.6%; and CLSI, 78.6%. When AmpC was coproduced, the sensitivities of the tests were as follows: ATM plus CA, 100%; CLSI plus BA, 93.3%; and CLSI, 60%. ATM plus CA also detected an ESBL in 90.1% of isolates that coproduced a metallo-β-lactamase. Falsely positive tests occurred only with the CLSI and CLSI plus BA tests. Overall, the ATM plus CA test detected ESBLs more accurately than the CLSI and CLSI plus BA tests, especially with isolates coproducing an AmpC or metallo-β-lactamase. Copyright © 2017 American Society for Microbiology.

Outbreak of IMI-1 Carbapenemase-producing colistin-resistant Enterobacter cloacae on the French island of Mayotte (Indian Ocean).

PubMed

Miltgen, Guillaume; Bonnin, Rémy A; Avril, Catherine; Benoit-Cattin, Thierry; Martak, Daniel; Leclaire, Alexandre; Traversier, Nicolas; Roquebert, Bénédicte; Jaffar-Bandjee, Marie-Christine; Lugagne, Nathalie; Filleul, Laurent; Subiros, Marion; de Montera, Anne-Marie; Cholley, Pascal; Thouverez, Michelle; Dortet, Laurent; Bertrand, Xavier; Naas, Thierry; Hocquet, Didier; Belmonte, Olivier

2018-05-25

The spread of carbapenemase-producing Enterobacteriaceae (CPE) in the Southwest Indian Ocean islands is poorly known. We describe here an outbreak of colistin-resistant Enterobacter cloacae harboring bla IMI-1 in the French overseas department of Mayotte. Between October 2015 and January 2017, all isolates of imipenem-non-susceptible E. cloacae at the Mayotte Medical Center and University Hospital of Reunion Island were screened for carbapenemase production. Positive isolates were typed by pulsed-field gel electrophoresis and WGS-based MLST and all β-lactamase genes identified by PCR and sequencing. The resistance profiles were determined by agar diffusion and E-tests. Genetic support of the bla IMI-1 gene was determined by WGS. We detected 18 isolates of E.cloacae harboring the bla IMI-1 gene in 17 patients from Mayotte. PFGE analysis showed 16 of the 18 strains to be clonally-related and belonging to sequence type ST820. Based on clinical data, this outbreak most likely had a community origin. The bla IMI-1 gene in the 18 isolates was carried by a new variant of an integrative mobile element involving the Xer recombinases, called EcloIMEX-8. The mcr-1 to mcr-5 genes were absent from the collection. These isolates belong to E. cloacae cluster XI, known to be colistin heteroresistant. We report here the first outbreak of IMI-1-producing Enterobacteriaceae. IMI-1 producers may be under-detected in microbiology laboratories because of their unusual antibiotic resistance profile (resistant to imipenem, but with intermediate resistance to ertapenem and susceptible to extended-spectrum cephalosporins) and the absence of bla IMI-1 in the panel of genes targeted by molecular diagnostic kits. Copyright © 2018. Published by Elsevier B.V.

Wedge-shaped microfluidic chip for circulating tumor cells isolation and its clinical significance in gastric cancer.

PubMed

Yang, Chaogang; Zhang, Nangang; Wang, Shuyi; Shi, Dongdong; Zhang, Chunxiao; Liu, Kan; Xiong, Bin

2018-05-23

Circulating tumor cells (CTCs) have great potential in both basic research and clinical application for the managements of cancer. However, the complicated fabrication processes and expensive materials of the existing CTCs isolation devices, to a large extent, limit their clinical translation and CTCs' clinical value. Therefore, it remains to be urgently needed to develop a new platform for achieving CTCs detection with low-cost, mass-producible but high performance. In the present study, we introduced a novel wedge-shaped microfluidic chip (named CTC-ΔChip) fabricated by two pieces of glass through wet etching and thermal bonding technique for CTCs isolation, which achieved CTCs enrichment by different size without cell surface expression markers and CTCs identification with three-color immunocytochemistry method (CK+/CD45-/Nucleus+). We validated the feasibility of CTC-ΔChip for detecting CTCs from different types of solid tumor. Furthermore, we applied the newly-developed platform to investigate the clinical significance of CTCs in gastric cancer (GC). Based on "label-free" characteristic, the capture efficiency of CTC-ΔChip can be as high as 93.7 ± 3.2% in DMEM and 91.0 ± 3.0% in whole blood sample under optimized conditions. Clinically, CTC-ΔChip exhibited the feasibility of detecting CTCs from different types of solid tumor, and it identified 7.30 ± 7.29 CTCs from 2 mL peripheral blood with a positive rate of 75% (30/40) in GC patients. Interestingly, we found that GC CTCs count was significantly correlated with multiple systemic inflammation indexes, including the lymphocyte count, platelet count, the level of neutrophil to lymphocyte ratio and platelet to lymphocyte ratio. In addition, we also found that both the positivity rate and CTCs count were significantly associated with multiple clinicopathology parameters. Our novel CTC-ΔChip shows high performance for detecting CTCs from less volume of blood samples of cancer patients and important clinical significance in GC. Owing to the advantages of low-cost and mass-producible, CTC-ΔChip holds great potential of clinical application for cancer therapeutic guidance and prognostic monitoring in the future.

Extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli and Acinetobacter baumannii among horses entering a veterinary teaching hospital: The contemporary "Trojan Horse".

PubMed

Walther, Birgit; Klein, Katja-Sophia; Barton, Ann-Kristin; Semmler, Torsten; Huber, Charlotte; Wolf, Silver Anthony; Tedin, Karsten; Merle, Roswitha; Mitrach, Franziska; Guenther, Sebastian; Lübke-Becker, Antina; Gehlen, Heidrun

2018-01-01

Pathogens frequently associated with multi-drug resistant (MDR) phenotypes, including extended-spectrum beta-lactamase (ESBL)-producing Enterobacteriaceae (ESBL-E) and Acinetobacter baumannii isolated from horses admitted to horse clinics, pose a risk for animal patients and personnel in horse clinics. To estimate current rates of colonization, a total of 341 equine patients were screened for carriage of zoonotic indicator pathogens at hospital admission. Horses showing clinical signs associated with colic (n = 233) or open wounds (n = 108) were selected for microbiological examination of nostril swabs, faecal samples and wound swabs taken from the open wound group. The results showed alarming carriage rates of Gram-negative MDR pathogens in equine patients: 10.7% (34 of 318) of validated faecal specimens were positive for ESBL-E (94%: ESBL-producing Escherichia coli), with recorded rates of 10.5% for the colic and 11% for the open wound group. 92.7% of the ESBL-producing E. coli were phenotypically resistant to three or more classes of antimicrobials. A. baumannii was rarely detected (0.9%), and all faecal samples investigated were negative for Salmonella, both directly and after two enrichment steps. Screening results for the equine nostril swabs showed detection rates for ESBL-E of 3.4% among colic patients and 0.9% in the open wound group, with an average rate of 2.6% (9/340) for both indications. For all 41 ESBL-producing E. coli isolated, a broad heterogeneity was revealed using pulsed-field gel electrophoresis (PFGE) patterns and whole genome sequencing (WGS) -analysis. However, a predominance of sequence type complex (STC)10 and STC1250 was observed, including several novel STs. The most common genes associated with ESBL-production were identified as blaCTX-M-1 (31/41; 75.6%) and blaSHV-12 (24.4%). The results of this study reveal a disturbingly large fraction of multi-drug resistant and ESBL-producing E. coli among equine patients, posing a clear threat to established hygiene management systems and work-place safety of veterinary staff in horse clinics.

Antibiotic susceptibility of enterococci isolated from traditional fermented meat products.

PubMed

Barbosa, J; Ferreira, V; Teixeira, P

2009-08-01

Antibiotic susceptibility was evaluated for 182 Enterococcus spp. isolated from Alheira, Chouriça de Vinhais and Salpicão de Vinhais, fermented meat products produced in the North of Portugal. Previously, a choice was made from a group of 1060 isolates, using phenotypic and genotypic tests. From these, 76 were previously identified as Enterococcus faecalis, 44 as Enterococcus faecium, one as Enterococcus casseliflavus and 61 as Enteroccocus spp. In order to encompass several of the known chemical and functional classes of antibiotics, resistance to ampicillin, penicillin G, ciprofloxacin, chloramphenicol, erythromycin, nitrofurantoin, rifampicin, tetracycline and vancomycin was evaluated. All the isolates were sensitive to antibiotics of clinical importance, such as penicillins and vancomycin. Some differences in Minimal Inhibitory Concentrations (MICs) of antibiotics, could be associated with the enterococcal species.

Comparison of methods for cultivation and isolation of Chlamydia trachomatis.

PubMed Central

Benes, S; McCormack, W M

1982-01-01

McCoy cells treated with cycloheximide, iododeoxyuridine, and DEAE-dextran and untreated McCoy cells were inoculated with two stock strains of Chlamydia trachomatis and with 231 urethral specimens from men, 53 (23%) of which contained C. trachomatis. Isolation rates, number and quality of inclusions, and quality of the cell monolayers were compared. There were no significant differences between the isolation rates in the four systems, although the most isolations were made in the untreated and cycloheximide-treated cells. Cycloheximide-treated cells produced, from both the clinical specimens and the two stock strains, significantly more inclusions than any of the other systems. The monolayer of the cycloheximide-treated cells and the inclusions that grew in these cells were optimal for examination and detection of C. trachomatis. PMID:6185530

Slime-producing Staphylococcus epidermidis and S. aureus in acute bacterial conjunctivitis in soft contact lens wearers.

PubMed

Catalanotti, Piergiorgio; Lanza, Michele; Del Prete, Antonio; Lucido, Maria; Catania, Maria Rosaria; Gallè, Francesca; Boggia, Daniela; Perfetto, Brunella; Rossano, Fabio

2005-10-01

In recent years, an increase in ocular pathologies related to soft contact lens has been observed. The most common infectious agents were Staphylococcus spp. Some strains produce an extracellular polysaccharidic slime that can cause severe infections. Polysaccharide synthesis is under genetic control and involves a specific intercellular adhesion (ica) locus, in particular, icaA and icaD genes. Conjunctival swabs from 97 patients with presumably bacterial bilateral conjunctivitis, wearers of soft contact lenses were examined. We determined the ability of staphylococci to produce slime, relating it to the presence of icaA and icaD genes. We also investigated the antibiotic susceptibility and Pulsed Field Gel Electrophoresis (PFGE) patterns of the clinical isolates. We found that 74.1% of the S. epidermidis strains and 61.1% of the S. aureus strains isolated were slime producers and showed icaA and icaD genes. Both S. epidermidis and S. aureus slime-producing strains exhibited more surface hydrophobicity than non-producing slime strains. The PFGE patterns overlapped in S. epidermidis strains with high hydrophobicity. The similar PFGE patterns were not related to biofilm production. We found scarce matching among the Staphylococcus spp. studied, slime production, surface hydrophobicity and antibiotic susceptibility.

Phenotypic and genotypic study of biofilm formation in Enterococci isolated from urinary tract infections.

PubMed

Fallah, F; Yousefi, M; Pourmand, M R; Hashemi, A; Nazari Alam, A; Afshar, D

2017-07-01

Urinary tract infection (UTI) is one of the most frequent types of nosocomial and community acquired infections in humans. Management of multidrug-resistant Enterococci UTI due to the limited therapeutic options is a great challenge for physicians and clinical microbiologists. The role of bacterial biofilms in recurrent urinary tract infections and antimicrobial resistance has great importance for public health. The aim of this study was to investigate the antibiotic susceptibility pattern as well as the phenotypic and genotypic biofilm formation ability of Enterococci isolates from patients with UTI. A total of 57 isolates of Enterococci were collected from patients with UTI. Enterococcus species were identified using conventional microbiological methods. The antibiotic susceptibility patterns of the isolates were determined by the Kirby-Bauer disk-diffusion. The Modified Congo red agar (MCRA) and Microtiter plate methods used to assess the ability of biofilm formation. All enterococcal isolates were examined for determination of biofilm-related genes, esp, asa1 and ebpR using PCR method. Of 57 enterococcal isolates, 85.9% were recognized as E. faecalis and 14.1% of them were E. faecium. According to our results, linezolid, chloramphenicol and nitrofurantoin were the most effective agents against Enterococcus species. Overall, 26.5% of E. faecalis and 75% of E. faecium isolates were biofilm producers, respectively. Resistance to some antibiotics including penicillin G, ampicillin, vancomycin, nitrofurantoin and chloramphenicol, and ciprofloxacin was significantly higher among biofilm producers than non-biofilm producers Enterococci. The esp, asa1 and ebpR genes were present in 84.2%, 91.2% and 100% isolates. In this study, there was not a significant relationship between presence of these genes and biofilm formation. Our findings reinforce the role of biofilm formation in resistance to antimicrobial agents. Quinupristin/dalfopristin, tetracycline and rifampin may be used as an effective treatment for UTI caused by biofilm producers Enterococci. Our results suggest that biofilm formation is complex and depends on various factors but not just esp, asa1 and ebpR genes in Enterococcus strains. Copyright © 2017 Elsevier Ltd. All rights reserved.

Appraisal of Biofilm Formation in Diabetic Foot Infections by Comparing Phenotypic Methods With the Ultrastructural Analysis.

PubMed

Pugazhendhi, Sugandhi; Dorairaj, Arvind Prasanth

Diabetic patients are more prone to the development of foot ulcers, because their underlying tissues are exposed to colonization by various pathogenic organisms. Hence, biofilm formation plays a vital role in disease progression by antibiotic resistance to the pathogen found in foot infections. The present study has demonstrated the correlation of biofilm assay with the clinical characteristics of diabetic foot infection. The clinical characteristics such as the ulcer duration, size, nature, and grade were associated with biofilm production. Our results suggest that as the size of the ulcer with poor glycemic control increased, the organism was more likely to be positive for biofilm formation. A high-degree of antibiotic resistance was exhibited by the biofilm-producing gram-positive isolates for erythromycin and gram-negative isolates for cefpodoxime. Comparisons of biofilm production using 3 different conventional methods were performed. The strong producers with the tube adherence method were able to produce biofilm using the cover slip assay method, and the weak producers in tube adherence method had difficulty in producing biofilm using the other 2 methods, indicating that the tube adherence method is the best method for assessing biofilm formation. The strong production of biofilm with the conventional method was further confirmed by scanning electron microscopy analysis, because bacteria attached as a distinct layer of biofilm. Thus, the high degree of antibiotic resistance was exhibited by biofilm producers compared with nonbiofilm producers. The tube adherence and cover slip assay were found to be the better method for biofilm evaluation. Copyright © 2018 The American College of Foot and Ankle Surgeons. Published by Elsevier Inc. All rights reserved.

[Results of a multicenter study investigating plasmid mediated colistin resistance genes (mcr-1 and mcr-2) in clinical Enterobacteriaceae ısolates from Turkey].

PubMed

Sarı, Ayşe Nur; Süzük, Serap; Karatuna, Onur; Öğünç, Dilara; Karakoç, Ayşe Esra; Çizmeci, Zeynep; Alışkan, Hikmet Eda; Cömert, Füsun; Bakıcı, Mustafa Zahir; Akpolat, Nezahat; Çilli, Fatma Feriha; Zer, Yasemin; Karataş, Aysel; Akgün Karapınar, Bahar; Bayramoğlu, Gülçin; Özdamar, Melda; Kalem, Fatma; Delialioğlu, Nuran; Aktaş, Elif; Yılmaz, Nisel; Gürcan, Şaban; Gülay, Zeynep

2017-07-01

Colistin is a polymyxin antibiotic which is considered as one of the last line agents against infections due to multidrug resistant or carbapenem resistant gram-negative pathogens. Colistin resistance is associated with chromosomal alterations which can usually cause mutations in genes coding specific two component regulator systems. The first plasmid-mediated colistin resistance gene, mcr-1 was described in Escherichia coli and Klebsiella pneumoniae isolates in December 2015 and followed by another plasmid-mediated colistin resistance gene mcr-2 in 2016. The rapid and interspecies dissemination of plasmid-mediated resistance mechanisms through horizontal gene transfer, have made these genes considerably threatening. After the first reports, although mcr-1/mcr-2 producing Enterobacteriaceae isolates have been reported from many countries, there have been no reports from Turkey. Thus, the aim of this study was to investigate the presence of mcr-1/mcr-2 in clinical Enterobacteriaceae isolates from different parts of our country. A total of 329 Enterobacteriaceae isolates from 22 laboratories were collected which were isolated between March, 2015 and February, 2016. mcr-1/mcr-2 were investigated by polymerase chain reaction during February-March, 2016. Two hundred and seventeen of Klebsiella pneumoniae (66%), 75 of Salmonella spp. (22.8%), 31 of Esherichia coli (9.4%), 3 of Enterobacter cloacae (0.9%), 2 of Klebsiella oxytoca (0.6%) and 1 of Enterobacter aerogenes (0.3%) isolates were included to the study. Agarose gel electrophoresis results of PCR studies have shown expected band sizes for positive control isolates as 309 bp for mcr-1 and 567 bp for mcr-2. However, the presence of mcr-1/mcr-2 genes was not detected among the tested study isolates of Enterobacteriaceae. Although mcr-1/mcr-2 were not detected in our study isolates, it is highly important to understand the mechanism of resistance dissemination and determine the resistant isolates by considering that colistin is a last-line antibiotic against infections of multidrug or carbapenem resistant gram-negative bacteria. Thus, it is suggested that these mechanisms should be followed-up in both clinical and non-clinical (e.g. isolates from food animals, raw meats and environment) isolates of special populations.

Four Main Virotypes among Extended-Spectrum-β-Lactamase-Producing Isolates of Escherichia coli O25b:H4-B2-ST131: Bacterial, Epidemiological, and Clinical Characteristics

PubMed Central

Mora, Azucena; Mamani, Rosalia; López, Cecilia; Blanco, Miguel; Dahbi, Ghizlane; Herrera, Alexandra; Marzoa, Juan; Fernández, Val; de la Cruz, Fernando; Martínez-Martínez, Luis; Alonso, María Pilar; Nicolas-Chanoine, Marie-Hélène; Johnson, James R.; Johnston, Brian; López-Cerero, Lorena; Pascual, Álvaro; Rodríguez-Baño, Jesús

2013-01-01

A total of 1,021 extended-spectrum-β-lactamase-producing Escherichia coli (ESBLEC) isolates obtained in 2006 during a Spanish national survey conducted in 44 hospitals were analyzed for the presence of the O25b:H4-B2-ST131 (sequence type 131) clonal group. Overall, 195 (19%) O25b-ST131 isolates were detected, with prevalence rates ranging from 0% to 52% per hospital. Molecular characterization of 130 representative O25b-ST131 isolates showed that 96 (74%) were positive for CTX-M-15, 15 (12%) for CTX-M-14, 9 (7%) for SHV-12, 6 (5%) for CTX-M-9, 5 (4%) for CTX-M-32, and 1 (0.7%) each for CTX-M-3 and the new ESBL enzyme CTX-M-103. The 130 O25b-ST131 isolates exhibited relatively high virulence scores (mean, 14.4 virulence genes). Although the virulence profiles of the O25b-ST131 isolates were fairly homogeneous, they could be classified into four main virotypes based on the presence or absence of four distinctive virulence genes: virotypes A (22%) (afa FM955459 positive, iroN negative, ibeA negative, sat positive or negative), B (31%) (afa FM955459 negative, iroN positive, ibeA negative, sat positive or negative), C (32%) (afa FM955459 negative, iroN negative, ibeA negative, sat positive), and D (13%) (afa FM955459 negative, iroN positive or negative, ibeA positive, sat positive or negative). The four virotypes were also identified in other countries, with virotype C being overrepresented internationally. Correspondingly, an analysis of XbaI macrorestriction profiles revealed four major clusters, which were largely virotype specific. Certain epidemiological and clinical features corresponded with the virotype. Statistically significant virotype-specific associations included, for virotype B, older age and a lower frequency of infection (versus colonization), for virotype C, a higher frequency of infection, and for virotype D, younger age and community-acquired infections. In isolates of the O25b:H4-B2-ST131 clonal group, these findings uniquely define four main virotypes, which are internationally distributed, correspond with pulsed-field gel electrophoresis (PFGE) profiles, and exhibit distinctive clinical-epidemiological associations. PMID:23926164

Molecular characterization of metallo β-lactamase producing multidrug resistant Pseudomonas aeruginosa from various clinical samples.

PubMed

Ramakrishnan, Kalaivani; Rajagopalan, Saranathan; Nair, Shashikala; Kenchappa, Prashanth; Chandrakesan, Sheela Devi

2014-01-01

Pseudomonas aeruginosa is a potent opportunistic nosocomial human pathogen among Gram-negative bacteria causing various life-threatening infections in patients from Intensive Care Units. This bacterium has become resistant to almost all commonly available antibiotics with limited treatment options. Multi drug resistant P. aeruginosa (MDRPA) is a major cause of concern among hospital acquired infections. It uses distinctive resistant mechanisms virtually to all the available antibiotics such as Metallo β-lactamases (MBL) production, extended spectrum β-lactamase production (ESBL), up regulation of efflux systems related genes and decreased outer membrane permeability. This study was carried out to find one the predominant resistance mechanisms among MDRPA and the prevalence of corresponding resistance genes. MDRPA isolates collected from various clinical samples for a period of 1-year (November 2009-Octo ber 2010) were included to detect the predominant mechanism of resistance using phenotypic and molecular methods. Molecular characterization of all these isolates was done by polymerase chain reaction (PCR) for the presence of blaVIM-₂, blaIMP-₁, blaOXA-₂₃, and blaNDM-₁ genes with specific primers. Among 75 MDRPA isolates 84% (63) were MBL producers. Molecular characterization studied by PCR showed the presence of blaVIM-₂ gene in 13% of MBL producers. The prevalence of MBLs has been increasing worldwide, particularly among P. aeruginosa, leading to severe limitations in the therapeutic options for the management. Thus, proper resistance screening measures and appropriate antibiotic policy can be strictly adopted by all the healthcare facility providers to overcome these superbugs.

Assessment and comparison of the pathogenicity of Sheeppox Virus strains isolated in Morocco

PubMed Central

Hajjou, Saida; Khataby, Khadija; Amghar, Souad; El Fahime, Mustapha; El Harrak, Mehdi; Fakiri, Malika; Loutfi, Chafiqa

2017-01-01

Background and Objectives: Sheeppox virus causes systemic disease in sheep that is often associated with high morbidity and mortality. Protection against sheep pox is mainly based on medical prophylaxis, vaccination being the only way. In Morocco, and up to now, there is no available information about local challenge strain to use for controlling the efficiency of vaccines produced against sheep pox. Hence, the objective of the present study was to evaluate and compare the pathogenicity of seven Sheeppox virus (SPVs) isolates from 1993–1995 in Morocco. Materials and Methods: These seven SPV isolates have undergone various tests to evaluate their pathogenicity: Passages and titration on cell culture, Experimental inoculation on sheep, Virus-neutralization, In vivo titration and viral re-isolation by real-time PCR assay. Results: All infected lambs showed severe clinical signs, while most of them have been reproduced on 5 dpi and persisted until 21 dpi. The lambs infected by Oj1P4, Oj2P4 and BerP5 appeared lethargic, reluctant to move compared to those infected by other isolates. The results also revealed that all isolates were able to induce serological response. Virus isolation from infected organs and blood and amplification of the viral DNA by real-time PCR proved the presence of the virus in tissues and blood of infected lambs. These Moroccan SPVs demonstrated that the three isolates Oj1P4, Oj2P4 and BerP5 have a high pathogenicity; especially the BerP5 isolate which has an important infectious titer. Conclusion: These results demonstrate that the Berkane isolate is the most pathogenic of the tested isolates and it can be an excellent challenge strain for the control of the efficiency of vaccines against sheep pox produced in Morocco. PMID:29487736

Laboratory Diagnostics of Botulism

PubMed Central

Lindström, Miia; Korkeala, Hannu

2006-01-01

Botulism is a potentially lethal paralytic disease caused by botulinum neurotoxin. Human pathogenic neurotoxins of types A, B, E, and F are produced by a diverse group of anaerobic spore-forming bacteria, including Clostridium botulinum groups I and II, Clostridium butyricum, and Clostridium baratii. The routine laboratory diagnostics of botulism is based on the detection of botulinum neurotoxin in the patient. Detection of toxin-producing clostridia in the patient and/or the vehicle confirms the diagnosis. The neurotoxin detection is based on the mouse lethality assay. Sensitive and rapid in vitro assays have been developed, but they have not yet been appropriately validated on clinical and food matrices. Culture methods for C. botulinum are poorly developed, and efficient isolation and identification tools are lacking. Molecular techniques targeted to the neurotoxin genes are ideal for the detection and identification of C. botulinum, but they do not detect biologically active neurotoxin and should not be used alone. Apart from rapid diagnosis, the laboratory diagnostics of botulism should aim at increasing our understanding of the epidemiology and prevention of the disease. Therefore, the toxin-producing organisms should be routinely isolated from the patient and the vehicle. The physiological group and genetic traits of the isolates should be determined. PMID:16614251

Virulence characteristics of Escherichia coli in nosocomial urinary tract infection.

PubMed

Ikäheimo, R; Siitonen, A; Kärkkäinen, U; Mäkelä, P H

1993-06-01

We examined 148 strains of Escherichia coli isolated from the urine from patients with nosocomial urinary tract infection (UTI). The prevalence of P fimbriation was only 11.5%. Of the strains, 17.6% expressed non-P M(R) adhesins (defined as strains expressing mannose-resistant but not P-specific hemagglutination); 33.1% produced hemolysin, and 15.2% expressed type 1C fimbriae. O6 was the most common group of O antigens (12.2%), closely followed by O75 (9.5%); both of these groups are relatively uncommon (4.5% and 1%, respectively) in fecal strains isolated from healthy adults. Of the strains with O6 and O75 antigens, 78.8% and 79% produced hemolysin, but of all other strains causing UTI, only 21% produced hemolysin. Of the strains with O6 antigens, 61% expressed non-P M(R) adhesins, but only 12% of all other strains causing UTI expressed non-P M(R) adhesins. There were no significant differences in the prevalence of virulence properties between strains isolated from patients with or without an underlying medical illness or between strains causing different clinical categories of UTI. We conclude that the prevalence of bacterial virulence factors is low among patients with nosocomial UTI.

Comparison of CHROMagar Salmonella Medium and Hektoen Enteric Agar for Isolation of Salmonellae from Stool Samples

PubMed Central

Gaillot, Olivier; Di Camillo, Patrick; Berche, Patrick; Courcol, René; Savage, Colette

1999-01-01

CHROMagar Salmonella (CAS), a new chromogenic medium, was retrospectively compared to Hektoen enteric agar (HEA) with 501 Salmonella stock isolates and was then prospectively compared to HEA for the detection and presumptive identification of Salmonella spp. with 508 stool samples before and after enrichment. All stock cultures (100%), including cultures of H2S-negative isolates, yielded typical mauve colonies on CAS, while 497 (99%) isolates produced typical lactose-negative, black-centered colonies on HEA. Following overnight incubation at 37°C, a total of 20 Salmonella strains were isolated from the 508 clinical samples. Sensitivities for primary plating and after enrichment were 95% (19 isolates) and 100% (20 isolates), respectively, for CAS and 80% (16 isolates) and 100% (20 isolates), respectively, for HEA. The specificity of CAS (88.9%) was significantly higher than that of HEA (78.5%; P < 0.0001). On the basis of its good sensitivity and specificity, CAS medium can be recommended for use for primary plating when human stool samples are screened for Salmonella spp. PMID:9986847

Virulence Factors of Staphylococcus aureus Isolates in an Iranian Referral Children's Hospital.

PubMed

Sabouni, Farah; Mahmoudi, Shima; Bahador, Abbas; Pourakbari, Babak; Sadeghi, Reihaneh Hosseinpour; Ashtiani, Mohammad Taghi Haghi; Nikmanesh, Bahram; Mamishi, Setareh

2014-04-01

The clinical importance of Staphylococcus aureus (S. aureus) is attributed to notable virulence factors, surface proteins, toxins, and enzymes as well as the rapid development of drug resistance. The aim of this study was to compare the occurrence of virulence factors produced by S. aureus strains isolated from children in an Iranian referral children's hospital. The presence of genes encoding for the enterotoxins A (sea), B (seb), C (sec), D (sed), TSST-1 (tsst), exfoliative toxin A (eta), and exfoliative toxin B (etb) were detected by Multiplex polymerase chain reaction (PCR) using specific primers. In addition, the standardized Kirby-Bauer disc-diffusion method was performed on Mueller-Hinton agar. In total, 133 S. aureus isolates were obtained from different patients. Of these S. aureus isolates, 64 (48%) were methicillin-resistant S. aureus (MRSA), and all of these tested positive for the mecA gene. Regarding the classical enterotoxin genes, sea gene (40.6%) was the most prevalent followed by seb (19.6%), tsst (12.8%), eta (11.3%), etb (9%), sed (4.5%), and sec (3%). Among methicillin-susceptible S. aureus (MSSA) isolates, seb and tsst were the more prevalent toxins in comparison with MRSA isolates (p < 0.05), while the frequency of sea, sed, eta, and etb genes were higher among MRSA isolates (p > 0.05). In our study enterotoxin A was produced by 40.6% of the isolates (48% from MRSA and 33% from MSSA isolates) which was higher than in previous reports. According to our results, strict hygiene and preventative measures during food processing are highly recommended.

Defining the Relationship Between Phenotypic and Genotypic Resistance Profiles of Multidrug-Resistant Enterobacterial Clinical Isolates.

PubMed

Galal, Lamis; Abdel Aziz, Neveen A; Hassan, Walaa M

2018-05-11

Fluoroquinolones and aminoglycosides offer effective therapy for extended-spectrum beta-lactamase (ESBL)-producing enterobacterial infections, but their usefulness is threatened by increasing resistant strains. This study was conducted to demonstrate the phenotypic outcomes of the coexistence of genetic determinants mediating resistance to extended-spectrum cephalosporins and quinolones in enterobacterial isolates collected from patients with health-care-associated infections in Egypt. ESBL phenotype was determined using double-disk synergy test (DDST). The PCR technique was used to detect the presence of the genes mediating quinolone resistance (qnr and aac(6')-Ib-cr) and coexistence with ESBL genes. We also examined the association between the genetic makeup of the isolates and their resistance profiles including effect on MIC results. Phenotypically ESBLs were detected in 60-82% of the enterobacterial isolates. ESBL, qnr and aac(6')-Ib-cr genes were detected with the following percentages in Citrobacter isolates (69%, 69%, and 43%, respectively), E.coli isolates (65%, 70%, and 45%, respectively), Enterobacter isolates (56%, 67%, and 33%, respectively), and finally Klebsiella isolates (42%, 66%, and 25%, respectively). The coexistence of these multiresistant genetic elements significantly increased the MIC values of the tested antibiotics from different classes. We suggest using blaTEM, blaCTX-M-15, qnr, and aac(6')-Ib-cr genes for better and faster prediction of suitable antibiotic therapy with effective doses against ESBL-producing isolates harboring plasmid-mediated quinolone resistance (PMQR) determinants. Amikacin, meropenem, gentamicin, and imipenem seem to be better choices of treatment for such life-threatening infections, because of their remaining highest activity.

Presence of qnr gene in Escherichia coli and Klebsiella pneumoniae resistant to ciprofloxacin isolated from pediatric patients in China.

PubMed

Wang, Aihua; Yang, Yonghong; Lu, Quan; Wang, Yi; Chen, Yuan; Deng, Li; Ding, Hui; Deng, Qiulian; Zhang, Hong; Wang, Chuanqing; Liu, Lan; Xu, Xiwei; Wang, Li; Shen, Xuzhuang

2008-05-22

Quinolone resistance in Enterobacteriaceae results mainly from mutations in type II DNA topoisomerase genes and/or changes in the expression of outer membrane and efflux pumps. Several recent studies have indicated that plasmid-mediated resistance mechanisms also play a significant role in fluoroquinolone resistance, and its prevalence is increasing worldwide. In China, the presence of the qnr gene in the clinical isolates of Enterobacteriaceae has been reported, but this transmissible quinolone resistance gene has not been detected in strains isolated singly from pediatric patients. Because quinolones associated with a variety of adverse side effects on children, they are not authorized for pediatric use. This study therefore aimed to investigate the presence of the qnr gene in clinical isolates of E. coli and K. pneumoniae from pediatric patients in China. A total 213 of non-repetitive clinical isolates resistant to ciprofloxacin from E. coli and K. pneumoniae were collected from hospitalized patients at five children's hospital in Beijing, Shanghai, Guangzhou, and Chongqing. The isolates were screened for the plasmid-mediated quinolone resistance genes of qnrA, qnrB, and qnrS by PCR. Transferability was examined by conjugation with the sodium azide-resistant E. coli J53. All qnr-positive were analyzed for clonality by enterobacterial repetitive intergenic consensus (ERIC)-PCR. The study found that 19 ciprofloxacin-resistant clinical isolates of E. coli and K. pneumoniae were positive for the qnr gene, and most of the qnr positive strains were ESBL producers. Conjugation experiments showed that quinolone resitance could be transferred to recipients. Apart from this, different DNA banding patterns were obtained by ERIC-PCR from positive strains, which means that most of them were not clonally related. This report on transferable fluoroquinolone resistance due to the qnr gene among E. coli and K. pneumoniae strains indicated that plasmid-mediated quinolone resistance has emerged in pediatric patients in China.

Performance of a PCR assay for the rapid identification of the Klebsiella pneumoniae ST258 epidemic clone in Latin American clinical isolates.

PubMed

Gomez, S A; Rapoport, M; Piergrossi, N; Faccone, D; Pasteran, F; De Belder, D; ReLAVRA-Group; Petroni, A; Corso, A

2016-10-01

The worldwide dissemination of Klebsiella pneumoniae carbapenemase (KPC)-producing Klebsiella pneumoniae ST258 demands a rapid PCR-based typing method to detect unique genes of the ST258 clone. This study evaluates a PCR developed by Adler et al. (2014) for the detection of ST258 in K. pneumoniae clinical isolates centered on the identification of the pilv-I and prp genes. We tested 143 clinical isolates from Argentina (n=109), Chile (n=1), Colombia (n=1), Costa Rica (n=2), Ecuador (n=5), El Salvador (n=2), Nicaragua (n=5), Panamá (n=2), Paraguay (n=2), Perú (n=3) and Trinidad and Tobago (n=11) recovered from 2006 to 2015. blaKPC, pilv-l and prp genes were detected by PCR and sequenced by standard procedures. ST258 and non-ST258 were defined by PFGE and/or MLST. Isolates were grouped according to PFGE patterns: 58 were compatible with ST258 (group 1) and 85 with non-ST258 (group 2). MLST study was done on an arbitrary selection of isolates. The pilv-l gene was present only in ST258 isolates, regardless of the presence of the blaKPC gene. Results for the prp gene were variable. Its presence did not define ST258. The pilv-I PCR had a sensitivity and specificity of 100%, respectively, for the detection of ST258 in the isolates under investigation. Given our findings, the pilv-I PCR could replace more time and resource consuming methods, allowing for more rapid detection of the circulating high risk K. pneumoniae clone ST258 in Latin American (LA) countries. Copyright © 2016 Elsevier B.V. All rights reserved.

Comparative Genotypic and Phenotypic Analysis of Cronobacter Species Cultured from Four Powdered Infant Formula Production Facilities: Indication of Pathoadaptation along the Food Chain

PubMed Central

Yan, Qiongqiong; Wang, Juan; Gangiredla, Jayanthi; Cao, Yu; Martins, Marta; Gopinath, Gopal R.; Stephan, Roger; Lampel, Keith; Tall, Ben D.

2015-01-01

Cronobacter species are opportunistic pathogens commonly found in the environment. Among the seven Cronobacter species, Cronobacter sakazakii sequence type 4 (ST-4) is predominantly associated with recorded cases of infantile meningitis. This study reports on a 26-month powdered infant formula (PIF) surveillance program in four production facilities located in distinct geographic regions. The objective was to identify the ST(s) in PIF production environments and to investigate the phenotypic features that support their survival. Of all 168 Cronobacter isolates, 133 were recovered from a PIF production environment, 31 were of clinical origin, and 4 were laboratory type strains. Sequence type 1 (n = 84 isolates; 63.9%) was the dominant type in PIF production environments. The majority of these isolates clustered with an indistinguishable pulsotype and persisted for at least an 18-month period. Moreover, DNA microarray results identified two phylogenetic lineages among ST-4 strains tested. Thereafter, the ST-1 and -4 isolates were phenotypically compared. Differences were noted based on the phenotypes expressed by these isolates. The ST-1 PIF isolates produced stronger biofilms at both 28°C and 37°C, while the ST-4 clinical isolates exhibited greater swimming activity and increased binding to Congo red dye. Given the fact that PIF is a low-moisture environment and that the clinical environment provides for an interaction between the pathogen and its host, these differences may be consistent with a form of pathoadaptation. These findings help to extend our current understanding of the epidemiology and ecology of Cronobacter species in PIF production environments. PMID:25911470

Prevalence of quinolone resistance determinant qnrA6 among broad- and extended-spectrum beta-lactam-resistant Proteus mirabilis and Morganella morganii clinical isolates with sul1-type class 1 integron association in a Tunisian Hospital.

PubMed

Mahrouki, Sihem; Perilli, Mariagrazia; Bourouis, Amel; Chihi, Hela; Ferjani, Mustapha; Ben Moussa, Mohamed; Amicosante, Gianfranco; Belhadj, Omrane

2013-08-01

The aim of this study was to investigate the prevalence and the emergence of plasmid-mediated quinolone resistance among broad-spectrum beta-lactam-resistant Proteus mirabilis and Morganella morganii clinical isolates recovered in the Military Hospital in Tunisia. Of 200 strains examined, 50 exhibited resistance to quinolones. Quinolone resistance determinants (qnr and aac(6')-Ib-cr) were characterized by multiplex PCR and sequencing. Chromosomal quinolone resistance mutations in the quinolone resistance-determining region (QRDR) and class 1 integron characterization were analysed by PCR and sequencing. The clonal relationship between the isolates was studied by pulsed-field gel electrophoresis (PFGE). Fourteen isolates harboured qnrA6 and among them 8 (57%) were extended-spectrum beta-lactamase (ESBL) producers, whilst 12 (85%) isolates harboured blaDHA-1. Mutations in the QRDR were detected in gyrA (Ser83Ile, Glu87Lys), gyrB (Ser464Phe), and parC (Ser80Ile). qnrA6 and blaDHA-1 genes were found embedded in complex sul1-type class 1 integrons. A gene cassette carrying aac(6')-Ib-cr was found located in the class 1 integron upstream of the qacEΔ1 gene. According to the PFGE analysis, the isolates were clonally unrelated. This is the first description in North Africa of class 1 integrons carrying blaDHA-1, qnrA6 gene, and aac(6')-Ib-cr determinants in clinical strains of Proteus mirabilis and Morganella morganii.

Multicenter Evaluation of the Bruker MALDI Biotyper CA System for the Identification of Clinical Aerobic Gram-Negative Bacterial Isolates

PubMed Central

Faron, Matthew L.; Buchan, Blake W.; Hyke, Josh; Madisen, Neil; Lillie, Jennifer L.; Granato, Paul A.; Wilson, Deborah A.; Procop, Gary W.; Novak-Weekley, Susan; Marlowe, Elizabeth; Cumpio, Joven; Griego-Fullbright, Christen; Kindig, Sandra; Timm, Karen; Young, Stephen; Ledeboer, Nathan A.

2015-01-01

The prompt and accurate identification of bacterial pathogens is fundamental to patient health and outcome. Recent advances in matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) have revolutionized bacterial identification in the clinical laboratory, but uniform incorporation of this technology in the U.S. market has been delayed by a lack of FDA-cleared systems. In this study, we conducted a multicenter evaluation of the MALDI Biotyper CA (MBT-CA) System (Bruker Daltonics Inc, Billerica, MA) for the identification of aerobic gram-negative bacteria as part of a 510(k) submission to the FDA. A total of 2,263 aerobic gram negative bacterial isolates were tested representing 23 genera and 61 species. Isolates were collected from various clinical sources and results obtained from the MBT-CA System were compared to DNA sequencing and/or biochemical testing. Isolates that failed to report as a "high confidence species ID" [log(score) ≥2.00] were re-tested using an extraction method. The MBT-CA System identified 96.8% and 3.1% of isolates with either a "high confidence" or a "low confidence" [log(score) value between 1.70 and <2.00] species ID, respectively. Two isolates did not produce acceptable confidence scores after extraction. The MBT-CA System correctly identified 99.8% (2,258/2,263) to genus and 98.2% (2,222/2,263) to species level. These data demonstrate that the MBT-CA System provides accurate results for the identification of aerobic gram-negative bacteria. PMID:26529504

Multicenter Evaluation of the Bruker MALDI Biotyper CA System for the Identification of Clinical Aerobic Gram-Negative Bacterial Isolates.

PubMed

Faron, Matthew L; Buchan, Blake W; Hyke, Josh; Madisen, Neil; Lillie, Jennifer L; Granato, Paul A; Wilson, Deborah A; Procop, Gary W; Novak-Weekley, Susan; Marlowe, Elizabeth; Cumpio, Joven; Griego-Fullbright, Christen; Kindig, Sandra; Timm, Karen; Young, Stephen; Ledeboer, Nathan A

2015-01-01

The prompt and accurate identification of bacterial pathogens is fundamental to patient health and outcome. Recent advances in matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) have revolutionized bacterial identification in the clinical laboratory, but uniform incorporation of this technology in the U.S. market has been delayed by a lack of FDA-cleared systems. In this study, we conducted a multicenter evaluation of the MALDI Biotyper CA (MBT-CA) System (Bruker Daltonics Inc, Billerica, MA) for the identification of aerobic gram-negative bacteria as part of a 510(k) submission to the FDA. A total of 2,263 aerobic gram negative bacterial isolates were tested representing 23 genera and 61 species. Isolates were collected from various clinical sources and results obtained from the MBT-CA System were compared to DNA sequencing and/or biochemical testing. Isolates that failed to report as a "high confidence species ID" [log(score) ≥2.00] were re-tested using an extraction method. The MBT-CA System identified 96.8% and 3.1% of isolates with either a "high confidence" or a "low confidence" [log(score) value between 1.70 and <2.00] species ID, respectively. Two isolates did not produce acceptable confidence scores after extraction. The MBT-CA System correctly identified 99.8% (2,258/2,263) to genus and 98.2% (2,222/2,263) to species level. These data demonstrate that the MBT-CA System provides accurate results for the identification of aerobic gram-negative bacteria.

Alfalfa endophytes as novel sources of antimicrobial compounds that inhibit the growth of human and plant pathogens

USDA-ARS?s Scientific Manuscript database

Fungal endophytes may contribute to plant health and disease protection, yet little is known about their various roles in alfalfa. Also, endophytes from several plant species produce novel antimicrobial compounds that may be useful clinically. We isolated endophytic fungi from over 50 samples from s...

[Phenotypic and molecular identification of extended-spectrum beta-lactamase (ESBL) TEM and SHV produced by clinical isolates Escherichia coli and Klebsiella spp. in hospitals].

PubMed

González Mesa, Leonora; Ramos Morí, Astrid; Nadal Becerra, Loreta; Morffi Figueroa, Janet; Hernández Robledo, Ernesto; Alvarez, Ana Berta; Marchena Bequer, Juan J; González Alemán, Mabel; Villain Plous, Carlos

2007-01-01

Nosocomial infections caused by gram-negative bacilli which produce extended spectrum beta-lactamase (ESBL) are associated with the increase of morbidity and mortality in hospitals. The objective of this study was to evaluate the frequency of ESBL, specifically the TEM and SHV type, produced by Escherichia coli and Klebsiella spp. strains, and also to determine the antimicrobial susceptibility of these isolates in comparison with other antibiotic families. A total of 326 strains were collected between 2002-2004 from hospitals in Havana City. The susceptibility tests were carried out according to the NCCLS guides and they were confirmed as. ESBL producers by the double disk diffusion method. The molecular characterization of these enzymes was determined by polymerase chain reaction (PCR), using two sets of oligonucleotides to amplify genes encoding TEM and SHV type beta-lactamase. The ESBL phenotype was detected in 31 (10%) Escherichia coli isolates, 19 of these strains (61%) carried the blaTEM genes, 5 (16%) blaSHV genes, 4 (12%) strains carried both genes and 11 strains (35%) carried the non-ESBL blaTEM and blaSHV genes. In Klebsiella spp. the ESBL phenotype was detected in 10 (36 %) isolates, only one strain carried the blaTEM gene. The most active antimicrobials against Escherichia coli were ciprofloxacin (64.5%) and gentamicin (58.07%); in the case of Klebsiella spp. the same antimicrobials were the most active with similar susceptibility (70%) for both. The carbapenems still remain the most active antibiotics against Escherichia coli and Klebsiella spp. strains, which are ESBL producers. However, their use should be closely controlled.

Differentiating Botulinum Neurotoxin-Producing Clostridia with a Simple, Multiplex PCR Assay.

PubMed

Williamson, Charles H D; Vazquez, Adam J; Hill, Karen; Smith, Theresa J; Nottingham, Roxanne; Stone, Nathan E; Sobek, Colin J; Cocking, Jill H; Fernández, Rafael A; Caballero, Patricia A; Leiser, Owen P; Keim, Paul; Sahl, Jason W

2017-09-15

Diverse members of the genus Clostridium produce botulinum neurotoxins (BoNTs), which cause a flaccid paralysis known as botulism. While multiple species of clostridia produce BoNTs, the majority of human botulism cases have been attributed to Clostridium botulinum groups I and II. Recent comparative genomic studies have demonstrated the genomic diversity within these BoNT-producing species. This report introduces a multiplex PCR assay for differentiating members of C. botulinum group I, C. sporogenes , and two major subgroups within C. botulinum group II. Coding region sequences unique to each of the four species/subgroups were identified by in silico analyses of thousands of genome assemblies, and PCR primers were designed to amplify each marker. The resulting multiplex PCR assay correctly assigned 41 tested isolates to the appropriate species or subgroup. A separate PCR assay to determine the presence of the ntnh gene (a gene associated with the botulinum neurotoxin gene cluster) was developed and validated. The ntnh gene PCR assay provides information about the presence or absence of the botulinum neurotoxin gene cluster and the type of gene cluster present ( ha positive [ ha + ] or orfX + ). The increased availability of whole-genome sequence data and comparative genomic tools enabled the design of these assays, which provide valuable information for characterizing BoNT-producing clostridia. The PCR assays are rapid, inexpensive tests that can be applied to a variety of sample types to assign isolates to species/subgroups and to detect clostridia with botulinum neurotoxin gene ( bont ) clusters. IMPORTANCE Diverse clostridia produce the botulinum neurotoxin, one of the most potent known neurotoxins. In this study, a multiplex PCR assay was developed to differentiate clostridia that are most commonly isolated in connection with human botulism cases: C. botulinum group I, C. sporogenes , and two major subgroups within C. botulinum group II. Since BoNT-producing and nontoxigenic isolates can be found in each species, a PCR assay to determine the presence of the ntnh gene, which is a universally present component of bont gene clusters, and to provide information about the type ( ha + or orfX + ) of bont gene cluster present in a sample was also developed. The PCR assays provide simple, rapid, and inexpensive tools for screening uncharacterized isolates from clinical or environmental samples. The information provided by these assays can inform epidemiological studies, aid with identifying mixtures of isolates and unknown isolates in culture collections, and confirm the presence of bacteria of interest. Copyright © 2017 Williamson et al.

In-vitro susceptibility of 400 isolates of Neisseria gonorrhoeae in Vancouver, 1982-84.

PubMed Central

Bowie, W R; Shaw, C E; Chan, D G; Jones, H D; Black, W A

1986-01-01

Consecutive isolates of Neisseria gonorrhoeae obtained at a sexually transmitted disease clinic in Vancouver between June 1982 and June 1984 were tested for in-vitro susceptibility to eight antimicrobial agents. Of the 400 isolates 6 (1.5%) were penicillinase-producing N. gonorrhoeae, and for 25 (6.2%) the minimum inhibitory concentrations (MICs) of penicillin were 1.0 to 4.0 micrograms/ml. Ceftriaxone sodium was the most active agent. The MICs were higher than those reported in a Canadian study in 1973-74, except for tetracycline hydrochloride. The patterns of susceptibility of the isolates to one antimicrobial agent correlated significantly with those to each other agent, although the relation was weakest for trimethoprim-sulfamethoxazole and spectinomycin. The results reinforce the need to evaluate local in-vitro susceptibility patterns, especially since the proportion of isolates with relative and absolute resistance to penicillin is increasing. PMID:3091234

High prevalence of plasmid-mediated quinolone resistance determinants qnr, aac(6')-Ib-cr, and qepA among ceftiofur-resistant Enterobacteriaceae isolates from companion and food-producing animals.

PubMed

Ma, Junying; Zeng, Zhenling; Chen, Zhangliu; Xu, Xiaogang; Wang, Xiaoying; Deng, Yuting; Lü, Dianhong; Huang, Liangzong; Zhang, Yunyuan; Liu, Jianhua; Wang, Minggui

2009-02-01

Three kinds of plasmid-mediated quinolone resistance (PMQR) determinants have been discovered and have been shown to be widely distributed among clinical isolates: qnr genes, aac(6')-Ib-cr, and qepA. Few data on the prevalence of these determinants in strains from animals are available. The presence of PMQR genes in isolates from animals was determined by PCR amplification and DNA sequencing. The production of extended-spectrum beta-lactamases (ESBLs) and AmpC beta-lactamases in the strains was detected, and their genotypes were determined. The genetic environment of PMQR determinants in selected plasmids was analyzed. All samples of ceftiofur-resistant (MICs > or = 8 microg/ml) isolates of the family Enterobacteriaceae were selected from 36 companion animals and 65 food-producing animals in Guangdong Province, China, between November 2003 and April 2007, including 89 Escherichia coli isolates, 9 Klebsiella pneumoniae isolates, and isolates of three other genera. A total of 68.3% (69/101) of the isolates produced ESBLs and/or AmpC beta-lactamases, mainly those of the CTX-M and CMY types. Of the 101 strains, PMQR determinants were present in 35 (34.7%) isolates, with qnr, aac(6')-Ib-cr, and qepA detected alone or in combination in 8 (7.9%), 19 (18.8%), and 16 (15.8%) strains, respectively. The qnr genes detected included one qnrB4 gene, four qnrB6 genes, and three qnrS1 genes. Five strains were positive for both aac(6')-Ib-cr and qepA, while one strain was positive for qnrS1, aac(6')-Ib-cr, and qepA. qnrB6 was flanked by two copies of ISCR1 with an intervening dfr gene downstream and sul1 and qacEDelta1 genes upstream. In another plasmid, aac(6')-Ib-cr followed intI1 and arr-3 was downstream. PMQR determinants are highly prevalent in ceftiofur-resistant Enterobacteriaceae strains isolated from animals in China. This is the first report of the occurrence of PMQR determinants among isolates from companion animals.

Frequency and expression of mutacin biosynthesis genes in isolates of Streptococcus mutans with different mutacin-producing phenotypes.

PubMed

Kamiya, Regianne Umeko; Höfling, José Francisco; Gonçalves, Reginaldo Bruno

2008-05-01

The aim of this study was to analyse the frequency and expression of biosynthesis genes in 47 Streptococcus mutans isolates with different mutacin-producing phenotypes. Detection of the frequency and expression of genes encoding mutacin types I, II, III and IV were carried out by PCR and semi-quantitative RT-PCR, respectively, using primers specific for each type of biosynthesis gene. In addition, a further eight genes encoding putative bacteriocins, designated bsm 283, bsm 299, bsm 423, bsm 1889c, bsm 1892c, bsm 1896, bsm 1906c and bsm 1914, were also screened. There was a high phenotypic diversity; some Streptococcus mutans isolates presented broad antimicrobial spectra against other Streptococcus mutans clinical isolates, including bacteria resistant to common antibiotics, as well as Staphylococcus aureus, Staphylococcus epidermidis, Enterococcus faecalis and Streptococcus pyogenes. The expression frequency of the bsm gene was higher than that of the previously characterized mutacins (I-IV). There was no positive correlation between the number of indicator strains inhibited (antimicrobial spectra) and the number of biosynthesis genes expressed (Spearman correlation test, r=-0.03, P>0.05). In conclusion, the high diversity of mutacin-producing phenotypes, associated with high frequency of expression of the biosynthesis genes screened, reveals a broad repertoire of genetic determinants encoding antimicrobial peptides that can act in different combinations.

Producing recombinant human milk proteins in the milk of livestock species.

PubMed

Bösze, Zsuzsanna; Baranyi, Mária; Whitelaw, C Bruce A

2008-01-01

Recombinant human proteins produced by the mammary glands of genetically modified transgenic livestock mammals represent a special aspect of milk bioactive components. For therapeutic applications, the often complex posttranslational modifications of human proteins should be recapitulated in the recombinant products. Compared to alternative production methods, mammary gland production is a viable option, underlined by a number of transgenic livestock animal models producing abundant biologically active foreign proteins in their milk. Recombinant proteins isolated from milk have reached different phases of clinical trials, with the first marketing approval for human therapeutic applications from the EMEA achieved in 2006.

In vitro Effectiveness of Commercial Bacteriophage Cocktails on Diverse Extended-Spectrum Beta-Lactamase Producing Escherichia coli Strains.

PubMed

Gundogdu, Aycan; Bolkvadze, Darajen; Kilic, Huseyin

2016-01-01

The objective of this study is to determine the in vitro susceptibility of Georgian bacteriophage cocktails on multidrug resistant (MDR) extended-spectrum beta-lactamase producing Escherichia coli (ESBL-EC) isolated from patients' blood and urine cultures. A total of 615 E. coli isolates were included in this study. Phene Plate (PhP)-typing and phylogenetic grouping were used for the typing. Antimicrobial resistance profiles and ESBL production of all isolates were confirmed according to Clinical and Laboratory Standards Institute (CLSI) criteria. The activities of four bacteriophage cocktails (Enko-phage, SES-bacteriophage, Pyo-bacteriophage, and Intesti-bacteriophage) were determined against 142 ESBL-EC using in vitro spot tests. According to this, Enko-phage were active against 87.3% of the tested strains while that ratio was 81.7% for Intesti-bacteriophage, 81.7% for Pyo-bacteriophage, and 59.2% for SES-bacteriophage cocktails. Based on the contingency tests, the phage cocktails were observed to be statistically significantly ( p < 0.001) more effective on ESBL-EC strains belonging to phylogenetic groups D and B2. The employed phage cocktails were found to be affective against all tested resistant types. These results are promising especially for the infections that are caused by MDR pathogens that are difficult to treat. As this is a preliminary step to the potential clinical trials to be designed for the country, in vitro confirmation of their success on a MDR ESBL-EC collection should be accepted as an initial action, which is encouraging to consider clinical trials of phage therapy especially in countries which are not introduce phage therapy.

Carriage of Class 1 and 2 Integrons in Quinolone, Extended-Spectrum-β-Lactamase-Producing and Multi Drug Resistant E.coli and K.pneumoniae: High Burden of Antibiotic Resistance.

PubMed

Shams, Froogh; Hasani, Alka; Ahangarzadeh Rezaee, Mohammad; Nahaie, Mohammad Reza; Hasani, Akbar; Soroush Bar Haghi, Mohammad Hossein; Pormohammad, Ali; Elli Arbatan, Asghar

2015-09-01

The study aimed at assessing any association between quinolone resistance, MDR and ESBL production and their relation with the presence of integrons in Esherichia coli and Klebsiella pneumoniae. E.coli and K.pneumoniae isolated from various clinical infections were fully identified and analyzed for being quinolone resistant. These isolates were further tested for ESBL production, multi drug resistance and carriage of integrons. In total, 135 isolates were confirmed as quinolone resistant. K.pneumoniae was observed as potent ESBL producer in comparison to E.coli. Ciprofloxacin resistance in both organisms was related significantly with the presence of integron class 1, co-presence of class 1 and 2 as well as to the presence of ESBL production (p< 0.001). However, nalidixic acid resistance was related significantly (p< 0.01) with only integron class 1 and to the presence of ESBL production. Class 1 and 2 integrons were found in 73.5% of MDR isolates with 13.2% of them possessing both intI1 and intI2 genes. Prevalence of quinolone resistance together with ESBL production and MDR in E.coli and K.pneumoniae has contributed to the emergence of antibacterial resistance burden. The higher integron prevalence in our isolates advocates the potentiality of these isolates as a source for dissemination of resistance determinants.

The first occurrence of a CTX-M ESBL-producing Escherichia coli outbreak mediated by mother to neonate transmission in an Irish neonatal intensive care unit.

PubMed

O'Connor, Ciara; Philip, Roy K; Kelleher, John; Powell, James; O'Gorman, Alan; Slevin, Barbara; Woodford, Neil; Turton, Jane F; McGrath, Elaine; Finnegan, Cathriona; Power, Lorraine; O'Connell, Nuala H; Dunne, Colum P

2017-01-05

Escherichia coli (E. coli) comprise part of the normal vaginal microflora. Transfer from mother to neonate can occur during delivery resulting, sometimes, in neonatal bacterial disease. Here, we aim to report the first outbreak of CTX-M ESBL-producing E. coli with evidence of mother-to-neonate transmission in an Irish neonatal intensive care unit (NICU) followed by patient-to-patient transmission. Investigation including molecular typing was conducted. Infection was defined by clinical and laboratory criteria and requirement for antimicrobial therapy with or without positive blood cultures. Colonisation was determined by isolation without relevant symptoms or indicators of infection. Index case was an 8-day-old baby born at 34 weeks gestation who developed ESBL-producing E. coli infections at multiple body sites. Screening confirmed their mother as colonised with ESBL-producing E. coli. Five other neonates, in the NICU simultaneously with the index case, also tested positive. Of these, four were colonised while one neonate developed sepsis, requiring antimicrobial therapy. The second infected neonate's mother was also colonised by ESBL-producing E. coli. Isolates from all eight positive patients (6 neonates, 2 mothers) were compared using pulsed-field gel electrophoresis (PFGE). Two distinct ESBL-producing strains were implicated, with evidence of transmission between mothers and neonates for both strains. All isolates were confirmed as CTX-M ESBL-producers. There were no deaths associated with the outbreak. Resources were directed towards control interventions focused on hand hygiene and antimicrobial stewardship, which ultimately proved successful. Since this incident, all neonates admitted to the NICU have been screened for ESBL-producers and expectant mothers are screened at their first antenatal appointment. To date, there have been no further outbreaks.

Improvement of glucose and lipid metabolism via mung bean protein consumption: clinical trials of GLUCODIA™ isolated mung bean protein in the USA and Canada.

PubMed

Kohno, Mitsutaka; Sugano, Hideo; Shigihara, Yuhko; Shiraishi, Yoshiaki; Motoyama, Takayasu

2018-01-01

The aim of the present study was to confirm the effects of a commercially available mung bean protein isolate (GLUCODIA™) on glucose and lipid metabolism. The main component of GLUCODIA™ is 8S globulin, which constitutes 80 % of the total protein. The overall structure of this protein closely resembles soyabean β-conglycinin, which accounts for 20 % of total soya protein (soya protein isolate; SPI). Many physiological beneficial effects of β-conglycinin have been reported. GLUCODIA™ is expected to produce beneficial effects with fewer intakes than SPI. We conducted two independent double-blind, placebo-controlled clinical studies. In the first (preliminary dose decision trial) study, mung bean protein was shown to exert physiological beneficial effects when 3·0 g were ingested per d. In the second (main clinical trial) study, mung bean protein isolate did not lower plasma glucose levels, although the mean insulin level decreased with consumption of mung bean protein. The homeostatic model assessment of insulin resistance (HOMA-IR) values significantly decreased with mung bean protein. The mean TAG level significantly decreased with consumption of mung bean protein isolate. A significant increase in serum adiponectin levels and improvement in liver function enzymes were observed. These findings suggest that GLUCODIA™ could be useful in the prevention of insulin resistance and visceral fat accumulation, which are known to trigger the metabolic syndrome, and in the prevention of liver function decline.

Characterization of Staphylococcus caprae Clinical Isolates Involved in Human Bone and Joint Infections, Compared with Goat Mastitis Isolates.

PubMed

d'Ersu, J; Aubin, G G; Mercier, P; Nicollet, P; Bémer, P; Corvec, S

2016-01-01

Staphylococcus caprae is an emerging microorganism in human bone and joint infections (BJI). The aim of this study is to describe the features of S. caprae isolates involved in BJI (H for human) compared with those of isolates recovered in goat mastitis (A for animal). Fourteen isolates of each origin were included. Identifications were performed using a Vitek 2 GP ID card, tuf gene sequencing, and matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) Vitek MS. Molecular typing was carried out using pulsed-field gel electrophoresis (PFGE) and DiversiLab technology. The crystal violet method was used to determine biofilm-forming ability. Virulence factors were searched by PCR. Vitek MS technology provides an accurate identification for the two types of isolates compared to that of gold-standard sequencing (sensitivity, 96.4%), whereas the Vitek 2 GP ID card was more effective for H isolates. Molecular typing methods revealed two distinct lineages corresponding to the origin despite few overlaps: H and A. In our experimental conditions, no significant difference was observed in biofilm production ability between H and A isolates. Nine isolates (5 H isolates and 4 A isolates) behaved as weak producers while one A isolate was a strong producer. Concerning virulence factors, the autolysin atlC and the serine aspartate adhesin (sdrZ) genes were detected in 24 isolates (86%), whereas the lipase gene was always detected, except in one H isolate (96%). The ica operon was present in 23 isolates (82%). Fibrinogen-binding (fbe) or collagen-binding (cna) genes were not detected by using primers designed for Staphylococcus aureus or Staphylococcus epidermidis, even in low stringency conditions. Although S. caprae probably remains underestimated in human infections, further studies are needed to better understand the evolution and the adaptation of this species to its host. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

The incidence and clinical implication of sputum with positive acid-fast bacilli smear but negative in mycobacterial culture in a tertiary referral hospital in South Korea.

PubMed

Lee, Jae Seok; Kim, Eui-Chong; Joo, Sei Ick; Lee, Sang-Min; Yoo, Chul-Gyu; Kim, Young Whan; Han, Sung Koo; Shim, Young-Soo; Yim, Jae-Joon

2008-10-01

Although it is not rare to find sputum that is positive acid-fast bacilli (AFB) smear but subsequent culture fails to isolate mycobacteria in clinical practice, the incidence and clinical implication of those sputa from new patients has not been clearly elucidated. The aim of this study was to determine the incidence and clinical implication of sputum with positive AFB smear but negative in mycobacterial culture. All sputa that were positive AFB smear requested during diagnostic work up for new patients visiting Seoul National University Hospital from 1 January 2005 through 31 December 2006 were included. Sputa producing a positive AFB smear but negative mycobacterial culture were classified into one of four categories: laboratory failure to isolate mycobacteria, false positive AFB smear, pathogen may show a positive AFB smear other than mycobacteria, and indeterminate results. Out of 447 sputa with a positive AFB smear, 29 (6.5%) failed to culture any organism. Among these 29 sputa, 18 were caused by laboratory failure to isolate mycobacteria, six were false positive smears, and five indeterminate. Although most sputum with a positive AFB smear but negative culture could be classified as a laboratory failure, clinicians should consider the possibility of false positive AFB smear.

Fermentation products in the cystic fibrosis airways induce aggregation and dormancy-associated expression profiles in a CF clinical isolate of Pseudomonas aeruginosa

PubMed Central

Phan, Joann; Gallagher, Tara; Oliver, Andrew; England, Whitney E; Whiteson, Katrine

2018-01-01

Abstract Pseudomonas aeruginosa is a well-known dominant opportunistic pathogen in cystic fibrosis (CF) with a wide range of metabolic capacities. However, P. aeruginosa does not colonize the airways alone, and benefits from the metabolic products of neighboring cells—especially volatile molecules that can travel between different parts of the airways easily. Here, we present a study that investigates the metabolic, gene expression profiles and phenotypic responses of a P. aeruginosa clinical isolate to fermentation products lactic acid and 2,3-butanediol, metabolites that are produced by facultative anaerobic members of the CF polymicrobial community and potential biomarkers of disease progression. Although previous studies have successfully investigated the metabolic and transcriptional profiles of P. aeruginosa, most have used common lab reference strains that may differ in important ways from clinical isolates. Using transcriptomics and metabolomics with gas chromatography time of flight mass spectrometry, we observe that fermentation products induce pyocyanin production along with the expression of genes involved in P. aeruginosa amino acid utilization, dormancy and aggregative or biofilm modes of growth. These findings have important implications for how interactions within the diverse CF microbial community influence microbial physiology, with potential clinical consequences. PMID:29617986

Optimizing identification of clinically relevant Gram-positive organisms by use of the Bruker Biotyper matrix-assisted laser desorption ionization-time of flight mass spectrometry system.

PubMed

McElvania Tekippe, Erin; Shuey, Sunni; Winkler, David W; Butler, Meghan A; Burnham, Carey-Ann D

2013-05-01

Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) can be used as a method for the rapid identification of microorganisms. This study evaluated the Bruker Biotyper (MALDI-TOF MS) system for the identification of clinically relevant Gram-positive organisms. We tested 239 aerobic Gram-positive organisms isolated from clinical specimens. We evaluated 4 direct-smear methods, including "heavy" (H) and "light" (L) smears, with and without a 1-μl direct formic acid (FA) overlay. The quality measure assigned to a MALDI-TOF MS identification is a numerical value or "score." We found that a heavy smear with a formic acid overlay (H+FA) produced optimal MALDI-TOF MS identification scores and the highest percentage of correctly identified organisms. Using a score of ≥2.0, we identified 183 of the 239 isolates (76.6%) to the genus level, and of the 181 isolates resolved to the species level, 141 isolates (77.9%) were correctly identified. To maximize the number of correct identifications while minimizing misidentifications, the data were analyzed using a score of ≥1.7 for genus- and species-level identification. Using this score, 220 of the 239 isolates (92.1%) were identified to the genus level, and of the 181 isolates resolved to the species level, 167 isolates (92.2%) could be assigned an accurate species identification. We also evaluated a subset of isolates for preanalytic factors that might influence MALDI-TOF MS identification. Frequent subcultures increased the number of unidentified isolates. Incubation temperatures and subcultures of the media did not alter the rate of identification. These data define the ideal bacterial preparation, identification score, and medium conditions for optimal identification of Gram-positive bacteria by use of MALDI-TOF MS.

Optimizing Identification of Clinically Relevant Gram-Positive Organisms by Use of the Bruker Biotyper Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry System

PubMed Central

McElvania TeKippe, Erin; Shuey, Sunni; Winkler, David W.; Butler, Meghan A.

2013-01-01

Matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) can be used as a method for the rapid identification of microorganisms. This study evaluated the Bruker Biotyper (MALDI-TOF MS) system for the identification of clinically relevant Gram-positive organisms. We tested 239 aerobic Gram-positive organisms isolated from clinical specimens. We evaluated 4 direct-smear methods, including “heavy” (H) and “light” (L) smears, with and without a 1-μl direct formic acid (FA) overlay. The quality measure assigned to a MALDI-TOF MS identification is a numerical value or “score.” We found that a heavy smear with a formic acid overlay (H+FA) produced optimal MALDI-TOF MS identification scores and the highest percentage of correctly identified organisms. Using a score of ≥2.0, we identified 183 of the 239 isolates (76.6%) to the genus level, and of the 181 isolates resolved to the species level, 141 isolates (77.9%) were correctly identified. To maximize the number of correct identifications while minimizing misidentifications, the data were analyzed using a score of ≥1.7 for genus- and species-level identification. Using this score, 220 of the 239 isolates (92.1%) were identified to the genus level, and of the 181 isolates resolved to the species level, 167 isolates (92.2%) could be assigned an accurate species identification. We also evaluated a subset of isolates for preanalytic factors that might influence MALDI-TOF MS identification. Frequent subcultures increased the number of unidentified isolates. Incubation temperatures and subcultures of the media did not alter the rate of identification. These data define the ideal bacterial preparation, identification score, and medium conditions for optimal identification of Gram-positive bacteria by use of MALDI-TOF MS. PMID:23426925

Metal Accumulation and Vanadium-Induced Multidrug Resistance by Environmental Isolates of Escherichia hermannii and Enterobacter cloacae

PubMed Central

Hernández, Alicia; Mellado, Rafael P.; Martínez, José L.

1998-01-01

Contaminated soils from an oil refinery were screened for the presence of microorganisms capable of accumulating either nickel, vanadium, or both metals. Three strains of bacteria that belonged to the family Enterobacteriaceae were selected. Two of them were Escherichia hermannii strains, and outer membrane profile (OMP) analysis showed that they were similar to a strain of clinical origin; the other one was an Enterobacter cloacae strain that differed from clinical isolates. The selected bacteria accumulated both nickel and vanadium. Growth in the presence of vanadium induced multidrug resistance phenotypes in E. hermannii and E. cloacae. Incubation with this metal changed the OMP profile of E. hermannii but did not produce variations in the expression of the major OMPs of E. cloacae. PMID:9797283

Pirbenicillin: Comparison with Carbenicillin and BL-P1654, Alone and with Gentamicin, Against Pseudomonas aeruginosa

PubMed Central

Lopez, Carlos E.; Standiford, Harold C.; Tatem, Beverly A.; Calia, Frank M.; Schimpff, Stephen C.; Snyder, Merrill J.; Hornick, Richard B.

1977-01-01

Minimum inhibitory concentrations (MIC) of pirbenicillin against 135 clinical isolates of Pseudomonas aeruginosa were one-fourth of those required for carbenicillin but two times higher than those for BL-P1654. Increasing the inoculum size produced an adverse effect on the bactericidal activity for all three antibiotics. This was more apparent for pirbenicillin than for carbenicillin, but less than the effect on BL-P1654. When concentrations of antibiotics likely to be achieved clinically were used, gentamicin increased the inhibitory and bactericidal effects of all three semisynthetic penicillins for the majority of isolates. Strains highly resistant to the aminoglycoside antibiotic, however, were inhibited no more by the penicillin-gentamicin combinations than by the most effective of the antibiotics alone. PMID:404963

Development of novel agar media for isolating guaiacol producing Alicyclobacillus spp.

PubMed

Chang, S S; Park, S H; Kang, D H

2013-06-03

The purpose of this study is to develop a selective and differential medium (SK2 agar) for isolating guaiacol producing Alicyclobacillus. Forty-one selected dyes and vanillic acid were incorporated in SK agar for screening selective and differential agents. Two guaiacol producing (1016, 1101) and two non-guaiacol producing (19220, C-GD 1-1) Alicyclobacillus isolates were streaked onto media and color differentiation of the isolates was assessed. Among 41 tested dyes, Chrome Azurol S (CAS) allowed color differentiation of the two types of Alicyclobacillus. Colonies of guaiacol producing Alicyclobacillus isolates appeared as dark purple to royal blue color with yellow background, whereas non-guaiacol producing Alicyclobacillus isolates produced cream colored colonies with yellow background. Vanillic acid not only served as a precursor for guaiacol formation but also inhibited non-guaiacol producing Alicyclobacillus. Non-guaiacol producing isolates did not grow on SK agar containing more than 70 ppm vanillic acid, whereas the recovery of guaiacol producing isolates was unaffected. When compared with other Alicyclobacillus isolation media, not only was SK2 agar capable of selectively recovering guaiacol-producing Alicyclobacillus, the degree of growth was also approximately equal if not better than orange serum agar, potato dextrose agar, and K agar. The development of SK2 agar provides the fruit juice industry with an inexpensive, simple to use alternative for the detection of guaiacol producing Alicyclobacillus. Copyright © 2013 Elsevier B.V. All rights reserved.

Stenotrophomonas maltophilia isolated from patients exposed to invasive devices in a university hospital in Argentina: molecular typing, susceptibility and detection of potential virulence factors.

PubMed

Alcaraz, Eliana; Garcia, Carlos; Papalia, Mariana; Vay, Carlos; Friedman, Laura; Passerini de Rossi, Beatriz

2018-05-25

The aim of this work was to investigate the presence of selected potential virulence factors, susceptibility and clonal relatedness among 63 Stenotrophomonas maltophilia isolates recovered from patients exposed to invasive devices in a university hospital in Argentina between January 2004 and August 2012. Genetic relatedness was assessed by enterobacterial repetitive intergenic consensus PCR (ERIC-PCR) and pulsed-field gel electrophoresis (PFGE). Isolates were characterized by antimicrobial resistance, the presence and/or expression of potential virulence determinants, and virulence in the Galleria mellonella model.Results/Key findings. ERIC-PCR generated 52 fingerprints, and PFGE added another pattern. Resistance to trimethoprim-sulfamethoxazole (6.35 %), levofloxacin (9.52 %) and ciprofloxacin (23.80 %) was detected. All isolates were susceptible to minocycline. All isolates were lipase, protease and siderophore producers, while all but Sm61 formed biofilms. However, 11/63 isolates did not amplify the major extracellular protease-coding gene (stmPr1). Sm61 is an stmPr1-negative isolate, and showed (as did Sm13 and the reference strain K279a) strong proteolysis and siderophore production, and high resistance to hydrogen peroxide. The three isolates were virulent in the G. mellonella model, while Sm10, a low-resistance hydrogen peroxide stmPr1-negative isolate, and weak proteolysis and siderophore producer, was not virulent. This is the first epidemiological study of the clonal relatedness of S. maltophilia clinical isolates in Argentina. Great genomic diversity was observed, and only two small clusters of related S. maltophilia types were found. Minocycline and trimethoprim-sulfamethoxazole were the most active agents. S. maltophilia virulence in the G. mellonella model is multifactorial, and further studies are needed to elucidate the role of each potential virulence factor.

Secondary metabolites produced by marine streptomyces as antibiofilm and quorum-sensing inhibitor of uropathogen Proteus mirabilis.

PubMed

Younis, Khansa Mohammed; Usup, Gires; Ahmad, Asmat

2016-03-01

Quorum-sensing regulates bacterial biofilm formation and virulence factors, thereby making it an interesting target for attenuating pathogens. In this study, we investigated anti-biofilm and anti-quorum-sensing compounds from secondary metabolites of halophiles marine streptomyces against urinary catheter biofilm forming Proteus mirabilis without effect on growth viability. A total of 40 actinomycetes were isolated from samples collected from different places in Iraq including marine sediments and soil samples. Fifteen isolates identified as streptomyces and their supernatant screened as anti-quorum-sensing by inhibiting quorum-sensing regulated prodigiosin biosynthesis of Serratia marcescens strain Smj-11 as a reporter strain. Isolate Sediment Lake Iraq (sdLi) showed potential anti-quorum-sensing activity. Out of 35 clinical isolates obtained from Urinary catheter used by patient at the Universiti Kebangsaan Malaysia Medical Center, 22 isolates were characterized and identified as Proteus mirabilis. Isolate Urinary Catheter B4 (UCB4) showed the highest biofilm formation with highest resistance to used antibiotic and was chosen for further studies. Ethyl acetate secondary metabolites extract was produced from sdLi isolate. First, we determined the Minimum Inhibitory Concentration (MIC) of sdLi crude extract against UCB4 isolate, and all further experiments used concentrations below the MIC. Tests of subinhibitory concentrations of sdLi crude extract showed good inhibition against UCB4 isolate biofilm formation on urinary catheter and cover glass using Scanning electron microscopy and light microscopy respectively. The influence of sub-MIC of sdLi crude extract was also found to attenuate the quorum sensing (QS)-dependent factors such as hemolysin activity, urease activity, pH value, and motility of UCB4 isolate. Evidence is presented that these nontoxic secondary metabolites may act as antagonists of bacterial quorum sensing by competing with quorum-sensing signals for receptor binding.

Evaluation of a surface plasmon resonance imaging-based multiplex O-antigen serogrouping for Escherichia coli using eleven major serotypes of Shiga -toxin-producing E. coli.

PubMed

Nakano, Satoshi; Nagao, Miki; Yamasaki, Tomomi; Morimura, Hiroyuki; Hama, Natsuki; Iijima, Yoshio; Shinomiya, Hiroto; Tanaka, Michio; Yamamoto, Masaki; Matsumura, Yasufumi; Miyake, Shiro; Ichiyama, Satoshi

2018-06-01

The early detection of Shiga toxin-producing Escherichia coli (STEC) is important for early diagnosis and preventing the spread of STEC. Although the confirmatory test for STEC should be based on the detection of Shiga toxin using molecular analysis, isolation permits additional characterization of STEC using a variety of methods, including O:H serotyping. The conventional slide agglutination O-antigen serogrouping used in many clinical laboratories is laborious and time-consuming. Surface plasmon resonance (SPR)-based immunosensors are commonly used to investigate a large variety of bio-interactions such as antibody/antigen, peptide/antibody, DNA/DNA, and antibody/bacteria interactions. SPR imaging (SPRi) is characterized by multiplexing capabilities for rapidly screening (approximately 100 to several hundred sensorgrams in parallel) molecules. SPRi-based O-antigen serogrouping method for STEC was recently developed by detecting the interactions between O-antigen-specific antibodies and bacterial cells themselves. The aim of this study was to evaluate its performance for E. coli serogrouping using clinical STEC isolates by comparing the results of slide agglutination tests. We tested a total of 188 isolates, including O26, O45, O91, O103, O111, O115, O121, O128, O145, O157, and O159. The overall sensitivity of SPRi-based O-antigen serogrouping was 98.9%. Only two O157 isolates were misidentified as nontypeable and O121. The detection limits of all serotypes were distributed between 1.1 × 10 6 and 17.6 × 10 6  CFU/ml. Pulsed-field gel electrophoresis (PFGE) revealed the heterogeneity of the examined isolates. In conclusion, SPRi is a useful method for the O-antigen serogrouping of STEC isolates, but the further evaluation of non-O157 minor serogroups is needed. Copyright © 2018 Japanese Society of Chemotherapy and The Japanese Association for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

Rapid lysostaphin test to differentiate Staphylococcus and Micrococcus species.

PubMed Central

Geary, C; Stevens, M

1986-01-01

A rapid, simple lysostaphin lysis susceptibility test to differentiate the genera Staphylococcus and Micrococcus was evaluated. Of 181 strains from culture collections, 95 of 95 Staphylococcus strains were lysed, and 79 of 79 Micrococcus strains were not lysed. The seven Planococcus strains were resistant. Clinical isolates (890) were tested with lysostaphin and for the ability to produce acid from glycerol in the presence of erythromycin. Overall agreement between the methods was 99.2%. All clinical Micrococcus strains (43) were resistant to lysostaphin, and all clinical Staphylococcus strains (847) were susceptible. Seven of the Staphylococcus strains did not produce acid from glycerol in the presence of erythromycin. This lysostaphin test provides results in 2 h. It is easier to perform than previously described lysostaphin lysis methods. It is also more rapid and accurate than the glycerol-erythromycin test. PMID:3519667

First report of OXA-143-lactamase producing Acinetobacter baumannii in Qom, Iran.

PubMed

Sarikhani, Zohreh; Nazari, Razieh; Nateghi Rostami, Mahmoud

2017-11-01

Antibiotic resistance in Acinetobacter baumannii and outbreaks caused by this organism have been reported from several areas of the world. The present study aimed at determining the antibiotic susceptibility profiles and the distribution of OXA-type beta-lactamases among Iranian Acinetobacter baumannii isolates from Qom of Iran. For this study, 108 non-duplicate A. baumannii isolates were obtained from clinical specimens in four teaching hospitals in Qom in the central of Iran. The antimicrobial susceptibility of isolates was tested by standard disk diffusion and prevalence of bla OXA genes was investigated by PCR method. Among 97 carbapenem non-susceptible isolates of A. baumannii , 90.72% (88 isolates) isolates showed extensive drug resistance to multiple antibiotics. Among carbapenem resistant isolates, 100% carried bla OXA-51-like , 82.47% carried bla OXA-23-like , 55.67% carried bla OXA-58-like , 22.68% carried bla OXA-40-like and 14.43% had bla OXA-143-like resistance genes. This study demonstrated high genetic diversity of OXA genes among isolates of A. baumannii in Qom, Iran.

First report of OXA-143-lactamase producing Acinetobacter baumannii in Qom, Iran

PubMed Central

Sarikhani, Zohreh; Nazari, Razieh; Nateghi Rostami, Mahmoud

2017-01-01

Objective(s): Antibiotic resistance in Acinetobacter baumannii and outbreaks caused by this organism have been reported from several areas of the world. The present study aimed at determining the antibiotic susceptibility profiles and the distribution of OXA-type beta-lactamases among Iranian Acinetobacter baumannii isolates from Qom of Iran. Materials and Methods: For this study, 108 non-duplicate A. baumannii isolates were obtained from clinical specimens in four teaching hospitals in Qom in the central of Iran. The antimicrobial susceptibility of isolates was tested by standard disk diffusion and prevalence of bla OXA genes was investigated by PCR method. Results: Among 97 carbapenem non-susceptible isolates of A. baumannii, 90.72% (88 isolates) isolates showed extensive drug resistance to multiple antibiotics. Among carbapenem resistant isolates, 100% carried blaOXA-51-like, 82.47% carried blaOXA-23-like, 55.67% carried blaOXA-58-like, 22.68% carried blaOXA-40-like and 14.43% had blaOXA-143-like resistance genes. Conclusion: This study demonstrated high genetic diversity of OXA genes among isolates of A. baumannii in Qom, Iran. PMID:29299207

Clinical Manifestations and Molecular Characterization of Pertactin-Deficient and Pertactin-Producing Bordetella pertussis in Children, Philadelphia 2007-2014.

PubMed

Vodzak, Jennifer; Queenan, Anne Marie; Souder, Emily; Evangelista, Alan T; Long, Sarah S

2017-01-01

 Bordetella pertussis strains lacking expression of pertactin, a bacterial adhesin and vaccine target, are emerging. There are limited data on disease manifestations of mutant strains in children. We sought to compare clinical manifestations of pertactin-deficient and pertactin-producing B. pertussis infection in infants and describe corresponding molecular characteristics.  Molecular characterization of archived B. pertussis isolates (collected January 2007 to March 2014) included Western blot analysis, pulsed-field gel electrophoresis (PFGE), polymerase chain reaction, and pertactin gene sequencing. Medical record review compared epidemiologic and clinical courses of pertactin-producing and pertactin-deficient B. pertussis infections.  Sixty of 72 B. pertussis isolates were viable for analysis. Within the cohort of infants, the median age was 95 days, 90% received ≤1 dose of vaccine, and 72% were hospitalized. Pertactin deficiency was first noted in 2008, and its prevalence increased over time (68% overall prevalence). There were no statistically significant differences in presenting symptoms or signs, hospitalization, intensive care, respiratory support, or laboratory results related to pertactin expression. Illness length was shorter in pertactin-deficient group (mean difference, 3.2 days; P = .04); no difference was noted in the subgroup of infants <4 months old. Molecular analyses identified 11 PFGE profiles (Centers for Disease Control and Prevention profile No. 002 predominant, 47%). In 41 pertactin-deficient strains, sequencing identified 2 stop codon and 3 IS481 locations disrupting the prn gene. Mutations and nucleotide positions were not unique to PFGE type, nor were they clustered in time.  In this cohort of predominantly unimmunized infants, clinical disease did not differ between infection with pertactin-deficient and those with pertactin-producing B. pertussis. Molecular analyses demonstrated remarkable PFGE strain diversity, with multiple mechanisms and molecular sites of pertactin inactivation. © The Author 2016. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail journals.permissions@oup.com.

Plasmid curing analysis of antibiotic resistance in beta-lactamase producing Staphylococci from wounds and burns patients.

PubMed

Ojo, S K S; Sargin, B O; Esumeh, F I

2014-01-01

Hospitals worldwide are facing unprecedented crisis due to increasingly rapid emergence and dissemination of antimicrobial resistant staphylococci in wounds and burns and its environs via plasmid mediation. This study was conducted to evaluate the plasmid-mediated or chromosomal-mediated resistance in staphylococci. One hundred clinical swabs from wounds and burns patients were demonstrated for presence of staphylococci using mannitol salt agar. Various biochemical, DNase and beta-lactamase test was carried out and the plasmid curing assay was demonstrated using 0.1 mg mL(-1) acridine orange on antibiotic resistant isolates. The results revealed S. aureus (47) and coagulase negative staphylococci (CoNS) (6). beta-lactamase producing species of S. aureus were 14 and CoNS was 1. Most isolates showed high resistance pattern to gentamicin, ciprofloxacin, norfloxacin, rifampicin, chloramphenicol, ampiclox and others. The antibiotic resistance isolates were highly indicative ofplasmid-borne and few are chromosomal-borne after the plasmid curing analysis. The plasmid-mediated resistance observed among various antibiotics poses difficulty in treatment for clinicians. This high plasmid-mediated resistance among the isolates and from other studies calls for an urgent surveillance and epidemiological studies to infection control.

Endophytic Paraconiothyrium sp. from Zingiber officinale Rosc. Displays Broad-Spectrum Antimicrobial Activity by Production of Danthron.

PubMed

Anisha, C; Sachidanandan, P; Radhakrishnan, E K

2018-03-01

The bioactivity spectrum of fungal endophytes isolated from Zingiber officinale was analyzed against clinical pathogens and against the phytopathogen Pythium myriotylum, which causes Pythium rot in ginger. One of the isolates GFM13 showed broad bioactivity against various pathogens tested including P. myriotylum. The spore suspension as well as the culture filtrate of the endophytic fungal isolate was found to effectively protect ginger rhizomes from Pythium rot. By molecular identification, the fungal endophyte was identified as Paraconiothyrium sp. The bioactive compound produced by the isolate was separated by bioactivity-guided fractionation and was identified by GC-MS as danthron, an anthraquinone derivative. PCR amplification showed the presence of non-reducing polyketide synthase gene (NR-PKS) in the endophyte GFM13, which is reported to be responsible for the synthesis of anthraquinones in fungi. This is the first report of danthron being produced as the biologically active component of Paraconiothyrium sp. Danthron is reported to have wide pharmaceutical and agronomic applications which include its use as a fungicide in agriculture. The broad-spectrum antimicrobial activity of danthron and the endophytic origin of Paraconiothyrium sp. offer immense applications of the study.

Genotyping by randomly amplified polymorphic DNA of bacteriocin producing Lactobacillus acidophilus strains from Nigeria.

PubMed

Alli, John Adeolu; Iwalokun, Bamidele A; Oluwadun, Afolabi; Okonko, Iheanyi Omezuruike

2015-01-01

Yogurt and starter culture producers are still searching strains of Lactobacillus acidophilus to produce healthier yogurt with a longer shelf life and better texture, taste, and quality. This study determined the genotyping of bacteriocin producing Lactobacillus acidophilus strains recovered from Nigerian yogurts. Yogurt samples were collected from four different states of South West regions of Nigeria. Isolates were obtained from MRS Medium and biochemically characterized. This was further confirmed by API50CH. The bacteriocin positivity and activity was determined. Genomic characterization of our Lactobacillus acidophilus strains was done with randomly amplified polymorphic DNA-PCR. All yogurt samples containing Lactobacillus acidophilus strains meet the probiotic requirement of ≥10(6) cfu/mL. The gel picture revealed 6 RAPD clonal types of Lactobacillus acidophilus strains with RAPD type C observed to be more common. Significant differences existed in the mean growth inhibition zone (t = -7.32, P < 0.05 for E. coli ATCC; t = -6.19, P < 0.05 for E. coli clinical isolates; t = -6.16, P < 0.05 for Enterobacter sp; t = -11.92, P < 0.05 for Salmonella typhi, t = -1.10, P > 0.05 Staphylococcus aureus). No correlation between the bacteriocin production, activity, and their RAPD clonal division (X(2) = 7.49, P = 0.1610, df = 5). In conclusion, L. acidophilus isolated in Nigeria samples met the probiotic requirements of ≥10(6) cfu/mL and produce bacteriocins with good spectrum of activity.

Cryptococcus gattii VGIII Isolates Causing Infections in HIV/AIDS Patients in Southern California: Identification of the Local Environmental Source as Arboreal

PubMed Central

Springer, Deborah J.; Billmyre, R. Blake; Filler, Elan E.; Voelz, Kerstin; Pursall, Rhiannon; Mieczkowski, Piotr A.; Larsen, Robert A.; Dietrich, Fred S.; May, Robin C.; Filler, Scott G.; Heitman, Joseph

2014-01-01

Ongoing Cryptococcus gattii outbreaks in the Western United States and Canada illustrate the impact of environmental reservoirs and both clonal and recombining propagation in driving emergence and expansion of microbial pathogens. C. gattii comprises four distinct molecular types: VGI, VGII, VGIII, and VGIV, with no evidence of nuclear genetic exchange, indicating these represent distinct species. C. gattii VGII isolates are causing the Pacific Northwest outbreak, whereas VGIII isolates frequently infect HIV/AIDS patients in Southern California. VGI, VGII, and VGIII have been isolated from patients and animals in the Western US, suggesting these molecular types occur in the environment. However, only two environmental isolates of C. gattii have ever been reported from California: CBS7750 (VGII) and WM161 (VGIII). The incongruence of frequent clinical presence and uncommon environmental isolation suggests an unknown C. gattii reservoir in California. Here we report frequent isolation of C. gattii VGIII MATα and MAT a isolates and infrequent isolation of VGI MATα from environmental sources in Southern California. VGIII isolates were obtained from soil debris associated with tree species not previously reported as hosts from sites near residences of infected patients. These isolates are fertile under laboratory conditions, produce abundant spores, and are part of both locally and more distantly recombining populations. MLST and whole genome sequence analysis provide compelling evidence that these environmental isolates are the source of human infections. Isolates displayed wide-ranging virulence in macrophage and animal models. When clinical and environmental isolates with indistinguishable MLST profiles were compared, environmental isolates were less virulent. Taken together, our studies reveal an environmental source and risk of C. gattii to HIV/AIDS patients with implications for the >1,000,000 cryptococcal infections occurring annually for which the causative isolate is rarely assigned species status. Thus, the C. gattii global health burden could be more substantial than currently appreciated. PMID:25144534

[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 Medicine are multidrug-resistant, and they mainly produce type B and D carbapenemases.

Iodometric and Molecular Detection of ESBL Production Among Clinical Isolates of E. coli Fingerprinted by ERIC-PCR: The First Egyptian Report Declares the Emergence of E. coli O25b-ST131clone Harboring blaGES.

PubMed

El-Badawy, Mohamed F; Tawakol, Wael M; Maghrabi, Ibrahim A; Mansy, Moselhy S; Shohayeb, Mohamed M; Ashour, Mohammed S

2017-09-01

The extensive use of β-lactam antibiotics has led to emergence and spread of extended-spectrum β-lactamases (ESBLs). This study was conducted to investigate the prevalence of 7 different ESBL genes (bla TEM , bla SHV , bla CTX-M , bla VEB , bla PER , bla GES , and bla OXA-10 ) and O25b-ST131 high-risk clone among 61 clinical isolates of Escherichia coli. Also, one broad-spectrum β-lactamase (bla OXA-1 ) was investigated. This study was also constructed to evaluate iodometric overlay method in detection of ESBL production. Phenotypic identification of E. coli isolates using API 20E revealed 18 distinct biotypes. DNA fingerprinting using enterobacterial repetitive intergenic consensus polymerase chain reaction (ERIC-PCR) differentiated all isolates into 2 main phylogenetic groups with 60 distinct genetic profiles. Elevated values of minimal inhibitory concentration (MIC) 50 and MIC 90 for third- and fourth-generation cephalosporins were observed. Phenotypic tests revealed that 85.24% of isolates were ESBL producers. The incidence rates of bla TEM , bla SHV , bla CTX-M , bla GES , bla OXA-1 , and bla OXA-10 among E. coli ESBL producer phenotype were 69.23%, 25%, 96.15%, 3.85%, 11.54%, and 48%, respectively. On the other hand, bla VEB and bla PER were not detected. Sequencing of bla TEM and bla SHV revealed that bla TEM-214 and bla SHV-11 were the most prevalent variants. Group characterization of bla CTX-M revealed that bla CTX-M-1 was the most prevalent group of bla CTX-M family. It was found that 30.77% of E. coli ESBL producers belonged to O25b-ST131 clone harboring bla CTX-M-15 . This study concluded that iodometric overlay method was 100% sensitive in detection of ESBL production. To our knowledge, this is the first Egyptian study that declares the emergence of E. coli O25b-ST131 harboring bla GES .

Biofilm Formation and Its Relationship with the Molecular Characteristics of Food-Related Methicillin-Resistant Staphylococcus aureus (MRSA).

PubMed

Vergara, Alberto; Normanno, Giovanni; Di Ciccio, Pierluigi; Pedonese, Francesca; Nuvoloni, Roberta; Parisi, Antonio; Santagada, Gianfranco; Colagiorgi, Angelo; Zanardi, Emanuela; Ghidini, Sergio; Ianieri, Adriana

2017-10-01

The capability to produce biofilm is an important persistence and dissemination mechanism of some foodborne bacteria. This paper investigates the relationship between some molecular characteristics (SCCmec, ST, spa-type, agr-type, cna, sarA, icaA, icaD, clfA, fnbA, fnbB, hla, hlb) of 22 food-related methicillin-resistant Staphylococcus aureus (MRSA) strains and their ability to form biofilm on stainless steel and polystyrene. Five (22.7%, 5/22) strains were able to synthesize biofilm on polystyrene, and one of these (4.5%, 1/22) strains was also able to synthesize biofilm on stainless steel. The largest amount of biofilm was formed on polystyrene by 2 MRSA strains isolated from cows' milk, thus raising concern about the dairy industry. The majority of MRSA biofilm producers carried SCCmec type IVa, suggesting that the presence of SCCmecIVa and/or agr type III could be related to the ability to form biofilm. In conclusion, in order to achieve an acceptable level of food safety, Good Hygiene Practices should be strictly implemented along the food chain to reduce the risk of colonization and dissemination of MRSA biofilm-producing strains in the food industry. In this study, some assayed isolates of food-related MRSA demonstrated the capacity to form biofilm. Biofilm formation differed according to surface characteristics and MRSA strains. A relationship was observed between some molecular characteristics and the ability to form biofilms. Few studies have investigated the ability of MRSA to form biofilms, and the majority of these studies have investigated clinical aspects. This work was performed to investigate whether or not there is a difference between MRSA food isolates and MRSA clinical isolates in their ability to form biofilm. These initial findings could provide information that will contribute to a better understanding of these aspects. © 2017 Institute of Food Technologists®.

Clinical and microbiologic characteristics of adult patients with recurrent bacteraemia caused by extended-spectrum β-lactamase-producing Escherichia coli or Klebsiella pneumoniae.

PubMed

Lee, C-H; Su, L-H; Chen, F-J; Tang, Y-F; Chien, C-C; Liu, J-W

2015-12-01

The characteristics of patients with recurrent bacteraemia caused by extended-spectrum β-lactamase (ESBL)-producing Escherichia coli or Klebsiella pneumoniae (EK) are rarely described. Flomoxef belongs to the cephamycins group and demonstrates in vitro activity against ESBL-producing organisms. Whether flomoxef may be used for the treatment of such infections remains controversial. This retrospective case-control study enrolled adult patients who had bacteraemia caused by ESBL-EK during 2005-2011. Case patients were those who had more than one episode of ESBL-EK bacteraemia. Controls were those who were matched for age and interval time of blood sampling and had only one episode of ESBL-EK bacteraemia with subsequent bacteraemia episodes caused by other non-ESBL-EK bacteria. Pulsed-field gel electrophoresis and microbiologic profiles of the initial and subsequent ESBL-EK isolates were analysed. During the study period, 424 patients were found to have at least one positive blood culture after the first ESBL-EK bacteraemia episode, and 67 (15.8%) had a second episode of ESBL-EK bacteraemia. Bacteraemia resulting from vascular catheter-related infection (odds ratio, 3.24; 95% confidence interval, 1.31-8.05), and definitive therapy with flomoxef (odds ratio, 2.99; 95% confidence interval, 1.10-8.15) were both independent risk factors for the recurrence. Among the 56 patients with available ESBL-EK isolates for analysis, 38 (67.8%) were infected by genetically similar strains. In three of these 38 recurrent ESBL-EK bacteraemia cases caused by an identical strain, the minimum inhibitory concentrations of carbapenem for the subsequent K. pneumoniae isolates were fourfold or higher than the initial isolates. Recurrent bacteraemia was not uncommon in our patients with ESBL-EK bacteraemia, and most of the episodes were caused by identical strains. Copyright © 2015 European Society of Clinical Microbiology and Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

Collateral damage of flomoxef therapy: in vivo development of porin deficiency and acquisition of blaDHA-1 leading to ertapenem resistance in a clinical isolate of Klebsiella pneumoniae producing CTX-M-3 and SHV-5 beta-lactamases.

PubMed

Lee, Chen-Hsiang; Chu, Chishih; Liu, Jien-Wei; Chen, Yi-Shung; Chiu, Chiung-Jung; Su, Lin-Hui

2007-08-01

The study aimed to characterize the genetic basis of flomoxef and collateral ertapenem resistance in a clinical isolate of extended-spectrum beta-lactamase-producing Klebsiella pneumoniae (ESBL-KP) after flomoxef exposure. Four ESBL-KP isolates (Lkp11-14) were recovered sequentially from four episodes of bacteraemia in an elderly patient. Laboratory investigations included genotyping by PFGE, resistance gene analysis by PCR and sequencing, and outer membrane protein analysis by SDS-PAGE. Plasmid analysis by DNA-DNA hybridization, electroporation and conjugation was also performed. Lkp14 was recovered after 21 days of flomoxef therapy. It demonstrated an indistinguishable PFGE pattern when compared with those produced by Lkp11-13. However, resistance to both flomoxef and ertapenem emerged in Lkp14. Molecular characterization revealed that, in addition to the pre-existing ESBL production (CTX-M-3 and SHV-5) and OmpK35 deficiency found in Lkp11-13, Lkp14 had acquired an extra plasmid-mediated AmpC beta-lactamase gene (blaDHA-1) and failed to express OmpK36, because of insertional inactivation by an insertion sequence IS5. Other resistance mechanisms, such as production of carbapenem-hydrolysing enzymes or expression of chromosomal efflux, were apparently not involved. Conjugational transfer of the plasmid-mediated blaDHA-1 gene into Lkp11 resulted in a significant increase in the MICs of cephamycins and beta-lactamase inhibitors, but not in those of carbapenems. Lkp14 was apparently derived from the previously flomoxef-susceptible isolates, Lkp11-13. After flomoxef exposure, the in vivo acquisition of the plasmid-mediated blaDHA-1 gene has led to flomoxef resistance in Lkp14, and the concomitant depletion of OmpK36 expression has resulted in a collateral effect of ertapenem resistance and diminished susceptibilities to imipenem and meropenem.

A multinational study of colonization with extended spectrum β-lactamase-producing Enterobacteriaceae in healthcare personnel and family members of carrier patients hospitalized in rehabilitation centres.

PubMed

Adler, A; Baraniak, A; Izdebski, R; Fiett, J; Salvia, A; Samso, J V; Lawrence, C; Solomon, J; Paul, M; Lerman, Y; Schwartzberg, Y; Mordechai, E; Rossini, A; Fierro, J; Lammens, C; Malhotra-Kumar, S; Goossens, H; Hryniewicz, W; Brun-Buisson, C; Gniadkowski, M; Carmeli, Y

2014-08-01

The study aims were: (i) to define the prevalence of and risk factors for colonization by extended spectrum β-lactamase (ESBL) -producing Enterobacteriaceae (EPE) among healthcare workers (HCWs) and family members (FMs) of EPE-colonized patients in rehabilitation units and (ii) to compare EPE isolates from these three groups. The study included 286 FMs of 194 EPE-carrying patients identified in five rehabilitation units located in Israel, Italy, France and Spain. The EPE were detected in rectal swabs from 26 (9%) of 286 FMs screened. In multivariate analyses, older age of FM, greater mean number of hours spent with the patient, being a daughter or a female spouse of a patient, and chronic lung disease of the patient were significantly associated with carriage in the FM. Escherichia coli was the most common organism (76%), followed by Klebsiella pneumoniae (19%). Isolates were typed by pulsed field gel electrophoresis and multilocus sequence typing, and ESBLs were identified by PCR sequencing. A comparison of paired species isolates from FMs and their respective patient showed that 17 of 23 strains were indistinguishable. EPE were detected in 35 (3.5%, E. coli = 34) of the 1001 HCWs screened. Feeding patients was associated with EPE carriage by HCWs. Only 7 of 23 E. coli subclones cultured from HCWs were also represented among 376 patient-derived ESBL-producing E. coli isolates from the same rehabilitation units. In Spain, a higher proportion of HCWs and FMs were ESBL carriers than elsewhere (p <0.05). In conclusion, the molecular and epidemiological data suggest that FMs are at higher risk of EPE acquisition from their relative patients than HCWs. © 2014 The Authors Clinical Microbiology and Infection © 2014 European Society of Clinical Microbiology and Infectious Diseases.

Towards proteomic species barcoding of fungi - An example using Scedosporium/Pseudallescheria complex isolates.

PubMed

Bernhard, Mareike; Zautner, Andreas Erich; Steinmann, Joerg; Weig, Michael; Groß, Uwe; Bader, Oliver

2016-02-01

MALDI-ToF mass spectrometry offers fast and reliable species identification for bacteria and yeasts under clinical routine conditions. Here, we produced mass spectra for identification of clinically important species of the Pseudallescheria/Scedosporium complex using the recently suggested new nomenclature and use this example to discuss to what extent the principle of DNA barcoding might be transferred to mass spectrometry. Copyright © 2015 The British Mycological Society. Published by Elsevier Ltd. All rights reserved.

Molecular mechanisms of membrane impermeability in clinical isolates of Enterobacteriaceae exposed to imipenem selective pressure.

PubMed

Pavez, Monica; Vieira, Camila; de Araujo, Maria Rita; Cerda, Alvaro; de Almeida, Lara Mendes; Lincopan, Nilton; Mamizuka, Elsa Masae

2016-07-01

Intrinsic mechanisms leading to carbapenem-induced membrane impermeability and multidrug resistance are poorly understood in clinical isolates of Enterobacteriaceae. In this study, molecular behaviours during the establishment of membrane impermeability in members of the Enterobacteriaceae family under imipenem selective pressure were investigated. Clinical isolates (n = 22) exhibiting susceptibility to multiple antibiotics or characterised as extended-spectrum β-lactamase (ESBL)- or AmpC-producers were submitted to progressive passages on Mueller-Hinton agar plates containing subclinical concentrations of imipenem [0.5 × the minimum inhibitory concentration (MIC)]. Changes in outer membrane permeability were evaluated by determination of antimicrobial MICs, sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE), and gene expression analysis related to membrane permeability (i.e. omp35-like, omp36-like and acrA) and regulatory mechanisms (i.e. marA and ompR) by quantitative reverse transcription PCR. Following imipenem induction, 73% of isolates showed increased carbapenem MICs by ≥2 doubling dilutions. At an early stage of treatment, imipenem modulated the expression of porins and efflux pump genes, represented by a reduction of 78% in omp36-like and a two-fold increase in acrA expression. Transcriptional factors marA and ompR were also affected by imipenem induction, increasing mRNA expression by 14- and 4-fold, respectively. High marA expression levels were associated with higher values of acrA expression. These results suggest that imipenem is an important factor in the development of an adaptive response to carbapenems by regulating key genes involved in the control of efflux pumps and porins, which could lead to a multidrug-resistant profile in clinical isolates, contributing to possible treatment failure. Copyright © 2016 Elsevier B.V. and International Society of Chemotherapy. All rights reserved.

Community-onset extended-spectrum-β-lactamase-producing Escherichia coli sequence type 131 at two Korean community hospitals: The spread of multidrug-resistant E. coli to the community via healthcare facilities.

PubMed

Kim, Young Ah; Kim, Jin Ju; Kim, Heejung; Lee, Kyungwon

2017-01-01

The recent molecular epidemiology of ESBL-producing Escherichia coli infection in two Korean community hospitals was evaluated in this prospective observational study. We collected non-duplicated E. coli isolates from consecutive, sequentially encountered patients with community-onset episodes between March and April 2016 in two community hospitals in Gyeonggi-do province, Korea. We studied the prevalence, clinical characteristics and molecular epidemiology of E. coli sequence type 131 (ST131) isolated from the community. From a total of 213 E. coli isolates collected from the community, 94 (44.1%) were community-onset healthcare-associated isolates and 119 (55.9%) were community-associated isolates, of which urinary tract infection was the majority. A total of 55 (25.8%) of the 213 E. coli isolates were confirmed to have ESBL genes, which were mainly CTX-M types such as CTX-M-14 and CTX-M-15. There was no difference in the proportion of globally epidemic ST131 clones or that of O25, O16, H30, or H30Rx subclones between community-associated and community-onset healthcare-associated isolates. In this study, considerable ST131 E. coli isolations in the community were observed and about half of them were related to the history of a visit to the healthcare facilities, indicating the spread of multidrug-resistant E. coli to the community via healthcare facilities. Copyright © 2016 The Author(s). Published by Elsevier Ltd.. All rights reserved.

Effects of Following National Committee for Clinical Laboratory Standards and Deutsche Industrie Norm-Medizinische Mikrobiologie Guidelines, Country of Isolate Origin, and Site of Infection on Susceptibility of Escherichia coli to Amoxicillin-Clavulanate (Augmentin)

PubMed Central

Simpson, I.; Durodie, J.; Knott, S.; Shea, B.; Wilson, J.; Machka, K.

1998-01-01

Amoxicillin-clavulanate (Augmentin), as a combination of two active agents, poses extra challenges over single agents in establishing clinically relevant breakpoints for in vitro susceptibility tests. Hence, reported differences in amoxicillin-clavulanate percent susceptibilities among Escherichia coli isolates may reflect localized resistance problems and/or methodological differences in susceptibility testing and breakpoint criteria. The objectives of the present study were to determine the effects of (i) methodology, e.g., those of the National Committee for Clinical Laboratory Standards (NCCLS) and the Deutsche Industrie Norm-Medizinische Mikrobiologie (DIN), (ii) country of origin (Spain, France, and Germany), and (iii) site of infection (urinary tract, intra-abdominal sepsis, or other site[s]) upon the incidence of susceptibility to amoxicillin-clavulanate in 185 clinical isolates of E. coli. Cefuroxime and cefotaxime were included for comparison. The use of NCCLS methodology resulted in different distribution of amoxicillin-clavulanate MICs than that obtained with the DIN methodology, a difference highlighted by the 10% more strains found to be within the 8- to 32-μg/ml MIC range. This difference reflects the differing amounts of clavulanic acid present. NCCLS and DIN methodologies also produce different MIC distributions for cefotaxime but not for cefuroxime. Implementation of NCCLS and DIN breakpoints produced markedly different incidences of strains that were found to be susceptible, intermediate or resistant to amoxicillin-clavulanate. A total of 86.5% strains were found to be susceptible to amoxicillin-clavulanate by the NCCLS methodology, whereas only 43.8% were found to be susceptible by the DIN methodology. Similarly, 4.3% of the strains were found to be resistant by NCCLS guidelines compared to 21.1% by the DIN guidelines. The use of DIN breakpoints resulted in a fivefold-higher incidence of strains categorized as resistant to cefuroxime. There were no marked differences due to country of origin upon the MIC distributions for amoxicillin-clavulanate, cefuroxime, or cefotaxime, as determined with the NCCLS guidelines. Isolates from urinary tract and intra-abdominal infections were generally more resistant to amoxicillin-clavulanate than were isolates from other sites of infection. PMID:9574706

Islet product characteristics and factors related to successful human islet transplantation from the Collaborative Islet Transplant Registry (CITR) 1999-2010.

PubMed

Balamurugan, A N; Naziruddin, B; Lockridge, A; Tiwari, M; Loganathan, G; Takita, M; Matsumoto, S; Papas, K; Trieger, M; Rainis, H; Kin, T; Kay, T W; Wease, S; Messinger, S; Ricordi, C; Alejandro, R; Markmann, J; Kerr-Conti, J; Rickels, M R; Liu, C; Zhang, X; Witkowski, P; Posselt, A; Maffi, P; Secchi, A; Berney, T; O'Connell, P J; Hering, B J; Barton, F B

2014-11-01

The Collaborative Islet Transplant Registry (CITR) collects data on clinical islet isolations and transplants. This retrospective report analyzed 1017 islet isolation procedures performed for 537 recipients of allogeneic clinical islet transplantation in 1999-2010. This study describes changes in donor and islet isolation variables by era and factors associated with quantity and quality of final islet products. Donor body weight and BMI increased significantly over the period (p<0.001). Islet yield measures have improved with time including islet equivalent (IEQ)/particle ratio and IEQs infused. The average dose of islets infused significantly increased in the era of 2007-2010 when compared to 1999-2002 (445.4±156.8 vs. 421.3±155.4×0(3) IEQ; p<0.05). Islet purity and total number of β cells significantly improved over the study period (p<0.01 and <0.05, respectively). Otherwise, the quality of clinical islets has remained consistently very high through this period, and differs substantially from nonclinical islets. In multivariate analysis of all recipient, donor and islet factors, and medical management factors, the only islet product characteristic that correlated with clinical outcomes was total IEQs infused. This analysis shows improvements in both quantity and some quality criteria of clinical islets produced over 1999-2010, and these parallel improvements in clinical outcomes over the same period. © 2014 The Authors. American Journal of Transplantation Published by Wiley Periodicals, Inc. on behalf of American Society of Transplant Surgeons.

Islet Product Characteristics and Factors Related to Successful Human Islet Transplantation From the Collaborative Islet Transplant Registry (CITR) 1999–2010

PubMed Central

Balamurugan, A N; Naziruddin, B; Lockridge, A; Tiwari, M; Loganathan, G; Takita, M; Matsumoto, S; Papas, K; Trieger, M; Rainis, H; Kin, T; Kay, T W; Wease, S; Messinger, S; Ricordi, C; Alejandro, R; Markmann, J; Kerr-Conti, J; Rickels, M R; Liu, C; Zhang, X; Witkowski, P; Posselt, A; Maffi, P; Secchi, A; Berney, T; O’Connell, P J; Hering, B J; Barton, F B

2014-01-01

The Collaborative Islet Transplant Registry (CITR) collects data on clinical islet isolations and transplants. This retrospective report analyzed 1017 islet isolation procedures performed for 537 recipients of allogeneic clinical islet transplantation in 1999–2010. This study describes changes in donor and islet isolation variables by era and factors associated with quantity and quality of final islet products. Donor body weight and BMI increased significantly over the period (p < 0.001). Islet yield measures have improved with time including islet equivalent (IEQ)/particle ratio and IEQs infused. The average dose of islets infused significantly increased in the era of 2007–2010 when compared to 1999–2002 (445.4 ± 156.8 vs. 421.3 ± 155.4 ×103 IEQ; p < 0.05). Islet purity and total number of β cells significantly improved over the study period (p < 0.01 and <0.05, respectively). Otherwise, the quality of clinical islets has remained consistently very high through this period, and differs substantially from nonclinical islets. In multivariate analysis of all recipient, donor and islet factors, and medical management factors, the only islet product characteristic that correlated with clinical outcomes was total IEQs infused. This analysis shows improvements in both quantity and some quality criteria of clinical islets produced over 1999–2010, and these parallel improvements in clinical outcomes over the same period. PMID:25278159

[Isolated lesion of the Edinger-Westphal nucleus in topographical relation with a post-traumatic mesencephalic hematoma].

PubMed

Guerrero, A L; Onzáin, J I; Martín, J A; Blanco, A; Moreta, J A

1996-08-01

From the relevant literature, it would seem that the commonest single cause of lesion of the third cranial nerves is indirect, accompanying intracranial traumas. From multiple clinical observations however, it seems that many of these cases may be due to lesions of the mesencephalum which nevertheless have rarely been identified by current imaging techniques. Clinical case. We describe the clinical observation of isolated pupil involvement, attributed to a lesion of the Edinger-Westphal nucleus as a consequence of a mesencephalic haematoma in the context of closed craneo-encephalic trauma. In our review of the literature, we have not found any other such case. We briefly review the most frequently involved mechanisms implicated in the genesis of lesions of the third cranial nerves at different sites and the different changes seen in the pupil in each case, together with the characteristics and pathogenesis of the lesions produced in the mesencephalum as a consequence of intracranial trauma. We emphasize the importance of our case as being the first time an isolated lesion of the Edinger-Westphal nucleus has been described in topographic relation to a mesencephalic haematoma.

Lactobacillus Species Identification, H2O2 Production, and Antibiotic Resistance and Correlation with Human Clinical Status

PubMed Central

Felten, Annie; Barreau, Claude; Bizet, Chantal; Lagrange, Philippe Henri; Philippon, Alain

1999-01-01

Lactobacilli recovered from the blood, cerebrospinal fluid, respiratory tract, and gut of 20 hospitalized immunocompromised septic patients were analyzed. Biochemical carbohydrate fermentation and total soluble cell protein profiles were used to identify the species. Hydrogen peroxide production was measured. Susceptibility to 19 antibiotics was tested by a diffusion method, and the MICs of benzylpenicillin, amoxicillin, imipenem, erythromycin, vancomycin, gentamicin, and levofloxacin were determined. A small number of species produced H2O2, and antibiotic susceptibilities were species related. Eighteen (90%) of the isolates were L. rhamnosus, one was L. paracasei subsp. paracasei, and one was L. crispatus. L. rhamnosus, L. paracasei subsp. paracasei isolates, and the type strains were neither H2O2 producers nor vancomycin susceptible (MICs, ≥256 μg/ml). L. crispatus, as well as most of the type strains of lactobacilli which belong to the L. acidophilus group, was an H2O2 producer and vancomycin susceptible (MICs, <4 μg/ml). PMID:9986841

Genomic Epidemiology of Global Carbapenemase-Producing Enterobacter spp., 2008-2014.

PubMed

Peirano, Gisele; Matsumura, Yasufumi; Adams, Mark D; Bradford, Patricia; Motyl, Mary; Chen, Liang; Kreiswirth, Barry N; Pitout, Johann D D

2018-06-01

We performed whole-genome sequencing on 170 clinical carbapenemase-producing Enterobacter spp. isolates collected globally during 2008-2014. The most common carbapenemase was VIM, followed by New Delhi metallo-β-lactamase (NDM), Klebsiella pneumoniae carbapenemase, oxacillin 48, and IMP. The isolates were of predominantly 2 species (E. xiangfangensis and E. hormaechei subsp. steigerwaltii) and 4 global clones (sequence type [ST] 114, ST93, ST90, and ST78) with different clades within ST114 and ST90. Particular genetic structures surrounding carbapenemase genes were circulating locally in various institutions within the same or between different STs in Greece, Guatemala, Italy, Spain, Serbia, and Vietnam. We found a common NDM genetic structure (NDM-GE-U.S.), previously described on pNDM-U.S. from Klebsiella pneumoniae ATCC BAA-214, in 14 different clones obtained from 6 countries spanning 4 continents. Our study highlights the importance of surveillance programs using whole-genome sequencing in providing insight into the molecular epidemiology of carbapenemase-producing Enterobacter spp.

Phenotypic and Molecular Characterization of Multidrug Resistant Klebsiella pneumoniae Isolated from Different Clinical Sources in Al-Najaf Province-Iraq.

PubMed

Aljanaby, Ahmed Abduljabbar Jaloob; Alhasnawi, Haneen Mohammed Reda Jaber

2017-01-01

Burns infections and urinary tract infections are the most important prevalent diseases in Asian countries, such as Iraq. Klebsiella pneumoniae is one of the most important bacteria cause this type of infections especially in hospitals. Therefore, the aim of this study was to investigate the prevalence of multi-drug resistance K. pneumoniae and extended-spectrum beta-lactamases producing K. pneumoniae isolates from inpatients with urinary tract infection and burns infections in Al-Kufa hospital in Al-Najaf province, Iraq. A total of 285 clinical samples were collected from in-patients infected with urinary tract infection (141 urine samples) and burns infections (144 burns swabs). Fourteen different antibiotics were used by disc diffusion method and 13 antimicrobials resistance genes were used by PCR technique. A total of 43 K. pneumoniae strains were isolated. The highest resistance rate was observed for amoxicillin 25 μg and amoxicillin+clavulanic acid 20+10 μg (97.67%) while the lowest resistance rate was observed for imipenem 10 μg (9.30%). The most common resistance associated-genes were blaSHV (86.04%) and at lower prevalence were IMP (9.30%). Klebsiella pneumoniae strains isolated from burns infections were more virulent than those isolated from urinary tract infections.

Biliary diseases as main causes of pyogenic liver abscess caused by extended-spectrum beta-lactamase-producing Enterobacteriaceae.

PubMed

Shi, Shao-Hua; Feng, Xiao-Ning; Lai, Ming-Chun; Kong, Hai-Shen; Zheng, Shu-Sen

2017-05-01

Little is known about aetiology and morbidity and clinical characteristics of pyogenic liver abscess caused by extended-spectrum beta-lactamase-producing Enterobacteriaceae. An analysis between pyogenic liver abscess patients caused by extended-spectrum beta-lactamase-producing Enterobacteriaceae isolates and those caused by non-extended-spectrum beta-lactamase-producing Enterobacteriaceae was performed. Among 817 pyogenic liver abscess patients, there were 176 patients (21.5%) with pyogenic liver abscess of biliary origin, and 67 pyogenic liver abscess patients (8.2%) caused by extended-spectrum beta-lactamase-producing Enterobacteriaceae isolates (mainly Escherichia coli and Klebsiella pneumoniae). Of 176 pyogenic liver abscess patients related to biliary disorders, there were 48 pyogenic liver abscess patients (27.3%) caused by extended-spectrum beta-lactamase-producing Enterobacteriaceae. Within 67 pyogenic liver abscess patients caused by Enterobacteriaceae expressing extended-spectrum beta-lactamases, the occurrences of 48 pyogenic liver abscess patients (71.6%) were associated with biliary disorders. When compared with pyogenic liver abscess patients caused by non-extended-spectrum beta-lactamase-producing Enterobacteriaceae, there were significantly greater incidences of polymicrobial infections, bacteremia, pulmonary infection, recurrence and death in pyogenic liver abscess patients caused by extended-spectrum beta-lactamase-producing Enterobacteriaceae. Carbapenems remain mainstay drugs against extended-spectrum beta-lactamase-producing E. coli and K. pneumoniae. Independent risk factors for occurrence of pyogenic liver abscess caused by extended-spectrum beta-lactamase-producing Enterobacteriaceae were biliary disorders including extra- and intrahepatic cholangiolithiasis and an abnormal bilioenteric communication between bile and gut, a treatment history of malignancy such as operation and chemotherapy, pulmonary infection, and diabetes mellitus. Pyogenic liver abscess caused by extended-spectrum beta-lactamase-producing Enterobacteriaceae isolates mainly occurs in patients with biliary disorders or with a treatment history of malignancy. The mainstay of treatment remains carbapenems in combination with adequate aspiration or drainage. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Prevalence and clonal relationship of ESBL-producing Salmonella strains from humans and poultry in northeastern Algeria.

PubMed

Djeffal, Samia; Bakour, Sofiane; Mamache, Bakir; Elgroud, Rachid; Agabou, Amir; Chabou, Selma; Hireche, Sana; Bouaziz, Omar; Rahal, Kheira; Rolain, Jean-Marc

2017-05-15

The aims of this study were to investigate Salmonella contamination in broiler chicken farms and slaughterhouses, to assess the antibiotic resistance profile in avian and human Salmonella isolates, and to evaluate the relationship between avian and human Extended Spectrum β-Lactamase (ESBL)-producing isolates. Salmonella was screened in different sample matrices collected at thirty-two chicken farms and five slaughterhouses. The human isolates were recovered from clinical specimens at the University Teaching Hospital of Constantine (UTH). All suspected colonies were confirmed by MALDI-TOF (Matrix Assisted Laser Desorption Ionization Time OF light) and serotyped. Susceptibility testing against 13 antibiotics including, amoxicillin/clavulanic acid, ticarcillin, cefoxitin, cefotaxime, aztreonam, imipenem, ertapenem, gentamicin, amikacin, ciprofloxacin, colistin, trimethoprim/sulfamethoxazole and fosfomycin, was performed using the disk diffusion method on Mueller-Hinton agar. ESBL-production was screened by the double-disk synergy test and confirmed by molecular characterization using PCR (polymerase chain reaction) amplification and sequencing of ESBL encoding genes. Clonality of the avian and human strains was performed using the Multi Locus Sequencing Typing method (MLST). Forty-five isolated avian Salmonella strains and 37 human collected ones were studied. Five S. enterica serotypes were found in avian isolates (mainly Kentucky) and 9 from human ones (essentially Infantis). 51.11% and 26.6% of the avian isolates were resistant to ciprofloxacin and cefotaxime, respectively, whereas human isolates were less resistant to these antibiotics (13.5% to ciprofloxacin and 16.2% to cefotaxime). Eighteen (12 avian and 6 human) strains were found to produce ESBLs, which were identified as bla CTX-M-1 (n = 12), bla CTX-M-15 (n = 5) and bla TEM group (n = 8). Interestingly, seven of the ESBL-producing strains (5 avian and 2 human) were of the same ST (ST15) and clustered together, suggesting a common origin. The results of the combined phenotypic and genotypic analysis found in this study suggest a close relationship between human and avian strains and support the hypothesis that poultry production may play a role in the spread of multidrug-resistant Salmonella in the human community within the study region.

Genetic Platforms of blaCTX-M in Carbapenemase-Producing Strains of K. pneumoniae Isolated in Chile

PubMed Central

Carrasco-Anabalón, Sergio; Vera-Leiva, Alejandra; Quezada-Aguiluz, Mario; Morales-Rivera, María F.; Lima, Celia A.; Fernández, Jorge; Ulloa, Soledad; Domínguez, Mariana; González-Rocha, Gerardo; Bello-Toledo, Helia

2018-01-01

Objective: To elucidate whether the genetic platforms of blaCTX-M contribute to the phenotypes of multi-drug-resistance (MDR) in the first carbapenemase-producing K. pneumoniae strains isolated in Chile. Method: Twenty-two carbapenemase-producing K. pneumoniae strains isolated from different Chilean patients and hospitals were studied. Their genetic relatedness was assessed by PFGE and MLST. The levels of antibiotic resistance were evaluated by determining the minimum inhibitory concentration of various antimicrobials. In addition, several antibiotic resistance genes of clinical relevance in Chile were investigated. The prevalence, allelic variants, and genetic platforms of blaCTX-M were determined by PCR and sequencing. Results: Out of the 22 strains studied, 20 carry KPC, one carries NDM-1, and one carries OXA-370. The PFGE analysis showed three clades with a genetic relatedness >85%, two formed by four strains and one by eight strains. The other strains are not genetically related, and a total of 17 different pulse types were detected. Ten different STs were identified, the main ones being ST258 (five strains) and ST1161 (seven strains). The isolates presented different percentages of resistance, and 82% were resistant to all the β-lactams tested, 91% to ciprofloxacin, 73% to colistin, 59% to gentamicin, 50% to amikacin, and only 9% to tigecycline. All isolates carried blaTEM and blaSHV, whereas 71% carried aac(6′)Ib-cr, and 57% one qnr gene (A, B, C, D, or S). The blaCTX-M gene was found in 10 of the isolates (4 blaCTX-M−15 and 6 blaCTX-M−2). The characterization of the platform, in seven selected strains, revealed that the gene is associated with unusual class 1 integrons and insertion sequences such as ISCR1, ISECp1, and IS26. Conclusion: In the first carbapenemase-producing K. pneumoniae strains isolated in Chile the genetic platform of blaCTX-M−2 corresponds to an unusual class 1 integron that can be responsible for the MDR phenotype, whereas the genetic platforms of blaCTX-M−15 are associated with different IS and do not contribute to multi-drug resistance. PMID:29593660

Prevalence and risk factors for colonization by Clostridium difficile and extended-spectrum β-lactamase-producing Enterobacteriaceae in rehabilitation clinics in Germany.

PubMed

Arvand, M; Ruscher, C; Bettge-Weller, G; Goltz, M; Pfeifer, Y

2018-01-01

Rehabilitation clinics may vary widely in terms of type of care provided, duration of hospital stay, and case severity. Few data are available on prevalence of Clostridium difficile or extended-spectrum β-lactamase-producing Enterobacteriaceae (ESBL-E) colonization in rehabilitation clinics in Germany. This study investigated the frequency of intestinal colonization by these pathogens among patients in rehabilitation clinics of different specialization. In the scope of a point prevalence study, faecal samples and demographic and clinical data were collected in five rehabilitation clinics. Samples were screened for C. difficile and ESBL-E by culture. Isolates were characterized by polymerase chain reaction for C. difficile toxins A and B, for β-lactamase genes, and by molecular typing including pulsed-field gel electrophoresis and PCR-based ribotyping. Of 305 patients screened, 11.1% were colonized by toxigenic C. difficile and 7.5% by ESBL-E. Colonization rates differed markedly between facilities, ranging from 1.6% to 26.3% for C. difficile and from zero to 23.7% for ESBL-E. Prevalence of colonization by C. difficile and ESBL-E was higher in neurological rehabilitation clinics than in clinics with other specialties (P<0.001). Molecular typing revealed six patients from one neurological rehabilitation clinic harbouring a unique C. difficile strain (ribotype 017). CTX-M-15 was the most prevalent ESBL type. We detected several indistinguishable pairs of ESBL-E isolates within some facilities. Significant differences were found in the prevalence of C. difficile and ESBL-E between rehabilitation clinics. Facilities providing specialized medical care for critically ill patients had higher prevalence rates. These results may help to delineate the requirements for infection prevention and control in rehabilitation clinics. Copyright © 2017 The Healthcare Infection Society. Published by Elsevier Ltd. All rights reserved.

Extended-Spectrum-β-Lactamase-Producing Enterobacteriaceae Isolated from Vegetables Imported from the Dominican Republic, India, Thailand, and Vietnam

PubMed Central

Zurfluh, Katrin; Nüesch-Inderbinen, Magdalena; Morach, Marina; Zihler Berner, Annina; Hächler, Herbert

2015-01-01

To examine to what extent fresh vegetables imported into Switzerland represent carriers of extended-spectrum-β-lactamase (ESBL)-producing Enterobacteriaceae, 169 samples of different types of fresh vegetables imported into Switzerland from the Dominican Republic, India, Thailand, and Vietnam were analyzed. Overall, 25.4% of the vegetable samples yielded one or more ESBL-producing Enterobacteriaceae, 78.3% of which were multidrug resistant. Sixty isolates were obtained: Escherichia coli, 26; Klebsiella pneumoniae, 26; Enterobacter cloacae, 6; Enterobacter aerogenes, 1; and Cronobacter sakazakii, 1. We found 29 isolates producing CTX-M-15, 8 producing CTX-M-14, 7 producing CTX-M-55, 3 producing CTX-M-65, 1 each producing CTX-M-1, CTX-M-3, CTX-M-27, and CTX-M-63, 5 producing SHV-2, 3 producing SHV-12, and 1 producing SHV-2a. Four of the E. coli isolates belonged to epidemiologically important clones: CTX-M-15-producing B2:ST131 (1 isolate), D:ST405 (1 isolate), and D:ST38 (2 isolates). One of the D:ST38 isolates belonged to the extraintestinal enteroaggregative E. coli (EAEC) D:ST38 lineage. Two of the K. pneumoniae isolates belonged to the epidemic clones sequence type 15 (ST15) and ST147. The occurrence of antibiotic-resistant pathogenic and commensal Enterobacteriaceae in imported agricultural foodstuffs constitutes a source of ESBL genes and a concern for food safety. PMID:25724954

Emergence of Carbapenemase-Producing Escherichia coli Isolated from Companion Animals in Algeria.

PubMed

Yousfi, Massilia; Touati, Abdelaziz; Mairi, Assia; Brasme, Lucien; Gharout-Sait, Alima; Guillard, Thomas; De Champs, Christophe

2016-06-01

The emergence and worldwide spread of carbapenemase-producing Enterobacteriaceae is of great concern to public health. The aim of this study was to investigate the occurrence of carbapenemase-producing Escherichia coli in companion animals in Algeria. Two hundred fecal samples were obtained from healthy and diseased dogs and cats in one veterinary office and private owners in Bejaia city, Algeria, during November 2014 to March 2015. Isolates were screened by polymerase chain reaction for the presence of carbapenemase, acquired plasmidic AmpC (pAmpC) and extended-spectrum beta-lactamase genes. Five carbapenemase-producing E. coli isolates were detected including four OXA-48-producing isolates and one isolate producing NDM-5. Coexpression of ESBL and pAmpC genes was observed in these isolates. Phylogenetic grouping revealed that these isolates belonged to A and D phylogroups. The results of this study show that carbapenemase-producing E. coli spread to the companion animals in Algeria.

Antimicrobial Activities of Methanol, Ethanol and Supercritical CO2 Extracts of Philippine Piper betle L. on Clinical Isolates of Gram Positive and Gram Negative Bacteria with Transferable Multiple Drug Resistance

PubMed Central

Valle, Demetrio L.; Cabrera, Esperanza C.; Puzon, Juliana Janet M.; Rivera, Windell L.

2016-01-01

Piper betle L. has traditionally been used in alternative medicine in different countries for various therapeutic purposes, including as an anti-infective agent. However, studies reported in the literature are mainly on its activities on drug susceptible bacterial strains. This study determined the antimicrobial activities of its ethanol, methanol, and supercritical CO2 extracts on clinical isolates of multiple drug resistant bacteria which have been identified by the Infectious Disease Society of America as among the currently more challenging strains in clinical management. Assay methods included the standard disc diffusion method and the broth microdilution method for the determination of the minimum inhibitory concentration (MIC) and the minimum bactericidal concentrations (MBC) of the extracts for the test microorganisms. This study revealed the bactericidal activities of all the P. betle leaf crude extracts on methicillin-resistant Staphylococcus aureus (MRSA), vancomycin-resistant Enterococcus (VRE), extended spectrum β-lactamase-producing Enterobacteriaceae, carbapenem-resistant Enterobacteriaceae, and metallo-β-lactamase-producing Pseudomonas aeruginosa and Acinetobacter baumannii, with minimum bactericidal concentrations that ranged from 19μg/ml to 1250 μg/ml. The extracts proved to be more potent against the Gram positive MRSA and VRE than for the Gram negative test bacteria. VRE isolates were more susceptible to all the extracts than the MRSA isolates. Generally, the ethanol extracts proved to be more potent than the methanol extracts and supercritical CO2 extracts as shown by their lower MICs for both the Gram positive and Gram negative MDRs. MTT cytotoxicity assay showed that the highest concentration (100 μg/ml) of P. betle ethanol extract tested was not toxic to normal human dermal fibroblasts (HDFn). Data from the study firmly established P. betle as an alternative source of anti-infectives against multiple drug resistant bacteria. PMID:26741962

Antimicrobial Activities of Methanol, Ethanol and Supercritical CO2 Extracts of Philippine Piper betle L. on Clinical Isolates of Gram Positive and Gram Negative Bacteria with Transferable Multiple Drug Resistance.

PubMed

Valle, Demetrio L; Cabrera, Esperanza C; Puzon, Juliana Janet M; Rivera, Windell L

2016-01-01

Piper betle L. has traditionally been used in alternative medicine in different countries for various therapeutic purposes, including as an anti-infective agent. However, studies reported in the literature are mainly on its activities on drug susceptible bacterial strains. This study determined the antimicrobial activities of its ethanol, methanol, and supercritical CO2 extracts on clinical isolates of multiple drug resistant bacteria which have been identified by the Infectious Disease Society of America as among the currently more challenging strains in clinical management. Assay methods included the standard disc diffusion method and the broth microdilution method for the determination of the minimum inhibitory concentration (MIC) and the minimum bactericidal concentrations (MBC) of the extracts for the test microorganisms. This study revealed the bactericidal activities of all the P. betle leaf crude extracts on methicillin-resistant Staphylococcus aureus (MRSA), vancomycin-resistant Enterococcus (VRE), extended spectrum β-lactamase-producing Enterobacteriaceae, carbapenem-resistant Enterobacteriaceae, and metallo-β-lactamase-producing Pseudomonas aeruginosa and Acinetobacter baumannii, with minimum bactericidal concentrations that ranged from 19μg/ml to 1250 μg/ml. The extracts proved to be more potent against the Gram positive MRSA and VRE than for the Gram negative test bacteria. VRE isolates were more susceptible to all the extracts than the MRSA isolates. Generally, the ethanol extracts proved to be more potent than the methanol extracts and supercritical CO2 extracts as shown by their lower MICs for both the Gram positive and Gram negative MDRs. MTT cytotoxicity assay showed that the highest concentration (100 μg/ml) of P. betle ethanol extract tested was not toxic to normal human dermal fibroblasts (HDFn). Data from the study firmly established P. betle as an alternative source of anti-infectives against multiple drug resistant bacteria.

Detection of qnrVC6, within a new genetic context, in a NDM-1-producing Citrobacter freundii clinical isolate from Uruguay.

PubMed

Bado, Inés; Ezdra, Romina Papa; Cordeiro, Nicolás; Outeda, Matilde; Caiata, Leticia; García-Fulgueiras, Virginia; Seija, Verónica; Vignoli, Rafael

2018-03-08

The objective of the present study was to characterise the mechanisms underlying quinolone and oxyimino-cephalosporin resistance in a Citrobacter freundii clinical isolate obtained from the ICU in Uruguay's University Hospital. Citrobacter freundii strain CF638 was isolated from a urine culture. Identification and susceptibility testing were performed using the VITEK ® 2 system, and MIC determination and disc diffusion assay, respectively. Resistance genes and mobile genetic elements were identified, by PCR and sequencing. Plasmid transfer was assessed by conjugation; plasmid, size was estimated by treatment with S1 and PFGE. Plasmid incompatibility, group and toxin-antitoxin systems were sought by PCR. Strain CF638 showed a multi-drug resistant profile, including, resistance to carbapenemes and quinolones. Transconjugant TcCF638, harbouring a ∼200 kb IncA/C plasmid, also showed resistance to all β-lactams, (except for aztreonam), and diminished susceptibility to ciprofloxacin. PCR, assays were positive for bla NDM-1 and qnrVC in CF638 and TcCF638. Two different class 1 integrons were detected, In127 and In907. In127, featured the genetic array: aadA2-ltr2. Conversely, complex In907 featured, two variable regions (VR); VR-1 consisted of aadB-bla OXA-10 -aadA1cc, whereas, VR-2, featured a gene qnrVC6 108 bp downstream from the ISCR1 and 45 bp, upstream from the qacEΔ1. Expression of qnrVC6 would be on account of a, putative promoter region, detected using the Neural Network Promoter, Prediction program. To the best of our knowledge this constitutes the first report of a qnrVC gene within a complex class 1 integron, and the first report as well of the occurrence of such gene in an NDM-1-producing enterobacterial clinical isolate. Copyright © 2018. Published by Elsevier Ltd.

Klebsiella pneumoniae Carbapenemase (KPC)-Producing K. pneumoniae at a Single Institution: Insights into Endemicity from Whole-Genome Sequencing

PubMed Central

Stoesser, Nicole; Sheppard, Anna E.; Pankhurst, Louise; Giess, Adam; Yeh, Anthony J.; Didelot, Xavier; Turner, Stephen D.; Sebra, Robert; Kasarskis, Andrew; Peto, Tim; Crook, Derrick; Sifri, Costi D.

2015-01-01

The global emergence of Klebsiella pneumoniae carbapenemase-producing K. pneumoniae (KPC-Kp) multilocus sequence type ST258 is widely recognized. Less is known about the molecular and epidemiological details of non-ST258 K. pneumoniae in the setting of an outbreak mediated by an endemic plasmid. We describe the interplay of blaKPC plasmids and K. pneumoniae strains and their relationship to the location of acquisition in a U.S. health care institution. Whole-genome sequencing (WGS) analysis was applied to KPC-Kp clinical isolates collected from a single institution over 5 years following the introduction of blaKPC in August 2007, as well as two plasmid transformants. KPC-Kp from 37 patients yielded 16 distinct sequence types (STs). Two novel conjugative blaKPC plasmids (pKPC_UVA01 and pKPC_UVA02), carried by the hospital index case, accounted for the presence of blaKPC in 21/37 (57%) subsequent cases. Thirteen (35%) isolates represented an emergent lineage, ST941, which contained pKPC_UVA01 in 5/13 (38%) and pKPC_UVA02 in 6/13 (46%) cases. Seven (19%) isolates were the epidemic KPC-Kp strain, ST258, mostly imported from elsewhere and not carrying pKPC_UVA01 or pKPC_UVA02. Using WGS-based analysis of clinical isolates and plasmid transformants, we demonstrate the unexpected dispersal of blaKPC to many non-ST258 lineages in a hospital through spread of at least two novel blaKPC plasmids. In contrast, ST258 KPC-Kp was imported into the institution on numerous occasions, with other blaKPC plasmid vectors and without sustained transmission. Instead, a newly recognized KPC-Kp strain, ST941, became associated with both novel blaKPC plasmids and spread locally, making it a future candidate for clinical persistence and dissemination. PMID:25561339

One-year surveillance of ESKAPE pathogens in an intensive care unit of Monterrey, Mexico.

PubMed

Llaca-Díaz, Jorge Martín; Mendoza-Olazarán, Soraya; Camacho-Ortiz, Adrian; Flores, Samantha; Garza-González, Elvira

2012-01-01

Bacterial species from the ESKAPE group (i.e. Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa and Enterobacter species) are frequently resistant to antibiotics. The purpose of this study was to monitor the incidence of ESKAPE pathogens at the intensive care unit (ICU) of a tertiary care hospital in Monterrey, Mexico. All clinically relevant organisms isolated from June 2011 to June 2012 were included. Identification and susceptibility testing was performed using panels from Sensititre. Resistance to oxacillin, for S. aureus, and the production of extended spectrum β-lactamases (ESBLs), for K. pneumonia, were determined as defined by the Clinical Laboratory Standards Institute. Also, the presence of vanA and vanB genes was determined in E. faecium vancomycin (VAN)-resistant isolates. The majority of pathogens (64.5%) isolated in the ICU unit were from the ESKAPE group. The organisms most frequently isolated were A. baumannii (15.8%) and P. aeruginosa (14.3%). A high resistance to carbapenems was detected for A. baumannii (75.3%) while 62% of S. aureus isolates were confirmed to be methicillin resistant. Of the K. pneumoniae isolates, 36.9% were ESBL producers. We detected three E. faecium VAN-resistant isolates, all of which contained the vanA gene. The presence of the ESKAPE group of pathogens is a major problem in the ICU setting. The results of this study support the implementation of special antimicrobial strategies to specifically target these microorganisms. Copyright © 2013 S. Karger AG, Basel.

Antimicrobial resistance levels amongst staphylococci isolated from clinical cases of bovine mastitis in Kosovo.

PubMed

Mehmeti, Ibrahim; Behluli, Behlul; Mestani, Mergim; Ademi, Arsim; Nes, Ingolf F; Diep, Dzung B

2016-10-31

Mastitis is one of the most frequent and costly disease in cattle. We studied milk samples from cattle with mastitis from farms in Kosovo to identify mastitis-causing pathogens and possible effective antibiotics. Our ultimate goal is to help implement adequate antibiotic management and treatment practices in Kosovo METHODOLOGY: A total of 152 milk samples were collected from cows with clinical mastitis from different farms in Kosovo. After identification of microorganisms, antibiotic susceptibility and the occurrence of enterotoxins was investigated. Staphylococci were found in 89 samples, of which 58 were coagulase negative and 31 coagulase positive. S. aureus was isolated from 27 samples, S. epidermidis from 25, and S. chromogenes from 15, while other species of staphylococci were isolated from the remaining 22 isolates. Interestingly, the bacterial diversity was different between cows in different periods of lactation and among different breeds. Most of the isolates (76/89) were resistant to two or more antibiotics. The highest resistance was to penicillin and ampicillin (> 65%), followed by tetracycline, oxacillin, streptomycin, chloramphenicol (> 23%), and less than 3% to erythromycin. Of the 89 isolates, 40 produced enterotoxins that were most frequently typed as A and C. We detected human bacterial pathogens in the cultures of milk samples from cows with mastitis. The isolates demonstrated resistance to two or more antibiotics, some of which are frequently used to treat animal and human infections. We recommend increased control and more stringent use of antibiotics in veterinary as well as human medicine.

Complete Genome Sequence of an Escherichia coli O121:H19 Strain from an Outbreak in Canada Associated with Flour

PubMed Central

Robertson, James; Lin, Janet; Levett, Paul N.; Nadon, Celine; Nash, John

2018-01-01

ABSTRACT Here, we present the first complete genome sequence of an Escherichia coli non-O157 Shiga-toxin producing isolate, 16-9255, from serotype O121:H19. This strain is notable as a clinical case recovered from a recent Canadian flour-associated outbreak event. PMID:29371368

Updated Campylobacter jejuni capsule PCR multiplex typing system and its application to clinical isolates from south and southeast Asia

USDA-ARS?s Scientific Manuscript database

Campylobacter jejuni produces a polysaccharide capsule that is the major determinant of the Penner serotyping scheme. This passive slide agglutination typing system was developed in the early 1980’s and was recognized for over two decades as gold standard for C. jejuni typing. A preliminary multiple...

CTX-M-15-non-ST131 Escherichia coli isolates are mainly responsible of faecal carriage with ESBL-producing Enterobacteriaceae in travellers, immigrants and those visiting friends and relatives.

PubMed

Valverde, A; Turrientes, M-C; Norman, F; San Martín, E; Moreno, L; Pérez-Molina, J A; López-Vélez, R; Cantón, R

2015-03-01

Prevalence of extended-spectrum β-lactamases (ESBL) and/or carbapenemase-producing Enterobacteriaceae (EPE and CPE) in stool samples from 75 travellers, 8 people visiting friends and relatives and 3 immigrants who had travelled or came from tropical or subtropical areas was determined. Thirty-one per cent (27/86) of the subjects were faecal carriers of EPE, and 37 EPE isolates were recovered (36 Escherichia coli, 1 Klebsiella pneumoniae). CTX-M-15 was the most prevalent enzyme (64.8%) mainly associated with E. coli belonging to phylogroup A and sequence type complex 10. Most of the ESBL-positive travellers (50%) had visited countries from Asia. Copyright © 2014 European Society of Clinical Microbiology and Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

Postantifungal effect of the combination of caspofungin with voriconazole and amphotericin B against clinical Candida krusei isolates.

PubMed

Oz, Yasemin; Kiremitci, Abdurrahman; Dag, Ilknur; Metintas, Selma; Kiraz, Nuri

2013-01-01

We evaluated the postantifungal effects (PAFEs) of caspofungin (CAS), voriconazole (VOR), amphotericin B (AmB), and the combinations of CAS + VOR and CAS + AmB against 30 clinical Candida krusei isolates at 0.25, 1 and 4 times the MIC of each individually and in the indicated combinations. Antifungals were removed after 1 hour and colony counts were performed at 0, 2, 6, 24, and 48 h. VOR did not display any measurable PAFE regardless of antifungal concentrations, while AmB and CAS exhibited dose-dependent PAFE. The most effective agent producing a prolonged PAFE in this study was CAS. Although the combination of CAS with VOR generated longer PAFEs at 0.25 and 1 times their respective MICs in comparison with CAS alone, this combination was indifferent rather than synergistic. However, the combination of CAS with AmB at 4 times their MICs exhibited the best performance, reducing the colony counts during the 48 h after removal of drugs and resulted in synergic interaction in respect to 20 (67%) isolates. Consequently, CAS has a prolonged PAFE in vitro against C. krusei isolates, and the combination of AmB + CAS may increase significantly the efficacy of CAS. Our data may be useful in optimizing dosing regimens for these agents and their combinations, although further studies are needed to explore the clinical usefulness of our results.

Pathogenesis of coxsackievirus B2 in mice: characterization of clinical isolates of the coxsackievirus B2 from patients with myocarditis and aseptic meningitis in Korea.

PubMed

Hong, Jiyoung; Kang, Bunghak; Yeo, Sanggu; Jee, Youngmee; Park, Jae-Hak

2017-12-31

Group B coxsackieviruses (CVBs) are a group of common human pathogens producing various clinical symptoms. Although the virology of CVB is well known, there is limited information on viral pathogenesis and the relationship between clinical symptoms and viral phenotype, particularly for CVB type 2 (CVB2). In 2004 in Korea, two CVB2 strains were isolated: CB2/04/279 from stool of an acute myocarditis patient with heart failure and CB2/04/243 from an aseptic meningitis patient. In this study, a high degree of homology was observed between the CB2/04/279 and CB2/04/243 full genome sequences. The two Korean CVB2 isolates had 93.1% homology compared to 82.1%-82.5% nucleotide sequence identity with the cardiovirulence-associated reference CVB strain Ohio-1 (CVB/O). CVB2-induced pathogenesis was analyzed, focusing on virus-induced pathology of various tissues in 4-week-old BALB/c inbred male mice. Myocarditis developed and extensive pancreatic inflammation was observed in all mice infected with CB2/04/279 or CVB/O, but not in animals infected with CB2/04/243. This is the first report of the full-genomic sequence and pathogenesis of the CVB2 strain isolated from an acute myocarditis patient in Korea.

[Examination of metallo-beta-lactamase-producing different types of Serratia marcescens detected in the same patient].

PubMed

Takamitsu, Ito; Fukui, Yasuo; Ono, Noriaki; Ikeda, Fumiaki; Kanayama, Akiko; Kobayashi, Intetsu

2013-03-01

Metallo-beta-lactamase (MBL) producing Serratia marcescens isolate was recovered from a study patient in September, 2007 in whom MBL non-producing S. marcescens had been isolated 2 months previously. Two S. marcescens isolates recovered from the study patient showed the same pulsed-field gel electrophoresis (PFGE) pattern. Seven S. marcescens isolates were recovered from other patients in our hospital during August, 2007 and November, 2007. Five of the seven isolates produced MBL. All of the MBL-producing isolates showed the same PFGE pattern and harbored plasmids of the same size and bla(IMP) genes. The bla(IMP) genes were easily transferred to Escherichia coli DH5alpha by transformation of a plasmid purified from the MBL-producing isolate. Those transformation experiments suggested that bla(IMP) genes were encoded by the plasmid. From these observations, it was speculated that the MBL non-producing S. marcescens isolate recovered from the study patient had acquired the plasmid which encoded bla(IMP) genes and a monoclone of MBL-producing S. marcescens spread horizontally in our hospital.

Agreement Assessment of Tigecycline Susceptibilities Determined by the Disk Diffusion and Broth Microdilution Methods among Commonly Encountered Resistant Bacterial Isolates: Results from the Tigecycline In Vitro Surveillance in Taiwan (TIST) Study, 2008 to 2010

PubMed Central

Liu, Jien-Wei; Ko, Wen-Chien; Huang, Cheng-Hua; Liao, Chun-Hsing; Lu, Chin-Te; Chuang, Yin-Ching; Tsao, Shih-Ming; Chen, Yao-Shen; Liu, Yung-Ching; Chen, Wei-Yu; Jang, Tsrang-Neng; Lin, Hsiu-Chen; Chen, Chih-Ming; Shi, Zhi-Yuan; Pan, Sung-Ching; Yang, Jia-Ling; Kung, Hsiang-Chi; Liu, Chun-Eng; Cheng, Yu-Jen; Chen, Yen-Hsu; Lu, Po-Liang; Sun, Wu; Wang, Lih-Shinn; Yu, Kwok-Woon; Chiang, Ping-Cherng; Lee, Ming-Hsun; Lee, Chun-Ming; Hsu, Gwo-Jong

2012-01-01

The Tigecycline In Vitro Surveillance in Taiwan (TIST) study, initiated in 2006, is a nationwide surveillance program designed to longitudinally monitor the in vitro activity of tigecycline against commonly encountered drug-resistant bacteria. This study compared the in vitro activity of tigecycline against 3,014 isolates of clinically important drug-resistant bacteria using the standard broth microdilution and disk diffusion methods. Species studied included methicillin-resistant Staphylococcus aureus (MRSA; n = 759), vancomycin-resistant Enterococcus faecium (VRE; n = 191), extended-spectrum β-lactamase (ESBL)-producing Escherichia coli (n = 602), ESBL-producing Klebsiella pneumoniae (n = 736), and Acinetobacter baumannii (n = 726) that had been collected from patients treated between 2008 and 2010 at 20 hospitals in Taiwan. MICs and inhibition zone diameters were interpreted according to the currently recommended U.S. Food and Drug Administration (FDA) criteria and the European Committee on Antimicrobial Susceptibility Testing (EUCAST) criteria. The MIC90 values of tigecycline against MRSA, VRE, ESBL-producing E. coli, ESBL-producing K. pneumoniae, and A. baumannii were 0.5, 0.125, 0.5, 2, and 8 μg/ml, respectively. The total error rates between the two methods using the FDA criteria were high: 38.4% for ESBL-producing K. pneumoniae and 33.8% for A. baumannii. Using the EUCAST criteria, the total error rate was also high (54.6%) for A. baumannii isolates. The total error rates between these two methods were <5% for MRSA, VRE, and ESBL-producing E. coli. For routine susceptibility testing of ESBL-producing K. pneumoniae and A. baumannii against tigecycline, the broth microdilution method should be used because of the poor correlation of results between these two methods. PMID:22155819

Detection of diarrheagenic Escherichia coli strains isolated from dogs and cats in Brazil.

PubMed

Puño-Sarmiento, Juan; Medeiros, Leonardo; Chiconi, Carolina; Martins, Fernando; Pelayo, Jacinta; Rocha, Sérgio; Blanco, Jorge; Blanco, Miguel; Zanutto, Marcelo; Kobayashi, Renata; Nakazato, Gerson

2013-10-25

Escherichia coli are gut microbiota bacteria that can cause disease in some humans and other animals, including dogs and cats that humans often keep as pets. Diarrheagenic E. coli (DEC) strains are classified into six categories: enteropathogenic (EPEC), enterotoxigenic (ETEC), Shiga toxin-producing (STEC), enteroinvasive (EIEC), enteroaggregative (EAEC), and diffuse-adhering E. coli (DAEC). In this study 144 and 163 E. coli colonies were isolated from the fecal samples of 50 dogs and 50 cats, respectively, with and without diarrhea from a Veterinary Hospital (clinical isolates). The virulence factors were determined using multiplex Polymerase Chain Reaction. Adherence assays, antibacterial susceptibility and serotyping (somatic or flagellar antigens) were performed on DEC isolates. We found 25 (17.4%) and 4 (2.5%) DEC strains isolated from dogs and cats, respectively. Only the EPEC and EAEC pathotypes were found in both animals. Meanwhile, genes from other pathotypes (STEC, EIEC, and ETEC) were not found in these clinical isolates. All of the DEC strains showed mannose-resistant adherence to HEp-2 and HeLa cells, and aggregative adherence was predominant in these isolates. Multiresistant strains to antimicrobials were found in most DEC strains including usual and unusual antimicrobials in veterinary practices. The serotypes of these DEC isolates were variable. The ONT serotype was predominant in these isolates. Some serotypes found in our study were described to human DEC. Here, we demonstrate that pets carry virulent DEC genes, which are mainly strains of EPECs and EAECs. The presence of these virulence factors in isolates from animals without diarrhea suggests that pets can act as a reservoir for human infection. Copyright © 2013 Elsevier B.V. All rights reserved.

Molecular characterization of Salmonella enterica serotype Enteritidis isolates from food and human samples by serotyping, antimicrobial resistance, plasmid profiling, (GTG)5-PCR and ERIC-PCR.

PubMed

Fardsanei, F; Nikkhahi, F; Bakhshi, B; Salehi, T Z; Tamai, I A; Soltan Dallal, M M

2016-11-01

In recent years, Salmonella enterica serovar Enteritidis has been a primary cause of human salmonellosis in many countries. The major objective of this study was to investigate genetic diversity among Salmonella Enteritidis strains from different origins (food and human) by Enterobacterial Repetitive Intergenic Consensus (ERIC) -PCR, as well as to assess their plasmid profiling and antimicrobial resistance. A total of 30 Salmonella Enteritidis isolates, 15 from food samples (chicken, lamb, beef and duck meats) and 15 from clinical samples were collected in Tehran. Identification of isolates as Salmonella was confirmed by using conventional standard biochemical and serological tests. Multiplex-PCR was used for serotyping of isolates to identify Salmonella Enteritidis. Antimicrobial susceptibility testing to 16 agents founds drug resistance patterns among Salmonella Enteritidis isolates. No resistance was observed to cephalexin, ceftriaxone, ceftazidime and cefotaxime, ciprofloxacin, imipenem or meropenem, chloramphenicol and gentamicin. The highest resistance (96.7%) was observed to nitrofurantoin. Seven plasmid profiles (P1-P7) were detected, and a 68-kb plasmid was found in all isolates. Two different primers; ERIC and (GTG)5 were used for genotyping, which each produced four profiles. The majority of clinical and food isolates fell into two separate common types (CTs) with a similar percentage of 95% by ERIC-PCR. Using primer (GTG)5, 29 isolates incorporated in three CTs with 70% of isolates showing a single banding pattern. Limited genetic diversity among human and food isolates of Salmonella Enteritidis may indicate that contaminated foods were possibly the source of human salmonellosis. These results confirmed that ERIC-PCR genotyping has limited discriminatory power for Salmonella Enteritidis of different origin.

[Isolation of a strain of M. tuberculosis which is considered to be rifampicin-dependent, from a patient with long-lasted smear positive and culture difficult (SPCD) mycobacteria].

PubMed

Nakamura, M; Harano, Y; Koga, T

1990-09-01

During the course of clinical examination of drug sensitivity tests for M. tuberculosis, a strain of M. tuberculosis which is considered to be rifampicin-dependent was isolated from a patient with persisting smear positive, culture negative (SPCN) or culture difficult (SPCD) mycobacteria status. The strain isolated produced a few tiny colonies on the control Ogawa-egg yolk medium, whereas it showed abundant growth like a bacteria plaque on the medium containing rifampicin 50 micrograms/ml. Furthermore, the growth of the strain on Ogawa medium containing rifampicin 50 micrograms/ml is much better than that on the medium containing rifampicin 10 micrograms/ml.

Detection of NDM-2-producing Acinetobacter baumannii and VIM-producing Pseudomonas aeruginosa in Palestine.

PubMed

Sjölander, Isabella; Hansen, Frank; Elmanama, Abdelraouf; Khayyat, Rasha; Abu-Zant, Alaeddin; Hussein, Ayman; Taha, Adham Abu; Hammerum, Anette M; Ciofu, Oana

2014-06-01

The aim of this study was to screen for carbapenem-resistant Gram-negative bacteria in Palestine and subsequently to identify and investigate the mechanisms of resistance. For a period of 6 weeks, all Gram-negative isolates were collected from six Palestinian hospital laboratories and were tested for susceptibility using 10μg meropenem disks. Isolates showing resistance to meropenem were further investigated. The presence of carbapenemases was assessed by PCR. In addition, antimicrobial susceptibility testing, an efflux pump inhibitor assay and pulsed-field gel electrophoresis (PFGE) were performed. Isolates producing carbapenemases were further investigated by multilocus sequence typing (MLST). In total, 248 Gram-negative isolates were collected from the six laboratories. Among the 248 tested isolates, 15 Acinetobacter baumannii and 6 Pseudomonas aeruginosa were resistant to meropenem. One A. baumannii from Gaza produced NDM-2 and belonged to ST103. Thirteen of the carbapenem-resistant A. baumannii isolates possessed the intrinsic upregulated bla OXA-66 gene and one isolate carried bla OXA-51 . All but one of the OXA-66-producing A. baumannii belonged to ST2; the remaining isolate belonged to ST183. One of the carbapenem-resistant P. aeruginosa was classified as VIM-4-producing and three were VIM-2-producing isolates. The three VIM-2-producing isolates belonged to three new sequences types (ST1562, ST1563 and ST1564). All of the carbapenemase-producing isolates were multiresistant non-fermenters. To the best of our knowledge, this is the first report on NDM-producing A. baumannii and VIM-producing P. aeruginosa from Palestine. Copyright © 2013 International Society for Chemotherapy of Infection and Cancer. Published by Elsevier Ltd. All rights reserved.

Detecting imipenem resistance in Acinetobacter baumannii by automated systems (BD Phoenix, Microscan WalkAway, Vitek 2); high error rates with Microscan WalkAway

PubMed Central

2009-01-01

Background Increasing reports of carbapenem resistant Acinetobacter baumannii infections are of serious concern. Reliable susceptibility testing results remains a critical issue for the clinical outcome. Automated systems are increasingly used for species identification and susceptibility testing. This study was organized to evaluate the accuracies of three widely used automated susceptibility testing methods for testing the imipenem susceptibilities of A. baumannii isolates, by comparing to the validated test methods. Methods Selected 112 clinical isolates of A. baumanii collected between January 2003 and May 2006 were tested to confirm imipenem susceptibility results. Strains were tested against imipenem by the reference broth microdilution (BMD), disk diffusion (DD), Etest, BD Phoenix, MicroScan WalkAway and Vitek 2 automated systems. Data were analysed by comparing the results from each test method to those produced by the reference BMD test. Results MicroScan performed true identification of all A. baumannii strains while Vitek 2 unidentified one strain, Phoenix unidentified two strains and misidentified two strains. Eighty seven of the strains (78%) were resistant to imipenem by BMD. Etest, Vitek 2 and BD Phoenix produced acceptable error rates when tested against imipenem. Etest showed the best performance with only two minor errors (1.8%). Vitek 2 produced eight minor errors(7.2%). BD Phoenix produced three major errors (2.8%). DD produced two very major errors (1.8%) (slightly higher (0.3%) than the acceptable limit) and three major errors (2.7%). MicroScan showed the worst performance in susceptibility testing with unacceptable error rates; 28 very major (25%) and 50 minor errors (44.6%). Conclusion Reporting errors for A. baumannii against imipenem do exist in susceptibility testing systems. We suggest clinical laboratories using MicroScan system for routine use should consider using a second, independent antimicrobial susceptibility testing method to validate imipenem susceptibility. Etest, whereever available, may be used as an easy method to confirm imipenem susceptibility. PMID:19291298

[Characterization of isolates of carbapenemase-producing Pseudomonas aeruginosa from seven Colombian provinces].

PubMed

Saavedra, Sandra Yamile; Duarte, Carolina; González, María Nilse; Realpe, María Elena

2014-04-01

Pseudomonas aeruginosa is an opportunistic pathogen that causes multiple infections in hospitalized patients. This microorganism has developed resistance to several antimicrobial agents, including carbapenems, which are considered to be the last therapeutic option against these infections. Carbapenem resistance of P. aeruginosa is mediated by different mechanisms: Carbapenemases class B (MBL) and A, alterations in the OprD expression and overexpression of the Mex efflux pump. To describe the presence of carbapenemases in P. aeruginosa isolates from seven Colombian provinces. A total of 57 P. aeruginosa isolates were collected between September 2012 and March 2013 from national surveillance in Colombia and were sent to the Grupo de Microbiología at the Instituto Nacional de Salud (INS) for evaluation. Iidentification and antimicrobial susceptibility were confirmed through automated method (Vitek ® 2) and disk diffusion (Kirby-Bauer) according to the Clinical and Laboratory Standards Institute, CLSI, 2013. Phenotypic and genotypic confirmation was determined using the modified Hodge test (MHT), a synergism test using imipenem, EDTA-SMA and meropenem, and conventional PCR to detect the bla KPC, bla VIM, bla IMP and bla NDM genes. Of the 57 isolates, two showed sensitivity to carbapenems. Forty-three isolates were positive for carbapenemases with a high percentage of sensitivity to colistin (76.4%, n=42). The 43 isolates producing carbapenemases showed multiple drug resistance: 72.1% were positive in the MHT and 79.1% showed MBL synergism. PCR amplification confirmed that 33 isolates were positive for bla VIM, nine were positive for bla KPC and one isolate expressed both KPC and VIM carbapenemases. No isolates showed amplified products with bla IMP and bla NDM primers. The most frequent carbapenemase was VIM, followed by KPC in an approximate ratio of 3:1.

Activity of fosfomycin against nosocomial multiresistant bacterial pathogens from Croatia: a multicentric study

PubMed Central

Bielen, Luka; Likić, Robert; Erdeljić, Viktorija; Mareković, Ivana; Firis, Nataša; Grgić-Medić, Marijana; Godan, Ana; Tomić, Ivan; Hunjak, Blaženka; Markotić, Alemka; Bejuk, Danijela; Tičić, Vladimira; Balzar, Silvana; Bedenić, Branka

2018-01-01

Aim To determine in vitro susceptibility of multiresistant bacterial isolates to fosfomycin. Methods In this prospective in vitro study (local non-random sample, level of evidence 3), 288 consecutively collected multiresistant bacterial isolates from seven medical centers in Croatia were tested from February 2014 until October 2016 for susceptibility to fosfomycin and other antibiotics according to Clinical and Laboratory Standards Institute methodology. Susceptibility to fosfomycin was determined by agar dilution method, while disc diffusion were performed for in vitro testing of other antibiotics. Polymerase chain reaction and sequencing was performed for the majority of extended spectrum β-lactamase (ESBL)-producing Klebsiella pneumoniae (K. pneumoniae) and carbapenem-resistant isolates. Results The majority of 288 multiresistant bacterial isolates (82.6%) were susceptible to fosfomycin. The 236 multiresistant Gram-negative isolates showed excellent susceptibility to fosfomycin. Susceptibility rates were as follows: Escherichia coli ESBL 97%, K. pneumoniae ESBL 80%, Enterobacter species 85.7%, Citrobacter freundii 100%, Proteus mirabilis 93%, and Pseudomonas aeruginosa 60%. Of the 52 multiresistant Gram-positive isolates, methicillin-resistant Staphylococcus aureus showed excellent susceptibility to fosfomycin (94.4%) and vancomycin-resistant enterococcus showed low susceptibility to fosfomycin (31%). Polymerase chain reaction analysis of 36/50 ESBL-producing K. pneumoniae isolates showed that majority of isolates had CTX-M-15 beta lactamase (27/36) preceded by ISEcp insertion sequence. All carbapenem-resistant Enterobacter and Citrobacter isolates had blaVIM-1 metallo-beta-lactamase gene. Conclusion With the best in vitro activity among the tested antibiotics, fosfomycin could be an effective treatment option for infections caused by multiresistant Gram-negative and Gram-positive bacterial strains in the hospital setting. PMID:29740989

Genetic and biochemical characterization of an acquired subgroup B3 metallo-β-lactamase gene, blaAIM-1, and its unique genetic context in Pseudomonas aeruginosa from Australia.

PubMed

Yong, Dongeun; Toleman, Mark A; Bell, Jan; Ritchie, Brett; Pratt, Rachael; Ryley, Henry; Walsh, Timothy R

2012-12-01

Three clinical Pseudomonas aeruginosa isolates (WCH2677, WCH2813, and WCH2837) isolated from the Women's and Children's Hospital, Adelaide, Australia, produced a metallo-β-lactamase (MBL)-positive Etest result. All isolates were PCR negative for known MBL genes. A gene bank was created, and an MBL gene, designated bla(AIM-1), was cloned and fully characterized. The encoded enzyme, AIM-1, is a group B3 MBL that has the highest level of identity to THIN-B and L1. It is chromosomal and flanked by two copies (one intact and one truncated) of an ISCR element, ISCR15. Southern hybridization studies indicated the movement of both ISCR15 and bla(AIM-1) within the three different clinical isolates. AIM-1 hydrolyzes most β-lactams, with the exception of aztreonam and, to a lesser extent, ceftazidime; however, it possesses significantly higher k(cat) values for cefepime and carbapenems than most other MBLs. AIM-1 was the first mobile group B3 enzyme detected and signals further problems for already beleaguered antimicrobial regimes to treat serious P. aeruginosa and other Gram-negative infections.

UV red fluorescence of Eubacterium lentum.

PubMed Central

Mosca, A; Strong, C A; Finegold, S M

1993-01-01

Twenty-nine clinical isolates of Eubacterium lentum and two type species were evaluated for the ability to fluoresce under UV light. Twenty-one of the 29 isolates and both of the reference strains showed orange-to-red fluorescence. This fluorescence did not require blood or hemin in the culture media and did not fade upon air exposure. The fluorescent pigment, after extraction by 1 N NaOH, showed peak excitation at a wavelength of around 400 nm. The capacity of E. lentum to produce fluorescence may be a useful and time-sparing laboratory aid for its identification. PMID:8463378

Clinical Features and Molecular Epidemiology of CMY-Type β-Lactamase-Producing Escherichia coli

PubMed Central

Sidjabat, Hanna E.; Paterson, David L.; Qureshi, Zubair A.; Adams-Haduch, Jennifer M.; O’Keefe, Alexandra; Pascual, Alvaro; Rodríguez-Baño, Jesús; Doi, Yohei

2009-01-01

Background Knowledge on the clinical features of infections caused by Escherichia coli producing plasmid-mediated AmpC β-lactamase is limited. Of the several groups of plasmid-mediated AmpC β-lactamase, CMY-type β-lactamase is the most common in the United States. Methods We prospectively identified E. coli producing CMY-type β-lactamase and collected clinical data over a seven-month period. A retrospective cohort study was performed to identify features associated with these cases, using cases due to extended-spectrum β-lactamase (ESBL)-producing E. coli as controls. Pulsed-field gel electrophoresis (PFGE), plasmid analysis and phylogenetic typing were performed. Results Twenty-two cases with CMY-producing E. coli and 25 cases with ESBL-producing E. coli were identified. The demographics of the patients were similar between the CMY and ESBL cohorts. CMY cases were significantly more likely to represent symptomatic infection compared with ESBL cases (P=0.028). The CMY-type β-lactamase was identified as CMY-2 or its variants. Ninety-four percent of the CMY-producing isolates belonged to E. coli phylogenetic groups B2 and D, which are associated with virulence. Many of them shared similar plasmid profiles, whereas the PFGE profiles were diverse. Co-resistance to non-β-lactam antimicrobials was common. Conclusion In Pittsburgh, CMY-producing E. coli is almost as common as ESBL-producing E. coli and causes symptomatic infection in the majority of cases. PMID:19187027

Assessment of Minimum Inhibitory Concentrations of Telavancin by Revised Broth Microdilution Method in Phase 3 Hospital-Acquired Pneumonia/Ventilator-Associated Pneumonia Clinical Isolates.

PubMed

Smart, Jennifer I; Corey, Gordon Ralph; Stryjewski, Martin E; Wang, Whedy; Barriere, Steven L

2016-12-01

The broth microdilution method (BMD) for testing telavancin minimum inhibitory concentrations (MICs) was revised (rBMD) in 2014 to improve the accuracy, precision, and reproducibility of the testing method. The aim of this study was to determine the effect of the revised method on telavancin MIC values for Staphylococcus aureus (S. aureus) clinical isolates obtained from hospital-acquired pneumonia (HAP) patients. Isolates from patients who participated in the phase 3 Assessment of Telavancin for Treatment of HAP Studies were retested using the rBMD method. Retesting of 647 isolates produced a range of telavancin MIC values from 0.015 µg/mL to 0.12 µg/mL with MIC 50/90 values of 0.06/0.06 µg/mL for the total pool of samples. For methicillin-resistant S. aureus (MRSA), MIC 50/90 values were 0.06/0.12 µg/mL. These values are up to 4-fold lower than MIC 50/90 values obtained using the original method. These results were used in part to justify lowering the telavancin breakpoints. All tested isolates remained susceptible to telavancin at the revised susceptibility breakpoint of ≤0.12 µg/mL. Overall, the clinical cure rate for microbiologically evaluable telavancin-treated patients was 78% for S. aureus, 76% for patients with MRSA, and 79% for patients with isolates with reduced susceptibility to vancomycin (MIC ≥1 µg/mL). Results from the rBMD method support the in vitro potency of telavancin against S. aureus. ATTAIN (NCT00107952 and NCT00124020). Theravance Biopharma Antibiotics, Inc.

Addiction of Hypertransformable Pneumococcal Isolates to Natural Transformation for In Vivo Fitness and Virulence

PubMed Central

Li, Guiling; Liang, Zhuowen; Wang, Xiatai; Yang, Yonghong; Shao, Zhujun; Li, Machao; Ma, Yueyun; Qu, Fen; Morrison, Donald A.

2016-01-01

Natural genetic transformation of Streptococcus pneumoniae, an important human pathogen, mediates horizontal gene transfer for the development of drug resistance, modulation of carriage and virulence traits, and evasion of host immunity. Transformation frequency differs greatly among pneumococcal clinical isolates, but the molecular basis and biological importance of this interstrain variability remain unclear. In this study, we characterized the transformation frequency and other associated phenotypes of 208 S. pneumoniae clinical isolates representing at least 30 serotypes. While the vast majority of these isolates (94.7%) were transformable, the transformation frequency differed by up to 5 orders of magnitude between the least and most transformable isolates. The strain-to-strain differences in transformation frequency were observed among many isolates producing the same capsule types, indicating no general association between transformation frequency and serotype. However, a statistically significant association was observed between the levels of transformation and colonization fitness/virulence in the hypertransformable isolates. Although nontransformable mutants of all the selected hypertransformable isolates were significantly attenuated in colonization fitness and virulence in mouse infection models, such mutants of the strains with relatively low transformability had no or marginal fitness phenotypes under the same experimental settings. This finding strongly suggests that the pneumococci with high transformation capability are “addicted” to a “hypertransformable” state for optimal fitness in the human host. This work has thus provided an intriguing hint for further investigation into how the competence system impacts the fitness, virulence, and other transformation-associated traits of this important human pathogen. PMID:27068094

A strain of Siniperca chuatsi rhabdovirus causes high mortality among cultured Largemouth Bass in South China.

PubMed

Ma, Dongmei; Deng, Guocheng; Bai, Junjie; Li, Shengjie; Yu, Lingyun; Quan, Yingchun; Yang, Xiaojing; Jiang, Xiaoyan; Zhu, Zemin; Ye, Xing

2013-09-01

In April 2011, 40% mortality of Largemouth Bass Micropterus salmoides juveniles occurred at a farm of Zhongshan City, Guangdong Province, China. Infected fish became lethargic, exhibited corkscrew and irregular swimming, and developed a distended abdomen and crooked body. Fish began to die within 2 d after the appearance of clinical signs. In order to analyze the pathogeny and diagnose the disease earlier, observation of clinical signs, cell infection, titer calculation, electron microscopy, immersion infection assay for fish, and nucleotide sequence analysis were carried out. Fathead minnow (FHM) cell cultures, inoculated with filtrate of liver and spleen homogenates from the diseased fish, developed the obvious cytopathic effect 46 h after inoculation in the primary culture and 24 h at the first passage. Typical rhabdovirus particles, 115-143 nm in length and 62-78 nm in diameter, were observed in infected FHM cells by direct transmission electron microscopy. The isolated virus produced a titer of 10(7.15) TCID50/mL. Immersion-Fish infected with the virus had similar clinical signs and 80% mortality with 10(2.5) LD50/mL. The data indicated that the rhabdovirus was the lethal pathogeny of the current disease. Based on nucleoprotein-gene nucleotide sequence multiple alignment analysis, the newly isolated virus is a strain of Siniperca chuatsi rhabdovirus (SCRV) under family Rhabdoviridae, which was initially isolated from Mandarin Fish Siniperca chuatsi. Up to the present, at least four virus strains have been isolated from diseased Largemouth Bass, which have had different clinical signs. Comparison of the clinical signs can help in an early diagnosis of the disease.

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.

Methylene blue induced morphological deformations in Plasmodium falciparum gametocytes: implications for transmission-blocking.

PubMed

Wadi, Ishan; Pillai, C Radhakrishna; Anvikar, Anupkumar R; Sinha, Abhinav; Nath, Mahendra; Valecha, Neena

2018-01-08

Malaria remains a global health problem despite availability of effective tools. For malaria elimination, drugs targeting sexual stages of Plasmodium falciparum need to be incorporated in treatment regimen along with schizonticidal drugs to interrupt transmission. Primaquine is recommended as a transmission blocking drug for its effect on mature gametocytes but is not extensively utilized because of associated safety concerns among glucose-6-phosphate dehydrogenase (G6PD) deficient patients. In present work, methylene blue, which is proposed as an alternative to primaquine is investigated for its gametocytocidal activity amongst Indian field isolates. An effort has been made to establish Indian field isolates of P. falciparum as in vitro model for gametocytocidal drugs screening. Plasmodium falciparum isolates were adapted to in vitro culture and induced to gametocyte production by hypoxanthine and culture was enriched for gametocyte stages using N-acetyl-glucosamine. Gametocytes were incubated with methylene blue for 48 h and stage specific gametocytocidal activity was evaluated by microscopic examination. Plasmodium falciparum field isolates RKL-9 and JDP-8 were able to reproducibly produce gametocytes in high yield and were used to screen gametocytocidal drugs. Methylene blue was found to target gametocytes in a concentration dependent manner by either completely eliminating gametocytes or rendering them morphologically deformed with mean IC 50 (early stages) as 424.1 nM and mean IC 50 (late stages) as 106.4 nM. These morphologically altered gametocytes appeared highly degenerated having shrinkage, distortions and membrane deformations. Field isolates that produce gametocytes in high yield in vitro can be identified and used to screen gametocytocidal drugs. These isolates should be used for validation of gametocytocidal hits obtained previously by using lab adapted reference strains. Methylene blue was found to target gametocytes produced from Indian field isolates and is proposed to be used as a gametocytocidal adjunct with artemisinin-based combination therapy. Further exploration of methylene blue in clinical studies amongst Indian population, including G6PD deficient patients, is recommended.

Prevalence of quinolone resistance mechanisms in Enterobacteriaceae producing acquired AmpC β-lactamases and/or carbapenemases in Spain.

PubMed

Machuca, Jesús; Agüero, Jesús; Miró, Elisenda; Conejo, María Del Carmen; Oteo, Jesús; Bou, Germán; González-López, Juan José; Oliver, Antonio; Navarro, Ferran; Pascual, Álvaro; Martínez-Martínez, Luis

2017-10-01

Quinolone resistance in Enterobacteriaceae species has increased over the past few years, and is significantly associated to beta-lactam resistance. The aim of this study was to evaluate the prevalence of chromosomal- and plasmid-mediated quinolone resistance in acquired AmpC β-lactamase and/or carbapenemase-producing Enterobacteriaceae isolates. The presence of chromosomal- and plasmid-mediated quinolone resistance mechanisms [mutations in the quinolone resistance determining region (QRDR) of gyrA and parC and qnr, aac(6')-Ib-cr and qepA genes] was evaluated in 289 isolates of acquired AmpC β-lactamase- and/or carbapenemase-producing Enterobacteriaceae collected between February and July 2009 in 35 Spanish hospitals. Plasmid mediated quinolone resistance (PMQR) genes were detected in 92 isolates (31.8%), qnr genes were detected in 83 isolates (28.7%), and the aac(6')-Ib-cr gene was detected in 20 isolates (7%). qnrB4 gene was the most prevalent qnr gene detected (20%), associated, in most cases, with DHA-1. Only 14.6% of isolates showed no mutations in gyrA or parC with a ciprofloxacin MIC of 0.5mg/L or higher, whereas PMQR genes were detected in 90% of such isolates. qnrB4 gene was the most prevalent PMQR gene detected, and was significantly associated with acquired AmpC β-lactamase DHA-1. PMQR determinants in association with other chromosomal-mediated quinolone resistance mechanisms, different to mutations in gyrA and parC (increased energy-dependent efflux, altered lipopolysaccharide or porin loss), could lead to ciprofloxacin MIC values that exceed breakpoints established by the main international committees to define clinical antimicrobial susceptibility breakpoints. Copyright © 2016 Elsevier España, S.L.U. y Sociedad Española de Enfermedades Infecciosas y Microbiología Clínica. All rights reserved.