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Sample records for enterococcus faecalis illustrated

  1. Genetic Diversity among Enterococcus faecalis

    PubMed Central

    McBride, Shonna M.; Fischetti, Vincent A.; LeBlanc, Donald J.; Moellering, Robert C.; Gilmore, Michael S.

    2007-01-01

    Enterococcus faecalis, a ubiquitous member of mammalian gastrointestinal flora, is a leading cause of nosocomial infections and a growing public health concern. The enterococci responsible for these infections are often resistant to multiple antibiotics and have become notorious for their ability to acquire and disseminate antibiotic resistances. In the current study, we examined genetic relationships among 106 strains of E. faecalis isolated over the past 100 years, including strains identified for their diversity and used historically for serotyping, strains that have been adapted for laboratory use, and isolates from previously described E. faecalis infection outbreaks. This collection also includes isolates first characterized as having novel plasmids, virulence traits, antibiotic resistances, and pathogenicity island (PAI) components. We evaluated variation in factors contributing to pathogenicity, including toxin production, antibiotic resistance, polymorphism in the capsule (cps) operon, pathogenicity island (PAI) gene content, and other accessory factors. This information was correlated with multi-locus sequence typing (MLST) data, which was used to define genetic lineages. Our findings show that virulence and antibiotic resistance traits can be found within many diverse lineages of E. faecalis. However, lineages have emerged that have caused infection outbreaks globally, in which several new antibiotic resistances have entered the species, and in which virulence traits have converged. Comparing genomic hybridization profiles, using a microarray, of strains identified by MLST as spanning the diversity of the species, allowed us to identify the core E. faecalis genome as consisting of an estimated 2057 unique genes. PMID:17611618

  2. Targeting Enterococcus faecalis biofilms with phage therapy.

    PubMed

    Khalifa, Leron; Brosh, Yair; Gelman, Daniel; Coppenhagen-Glazer, Shunit; Beyth, Shaul; Poradosu-Cohen, Ronit; Que, Yok-Ai; Beyth, Nurit; Hazan, Ronen

    2015-04-01

    Phage therapy has been proven to be more effective, in some cases, than conventional antibiotics, especially regarding multidrug-resistant biofilm infections. The objective here was to isolate an anti-Enterococcus faecalis bacteriophage and to evaluate its efficacy against planktonic and biofilm cultures. E. faecalis is an important pathogen found in many infections, including endocarditis and persistent infections associated with root canal treatment failure. The difficulty in E. faecalis treatment has been attributed to the lack of anti-infective strategies to eradicate its biofilm and to the frequent emergence of multidrug-resistant strains. To this end, an anti-E. faecalis and E. faecium phage, termed EFDG1, was isolated from sewage effluents. The phage was visualized by electron microscopy. EFDG1 coding sequences and phylogeny were determined by whole genome sequencing (GenBank accession number KP339049), revealing it belongs to the Spounavirinae subfamily of the Myoviridae phages, which includes promising candidates for therapy against Gram-positive pathogens. This analysis also showed that the EFDG1 genome does not contain apparent harmful genes. EFDG1 antibacterial efficacy was evaluated in vitro against planktonic and biofilm cultures, showing effective lytic activity against various E. faecalis and E. faecium isolates, regardless of their antibiotic resistance profile. In addition, EFDG1 efficiently prevented ex vivo E. faecalis root canal infection. These findings suggest that phage therapy using EFDG1 might be efficacious to prevent E. faecalis infection after root canal treatment. PMID:25662974

  3. Targeting Enterococcus faecalis Biofilms with Phage Therapy

    PubMed Central

    Khalifa, Leron; Brosh, Yair; Gelman, Daniel; Coppenhagen-Glazer, Shunit; Beyth, Shaul; Poradosu-Cohen, Ronit; Que, Yok-Ai; Beyth, Nurit

    2015-01-01

    Phage therapy has been proven to be more effective, in some cases, than conventional antibiotics, especially regarding multidrug-resistant biofilm infections. The objective here was to isolate an anti-Enterococcus faecalis bacteriophage and to evaluate its efficacy against planktonic and biofilm cultures. E. faecalis is an important pathogen found in many infections, including endocarditis and persistent infections associated with root canal treatment failure. The difficulty in E. faecalis treatment has been attributed to the lack of anti-infective strategies to eradicate its biofilm and to the frequent emergence of multidrug-resistant strains. To this end, an anti-E. faecalis and E. faecium phage, termed EFDG1, was isolated from sewage effluents. The phage was visualized by electron microscopy. EFDG1 coding sequences and phylogeny were determined by whole genome sequencing (GenBank accession number KP339049), revealing it belongs to the Spounavirinae subfamily of the Myoviridae phages, which includes promising candidates for therapy against Gram-positive pathogens. This analysis also showed that the EFDG1 genome does not contain apparent harmful genes. EFDG1 antibacterial efficacy was evaluated in vitro against planktonic and biofilm cultures, showing effective lytic activity against various E. faecalis and E. faecium isolates, regardless of their antibiotic resistance profile. In addition, EFDG1 efficiently prevented ex vivo E. faecalis root canal infection. These findings suggest that phage therapy using EFDG1 might be efficacious to prevent E. faecalis infection after root canal treatment. PMID:25662974

  4. Characterization of Enterococcus faecalis and Enterococcus faecium from wild flowers.

    PubMed

    Sánchez Valenzuela, Antonio; Benomar, Nabil; Abriouel, Hikmate; Pérez Pulido, Rubén; Martínez Cañamero, Magdalena; Gálvez, Antonio

    2012-05-01

    Wild flowers in the South of Spain were screened for Enterococcus faecalis and Enterococcus faecium. Enterococci were frequently associated with prickypear and fieldpoppy flowers. Forty-six isolates, from 8 different flower species, were identified as E. faecalis (28 isolates) or E. faecium (18 isolates) and clustered in well-defined groups by ERIC-PCR fingerprinting. A high incidence of antibiotic resistance was detected among the E. faecalis isolates, especially to quinupristin/dalfopristin (75%), rifampicin (68%) and ciprofloxacin (57%), and to a lesser extent to levofloxacin (35.7%), erythromycin (28.5%), tetracycline (3.5%), chloramphenicol (3.5%) and streptomycin (3.5%). Similar results were observed for E. faecium isolates, except for a higher incidence of resistance to tetracycline (17%) and lower to erythromycin (11%) or quinupristin/dalfopristin (22%). Vancomycin or teicoplanin resistances were not detected. Most isolates (especially E. faecalis) were proteolytic and carried the gelatinase gene gelE. Genes encoding other potential virulence factors (ace, efaA (fs), ccf and cpd) were frequently detected. Cytolysin genes were mainly detected in a few haemolytic E. faecium isolates, three of which also carried the collagen adhesin acm gene. Hyaluronidase gene (hyl ( Efm )) was detected in two isolates. Many isolates produced bacteriocins and carried genes for enterocins A, B, and L50 mainly. The similarities found between enterococci from wild flowers and those from animal and food sources raise new questions about the puzzling lifestyle of these commensals and opportunistic pathogens. PMID:22183298

  5. The stress proteome of Enterococcus faecalis.

    PubMed

    Giard, J C; Laplace, J M; Rincé, A; Pichereau, V; Benachour, A; Leboeuf, C; Flahaut, S; Auffray, Y; Hartke, A

    2001-08-01

    Enterococcus faecalis is a resident bacterium of the intestinal tract of humans and animals. This bacterium can be responsible for serious diseases and is one of the largest causes of hospital-based infections. This hardy organism resists many kinds of stresses and is used as a major indicator of the hygienic quality of food, milk, and drinking water. On the other side, enterococci seem to have beneficial role in the development of cheese aroma and are added in certain starter cultures. Since ten years, our laboratory has used the two-dimensional electrophoresis (2-DE) technique to study the response of E. faecalis to physical or chemical stresses as well as to glucose and total starvation. Twenty-seven protein spots on 2-D gels have been identified by N-terminal sequencing or Western blotting which make up the first proteome database of this species. The proteins were classified in four different groups according to their function and their regulation. The first group comprises well-characterized proteins with known protective functions towards stresses. The second group contains enzymes of catabolic pathways. Their implication in stress resistance seems not obvious. A third group are proteins induced in glucose-starved cells belonging to the CcpA regulon. Induction of these enzymes under starvation may serve to increase the scavenging capacity of the cells for nutrients or may be important to mobilize endogenous energetic reserves. Lastly, nine N-terminal amino acid sequences or open reading frames (ORF) showed no homologies with sequences in databases. A comprehensive description of stress proteins of E. faecalis and analysis of their patterns of expression under different environmental conditions would greatly increase our understanding of the molecular mechanisms underlying the extraordinary capacity of this bacterium to survive under hostile conditions. PMID:11565789

  6. Antimicrobial Susceptibility Patterns of Enterococcus faecalis and Enterococcus faecium Isolated from Poultry Flocks in Germany.

    PubMed

    Maasjost, J; Mühldorfer, K; Cortez de Jäckel S; Hafez, H M

    2015-03-01

    Between 2010 and 2011, 145 Enterococcus isolates (Enterococcus faecalis, n = 127; Enterococcus faecium, n = 18) were collected during routine bacteriologic diagnostics from broilers, layers, and fattening turkeys in Germany showing various clinical signs. The susceptibility to 24 antimicrobial agents was investigated by broth microdilution test to determine minimum inhibitory concentrations (MICs). All E. faecalis isolates (n = 127) were susceptible to the beta-lactam antibiotics ampicillin, amoxicillin-clavulanic acid, and penicillin. Corresponding MIC with 50% inhibition (MIC50) and MIC with 90% inhibition (MIC90) values of these antimicrobial agents were at the lower end of the test range (≤ 4 μg/ml). In addition, no vancomycin-resistant enterococci (VRE) were found. High resistance rates were identified in both Enterococcus species for lincomycin (72%-99%) and tetracycline (67%-82%). Half or more than half of Enterococcus isolates were resistant to gentamicin (54%-72%) and the macrolide antibiotics erythromycin (44%-61%) and tylosin-tartate (44%-56%). Enterococcus faecalis isolated from fattening turkeys showed the highest prevalence of antimicrobial resistance compared to other poultry production systems. Eighty-nine out of 145 Enterococcus isolates were resistant to three or more antimicrobial classes. Again, turkeys stood out with 42 (8 1%) multiresistant isolates. The most-frequent resistance patterns of E. faecalis were gentamicin, lincomycin, and tetracycline in all poultry production systems. PMID:26292548

  7. An antimicrobial peptidoglycan hydrolase for treating Enterococcus faecalis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Enterococcus faecalis is an intestinal bacteria species that can become an opportunistic pathogen in humans and farm animals with antibiotic resistant strains becoming increasingly common. In farm animals, strong antimicrobials, such as Vancomycin, should not be used due to the risk of propagation ...

  8. Transcriptome analysis of Enterococcus faecalis in response to alkaline stress

    PubMed Central

    Ran, Shujun; Liu, Bin; Jiang, Wei; Sun, Zhe; Liang, Jingping

    2015-01-01

    Enterococcus faecalis is the most commonly isolated species from endodontic failure root canals; its persistence in treated root canals has been attributed to its ability to resist high pH stress. The goal of this study was to characterize the E. faecalis transcriptome and to identify candidate genes for response and resistance to alkaline stress using Illumina HiSeq 2000 sequencing. We found that E. faecalis could survive and form biofilms in a pH 10 environment and that alkaline stress had a great impact on the transcription of many genes in the E. faecalis genome. The transcriptome sequencing results revealed that 613 genes were differentially expressed (DEGs) for E. faecalis grown in pH 10 medium; 211 genes were found to be differentially up-regulated and 402 genes differentially down-regulated. Many of the down-regulated genes found are involved in cell energy production and metabolism and carbohydrate and amino acid metabolism, and the up-regulated genes are mostly related to nucleotide transport and metabolism. The results presented here reveal that cultivation of E. faecalis in alkaline stress has a profound impact on its transcriptome. The observed regulation of genes and pathways revealed that E. faecalis reduced its carbohydrate and amino acid metabolism and increased nucleotide synthesis to adapt and grow in alkaline stress. A number of the regulated genes may be useful candidates for the development of new therapeutic approaches for the treatment of E. faecalis infections. PMID:26300863

  9. Comparative genomics of Enterococcus faecalis from healthy Norwegian infants

    PubMed Central

    Solheim, Margrete; Aakra, Ågot; Snipen, Lars G; Brede, Dag A; Nes, Ingolf F

    2009-01-01

    Background Enterococcus faecalis, traditionally considered a harmless commensal of the intestinal tract, is now ranked among the leading causes of nosocomial infections. In an attempt to gain insight into the genetic make-up of commensal E. faecalis, we have studied genomic variation in a collection of community-derived E. faecalis isolated from the feces of Norwegian infants. Results The E. faecalis isolates were first sequence typed by multilocus sequence typing (MLST) and characterized with respect to antibiotic resistance and properties associated with virulence. A subset of the isolates was compared to the vancomycin resistant strain E. faecalis V583 (V583) by whole genome microarray comparison (comparative genomic hybridization (CGH)). Several of the putative enterococcal virulence factors were found to be highly prevalent among the commensal baby isolates. The genomic variation as observed by CGH was less between isolates displaying the same MLST sequence type than between isolates belonging to different evolutionary lineages. Conclusion The variations in gene content observed among the investigated commensal E. faecalis is comparable to the genetic variation previously reported among strains of various origins thought to be representative of the major E. faecalis lineages. Previous MLST analysis of E. faecalis have identified so-called high-risk enterococcal clonal complexes (HiRECC), defined as genetically distinct subpopulations, epidemiologically associated with enterococcal infections. The observed correlation between CGH and MLST presented here, may offer a method for the identification of lineage-specific genes, and may therefore add clues on how to distinguish pathogenic from commensal E. faecalis. In this work, information on the core genome of E. faecalis is also substantially extended. PMID:19393078

  10. Identification of Enterococcus faecalis antigens specifically expressed in vivo

    PubMed Central

    Shet, Uttom K.; Park, Sang-Won; Lim, Hyun-Pil; Yun, Kwi-Dug; Kang, Seong Soo; Kim, Se Eun

    2015-01-01

    Objectives Molecular mechanism of the pathogenicity of Enterococcus faecalis (E. faecalis), a suspected endodontic pathogen, has not yet been adequately elucidated due to limited information on its virulence factors. Here we report the identification of in vivo expressed antigens of E. faecalis by using a novel immunoscreening technique called change-mediated antigen technology (CMAT) and an experimental animal model of endodontic infection. Materials and Methods Among 4,500 E. coli recombinant clones screened, 19 positive clones reacted reproducibly with hyperimmune sera obtained from rabbits immunized with E. faecalis cells isolated from an experimental endodontic infection. DNA sequences from 16 of these in vivo-induced (IVI) genes were determined. Results Identified protein antigens of E. faecalis included enzymes involved in housekeeping functions, copper resistance protein, putative outer membrane proteins, and proteins of unknown function. Conclusions In vivo expressed antigens of E. faecalis could be identified by using a novel immune-screening technique CMAT and an experimental animal model of endodontic infection. Detailed analysis of these IVI genes will lead to a better understanding of the molecular mechanisms involved in the endodontic infection of E. faecalis. PMID:26587417

  11. Postneurosurgical Central Nervous System Infection Due to Enterococcus faecalis Successfully Treated With Intraventricular Vancomycin

    PubMed Central

    Patel, Trisha; Lewis, Mark E.; Niesley, Michelle L.; Chowdhury, Mashiul

    2016-01-01

    Abstract Infections from Enterococcus faecalis and Enterococcus faecium are uncommon in the post-neurosurgical intervention setting., [1, 2, 3, 4] Intraventricular antibiotics are recommended when standard intravenous therapy fails. [5] Here we present a case of post-neurosurgical ventriculitis, meningitis, and cerebritis in an oncology patient caused by refractory Enterococcus faecalis successfully treated with intraventricular vancomycin. PMID:27226704

  12. Endocarditis and biofilm-associated pili of Enterococcus faecalis

    PubMed Central

    Nallapareddy, Sreedhar R.; Singh, Kavindra V.; Sillanpää, Jouko; Garsin, Danielle A.; Höök, Magnus; Erlandsen, Stanley L.; Murray, Barbara E.

    2006-01-01

    Increasing multidrug resistance in Enterococcus faecalis, a nosocomial opportunist and common cause of bacterial endocarditis, emphasizes the need for alternative therapeutic approaches such as immunotherapy or immunoprophylaxis. In an earlier study, we demonstrated the presence of antibodies in E. faecalis endocarditis patient sera to recombinant forms of 9 E. faecalis cell wall–anchored proteins; of these, we have now characterized an in vivo–expressed locus of 3 genes and an associated sortase gene (encoding sortase C; SrtC). Here, using mutation analyses and complementation, we demonstrated that both the ebp (encoding endocarditis and biofilm-associated pili) operon and srtC are important for biofilm production of E. faecalis strain OG1RF. In addition, immunogold electron microscopy using antisera against EbpA–EbpC proteins as well as patient serum demonstrated that E. faecalis produces pleomorphic surface pili. Assembly of pili and their cell wall attachment appeared to occur via a mechanism of cross-linking of the Ebp proteins by the designated SrtC. Importantly, a nonpiliated, allelic replacement mutant was significantly attenuated in an endocarditis model. These biologically important surface pili, which are antigenic in humans during endocarditis and encoded by a ubiquitous E. faecalis operon, may be a useful immunotarget for studies aimed at prevention and/or treatment of this pathogen. PMID:17016560

  13. Ecology of Enterococcus faecalis and niche adapted or non-niche-adapted Enterococcus faecium in continuous-flow anaerobic cultures

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Objectives: To study the survivability of niche adapted Enterococcus faecium I.3rif (I.3rif) vs. non-niche adapted Enterococcus faecium (GRE47) in cultures that contain Enterococcus faecalis I.2. Methods: An anaerobic continuous-flow culture of chicken microflora (CCF) that models the chicken gastr...

  14. Antibacterial Effect of Diclofenac Sodium on Enterococcus faecalis

    PubMed Central

    Salem-Milani, Amin; Balaei-Gajan, Esrafil; Rahimi, Saeed; Moosavi, Zohreh; Abdollahi, Ardalan; Zakeri-Milani, Parvin; Bolourian, Mehrdad

    2013-01-01

    Objective: Non-steroidal anti-inflammatory drugs (NSAIDs) have shown antibacterial activity in some recent studies. The aim of this study was to evaluate the antibacterial effect of diclofenac against Enterococcus faecalis (E. faecalis) as a resistant endodontic bacterium in comparison with ibuprofen, calcium hydroxide and amoxicillin. Materials and Methods: The antibacterial activity of materials was evaluated using agar diffusion test and tube dilution method. Mixtures of 400 mg/ml of materials were prepared. The bacteria were seeded on 10 Muller-Hinton agar culture plates. Thirty microliter of each test material was placed in each well punched in agar plates. After incubation, the zone of bacterial inhibition was measured. Minimum inhibitory concentration (MIC) of the test materials was determined by agar dilution method. One-way Analysis of Variance (ANOVA) followed by Sidak post hoc test was used to compare the mean zone of microbial growth in the groups. Results: There were significant differences between the two groups (p< 0.05). Results of the agar diffusion test showed that antibiotics (amoxicillin, gentamycin) had the greatest antibacterial activity followed by NSAIDs (ibuprofen, diclofenac). Ca(OH)2 failed to show antibacterial activity. Diclofenac and ibuprofen showed distinct antibacterial activity against E. faecalis in 50 μg/ml and above concentrations. Conclusion: Within the limitations of this in vitro study, it is concluded that diclofenac and ibuprofen have significantly more pronounced antibacterial activity against E. faecalis in comparison with Ca(OH)2. PMID:23724199

  15. Genes Important for Catalase Activity in Enterococcus faecalis

    PubMed Central

    Baureder, Michael; Hederstedt, Lars

    2012-01-01

    Little in general is known about how heme proteins are assembled from their constituents in cells. The Gram-positive bacterium Enterococcus faecalis cannot synthesize heme and does not depend on it for growth. However, when supplied with heme in the growth medium the cells can synthesize two heme proteins; catalase (KatA) and cytochrome bd (CydAB). To identify novel factors important for catalase biogenesis libraries of E. faecalis gene insertion mutants were generated using two different types of transposons. The libraries of mutants were screened for clones deficient in catalase activity using a colony zymogram staining procedure. Analysis of obtained clones identified, in addition to katA (encoding the catalase enzyme protein), nine genes distributed over five different chromosomal loci. No factors with a dedicated essential role in catalase biogenesis or heme trafficking were revealed, but the results indicate the RNA degradosome (srmB, rnjA), an ABC-type oligopeptide transporter (oppBC), a two-component signal transducer (etaR), and NADH peroxidase (npr) as being important for expression of catalase activity in E. faecalis. It is demonstrated that catalase biogenesis in E. faecalis is independent of the CydABCD proteins and that a conserved proline residue in the N-terminal region of KatA is important for catalase assembly. PMID:22590595

  16. Candida albicans and Enterococcus faecalis in the gut

    PubMed Central

    Garsin, Danielle A; Lorenz, Michael C

    2013-01-01

    The fungus Candida albicans and the gram-positive bacterium Enterococcus faecalis are both normal residents of the human gut microbiome and cause opportunistic disseminated infections in immunocompromised individuals. Using a nematode infection model, we recently showed that co-infection resulted in less pathology and less mortality than infection with either species alone and this was partly explained by an interkingdom signaling event in which a bacterial-derived product inhibits hyphal morphogenesis of C. albicans. In this addendum we discuss these findings in the contest of other described bacterial-fungal interactions and recent data suggesting a potentially synergistic relationship between these two species in the mouse gut as well. We suggest that E. faecalis and C. albicans promote a mutually beneficial association with the host, in effect choosing a commensal lifestyle over a pathogenic one. PMID:23941906

  17. In vitro alkaline pH resistance of Enterococcus faecalis.

    PubMed

    Weckwerth, Paulo Henrique; Zapata, Ronald Ordinola; Vivan, Rodrigo Ricci; Tanomaru Filho, Mário; Maliza, Amanda Garcia Alves; Duarte, Marco Antonio Hungaro

    2013-01-01

    Enterococcus faecalis is a bacterial species often found in root canals with failed endodontic treatment. Alkaline pastes are widely used in Endodontics because of their biocompatibility and antimicrobial activity, but this microorganism can resist alkalinity. The purpose of this study was to evaluate in vitro the alkaline pH resistance of E. faecalis for different periods up to 14 days. Samples were obtained from the oral cavity of 150 patients from the Endodontic clinic. The pH of the experimental tubes (n=84) was first adjusted with 6M NaOH to pH values of 9.5, 10.5, 11.5 and 12.5 (21 tubes per pH). Twenty clinical isolates and the ATCC 29212 strain were tested. The 5 positive controls and experimental tubes of each pH were inoculated with 10 µL of bacterial suspension and incubated at 36 °C for 24, 48 and 72 h, 7 and 14 days. For each period, the turbidity of the medium was visually compared with a 0.5 McFarland standard. The presence of the microorganism was confirmed by seeding on M-Enterococcus agar. Four tubes containing BHI broth adjusted to the tested pHs were incubated for 14 days to verify if pH changes occurred. The pH of inoculated BHI broth was also measured on day 14 to determine if the microorganism acidified the medium. The growth of all E. faecalis strains occurred at pH 9.5 to 11.5 in all periods. Although turbidity was not observed at pH 12.5, there was growth of 13 and 2 strains at 24 and 48 h, respectively, on M-Enterococcus agar. No tube showed growth at pH 12.5 after 72 h. It was concluded that E. faecalis can survive in highly alkaline pH, and some clinical isolates require 72 h at pH 12.5 to be killed. PMID:24474287

  18. Enterococcus faecalis prophage dynamics and contributions to pathogenic traits.

    PubMed

    Matos, Renata C; Lapaque, Nicolas; Rigottier-Gois, Lionel; Debarbieux, Laurent; Meylheuc, Thierry; Gonzalez-Zorn, Bruno; Repoila, Francis; Lopes, Maria de Fatima; Serror, Pascale

    2013-06-01

    Polylysogeny is frequently considered to be the result of an adaptive evolutionary process in which prophages confer fitness and/or virulence factors, thus making them important for evolution of both bacterial populations and infectious diseases. The Enterococcus faecalis V583 isolate belongs to the high-risk clonal complex 2 that is particularly well adapted to the hospital environment. Its genome carries 7 prophage-like elements (V583-pp1 to -pp7), one of which is ubiquitous in the species. In this study, we investigated the activity of the V583 prophages and their contribution to E. faecalis biological traits. We systematically analyzed the ability of each prophage to excise from the bacterial chromosome, to replicate and to package its DNA. We also created a set of E. faecalis isogenic strains that lack from one to all six non-ubiquitous prophages by mimicking natural excision. Our work reveals that prophages of E. faecalis V583 excise from the bacterial chromosome in the presence of a fluoroquinolone, and are able to produce active phage progeny. Intricate interactions between V583 prophages were also unveiled: i) pp7, coined EfCIV583 for E. faecalis chromosomal island of V583, hijacks capsids from helper phage 1, leading to the formation of distinct virions, and ii) pp1, pp3 and pp5 inhibit excision of pp4 and pp6. The hijacking exerted by EfCIV583 on helper phage 1 capsids is the first example of molecular piracy in Gram positive bacteria other than staphylococci. Furthermore, prophages encoding platelet-binding-like proteins were found to be involved in adhesion to human platelets, considered as a first step towards the development of infective endocarditis. Our findings reveal not only a role of E. faecalis V583 prophages in pathogenicity, but also provide an explanation for the correlation between antibiotic usage and E. faecalis success as a nosocomial pathogen, as fluoriquinolone may provoke release of prophages and promote gene dissemination among

  19. Enterococcus faecalis Prophage Dynamics and Contributions to Pathogenic Traits

    PubMed Central

    Matos, Renata C.; Lapaque, Nicolas; Rigottier-Gois, Lionel; Debarbieux, Laurent; Meylheuc, Thierry; Gonzalez-Zorn, Bruno; Repoila, Francis; Lopes, Maria de Fatima; Serror, Pascale

    2013-01-01

    Polylysogeny is frequently considered to be the result of an adaptive evolutionary process in which prophages confer fitness and/or virulence factors, thus making them important for evolution of both bacterial populations and infectious diseases. The Enterococcus faecalis V583 isolate belongs to the high-risk clonal complex 2 that is particularly well adapted to the hospital environment. Its genome carries 7 prophage-like elements (V583-pp1 to -pp7), one of which is ubiquitous in the species. In this study, we investigated the activity of the V583 prophages and their contribution to E. faecalis biological traits. We systematically analyzed the ability of each prophage to excise from the bacterial chromosome, to replicate and to package its DNA. We also created a set of E. faecalis isogenic strains that lack from one to all six non-ubiquitous prophages by mimicking natural excision. Our work reveals that prophages of E. faecalis V583 excise from the bacterial chromosome in the presence of a fluoroquinolone, and are able to produce active phage progeny. Intricate interactions between V583 prophages were also unveiled: i) pp7, coined EfCIV583 for E. faecalis chromosomal island of V583, hijacks capsids from helper phage 1, leading to the formation of distinct virions, and ii) pp1, pp3 and pp5 inhibit excision of pp4 and pp6. The hijacking exerted by EfCIV583 on helper phage 1 capsids is the first example of molecular piracy in Gram positive bacteria other than staphylococci. Furthermore, prophages encoding platelet-binding-like proteins were found to be involved in adhesion to human platelets, considered as a first step towards the development of infective endocarditis. Our findings reveal not only a role of E. faecalis V583 prophages in pathogenicity, but also provide an explanation for the correlation between antibiotic usage and E. faecalis success as a nosocomial pathogen, as fluoriquinolone may provoke release of prophages and promote gene dissemination among

  20. In vitro effectiveness of Brazilian brown propolis against Enterococcus faecalis.

    PubMed

    Pimenta, Hévelin Couto; Violante, Ivana Maria Povoa; Musis, Carlo Ralph de; Borges, Álvaro Henrique; Aranha, Andreza Maria Fábio

    2015-01-01

    The aim of this study was to evaluate the in vitro antimicrobial activity of Brazilian brown propolis as an intracanal medication against Enterococcus faecalis. Thirty dentin discs prepared from intact freshly extracted bovine maxillary central incisors were infected with E. faecalis for 21 days. The specimens were distributed into six groups according to the medicament used as follows: G1- calcium hydroxide paste; G2- Carbowax 400 (control group); G3- 20% brown propolis paste; G4- 40% brown propolis paste; G5- 20% brown propolis paste + calcium hydroxide paste; and G6- 40% brown propolis paste + calcium hydroxide paste. The experimental pastes were placed into the canal lumen and left for 14 days. After each period, irrigation was performed with sterile saline to remove the medicament, and the canals were dried with sterile paper points. The dentin chips were removed from the canals with sequential sterile round burs at low speed and were immediately collected in separate test tubes containing BHI broth. The tubes were incubated at 37°C, and microbial growth was analyzed by spectrophotometry after 15 days. All the experimental medications significantly reduced the number of viable bacteria. The G4 and G5 pastes were more effective than the G1 paste, with 35.8%, 41%, and 21.3% antibacterial activity, respectively. Brazilian brown propolis shows antibacterial capacity against E. faecalis. PMID:25992787

  1. Antimicrobial Action of Oleanolic Acid on Listeria monocytogenes, Enterococcus faecium, and Enterococcus faecalis

    PubMed Central

    Kim, Sejeong; Lee, Heeyoung; Lee, Soomin; Yoon, Yohan; Choi, Kyoung-Hee

    2015-01-01

    This study investigated the antimicrobial action of oleanolic acid against Listeria monocytogenes, Enterococcus faecium, and Enterococcus faecalis. To determine the cytotoxicity of oleanolic acid, HEp-2 cells were incubated with oleanolic acid at 37oC. MICs (minimal inhibition concentrations) for L. monocytogenes, E. faecium, and E. faecalis were determined using two-fold microdilutions of oleanolic acid, and bacterial cell viability was then assessed by exposing the bacteria to oleanolic acid at 2 × MIC. To investigate the mode of antimicrobial action of oleanolic acid, we measured leakage of compounds absorbing at 280 nm, along with propidium iodide uptake. Scanning electron microscope (SEM) images were also analysed. The viability of HEp-2 cells decreased (P < 0.05) at oleanolic acid concentrations greater than 128 μg mL-1. The MICs were 16-32 μg mL-1 for L. monocytogenes and 32-64 μg mL-1 for E. faecium and E. faecalis, and bacterial cell viability decreased (P < 0.05) about 3-4 log CFU mL-1 after exposure to 2 × MIC of oleanolic acid. Leakage of 280 nm absorbing materials and propidium iodide uptake was higher in oleanolic acid –treated cells than in the control. The cell membrane was damaged in oleanolic acid-treated cells, but the control group had intact cell membrane in SEM images. The results indicate that oleanolic acid can kill L. monocytogenes, E. faecium, and E. faecalis by destroying the bacterial cell membrane. PMID:25756202

  2. Effects of photodynamic therapy on Enterococcus faecalis biofilms.

    PubMed

    López-Jiménez, L; Fusté, E; Martínez-Garriga, B; Arnabat-Domínguez, J; Vinuesa, T; Viñas, M

    2015-07-01

    Microbial biofilms are involved in almost all infectious pathologies of the oral cavity. This has led to the search for novel therapies specifically aimed at biofilm elimination. In this study, we used atomic force microscopy (AFM) to visualize injuries and to determine surface roughness, as well as confocal laser scanning microscopy (CLSM) to enumerate live and dead bacterial cells, to determine the effects of photodynamic therapy (PDT) on Enterococcus faecalis biofilms. The AFM images showed that PDT consisting of methylene blue and a 670-nm diode laser (output power 280 mW during 30 s) or toluidine blue and a 628-nm LED light (output power 1000 mW during 30 s) induced severe damage, including cell lysis, to E. faecalis biofilms, with the former also causing an important increase in surface roughness. These observations were confirmed by the increase in dead cells determined using CLSM. Our results highlight the potential of PDT as a promising method to achieve successful oral disinfection. PMID:25917515

  3. In vitro inactivation of Enterococcus faecalis with a led device.

    PubMed

    D'Ercole, S; Spoto, G; Trentini, P; Tripodi, D; Petrini, M

    2016-07-01

    Non-coherent light-emitting diodes (LEDs) are effective in a large variety of clinical indications; however, the bactericidal activity of LEDs is unclear, although the effectiveness of such lights is well known. Currently, no studies have examined the effects of NIR-LED on bacteria. The aims of this study were to verify the antibacterial activity of 880-nm LED irradiation on a bacterial suspension of Enterococcus faecalis and to compare it with the actions of sodium hypochlorite (NaOCl) and the concurrent use of both treatments. Before we proceeded with the main experiment, we first performed preliminary tests to evaluate the influence of such parameters as the distance of irradiation, the energy density, the irradiation time and the presence of photosensitizers on the antimicrobial effects of LEDs. After treatment, the colony forming units per milliliter (CFU/mL) was recorded and the data were submitted to ANOVA and Bonferroni post hoc tests at a level of significance of 5%. The results showed that LED irradiation, at the parameters used, is able to significantly decrease E. faecalis viability in vitro. The total inhibition of E. faecalis was obtained throughout concurrent treatment of LED and NaOCl (1%) for 5min. The same antimicrobial activity was confirmed in all of the experiments (p<0.05), but no statistically significant differences were found by varying such parameters as the distance of irradiation (from 0.5mm to 10mm), energy density (from 2.37 to 8.15mJ/s), irradiation time (from 5min to 20min) or by adding toluidine blue O (TBO). PMID:27107704

  4. Draft Genome Sequence of an Enterococcus faecalis Strain Isolated from a Neonatal Blood Sepsis Patient.

    PubMed

    Kropp, K A; Lucid, A; Carroll, J; Belgrudov, V; Walsh, P; Kelly, B; Smith, C; Dickinson, P; O'Driscoll, A; Templeton, K; Ghazal, P; Sleator, R D

    2014-01-01

    Herein, we report the draft genome sequence of Enterococcus faecalis ED-NGS-1009, cultivated from a blood sample taken from a neonatal sepsis patient at the Royal Infirmary in Edinburgh, Scotland, United Kingdom. PMID:25212626

  5. Antimicrobial resistance and virulence of Enterococcus faecalis isolated from retail food

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Although enterococci are considered opportunistic nosocomial pathogens, their contribution to food-borne illnesses via dissemination through retail food remains undefined. In this study, prevalence and association of antimicrobial resistance and virulence factors of 80 Enterococcus faecalis isolate...

  6. The effect of berberine hydrochloride on Enterococcus faecalis biofilm formation and dispersion in vitro.

    PubMed

    Chen, Lihua; Bu, Qianqian; Xu, Huan; Liu, Yuan; She, Pengfei; Tan, Ruichen; Wu, Yong

    2016-01-01

    Enterococcus faecalis (E. faecalis) is one of the major causes of biofilm infections. Berberine hydrochloride (BBH) has diverse pharmacological effects; however, the effects and mechanisms of BBH on E. faecalis biofilm formation and dispersion have not been reported. In this study, 99 clinical isolates from the urine samples of patients with urinary tract infections (UTIs) were collected and identified. Ten strains of E. faecalis with biofilm formation ability were studied. BBH inhibited E. faecalis biofilm formation and promoted the biofilm dispersion of E. faecalis. In addition, sortase A and esp expression levels were elevated during early E. faecalis biofilm development, whereas BBH significantly reduced their expression levels. The results of this study indicated that BBH effectively prevents biofilm formation and promotes biofilm dispersion in E. faecalis, most likely by inhibiting the expressions of sortase A and esp. PMID:27242142

  7. Complete Genome Assembly of Enterococcus faecalis 29212, a Laboratory Reference Strain.

    PubMed

    Minogue, T D; Daligault, H E; Davenport, K W; Broomall, S M; Bruce, D C; Chain, P S; Coyne, S R; Chertkov, O; Freitas, T; Gibbons, H S; Jaissle, J; Koroleva, G I; Ladner, J T; Palacios, G F; Rosenzweig, C N; Xu, Y; Johnson, S L

    2014-01-01

    Enterococcus faecalis is a nonmotile Gram-positive coccus, found both as a commensal organism in healthy humans and animals and as a causative agent of multiple diseases, in particular endocarditis. We sequenced the genome of E. faecalis ATCC 29212, a commonly used reference strain in laboratory studies, to complete "finished" annotated assembly (3 Mb). PMID:25291775

  8. Complete Genome Assembly of Enterococcus faecalis 29212, a Laboratory Reference Strain

    PubMed Central

    Minogue, T. D.; Daligault, H. E.; Davenport, K. W.; Broomall, S. M.; Bruce, D. C.; Chain, P. S.; Coyne, S. R.; Chertkov, O.; Freitas, T.; Gibbons, H. S.; Jaissle, J.; Koroleva, G. I.; Ladner, J. T.; Palacios, G. F.; Rosenzweig, C. N.; Xu, Y.

    2014-01-01

    Enterococcus faecalis is a nonmotile Gram-positive coccus, found both as a commensal organism in healthy humans and animals and as a causative agent of multiple diseases, in particular endocarditis. We sequenced the genome of E. faecalis ATCC 29212, a commonly used reference strain in laboratory studies, to complete “finished” annotated assembly (3 Mb). PMID:25291775

  9. REAL-TIME PCR METHOD TO DETECT ENTEROCOCCUS FAECALIS IN WATER

    EPA Science Inventory

    A 16S rDNA real-time PCR method was developed to detect Enterococcus faecalis in water samples. The dynamic range for cell detection spanned five logs and the detection limit was determined to be 6 cfu/reaction. The assay was capable of detecting E. faecalis cells added to biof...

  10. Specificity of induction of glycopeptide resistance genes in Enterococcus faecalis.

    PubMed Central

    Baptista, M; Depardieu, F; Courvalin, P; Arthur, M

    1996-01-01

    Regulation of VanA- and VanB-type glycopeptide resistance in enterococci is mediated by related two-component regulatory systems (VanR-VanS and VanRB-VanSB). The transglycosylase inhibitors vancomycin, teicoplanin, and moenomycin induced synthesis of the VanX D,D-dipeptidase in a VanA-type Enterococcus faecalis harboring transposon Tn1546. Inhibitors of reactions immediately preceding (ramoplanin) or following (penicillin G and bacitracin) transglycosylation were not inducers. These results identify accumulation of membrane-bound lipid intermediate II as a potential signal for induction of VanA-type resistance. In E.faecalis BM4281 harboring a wild vanB genetic element, D,D-dipeptidase synthesis was only inducible by vancomycin. Induction of the production of the VanB ligase by vancomycin was required for growth of a vancomycin-dependent derivative of BM4281, since introduction of a plasmid coding for constitutive synthesis of the VanA ligase eliminated the requirement of glycopeptide for growth. Both vancomycin and teicoplanin were able to induce D,D-dipeptidase synthesis in BM4281 derivatives that were vancomycin and teicoplanin resistant or vancomycin and teicoplanin dependent. Acquisition of teicoplanin resistance in the latter types of strains was due to alteration in induction specificity associated with an increase in the sensitivity of the regulatory system to vancomycin. Thus, the wild VanRB-VanSB system is unable or not sensitive enough to sense teicoplanin, although mutations can lead to recognition of this antibiotic. PMID:8891132

  11. In vitro activity of Amazon plant extracts against Enterococcus faecalis

    PubMed Central

    de Castilho, Adriana Lígia; da Silva, Juliana Paola Correa; Saraceni, Cintia Helena Coury; Díaz, Ingrit Elida Collantes; Paciencia, Mateus Luís Barradas; Varella, Antonio Drauzio; Suffredini, Ivana Barbosa

    2014-01-01

    Previous studies analyzing 2,200 plant extracts indicated anti-enterococcal activity in 25 extracts obtained from Brazilian forests’ plants. In the present study, these extracts were subjected to microdilution broth assay (MDBA) and disk diffusion assay (DDA) using planktonic Enterococcus faecalis ATCC® 29212™ and were submitted to phytochemical analysis in TLC and HPLC. Three extracts obtained from Ipomoea alba (MIC < 40 μg/mL), Diclinanona calycina (MIC ≤ 40 μg/mL) and Moronobea coccinea (40 < MIC < 80 μg/mL; MBC = 80 μg/mL) showed significant bactericidal activity in the MDBA and four extracts obtained from I. alba (14.04 ± 0.55 mm diameter) S. globulifera (14.43 ± 0.33 mm and 12.18 ± 0.28 mm diameter) and Connarus ruber var. ruber (13.13 ± 0.18 mm diameter) were active in DDA. Residues H2O obtained from Psidium densicomum (mean of 16.78 mm diameter) and from Stryphnodendron pulcherrimum (mean of 15.97 mm diameter) have shown an improved antibacterial activity after fractionation if compared to that obtained from the respective crude extracts. Antioxidant activity was observed in some residues of the active extracts. TLC analysis showed that phenolic compounds are likely to be found in active extracts. Three molecules were isolated from S. globulifera and were identified by 13C NMR lupeol, α-amyrin and 3β-hydroxyglutin-5-ene. The present chemical and biological findings suggest that these extracts are a potential source of new anti-Enterococcus compounds to be introduced in endodontic therapy. PMID:25477906

  12. Rapid kill-novel endodontic sealer and Enterococcus faecalis.

    PubMed

    Beyth, Nurit; Kesler Shvero, Dana; Zaltsman, Nathan; Houri-Haddad, Yael; Abramovitz, Itzhak; Davidi, Michael Perez; Weiss, Ervin I

    2013-01-01

    With growing concern over bacterial resistance, the identification of new antimicrobial means is paramount. In the oral cavity microorganisms are essential to the development of periradicular diseases and are the major causative factors associated with endodontic treatment failure. As quaternary ammonium compounds have the ability to kill a wide array of bacteria through electrostatic interactions with multiple anionic targets on the bacterial surface, it is likely that they can overcome bacterial resistance. Melding these ideas, we investigated the potency of a novel endodontic sealer in limiting Enterococcus faecalis growth. We used a polyethyleneimine scaffold to synthesize nano-sized particles, optimized for incorporation into an epoxy-based endodontic sealer. The novel endodontic sealer was tested for its antimicrobial efficacy and evaluated for biocompatibility and physical eligibility. Our results show that the novel sealer foundation affixes the nanoparticles, achieving surface bactericidal properties, but at the same time impeding nanoparticle penetration into eukaryotic cells and thereby mitigating a possible toxic effect. Moreover, adequate physical properties are maintained. The nanosized quaternary amine particles interact within minutes with bacteria, triggering cell death across wide pH values. Throughout this study we demonstrate a new antibacterial perspective for endodontic sealers; a novel antibacterial, effective and safe antimicrobial means. PMID:24223159

  13. Biocide and antibiotic resistance of Enterococcus faecalis and Enterococcus faecium isolated from the swine meat chain.

    PubMed

    Rizzotti, Lucia; Rossi, Franca; Torriani, Sandra

    2016-12-01

    In this study nine strains of Enterococcus faecalis and 12 strains of Enterococcus faecium, isolated from different sample types in the swine meat chain and previously characterized for the presence of antibiotic resistance genes, were examined for phenotypic tolerance to seven biocides (chlorexidine, benzalkonium chloride, triclosan, sodium hypochlorite, 2-propanol, formaldehyde and hydrogen peroxide) and resistance to nine antibiotics (ampicillin, vancomycin, gentamicin, kanamycin, streptomycin, erythromycin, clindamycin, tetracycline and chloramphenicol). Moreover, the presence of efflux system encoding genes qacA/B, qacC, qacE, qacEΔ1, emeA, and stress response genes, sigV and gsp65, involved in the tolerance to biocides, was analysed. Most strains were not tolerant to the biocides, but showed minimum inhibitory concentrations (MICs) higher than the recommended cut-off values for all the antibiotics tested, except for vancomycin and chloramphenicol. Only weak correlations, if any, were found between biocide and antibiotic resistance data. One E. faecalis strain was tolerant to triclosan and one E. faecium strain, with higher tolerance to chlorexidine than the other strains tested, was found to carry a qacA/B gene. Our results indicated that phenotypic resistance to antibiotics is very frequent in enterococcal isolates from the swine meat chain, but phenotypic tolerance to biocides is not common. On the other hand, the gene qacA/B was found for the first time in the species E. faecium, an indication of the necessity to adopt measures suitable to control the spread of biocide resistance determinants among enterococci. PMID:27554158

  14. Identification and Imaging of Peptides and Proteins on Enterococcus faecalis Biofilms by Matrix Assisted Laser Desorption Ionization Mass Spectrometry

    PubMed Central

    Melvin Blaze, M. T.; Aydin, Berdan; Carlson, Ross; Hanley, Luke

    2013-01-01

    The heptapeptide ARHPHPH was identified from biofilms and planktonic cultures of two different strains of Enterococcus faecalis, V583 and ATCC 29212, using matrix assisted laser desorption ionization mass spectrometry (MALDI-MS). ARHPHPH was also imaged at the boundary of cocultured, adjacent E. faecalis and Escherichia coli (ATCC 25922) biofilms, appearing only on the E. faecalis side. ARHPHPH was proteolyzed from κ-casein, a component in the growth media, by E. faecalis microbes. Additionally, top down and bottom up proteomic approaches were combined to identify and spatially locate multiple proteins within intact E. faecalis V583 biofilms by MALDI-MS. The resultant tandem MS data were searched against the NCBInr E. faecalis V583 database to identify thirteen cytosolic and membrane proteins which have functional association with the cell surface. Two of these proteins, enolase and GAPDH, are glycolytic enzymes known to display multiple functions in bacterial virulence in related bacterial strains. This work illustrates a powerful approach for discovering and localizing multiple peptides and proteins within intact biofilms. PMID:22962657

  15. Role of (p)ppGpp in Biofilm Formation by Enterococcus faecalis

    PubMed Central

    Lemos, José A.; Wickström, Claes; Sedgley, Christine M.

    2012-01-01

    Enterococcus faecalis strain OG1RF and its (p)ppGpp-deficient ΔrelA, ΔrelQ, and ΔrelA ΔrelQ mutants were grown in biofilms and evaluated for growth profiles, biofilm morphology, cell viability, and proteolytic activity. E. faecalis lacking (p)ppGpp had a diminished capacity to sustain biofilm formation over an extended period of time and expressed abundant proteolytic activity. PMID:22179256

  16. Differences in Antibiotic Resistance Patterns of Enterococcus faecalis and Enterococcus faecium Strains Isolated from Farm and Pet Animals

    PubMed Central

    Butaye, Patrick; Devriese, Luc A.; Haesebrouck, Freddy

    2001-01-01

    The prevalence of acquired resistance in 146 Enterococcus faecium and 166 Enterococcus faecalis strains from farm and pet animals, isolated in 1998 and 1999 in Belgium, against antibiotics used for growth promotion and for therapy was determined. Acquired resistance against flavomycin and monensin, two antibiotics used solely for growth promotion, was not detected. Avoparcin (glycopeptide) resistance was found sporadically in E. faecium only. Avilamycin resistance was almost exclusively seen in strains from farm animals. Resistance rates were higher in E. faecium strains from broiler chickens than in strains from other animal groups with tylosin and virginiamycin and in E. faecalis as well as in E. faecium strains with narasin and bacitracin. Resistance against ampicillin was mainly found among E. faecium strains from pets and was absent in E. faecalis. Tetracycline resistance occurred most often in strains from farm animals, while enrofloxacin resistance, only found in E. faecalis, occurred equally among strains from all origins. Resistance against gentamicin was very rare in broiler strains, whereas resistance rates were high in strains from other origins. It can be concluded that resistance against antibiotics used solely for growth promotion was more prevalent in E. faecium strains than in E. faecalis strains. With few exceptions, resistance against the different categories of antibiotics was more prevalent in strains from farm animals than in those from pets. PMID:11302798

  17. Differences in antibiotic resistance patterns of Enterococcus faecalis and Enterococcus faecium strains isolated from farm and pet animals.

    PubMed

    Butaye, P; Devriese, L A; Haesebrouck, F

    2001-05-01

    The prevalence of acquired resistance in 146 Enterococcus faecium and 166 Enterococcus faecalis strains from farm and pet animals, isolated in 1998 and 1999 in Belgium, against antibiotics used for growth promotion and for therapy was determined. Acquired resistance against flavomycin and monensin, two antibiotics used solely for growth promotion, was not detected. Avoparcin (glycopeptide) resistance was found sporadically in E. faecium only. Avilamycin resistance was almost exclusively seen in strains from farm animals. Resistance rates were higher in E. faecium strains from broiler chickens than in strains from other animal groups with tylosin and virginiamycin and in E. faecalis as well as in E. faecium strains with narasin and bacitracin. Resistance against ampicillin was mainly found among E. faecium strains from pets and was absent in E. faecalis. Tetracycline resistance occurred most often in strains from farm animals, while enrofloxacin resistance, only found in E. faecalis, occurred equally among strains from all origins. Resistance against gentamicin was very rare in broiler strains, whereas resistance rates were high in strains from other origins. It can be concluded that resistance against antibiotics used solely for growth promotion was more prevalent in E. faecium strains than in E. faecalis strains. With few exceptions, resistance against the different categories of antibiotics was more prevalent in strains from farm animals than in those from pets. PMID:11302798

  18. Ampicillin in Combination with Ceftaroline, Cefepime, or Ceftriaxone Demonstrates Equivalent Activities in a High-Inoculum Enterococcus faecalis Infection Model.

    PubMed

    Luther, Megan K; Rice, Louis B; LaPlante, Kerry L

    2016-05-01

    Ampicillin-ceftriaxone combination therapy has become a predominant treatment for serious Enterococcus faecalis infections, such as endocarditis. Unfortunately, ceftriaxone use is associated with future vancomycin-resistant enterococcus colonization. We evaluated E. faecalis in an in vitro pharmacodynamic model against simulated human concentration-time profiles of ampicillin plus ceftaroline, cefepime, ceftriaxone, or gentamicin. Ampicillin-cefepime and ampicillin-ceftaroline demonstrated activities similar to those of ampicillin-ceftriaxone against E. faecalis. PMID:26926624

  19. Riboflavin-shuttled extracellular electron transfer from Enterococcus faecalis to electrodes in microbial fuel cells.

    PubMed

    Zhang, Enren; Cai, Yamin; Luo, Yue; Piao, Zhe

    2014-11-01

    Great attention has been focused on Gram-negative bacteria in the application of microbial fuel cells. In this study, the Gram-positive bacterium Enterococcus faecalis was employed in microbial fuel cells. Bacterial biofilms formed by E. faecalis ZER6 were investigated with respect to electricity production through the riboflavin-shuttled extracellular electron transfer. Trace riboflavin was shown to be essential for transferring electrons derived from the oxidation of glucose outside the peptidoglycan layer in the cell wall of E. faecalis biofilms formed on the surface of electrodes, in the absence of other potential electron mediators (e.g., yeast extract). PMID:25345758

  20. Outbreak of mastitis in sheep caused by multi-drug resistant Enterococcus faecalis in Sardinia, Italy.

    PubMed

    Sanciu, G; Marogna, G; Paglietti, B; Cappuccinelli, P; Leori, G; Rappelli, P

    2013-03-01

    An outbreak of infective mastitis due to Enterococcus faecalis occurred in an intensive sheep farm in north Sardinia (Italy). E. faecalis, which is only rarely isolated from sheep milk, was unexpectedly found in 22·3% of positive samples at microbiological examination. Forty-five out of the 48 E. faecalis isolates showed the same multi-drug resistance pattern (cloxacillin, streptomycin, kanamycin, clindamycin, oxytetracycline). E. faecalis isolates were analysed by pulsed-field gel electrophoresis, and all 45 multi-drug resistant strains showed an indistinguishable macrorestiction profile, indicating their clonal origin. To our knowledge, this is the first report of an outbreak of mastitis in sheep caused by E. faecalis. PMID:22595402

  1. Phenotypic and molecular antibiotic resistance profile of Enterococcus faecalis and Enterococcus faecium isolated from different traditional fermented foods.

    PubMed

    Sánchez Valenzuela, Antonio; Lavilla Lerma, Leyre; Benomar, Nabil; Gálvez, Antonio; Pérez Pulido, Rubén; Abriouel, Hikmate

    2013-02-01

    A collection of 55 enterococci (41 Enterococcus faecium and 14 E. faecalis strains) isolated from various traditional fermented foodstuffs of both animal and vegetable origins, and water was evaluated for resistance against 15 antibiotics. Lower incidence of resistance was observed with gentamicin, ampicillin, penicillin and teicoplanin. However, a high incidence of antibiotic resistance was detected for rifampicin (12 out of 14 of isolates), ciprofloxacin (9/14), and quinupristin/dalfopristin (8/14) in E. faecalis strains. Enterococcus faecium isolates were resistant to rifampicin (25/41), ciprofloxacin (23/41), erythromycin (18/41), levofloxacin (16/41), and nitrofurantoin (15/41). One Enterococcus faecalis and two E. faecium strains were resistant to vancomycin (MIC>16 μg/mL). Among 55 isolates, 27 (19 E. faecium and eight E. faecalis) were resistant to at least three antibiotics. High level of multidrug resistance to clinically important antibiotics was detected in E. faecalis strains (57% of E. faecalis versus 46% of E. faecium), which showed resistance to six to seven antibiotics, especially those isolated from foods of animal origin. So, it is necessary to re-evaluate the use of therapeutic antibiotics in stock farms at both regional and international levels due to the high number of multiple resistant (MR) bacteria. Fifty-six MR E. faecalis and E. faecium strains selected from this and previous studies (Valenzuela et al., 2008, 2010) were screened by polymerase chain reaction for antibiotic resistance genes, revealing the presence of tet(L), tet(M), ermB, cat, efrA, efrB, mphA, or msrA/B genes. The ABC Multidrug Efflux Pump EfrAB was detected in 96% of E. faecalis strains and also in 13% of E. faecium strains; this is the first report describing EfrAB in this enterococcal species. The efflux pump-associated msrA/B gene was detected in 66.66% of E. faecium strains, but not in E. faecalis strains. PMID:23259502

  2. The opportunistic pathogen Enterococcus faecalis resists phagosome acidification and autophagy to promote intracellular survival in macrophages.

    PubMed

    Zou, Jun; Shankar, Nathan

    2016-06-01

    While many strains of Enterococcus faecalis have been reported to be capable of surviving within macrophages for extended periods, the exact mechanisms involved are largely unknown. In this study, we found that after phagocytosis by macrophages, enterococci-containing vacuoles resist acidification, and E. faecalis is resistant to low pH. Ultrastructural examination of the enterococci-containing vacuole by transmission electron microscopy revealed a single membrane envelope, with no evidence of the classical double-membraned autophagosomes. Western blot analysis further confirmed that E. faecalis could trigger inhibition of the production of LC3-II during infection. By employing cells transfected with RFP-LC3 plasmid and infected with GFP-labelled E. faecalis, we also observed that E. faecalis was not delivered into autophagosomes during macrophage infection. While these observations indicated no role for autophagy in elimination of intracellular E. faecalis, enhanced production of reactive oxygen species and nitric oxide were keys to this process. Stimulation of autophagy suppressed the intracellular survival of E. faecalis in macrophages in vitro and decreased the burden of E. faecalis in vivo. In summary, the results from this study offer new insights into the interaction of E. faecalis with host cells and may provide a new approach to treatment of enterococcal infections. PMID:26663775

  3. Effects of ionophores on Enterococcus faecalis and E. faecium growth in pure and mixed ruminal culture

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Enterococcus faecalis and faecium are Gram-positive human pathogens that can live in the gastrointestinal tract of food animals. Vancomycin-resistant enterococci (VRE) are an increasing threat to humans as a nosocomial infection, as well as a reservoir of antibiotic resistance genes. Ionophores ar...

  4. An indigenous gut bacterium, Enterococcus faecalis (Lactobacillales: Enterococcaceae), increases seed consumption by Harpalus pensylvanicus (Coleoptera: Carabidae)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Harpalus pensylvanicus is a beneficial beetle contributing to insect control and seed predation in North American cropland. The bacterial endosymbiont Enterococcus faecalis is found in the intestinal tract of H. pensylvanicus and is thought to contribute to the digestion of the insect's seed diet. W...

  5. Draft Genome Sequence for a Clinical Isolate of Vancomycin-Resistant Enterococcus faecalis.

    PubMed

    Erickson, Keesha E; Madinger, Nancy E; Chatterjee, Anushree

    2016-01-01

    We report here the draft genome sequence of a multidrug-resistant Enterococcus faecalis strain, isolated from a patient at the University of Colorado Hospital. The genome assembly is 3,040,186 bp in length with 37.6% GC content. This isolate encodes eleven resistance genes, including those for glycopeptide, aminoglycoside, macrolide-lincosamide-streptogramin, and tetracycline resistance. PMID:27340066

  6. Draft Genome Sequence for a Clinical Isolate of Vancomycin-Resistant Enterococcus faecalis

    PubMed Central

    Erickson, Keesha E.; Madinger, Nancy E.

    2016-01-01

    We report here the draft genome sequence of a multidrug-resistant Enterococcus faecalis strain, isolated from a patient at the University of Colorado Hospital. The genome assembly is 3,040,186 bp in length with 37.6% GC content. This isolate encodes eleven resistance genes, including those for glycopeptide, aminoglycoside, macrolide-lincosamide-streptogramin, and tetracycline resistance. PMID:27340066

  7. Efficacy of Atmospheric Pressure Plasma as an Antibacterial Agent Against Enterococcus Faecalis in Vitro

    NASA Astrophysics Data System (ADS)

    Cao, Yingguang; Yang, Ping; Lu, Xinpei; Xiong, Zilan; Ye, Tao; Xiong, Qing; Sun, Ziyong

    2011-02-01

    Enterococcus faecalis (E. faecalis) is a microorganism that can survive extreme challenges in obturated root canals. The aim of this study was to evaluate the efficacy of a non-thermal atmospheric pressure plasma plume against E. faecalis in vitro. A non-thermal atmospheric pressure plasma jet device which could generate a cold plasma plume carrying a peak current of 300 mA was used. The antibacterial efficacy of this device against E. faecalis and its biofilm under different conditions was detected. The antibacterial efficacy of the plasma against E. faecalis and Staphylococcus aureus (S. aureus) was also evaluated. After plasma treatment, the average diameter of inhibition zone on S. aureus and E. faecalis was 2.62±0.26 cm and 1.06±0.30 cm, respectively (P < 0.05). The diameter was increased with prolongation of the treatment duration. The diameters of inhibition zone of the sealed Petri dishes were larger than those of the uncovered Petri dishes. There was significant difference in colony-forming units between plasma group and control group on E. faecalis biofilm (P < 0.01). The transmission electron microscopy revealed that the ultrastructural changes cytoderm of E. faecalis were observed after treatment for 2 min. It is concluded that the non-thermal atmospheric pressure plasma could serve as an effective adjunct to standard endodontic microbial treatment.

  8. New intracanal formulations containing doxycycline or chlorhexidine against Enterococcus faecalis.

    PubMed

    Silva, Ana Rita Marques da; Pinto, Shelon Cristina Souza; Santos, Elizabete Brasil dos; Santos, Fábio André dos; Farago, Paulo Vitor; Gomes, João Carlos; Pina-Vaz, Irene; Carvalho, Manuel Fontes

    2014-01-01

    The present study aims to evaluate the antimicrobial effect of two new intracanal preparations against E. faecalis. Thirty single-rooted human canine teeth were used. The crowns were removed and the roots were instrumented using a conventional technique. Three groups of ten teeth each were infected with 108 CFU/ ml of E. faecalis for 21 days. The root canals were flled with new intracanal medications containing 3% doxycycline hydrochloride (DX) or 2% chlorhexidine digluconate (CHX). Ten teeth received no medication (NM)-negative control. Microbial samples were obtained 21 days after contamination: 14 days under the effect of the intracanal medications and 7 days after replacing the medications by BHI broth. The samples were homogenized, diluted, seeded on BHI agar and incubated for 48h/36°C. The number of colony forming units (CFU/ml) was obtained and analyzed statistically. All intracanal dressings significantly reduced the number of bacterial cells in the root canal after 14 days with medication. After the period with 7 days with BHI broth, the CFU counts of E. faecalis remained at low values. However, the NM group showed a significant increase of CFU in this period to similar values of the initial contamination. 3% doxycycline hydrochloride gel and 2% CHX gel were effective to eliminate E. faecalis from the root canal system. PMID:24939266

  9. Identification of Polyketide Inhibitors Targeting 3-Dehydroquinate Dehydratase in the Shikimate Pathway of Enterococcus faecalis

    PubMed Central

    Hernandez-Valladares, Maria; Go, Maybelle Kho; Tung, Alvin; Aguda, Adeleke H.; Robinson, Robert C.; Yew, Wen Shan

    2014-01-01

    Due to the emergence of resistance toward current antibiotics, there is a pressing need to develop the next generation of antibiotics as therapeutics against infectious and opportunistic diseases of microbial origins. The shikimate pathway is exclusive to microbes, plants and fungi, and hence is an attractive and logical target for development of antimicrobial therapeutics. The Gram-positive commensal microbe, Enterococcus faecalis, is a major human pathogen associated with nosocomial infections and resistance to vancomycin, the “drug of last resort”. Here, we report the identification of several polyketide-based inhibitors against the E. faecalis shikimate pathway enzyme, 3-dehydroquinate dehydratase (DHQase). In particular, marein, a flavonoid polyketide, both inhibited DHQase and retarded the growth of Enterococcus faecalis. The purification, crystallization and structural resolution of recombinant DHQase from E. faecalis (at 2.2 Å resolution) are also reported. This study provides a route in the development of polyketide-based antimicrobial inhibitors targeting the shikimate pathway of the human pathogen E. faecalis. PMID:25072253

  10. Genetic relationships among Enterococcus faecalis isolates from different sources as revealed by multilocus sequence typing.

    PubMed

    Chen, X; Song, Y Q; Xu, H Y; Menghe, B L G; Zhang, H P; Sun, Z H

    2015-08-01

    Enterococcus faecalis is part of the natural gut flora of humans and other mammals; some isolates are also used in food production. So, it is important to evaluate the genetic diversity and phylogenetic relationships among E. faecalis isolates from different sources. Multilocus sequence typing protocol was used to compare 39 E. faecalis isolates from Chinese traditional food products (including dairy products, acidic gruel) and 4 published E. faecalis isolates from other sources including human-derived isolates employing 5 housekeeping genes (groEL, clpX, recA, rpoB, and pepC). A total of 23 unique sequence types were identified, which were grouped into 5 clonal complexes and 10 singletons. The value of standardized index of association of the alleles (IA(S)=0.1465) and network structure indicated a high frequency of intraspecies recombination across these isolates. Enterococcus faecalis lineages also exhibited clearly source-clustered distributions. The isolates from dairy source were clustered together. However, the relationship between isolates from acidic gruel and one isolate from a human source was close. The MLST scheme presented in this study provides a sharable and continuously growing sequence database enabling global comparison of strains from different sources, and will further advance our understanding of the microbial ecology of this important species. PMID:26074239

  11. Synergistic Antibacterial Effect of the Combination of ε-Polylysine and Nisin against Enterococcus faecalis.

    PubMed

    Liu, Fang; Liu, Mei; Du, Lihui; Wang, Daoying; Geng, Zhiming; Zhang, Muhan; Sun, Chong; Xu, Xiaoxi; Zhu, Yongzhi; Xu, Weimin

    2015-12-01

    This study evaluated the antibacterial effect of the combination of ε-polylysine (ε-PL) and nisin against Enterococcus faecalis strains. The combination of ε-PL and nisin showed synergistic antibacterial activity against three Enterococcus strains. Scanning electron microscopy and a membrane permeability assay revealed that the combined treatment with ε-PL and nisin synergistically damaged the cell morphology of E. faecalis strain R612Z1 cells. Both ε-PL and nisin can dissipate the transmembrane electric potential of E. faecalis R612Z1 cells, but these peptides did not affect the transmembrane pH gradient. The combination of ε-PL and nisin can produce a high reactive oxygen species level in E. faecalis R612Z1 cells. The results indicated that the uptake of ε-PL into cells was promoted through nisin and that the combination of ε-PL and nisin could produce a high reactive oxygen species level in E. faecalis R612Z1 cells, leading to cell growth inhibition. PMID:26613915

  12. Role of house flies in the ecology of Enterococcus faecalis from wastewater treatment facilities.

    PubMed

    Doud, C W; Scott, H M; Zurek, L

    2014-02-01

    Enterococci are important nosocomial pathogens, with Enterococcus faecalis most commonly responsible for human infections. In this study, we used several measures to test the hypothesis that house flies, Musca domestica (L.), acquire and disseminate antibiotic-resistant and potentially virulent E. faecalis from wastewater treatment facilities (WWTF) to the surrounding urban environment. House flies and sludge from four WWTF (1-4) as well as house flies from three urban sites close to WWTF-1 were collected and cultured for enterococci. Enterococci were identified, quantified, screened for antibiotic resistance and virulence traits, and assessed for clonality. Of the 11 antibiotics tested, E. faecalis was most commonly resistant to tetracycline, doxycycline, streptomycin, gentamicin, and erythromycin, and these traits were intra-species horizontally transferrable by in vitro conjugation. Profiles of E. faecalis (prevalence, antibiotic resistance, and virulence traits) from each of WWTF sludge and associated house flies were similar, indicating that flies successfully acquired these bacteria from this substrate. The greatest number of E. faecalis with antibiotic resistance and virulence factors (i.e., gelatinase, cytolysin, enterococcus surface protein, and aggregation substance) originated from WWTF-1 that processed meat waste from a nearby commercial meat-processing plant, suggesting an agricultural rather than human clinical source of these isolates. E. faecalis from house flies collected from three sites 0.7-1.5 km away from WWTF-1 were also similar in their antibiotic resistance profiles; however, antibiotic resistance was significantly less frequent. Clonal diversity assessment using pulsed-field gel electrophoresis revealed the same clones of E. faecalis from sludge and house flies from WWTF-1 but not from the three urban sites close to WWTF-1. This study demonstrates that house flies acquire antibiotic-resistant enterococci from WWTF and potentially

  13. Dichotomous Metabolism of Enterococcus faecalis Induced by Hematin Starvation Modulates Colonic Gene Expression

    PubMed Central

    Allen, Toby D.; Moore, Danny R.; Wang, Xingmin; Casu, Viviana; May, Randal; Lerner, Megan R.; Houchen, Courtney; Brackett, Daniel J.; Huycke, Mark M.

    2009-01-01

    Summary Enterococcus faecalis is an intestinal commensal that cannot synthesize porphyrins and only expresses a functional respiratory chain when provided exogenous hematin. In the absence of hematin, E. faecalis reverts to fermentative metabolism and produces extracellular superoxide that can damage epithelial cell DNA. The acute response of the colonic mucosa to hematin-starved E. faecalis was identified by gene array. E. faecalis was inoculated into murine colons using a surgical ligation model that preserved tissue architecture and homeostasis. The mucosa was exposed to hematin-starved E. faecalis and compared to a control consisting of the same strain grown with hematin. At 1 hour post-inoculation six mucosal genes were differentially regulated and this increased to 42 genes at 6 hours. At 6 hours a highly significant biological interaction network was identified with functions that included NF-κB signaling, apoptosis, and cell cycle regulation. Colon biopsies showed no histological abnormalities by hematoxylin and eosin staining. Immunohistochemical staining, however, detected NF-κB activation in tissue macrophages using antibodies to the nuclear localization sequence for p65 and the F4/80 marker for murine macrophages. Similarly, hematin-starved E. faecalis strongly activated NF-κB in murine macrophages in vitro. Furthermore, primary and transformed colonic epithelial cells activated the G2/M checkpoint in vitro following exposure to hematin-starved E. faecalis. Modulation of this cell cycle checkpoint was due to extracellular superoxide produced as a result of the respiratory block in hematin-starved E. faecalis. These results demonstrate that the uniquely dichotomous metabolism of E. faecalis can significantly modulate gene expression in the colonic mucosa for pathways associated with inflammation, apoptosis, and cell cycle regulation. PMID:18809545

  14. Different extracts of Zingiber officinale decrease Enterococcus faecalis infection in Galleria mellonella.

    PubMed

    Maekawa, Lilian Eiko; Rossoni, Rodnei Dennis; Barbosa, Júnia Oliveira; Jorge, Antonio Olavo Cardoso; Junqueira, Juliana Campos; Valera, Marcia Carneiro

    2015-01-01

    Dried, fresh and glycolic extracts of Zingiber officinale were obtained to evaluate the action against G. mellonella survival assay against Enterococcus faecalis infection. Eighty larvae were divided into: 1) E. faecalis suspension (control); 2) E. faecalis + fresh extract of Z. officinale (FEO); 3) E. faecalis + dried extract of Z. officinale (DEO); 4) E. faecalis + glycolic extract of Z. officinale (GEO); 5) Phosphate buffered saline (PBS). For control group, a 5 μL inoculum of standardized suspension (107 cells/mL) of E. faecalis (ATCC 29212) was injected into the last left proleg of each larva. For the treatment groups, after E. faecalis inoculation, the extracts were also injected, but into the last right proleg. The larvae were stored at 37 °C and the number of dead larvae was recorded daily for 168 h (7 days) to analyze the survival curve. The larvae were considered dead when they did not show any movement after touching. E. faecalis infection led to the death of 85% of the larvae after 168 h. Notwithstanding, in treatment groups with association of extracts, there was an increase in the survival rates of 50% (GEO), 61% (FEO) and 66% (DEO) of the larvae. In all treatment groups, the larvae exhibited a survival increase with statistically significant difference in relation to control group (p=0.0029). There were no statistically significant differences among treatment groups with different extracts (p=0.3859). It may be concluded that the tested extracts showed antimicrobial activity against E. faecalis infection by increasing the survival of Galleria mellonella larvae. PMID:25831098

  15. Enterococcus faecalis 6-Phosphogluconolactonase Is Required for Both Commensal and Pathogenic Interactions with Manduca sexta

    PubMed Central

    Holt, Jonathan F.; Frank, Kristi L.; Du, Jing; Guan, Changhui; Handelsman, Jo

    2014-01-01

    Enterococcus faecalis is a commensal and pathogen of humans and insects. In Manduca sexta, E. faecalis is an infrequent member of the commensal gut community, but its translocation to the hemocoel results in a commensal-to-pathogen switch. To investigate E. faecalis factors required for commensalism, we identified E. faecalis genes that are upregulated in the gut of M. sexta using recombinase-based in vivo expression technology (RIVET). The RIVET screen produced 113 clones, from which we identified 50 genes that are more highly expressed in the insect gut than in culture. The most frequently recovered gene was locus OG1RF_11582, which encodes a 6-phosphogluconolactonase that we designated pglA. A pglA deletion mutant was impaired in both pathogenesis and gut persistence in M. sexta and produced enhanced biofilms compared with the wild type in an in vitro polystyrene plate assay. Mutation of four other genes identified by RIVET did not affect persistence in caterpillar guts but led to impaired pathogenesis. This is the first identification of genetic determinants for E. faecalis commensal and pathogenic interactions with M. sexta. Bacterial factors identified in this model system may provide insight into colonization or persistence in other host-associated microbial communities and represent potential targets for interventions to prevent E. faecalis infections. PMID:25385794

  16. Efflux pump inhibitor potentiates antimicrobial photodynamic inactivation of Enterococcus faecalis biofilm.

    PubMed

    Kishen, Anil; Upadya, Megha; Tegos, George P; Hamblin, Michael R

    2010-01-01

    Microbial biofilm architecture contains numerous protective features, including extracellular polymeric material that render biofilms impermeable to conventional antimicrobial agents. This study evaluated the efficacy of antimicrobial photodynamic inactivation (aPDI) of Enterococcus faecalis biofilms. The ability of a cationic, phenothiazinium photosensitizer, methylene blue (MB) and an anionic, xanthene photosensitizer, rose bengal (RB) to inactivate biofilms of E. faecalis (OG1RF and FA 2-2) and disrupt the biofilm structure was evaluated. Bacterial cells were tested as planktonic suspensions, intact biofilms and biofilm-derived suspensions obtained by the mechanical disruption of biofilms. The role of a specific microbial efflux pump inhibitor (EPI), verapamil hydrochloride in the MB-mediated aPDI of E. faecalis biofilms was also investigated. The results showed that E. faecalis biofilms exhibited significantly higher resistance to aPDI when compared with E. faecalis in suspension (P < 0.001). aPDI with cationic MB produced superior inactivation of E. faecalis strains in a biofilm along with significant destruction of biofilm structure when compared with anionic RB (P < 0.05). The ability to inactivate biofilm bacteria was further enhanced when the EPI was used with MB (P < 0.001). These experiments demonstrated the advantage of a cationic phenothiazinium photosensitizer combined with an EPI to inactivate biofilm bacteria and disrupt biofilm structure. PMID:20860692

  17. Antimicrobial Effects of Four Intracanal Medicaments on Enterococcus Faecalis: An in Vitro Study

    PubMed Central

    Mozayeni, Mohammad Ali; Haeri, Ali; Dianat, Omid; Jafari, Ali Reza

    2014-01-01

    Introduction: The aim of this in vitro study was to evaluate the antimicrobial activity of four intracanal medicaments on Enterococcus Faecalis (E. Faecalis). Methods and Materials: Fifty extracted single-rooted human teeth were prepared with standard method. After contaminating the canals with E. Faecalis, the samples were divided into one control and four experimental groups (n=10). The teeth in each group were treated with one of the experimental medicaments, including calcium hydroxide (CH), 2% chlorhexidine gel (CHX), triple antibiotic paste (TAP) and nanosilver (NS). In control group, canals were filled with a neutral gel. Microbial samples were obtained from the roots after 7 days and optical density of the cultures was determined after 24 h of incubation. Optical density values were analyzed with one-way analysis of variance and Tukey’s post hoc tests. Results: CHX gel and TAP were significantly more effective against E. Faecalis than CH, which was also significantly more efficient than NS and normal saline. In the paper cone samples, CHX gel was more effective than TAP; however, samples obtained with sizes 2 and 4 Gate Glidden drills, indicated that TAP was much more efficient than CHX. Normal saline and NS had similar effects on E. Faecalis. Conclusion: NS gel was not efficient enough against E. Faecalis; however, TAP and CHX gel showed better antibacterial efficacy than CH and can be used as an alternative intracanal medicaments in root canal therapies. PMID:25031593

  18. Detection of Enterococcus faecalis in Necrotic Teeth Root Canals by Culture and Polymerase Chain Reaction Methods

    PubMed Central

    Cogulu, Dilsah; Uzel, Atac; Oncag, Ozant; Aksoy, Semiha C.; Eronat, Cemal

    2007-01-01

    Objectives The aim of this study was to investigate the presence of Enterococcus faecalis in endodontic infections in both deciduous and permanent teeth by culture and polymerase chain reaction (PCR) methods. Methods A total of 145 children aged 5–13 years old were involved in this study. The presence of E. faecalis in necrotic deciduous and permanent teeth root canals was studied using culture and polymerase chain reaction methods. Results Among 145 molar teeth, 57% (n=83) presented necrotic asymptomatic pulp tissues and were included in this study. Culture and PCR methods detected the test species in 18 and 22 of 83 teeth involved, respectively. E. faecalis was cultured from 8 (18%) of 45 necrotic deciduous teeth and from 10 (26%) of 38 necrotic permanent teeth. PCR detection identified the target species in 10 (22%) and 12 (32%) of necrotic deciduous and permanent teeth respectively. Statistically significant difference in the presence of E. faecalis in deciduous and permanent teeth was found by culture and PCR methods (P=0.03 and 0.02, respectively). The difference in the presence of E. faecalis between two different methods was not statistically significant (P>.05). Conclusions The results of the present study confirm that both culture and PCR methods are sensitive to detect E. faecalis in root canals. PMID:19212470

  19. Could β-hemolytic, group B Enterococcus faecalis be mistaken for Streptococcus agalactiae?

    PubMed

    Savini, Vincenzo; Gherardi, Giovanni; Marrollo, Roberta; Franco, Alessia; Pimentel De Araujo, Fernanda; Dottarelli, Samuele; Fazii, Paolo; Battisti, Antonio; Carretto, Edoardo

    2015-05-01

    A β-hemolytic Enterococcus faecalis strain agglutinating Lancefield group A, B, C, D, F, and G antisera was observed from a rectovaginal swab, in the context of antenatal screening for Streptococcus agalactiae (group B Streptococcus [GBS]). This is the first multi-Lancefield antisera-agglutinating isolate of this species, and it raised particular concern, as it may mimic GBS, leading to false reporting and useless receipt of intrapartum antibiotics. PMID:25766004

  20. Deactivation of Enterococcus Faecalis Bacteria by an Atmospheric Cold Plasma Brush

    NASA Astrophysics Data System (ADS)

    Chen, Wei; Huang, Jun; Du, Ning; Liu, Xiao-Di; Lv, Guo-Hua; Wang, Xing-Quan; Zhang, Guo-Ping; Guo, Li-Hong; Yang, Si-Ze

    2012-07-01

    An atmospheric cold plasma brush suitable for large area and low-temperature plasma-based sterilization is designed and used to treat enterococcus faecalis bacteria. The results show that the efficiency of the inactivation process by helium plasma is dependent on applied power and exposure time. After plasma treatments, the cell structure and morphology changes can be observed by scanning electron microscopy. Optical emission measurements indicate that reactive species such as O and OH play a significant role in the sterilization process.

  1. Serum as a Factor Influencing Adhesion of Enterococcus faecalis to Glass and Silicone

    PubMed Central

    Gallardo-Moreno, Amparo M.; González-Martín, M. Luisa; Pérez-Giraldo, Ciro; Bruque, José M.; Gómez-García, Antonio C.

    2002-01-01

    The purpose of this work was to analyze the effect of serum on the physicochemical surface properties and adhesion to glass and silicone of Enterococcus faecalis ATCC 29212 at 37°C. As is presented using thermodynamics analysis, serum minimizes the interaction of cells with water, which correlates well with the increase in hydrophobicity and in bacterial adhesion to glass and silicone. PMID:12406782

  2. Complete genome sequence of Enterococcus faecalis LD33, a bacteriocin-producing strain.

    PubMed

    Yuehua, Jiao; Lanwei, Zhang; Fei, Liu; Huaxi, Yi; Xue, Han

    2016-06-10

    Enterococcus faecalis LD33 strain was originally isolated from traditional naturally fermented cream in Inner Mongolia of China. Its complete genome sequence was carried out using the Illumina Hiseq and the PacBio RSII platform. The genome only has a circular chromosome and a GC content of 37.58%. Other core information shown in the genome sequencing results further insight on this bacterium's genetic elements for bacteriocin production and the genes related to respiratory chain. PMID:27090021

  3. Bactericidal Effects of Diode Laser Irradiation on Enterococcus faecalis Using Periapical Lesion Defect Model

    PubMed Central

    Nagayoshi, Masato; Nishihara, Tatsuji; Nakashima, Keisuke; Iwaki, Shigetsugu; Chen, Ker-Kong; Terashita, Masamichi; Kitamura, Chiaki

    2011-01-01

    Objective. Photodynamic therapy has been expanded for use in endodontic treatment. The aim of this study was to investigate the antimicrobial effects of diode laser irradiation on endodontic pathogens in periapical lesions using an in vitro apical lesion model. Study Design. Enterococcus faecalis in 0.5% semisolid agar with a photosensitizer was injected into apical lesion area of in vitro apical lesion model. The direct effects of irradiation with a diode laser as well as heat produced by irradiation on the viability of microorganisms in the lesions were analyzed. Results. The viability of E. faecalis was significantly reduced by the combination of a photosensitizer and laser irradiation. The temperature caused by irradiation rose, however, there were no cytotoxic effects of heat on the viability of E. faecalis. Conclusion. Our results suggest that utilization of a diode laser in combination with a photosensitizer may be useful for clinical treatment of periapical lesions. PMID:21991489

  4. Antimicrobial activity of essential oils and chloroform alone and combinated with cetrimide against Enterococcus faecalis biofilm

    PubMed Central

    Ferrer Luque, Carmen Maria; González-Rodríguez, Maria Paloma; Arias-Moliz, Maria Teresa; Baca, Pilar

    2013-01-01

    Abstract The Enterococcus faecalis bacteria have been identified as the most commonly recovered species from teeth with persistent endodontic infections. The antimicrobial activity of essential oils and chloroform (CHL), alone and in association with various concentrations of cetrimide (CTR), against biofilm of Enterococcus faecalis was investigated. Solutions of CHL, eucalyptus oil (EO) and orange oil (OO) associated with CTR at 0.3%, 0.2%, 0.1%, and 0.05% were used to determine antimicrobial activity by exposing treated bovine dentine blocks to E. faecalis. Biofilms grown in the dentine blocks for 7 days were exposed to solutions for 2 and 5 min. Biofilm reduction between OO and EO at 2 min did not show any significant differences; however, OO had a higher kill percentage of biofilms than did the eucalyptus oil at 5 min (p < 0.01). Combinations with CTR at all concentrations achieved a 100% kill rate at 2 and 5 min. The association of CTR with solvent agents achieved the maximum antimicrobial activity against E. faecalis biofilms in dentine. PMID:24265917

  5. Antibacterial Effect of All-in-one Self-etch Adhesives on Enterococcus faecalis

    PubMed Central

    Ebrahimi Chaharom, Mohammad Esmaeel; Ajami, Amir Ahmad; Abed Kahnamouei, Mehdi; Jafari Navimipour, Elmira; Tehranchi, Pardis; Zand, Vahid; Sadeghi, Mohammad Reza; Sohrabi, Aydin

    2014-01-01

    Background and aims. The aim of this study was to evaluate the antibacterial activity of one-step self-etch adhesives on Enterococcus faecalis on days 1, 7 and 14 with the use of modified direct contact test. Materials and methods. The modified directcontact test was used to evaluate the antibacterial effect of Adper Easy One, Bond Force, Clearfil S3 Bond, Futurabond M, G-Bond, iBond and OptiBond All-in-one adhesives on Enterococcus faecalis after aging the samples in phosphate-buffered saline for one, seven and fourteen days. Data were analyzed using one-way ANOVA and post hoc Tukey tests. Aging effect of each adhesive was evaluated by paired-sample test. In this study, P<0.05 was considered significant. Results. All the tested adhesives exhibited antibacterial activity after one day and had significant differences with the positive control group (P<0.05). After one week, OptiBond All-in-one, iBond and Futurabond M exhibited significant differences in bacterial growth from other groups (P<0.05). There were no significant differences between the groups in two weeks (P>0.05). Conclusion. iBond exhibited the highest antibacterial effect on E. faecalis after one week. Futurabond and OptiBond All-in-one exhibited antibacterial effects against E. faecalis for one week. PMID:25587384

  6. Multiple Roles for Enterococcus faecalis Glycosyltransferases in Biofilm-Associated Antibiotic Resistance, Cell Envelope Integrity, and Conjugative Transfer

    PubMed Central

    Dale, Jennifer L.; Cagnazzo, Julian; Phan, Chi Q.; Barnes, Aaron M. T.

    2015-01-01

    The emergence of multidrug-resistant bacteria and the limited availability of new antibiotics are of increasing clinical concern. A compounding factor is the ability of microorganisms to form biofilms (communities of cells encased in a protective extracellular matrix) that are intrinsically resistant to antibiotics. Enterococcus faecalis is an opportunistic pathogen that readily forms biofilms and also has the propensity to acquire resistance determinants via horizontal gene transfer. There is intense interest in the genetic basis for intrinsic and acquired antibiotic resistance in E. faecalis, since clinical isolates exhibiting resistance to multiple antibiotics are not uncommon. We performed a genetic screen using a library of transposon (Tn) mutants to identify E. faecalis biofilm-associated antibiotic resistance determinants. Five Tn mutants formed wild-type biofilms in the absence of antibiotics but produced decreased biofilm biomass in the presence of antibiotic concentrations that were subinhibitory to the parent strain. Genetic determinants responsible for biofilm-associated antibiotic resistance include components of the quorum-sensing system (fsrA, fsrC, and gelE) and two glycosyltransferase (GTF) genes (epaI and epaOX). We also found that the GTFs play additional roles in E. faecalis resistance to detergent and bile salts, maintenance of cell envelope integrity, determination of cell shape, polysaccharide composition, and conjugative transfer of the pheromone-inducible plasmid pCF10. The epaOX gene is located in a variable extended region of the enterococcal polysaccharide antigen (epa) locus. These data illustrate the importance of GTFs in E. faecalis adaptation to diverse growth conditions and suggest new targets for antimicrobial design. PMID:25918141

  7. Confocal microscopy evaluation of the effect of irrigants on Enterococcus faecalis biofilm: An in vitro study.

    PubMed

    Flach, Nicole; Böttcher, Daiana Elisabeth; Parolo, Clarissa Cavalcanti Fatturi; Firmino, Luciana Bitello; Malt, Marisa; Lammers, Marcelo Lazzaron; Grecca, Fabiana Soares

    2016-01-01

    The purpose of this study was to evaluate in vitro the effectiveness of two endodontic irrigants and their association against Enterococcus faecalis (E. faecalis) by confocal laser scanning microscope (CLSM). Twenty-four bovine incisors were inoculated in a monoculture of E. faecalis for 21 days. After this period, the teeth were divided into three test groups (n = 5) according to the chemical used. Group 1: 2.5% sodium hypochlorite (NaOCl), group 2: 2% chlorhexidine gel (CHX), group 3: 2.5% NaOCl + 2% CHX gel, and two control groups (n = 3): negative control group (NCG)-sterile and without root canals preparation and positive control group (PCG)-saline. Then, the samples were stained with SYTO9 and propidium iodide and subjected to analysis by CLSM. Bacterial viability was quantitatively analyzed by the proportions of dead and live bacteria in the biofilm remnants. Statistical analysis was performed by the One-way ANOVA test (p = 0.05). No statistical differences were observed to bacterial viability. According to CLSM analysis, none of the tested substances could completely eliminate E. faecalis from the root canal space. Until now, there are no irrigant solutions able to completely eliminate E. faecalis from the root canal. In this regard, the search for irrigants able to intensify the antimicrobial action is of paramount importance. SCANNING 38:57-62, 2016. © 2015 Wiley Periodicals, Inc. PMID:26153228

  8. The in vitro Effect of Irrigants with Low Surface Tension on Enterococcus faecalis

    PubMed Central

    Giardino, Luciano; Estrela, Carlos; Generali, Luigi; Mohammadi, Zahed; Asgary, Saeed

    2015-01-01

    Introduction: Due to the complex anatomy of the root canal system and high surface tension of common root canal irrigants (RCI), conducting an investigation on RCIs containing surfactants is a priority. The aim of this in vitro study was to verify the antibacterial potential of RCI with low surface tension in root canals infected with Enterococcus faecalis (E. faecalis). Methods and Materials: Thirty-five extracted human maxillary anterior teeth were prepared and inoculated with E. faecalis for 60 days. After root canal preparation, the teeth were randomly divided to one positive and one negative control groups and 5 experimental groups: Hypoclean/Tetraclean NA, Hypoclean, Tetraclean, NaOCl/Tetraclean and NaOCl. Bacterial growth was observed by turbidity of culture medium and then measured using a UV spectrophotometer. Data were analyzed in three time intervals (pre-instrumentation and, 20 min and 72 h after canal preparation) using the ANOVA and post hoc Tukey’s tests. The level of significance was set at 0.05. Results: The results indicated the presence of E. faecalis in all post-irrigation samples irrespective of the RCI. However, the optical densities in both post-irrigation periods showed bacterial reduction and significant differences between groups. Conclusion: RCI with low surface tension showed antibacterial potential in E. faecalis infected roots. PMID:26229541

  9. Screening of Bacteriocin-producing Enterococcus faecalis Strains for Antagonistic Activities against Clostridium perfringens

    PubMed Central

    Kim, So-Young

    2014-01-01

    This study was conducted to isolate and characterize bacteriocin-producing bacteria against Clostridium perfringens (C. perfringens) from domestic animals to determine their usefulness as probiotics. Bacteriocin-producing bacteria were isolated from pig feces by the spot-on-lawn method. A total of 1,370 bacterial stains were isolated, and six were tentatively selected after identifying the inhibitory activity against the pathogenic indicator C. perfringens KCTC 3269 and KCTC 5100. The selected strains were identified as Enterococcus faecalis (E. faecalis) by 16s rRNA sequencing. Most of the isolated bacterial strains were resistant to 0.5% bile salts for 48 h and remained viable after 2 h at pH 3.0. Some E. faecalis also showed strong inhibitory activity against Listeria monocytogenes KCTC 3569, KCTC 3586 and KCTC 3710. In the present study, we finally selected E. faecalis AP 216 and AP 45 strain based on probiotic selection criteria such as antimicrobial activity against C. perfringens and tolerance to acid and bile salts. The bacteriocins of E. faecalis AP 216 and AP 45 strains were highly thermostable, showing anticlostridial activities even after incubation at 121℃ for 15 min. These bacteriocinproducing bacteria and/or bacteriocins could be used in feed manufacturing as probiotics as an alternative to antibiotics in the livestock industry. PMID:26761495

  10. Proteolytic activity of Enterococcus faecalis VB63F for reduction of allergenicity of bovine milk proteins.

    PubMed

    Biscola, V; Tulini, F L; Choiset, Y; Rabesona, H; Ivanova, I; Chobert, J-M; Todorov, S D; Haertlé, T; Franco, B D G M

    2016-07-01

    With the aim of screening proteolytic strains of lactic acid bacteria to evaluate their potential for the reduction of allergenicity of the major bovine milk proteins, we isolated a new proteolytic strain of Enterococcus faecalis (Ent. faecalis VB63F) from raw bovine milk. The proteases produced by this strain had strong activity against caseins (αS1-, αS2-, and β-casein), in both skim milk and sodium caseinate. However, only partial hydrolysis of whey proteins was observed. Proteolysis of Na-caseinate and whey proteins, observed after sodium dodecyl sulfate-PAGE, was confirmed by analysis of peptide profiles by reversed-phase HPLC. Inhibition of proteolysis with EDTA indicated that the proteases produced by Ent. faecalis VB63F belonged to the group of metalloproteases. The optimal conditions for their activity were 42°C and pH 6.5. The majority of assessed virulence genes were absent in Ent. faecalis VB63F. The obtained results suggest that Ent. faecalis VB63F could be efficient in reducing the immunoreactivity of bovine milk proteins. PMID:27179865

  11. Monoclonal antibodies recognizing the Enterococcus faecalis collagen-binding MSCRAMM Ace: conditional expression and binding analysis.

    PubMed

    Hall, Andrea E; Gorovits, Elena L; Syribeys, Peter J; Domanski, Paul J; Ames, Brenda R; Chang, Cathy Y; Vernachio, John H; Patti, Joseph M; Hutchins, Jeff T

    2007-01-01

    Enterococci are opportunistic pathogens known to cause numerous clinical infections and complications in humans. Adhesin-mediated binding to extracellular matrix (ECM) proteins of the host is thought to be a crucial step in the pathogenesis of these bacterial infections. Adhesin of collagen from Enterococcus faecalis (Ace) is a cell-wall anchored protein of E. faecalis that has been shown to be important for bacterial binding to the ECM. In this report, we characterize the conditions for Ace expression and demonstrate Ace binding to mammalian epithelial and endothelial cells as well as to collagens found in the ECM. To further characterize Ace expression and function, we report the generation of a panel of monoclonal antibodies (mAbs) directed against this important E. faecalis virulence factor. Through the use of multiple in vitro assays, surface plasmon resonance and flow cytometry, we have characterized this panel of mAbs which may prove to be not only beneficial in studies that address the precise biological role of adhesion of E. faecalis, but may also serve as beneficial therapeutic agents against E. faecalis infections. PMID:17521860

  12. A Rex Family Transcriptional Repressor Influences H2O2 Accumulation by Enterococcus faecalis

    PubMed Central

    Vesić, Dušanka

    2013-01-01

    Rex factors are bacterial transcription factors thought to respond to the cellular NAD+/NADH ratio in order to modulate gene expression by differentially binding DNA. To date, Rex factors have been implicated in regulating genes of central metabolism, oxidative stress response, and biofilm formation. The genome of Enterococcus faecalis, a low-GC Gram-positive opportunistic pathogen, encodes EF2638, a putative Rex factor. To study the role of E. faecalis Rex, we purified EF2638 and evaluated its DNA binding activity in vitro. EF2638 was able to bind putative promoter segments of several E. faecalis genes in an NADH-responsive manner, indicating that it represents an authentic Rex factor. Transcriptome analysis of a ΔEF2638 mutant revealed that genes likely to be involved in anaerobic metabolism were upregulated during aerobic growth, and the mutant exhibited an altered NAD+/NADH ratio. The ΔEF2638 mutant also exhibited a growth defect when grown with aeration on several carbon sources, suggesting an impaired ability to cope with oxidative stress. Inclusion of catalase in the medium alleviated the growth defect. H2O2 measurements revealed that the mutant accumulates significantly more H2O2 than wild-type E. faecalis. In summary, EF2638 represents an authentic Rex factor in E. faecalis that influences the production or detoxification of H2O2 in addition to its more familiar role as a regulator of anaerobic gene expression. PMID:23417491

  13. Characterization of Enterococcus faecalis Phage IME-EF1 and Its Endolysin

    PubMed Central

    Fan, Hang; An, Xiaoping; Zhang, Zhiyi; Chen, Jiankui; Tong, Yigang

    2013-01-01

    Enterococcus faecalis is increasingly becoming an important nosocomial infection opportunistic pathogen. E. faecalis can easily obtain drug resistance, making it difficult to be controlled in clinical settings. Using bacteriophage as an alternative treatment to drug-resistant bacteria has been revitalized recently, especially for fighting drug-resistant bacteria. In this research, an E. faecalis bacteriophage named IME-EF1 was isolated from hospital sewage. Whole genomic sequence analysis demonstrated that the isolated IME-EF1 belong to the Siphoviridae family, and has a linear double-stranded DNA genome consisting of 57,081 nucleotides. The IME-EF1 genome has a 40.04% G+C content and contains 98 putative coding sequences. In addition, IME-EF1 has an isometric head with a width of 35 nm to 60 nm and length of 75 nm to 90 nm, as well as morphology resembling a tadpole. IME-EF1 can adsorb to its host cells within 9 min, with an absorbance rate more than 99% and a latent period time of 25 min. The endolysin of IME-EF1 contains a CHAP domain in its N-terminal and has a wider bactericidal spectrum than its parental bacteriophage, including 2 strains of vancomycin-resistant E. faecalis. When administrated intraperitoneally, one dose of IME-EF1 or its endolysin can reduce bacterial count in the blood and protected the mice from a lethal challenge of E. faecalis, with a survival rate of 60% or 80%, respectively. Although bacteriophage could rescue mice from bacterial challenge, to the best of our knowledge, this study further supports the potential function of bacteriophage in dealing with E. faecalis infection in vivo. The results also indicated that the newly isolated bacteriophage IME-EF1 enriched the arsenal library of lytic E. faecalis bacteriophages and presented another choice for phage therapy in the future. PMID:24236180

  14. Transmission and genetic diversity of Enterococcus faecalis among layer chickens during hatch

    PubMed Central

    2011-01-01

    Background Studies on transmission of Enterococcus faecalis among chickens during hatch have not been carried out so far. Information about vertical transmission and subsequent spreading and colonization of the cloacal mucosa through cloacal 'drinking' during hatch are important to understand the epidemiology of E. faecalis infections. In the present investigation vertical transmission and subsequent spreading and colonization of the cloacal mucosa of chickens by E. faecalis through cloacal 'drinking' were examined. Methods Two different batches of layer chickens originating from 45 weeks old Brown and White Lohmann parents, respectively from the same farm were sampled in the hatcher. Isolates were confirmed to be E. faecalis by polymerase chain reaction (PCR) and further by multilocus sequence typing (MLST) to state their population structure and comparison made to sequence types previously obtained from chicken. Results A total of 480 chickens were swabbed from the cloacae just after hatch and after 24 hours. A total of 101 isolates were confirmed as E. faecalis by a species specific PCR. The prevalence of E. faecalis increased from 14% at 0 h to 97% after 24 h for the Brown Lohmann chickens and from 0.5% to 23% for the White Lohmann flock. The 84 isolates analysed by MLST were distributed on 14 sequence types (ST). Three ST (401, 82 and 249) accounted for 64% of all isolates analysed by MLST after 24 h. ST 82 has previously been reported from amyloid arthropathy and other lesions in poultry. Conclusions The present findings demonstrated a high potential of a few contaminated eggs or embryos to rapidly facilitate the spread of E. faecalis to almost all chickens during hatch. PMID:22017822

  15. First detection of the antiseptic resistance gene qacA/B in Enterococcus faecalis.

    PubMed

    Bischoff, Meike; Bauer, Johann; Preikschat, Petra; Schwaiger, Karin; Mölle, Gabriele; Hölzel, Christina

    2012-02-01

    Resistance to disinfectants is well investigated in staphylococci and pseudomonads but nearly unexplored in bacteria of the genus Enterococcus, despite their rising significance as nosocomial pathogens. In this study, Enterococcus faecalis (n=585) from blood (n=42) and stool (n=109) of hospitalized humans, from faeces of farm animals (n=226), and from food (milk and dairy products, n=96; meat and meat products, n=112) were screened for the presence of qac-genes (qacA, qacB, qacC, smr [qacC+qacD], qacEΔ1, qacG, qacH, qacJ) via PCR. The isolates' susceptibility to a quaternary ammonium compound (didecyldimethylammoniumchloride, DDAC) and antibiotics was assessed by microdilution. Four E. faecalis strains were positive for qac-genes: qacA/B was found in one isolate from cattle and one isolate from human blood; smr (qacC+qacD) was detected in one isolate from human stool and in one isolate from cheese ("Camembert"). The sequences of the qacA/B-amplicons differed in two basepairs. DDAC had an elevated minimum inhibitory concentration (MIC) of 2.45-3.5 mg/L in one qacA/B-positive strain from human blood, whereas the other qac-gene carriers had wild-type MIC-values for DDAC (1.05 mg/L). This is the first detection of qacA/B in the genus Enterococcus. PMID:22017402

  16. Enterococcus faecalis sufCDSUB complements Escherichia coli sufABCDSE.

    PubMed

    Riboldi, Gustavo P; Larson, Timothy J; Frazzon, Jeverson

    2011-07-01

    Iron-sulfur [Fe-S] clusters are inorganic prosthetic groups that play essential roles in all living organisms. Iron and sulfur mobilization, formation of [Fe-S] clusters, and delivery to its final protein targets involves a complex set of specific protein machinery. Proteobacteria has three systems of [Fe-S] biogenesis, designated NIF, ISC, and SUF. In contrast, the Firmicutes system is not well characterized and has only one system, formed mostly by SUF homologs. The Firmicutes phylum corresponds to a group of pathological bacteria, of which Enterococcus faecalis is a clinically relevant representative. Recently, the E. faecalis sufCDSUB [Fe-S] cluster biosynthetic machinery has been identified, although there is no further information available about the similarities and/or variations of Proteobacteria and Firmicutes systems. The aim of the present work was to compare the ability of the different Proteobacteria and Firmicutes systems to complement the Azotobacter vinelandii and Escherichia coli ISC and SUF systems. Indeed, E. faecalis sufCDSUB is able to complement the E. coli SUF system, allowing viable mutants of both sufABCDSE and iscRSU-hscBA-fdx systems. The presence of all E. faecalis SUF factors enables proper functional interactions, which would not otherwise occur in proteins from different systems. PMID:21480963

  17. Genetic analysis of faropenem-resistant Enterococcus faecalis in urinary isolates.

    PubMed

    Hiraga, Noriyuki; Muratani, Tetsuro; Naito, Seiji; Matsumoto, Tetsuro

    2008-04-01

    We isolated faropenem-resistant Enterococcus faecalis in urine specimens and studied the mechanisms of resistance to faropenem in these isolates. Three mechanisms of penicillin resistance have been reported in E. faecalis; (1) beta-lactamase production, (2) overproduction of penicillin-binding protein (PBP) 4 or PBP5, and (3) decreasing affinities of penicillins for PBP4 by the occurrence of point mutations of the penicillin-binding domain. None of the E. faecalis isolates examined produced beta-lactamase or overproduced any PBPs, but the affinities of faropenem for PBP4 were decreased in faropenem-insensitive and -resistant strains. We found single amino acid substitutions at positions 475, 520 or 605 in PBP4 in the insensitive strains and two amino acid substitutions at positions 520 and 605 in PBP4 in the resistant strains by sequencing the entire pbp4 gene from each isolate. We conclude that development of resistance to faropenem in E. faecalis is due to decreasing affinities for PBP4 that are the result of the occurrence of one or two point mutations. PMID:18503200

  18. Effect of the quorum-sensing luxS gene on biofilm formation by Enterococcus faecalis.

    PubMed

    He, Zhiyan; Liang, Jingping; Zhou, Wei; Xie, Qian; Tang, Zisheng; Ma, Rui; Huang, Zhengwei

    2016-06-01

    Enterococcus faecalis is the species of bacterium most frequently isolated from the root canals of teeth that exhibit chronic apical periodontitis refractory to endodontic treatment. In this study, we evaluated the effect of the S-ribosylhomocysteine lyase (luxS) quorum-sensing gene on E. faecalis biofilm formation by constructing a knockout mutant. The biofilms formed by both E. faecalis and its luxS mutant strain were evaluated using the MTT method. Important parameters that influence biofilm formation, including cell-surface hydrophobicity and the nutrient content of the growth medium, were also studied. Biofilm structures were observed using confocal laser scanning microscopy (CLSM), and expression of biofilm-related genes was investigated using RT-PCR. The results showed that the luxS gene can affect biofilm formation, whereas it does not affect the bacterial growth rate. Deletion of the luxS gene also increased cell-surface hydrophobicity. Biofilm formation was accelerated by the addition of increasing concentrations of glucose. The CLSM images revealed that the luxS mutant strain tends to aggregate into distinct clusters and relatively dense structures, whereas the wild-type strain appears confluent and more evenly distributed. All genes examined were up-regulated in the biofilms formed by the luxS mutant strain. The quorum-sensing luxS gene can affect E. faecalis biofilm formation. PMID:27080421

  19. Antimicrobial Activity of Calcium Hydroxide and Betamethasone on Enterococcus faecalis; An in vitro Assessment

    PubMed Central

    Tabrizizadeh, Mahdi; Rasti, Mojtaba; Ayatollahi, Fatemeh; Mossadegh, Mohammad Hossein; Zandi, Hengameh; Dehghan, Farzad; Mousavi, Zohreh

    2015-01-01

    Introduction: Calcium hydroxide (CH) is one of the most common intracanal medications. Corticosteroids (CS) are used in endodontics because of their anti-inflammatory activity. This study aimed to evaluate the antimicrobial effect of CH+betamethasone and CH+saline against Enterococcus faecalis (E. faecalis) using agar diffusion test and measuring the microbial zone of inhibition (ZOI). Methods and Materials: Four plates containing Mueller-Hinton broth and E. faecalis culture media, were prepared. In each plate, 5 holes (5×3 mm) were created and a creamy mixture of CH+betamethasone was inserted into the holes (10 holes for each material). Two holes with ampicillin disks and two empty holes were used as negative and positive controls, respectively. Plates were incubated for 24 h and then the diameter of microbial ZOI was measured. The pH of each mixture was measured by pH meter. Data were analyzed using the Mann-Whitney U test. Results: The mean diameter of ZOI for CH+betamethasone and CH+saline was 3.4 and 3 mm, respectively. The difference was not significant (P=0.143). The pH was 12.5 for CH+saline and 12.3 CH+betamethasone, respectively. Conclusion: The mixture of CH+betamethasone had good antimicrobial effects against E. faecalis. Further studies are needed to confirm the value of this mixture in clinical settings. PMID:26213541

  20. Evaluation of the presence of Enterococcus Faecalis in root cementum: A confocal laser scanning microscope analysis

    PubMed Central

    Halkai, Rahul; Hegde, Mithra N; Halkai, Kiran

    2014-01-01

    Aim: The aim of this study is to address the cause of persistent infection of root cementum by Enterococcus faecalis. Materials and Methods: A sample of 60 human single-rooted teeth were divided into three groups. Group I (control group) had no access opening and one-third of the apical root cementum was sealed using varnish. Group II had no preparation of teeth samples. In group III, apical root cementum was exposed to organic acid and roughened using diamond point to mimic apical resorption. After access opening in groups II and III, all teeth samples were sterilized using gamma irradiation (25 kGy). E. faecalis broth was placed in the root canal and apical one-third of the tooth was immersed in the broth for 8 weeks with alternate day refreshment followed by biomechanical preparation, obturation and coronal seal. Apical one-third of all teeth samples were again immersed in the broth for 8 weeks with alternate day refreshment to mimic secondary infection. The samples were observed under a confocal microscope after splitting the teeth into two halves. Results: E. faecalis penetrated 160 μm deep into the root cementum in group III samples and only showed adhesion in group II samples. Conclusion: Penetration and survival of E. faecalis deep inside the cementum in extreme conditions could be the reason for persistent infection. PMID:24778505

  1. Effectiveness of a high purity chlorine dioxide solution in eliminating intracanal Enterococcus faecalis biofilm.

    PubMed

    Herczegh, Anna; Ghidan, Agoston; Friedreich, Dóra; Gyurkovics, Milán; Bendő, Zsolt; Lohinai, Zsolt

    2013-03-01

    We investigated the effectiveness of chlorine dioxide (ClO2) solution in comparison to sodium hypochlorite (NaOCl) and chlorhexidine gluconate (CHX) in the elimination of intracanal Enterococcus faecalis biofilm. Extracted human teeth were inoculated with E. faecalis. After preparation the canals were irrigated with ClO2, NaOCl, CHX or physiologic saline for control. Two and five days later bacterial samples were collected and streaked onto Columbia agar. CFU/mL were counted. The canal walls were investigated by scanning electron microscopy (SEM). The gas phase was investigated in an upside down Petri dish where E. faecalis was inoculated onto blood agar. The irrigants were placed on absorbent paper into the cover. Bacteria were detectable in the control group, but not in any of the irrigants groups. There was a massive reinfection 2 or 5 days after irrigation in the control group. The lowest reinfection was found after the ClO2 treatment. These findings were confirmed by SEM images. We observed an antibacterial effect of ClO2 and NaOCl gas phases on E. faecalis growth, but not of CHX. ClO2 eliminates intracanal biofilm and keeps canal nearly free from bacteria. We suggest the use of high purity ClO2 as a root canal irrigant in clinical practice. PMID:23529300

  2. Epidemiology of Enterococcus faecalis urinary tract infection in a teaching hospital in London, United Kingdom.

    PubMed Central

    Hall, L M; Duke, B; Urwin, G; Guiney, M

    1992-01-01

    Enterococcus faecalis is a frequent cause of urinary tract infection in hospitalized patients. Recent reports have suggested that the organism may frequently be acquired by cross-infection from other patients. In this study, we used total DNA restriction patterns to type 135 urine isolates of E. faecalis from four sets of patients. Isolates were placed into types (all bands identical) and into groups (most bands identical). Most isolates were discriminated by the typing method, and the results suggested that direct cross-infection occurred rarely if at all. However, two groups of clonally related isolates occurred frequently in the urine specimens and also in feces from hospital-associated patients and were often associated with antibiotic resistance. Isolates from these two groups were found less frequently in feces from people not associated with the hospital. Images PMID:1500498

  3. Survival of Enterococcus faecalis in seawater microcosms is limited in the presence of bacterivorous zooflagellates.

    PubMed

    Hartke, A; Lemarinier, S; Pichereau, V; Auffray, Y

    2002-05-01

    The survival and persistence of growing and starved cells of Enterococcus faecalis in untreated and differentially filtered (20 microm, 5 microm, 3 microm, 1.2 microm, and 0.1 microm) seawater was analyzed in samples taken at different times over a 1-year period by plate counts and scanning electron microscopy. Whereas seawater filtered through a 0.1-microm mesh was not at all or only slightly bactericidal during incubation at 16 degrees C in the dark, culturability of E. faecalis in the other systems decreased as a function of increasing pore size of the filters. Recovery of culturable, glucose pre-starved cells was always higher than that of cells harvested from the exponential growth phase. Electron microscopic analysis showed that the disappearance of enterococci appeared related to the presence and multiplication of various zooflagellates. PMID:11927983

  4. Cloning, purification, crystallization and preliminary crystallographic analysis of SecA from Enterococcus faecalis

    SciTech Connect

    Meining, Winfried; Scheuring, Johannes; Fischer, Markus; Weinkauf, Sevil

    2006-06-01

    SecA ATPase from E. faecalis has been cloned, overexpressed, purified and crystallized. Crystals belong to space group C2 and diffract to 2.4 Å resolution. The gene coding for SecA from Enterococcus faecalis was cloned and overexpressed in Escherichia coli. In this protein, the lysine at position 6 was replaced by an asparagine in order to reduce sensitivity towards proteases. The modified protein was purified and crystallized. Crystals diffracting to 2.4 Å resolution were obtained using the vapour-diffusion technique. The crystals belong to the monoclinic space group C2, with unit-cell parameters a = 203.4, b = 49.8, c = 100.8 Å, α = γ = 90.0, β = 119.1°. A selenomethionine derivative was prepared and is currently being tested in crystallization trials.

  5. Enterococcus faecalis cytolysin without effect on the intestinal growth of susceptible enterococci in mice.

    PubMed

    Huycke, M M; Joyce, W A; Gilmore, M S

    1995-07-01

    A murine model was developed to determine whether the Enterococcus faecalis cytolysin, through its bacteriolytic action on gram-positive bacteria, could promote intestinal overgrowth of cytolytic strains. Sets of E. faecalis strains with varying cytolytic production and susceptibility to cytolytic activity were mixed 1:1 and allowed to compete in vitro in broth or in vivo after orogastric administration in mice pretreated with antibiotics. In general, cytolytic strains outgrew, by as much as 2000-fold, competing cytolysin-susceptible or -hypersusceptible strains in vitro. In contrast, no growth advantage was observed in vivo, despite similar transient colonization of the murine intestinal tract by both cytolytic and cytolysin-susceptible strains. These data suggest that cytolysin plays little role in promoting intestinal overgrowth of enterococci through bacteriolytic activity. PMID:7797930

  6. Characterisation of the Plasmidome within Enterococcus faecalis Isolated from Marginal Periodontitis Patients in Norway

    PubMed Central

    Mikalsen, Theresa; Roberts, Adam P.; Sundsfjord, Arnfinn

    2013-01-01

    The present study aimed to identify and characterize plasmids in a national collection of oral Enterococcus faecalis (n = 106) isolated from patients with marginal periodontitis. Plasmid replicon typing was performed by multiplex-PCR and sequencing with specific primers for 18 rep-families and 1 unique sequence. Additional plasmid analysis by S1-PFGE was performed for comparison. Totally 120 plasmid replicon amplicons of seven rep-families were identified in 93 E. faecalis strains, e.g. rep9 (prototype pCF10), rep6 (prototype pS86), rep2 (prototype pRE25/pEF1), and rep8 (prototype pAM373). Rep9 was the most predominant rep-family being detected in 81 (76.4%) strains. Forty of these strains were tetracycline resistant and three were erythromycin resistant. Rep6 was the second predominant rep-family being detected in 22 (20.8%) strains. Rep2 was detected in eight (7.5%) strains. All rep2-positive strains were resistant to tetracycline and/or erythromycin and six of them contained Tn916/Tn1545 genes. The rep-positive E. faecalis exhibited divergence in multilocus sequence types (STs). There was a significant correlation between rep9 and ST21, while multiple rep-families appeared in ST40. Totally 145 plasmid bands were identified in 95 E. faecalis strains by S1-PFGE, 59 strains carrying one plasmid, 27 carrying two, five carrying three, three carrying four, and one strain carrying five plasmids. Plasmid sizes varied between 5–150 kbp. There was a significant correlation between the number of plasmids identified by PCR rep-typing and by S1-PFGE. The results indicate that the majority of E. faecalis of marginal periodontitis are likely to be a reservoir for diverse mobile genetic elements and associated antimicrobial resistance determinants. PMID:23646122

  7. Enterococcus faecalis subverts and invades the host urothelium in patients with chronic urinary tract infection.

    PubMed

    Horsley, Harry; Malone-Lee, James; Holland, David; Tuz, Madeleine; Hibbert, Andrew; Kelsey, Michael; Kupelian, Anthony; Rohn, Jennifer L

    2013-01-01

    Bacterial urinary tract infections (UTI) are a major growing concern worldwide. Uropathogenic Escherichia coli has been shown to invade the urothelium during acute UTI in mice and humans, forming intracellular reservoirs that can evade antibiotics and the immune response, allowing recurrence at a later date. Other bacterial species, such as Staphylococcus saprophyticus, Klebsiella pneumonia and Salmonella enterica have also been shown to be invasive in acute UTI. However, the role of intracellular infection in chronic UTI causing more subtle lower urinary tract symptoms (LUTS), a particular problem in the elderly population, is poorly understood. Moreover, the species of bacteria involved remains largely unknown. A previous study of a large cohort of non-acute LUTS patients found that Enterococcus faecalis was frequently found in urine specimens. E. faecalis accounts for a significant proportion of chronic bladder infections worldwide, although the invasive lifestyle of this uropathogen has yet to be reported. Here, we wanted to explore this question in more detail. We harvested urothelial cells shed in response to inflammation and, using advanced imaging techniques, inspected them for signs of bacterial pathology and invasion. We found strong evidence of intracellular E. faecalis harboured within urothelial cells shed from the bladder of LUTS patients. Furthermore, using a culture model system, these patient-isolated strains of E. faecalis were able to invade a transitional carcinoma cell line. In contrast, we found no evidence of cellular invasion by E. coli in the patient cells or the culture model system. Our data show that E. faecalis is highly competent to invade in this context; therefore, these results have implications for both the diagnosis and treatment of chronic LUTS. PMID:24363814

  8. Sub-lethal stress effects on virulence gene expression in Enterococcus faecalis.

    PubMed

    Lenz, Christian A; Hew Ferstl, Carrie M; Vogel, Rudi F

    2010-05-01

    Enterococci are ubiquitous lactic acid bacteria commonly associated with the human digestive tract as commensal organisms. Additionally, these organisms have a long history of use in foods improving flavor as well as providing protective mechanisms as either a probiotic or antimicrobial additive. However, Enterococcus faecalis accounts for up to 10% of all nosocomial infections of the bloodstream, wounds, urinary tract and heart. Knowledge about the regulation of virulence factors is limited and the involvement of environmental signals contributing to E. faecalis pathogenicity is poorly documented. In this study, two clinical E. faecalis isolates, TMW 2.63 and OG1RF, as well as one food isolate, TMW 2.629, were subjected to six sub-lethal food- and host-related stresses including 6.8% NaCl, 200 ppm nitrite, 51 degrees C, 80 MPa, pH 4.1 and 0.08% bile salts (cholic acid:chenodeoxycholic acid 1:1), respectively, reducing their growth rate to 10%. Relative gene expression of 15 stress and virulence-associated genes including dnaK, groEL, ctsR, clpPBCEX, gls24, efaAfs, ace, fsrB, gelE, sprE and cylB, was quantified by using real time PCR and Lightcycler((R)) technology (reference conditions: BHI broth, 37 degrees C, pH = 7.4). Apart from strain-dependent differences, sub-lethal environmental stress was capable of provoking significant alterations in the expression of virulence-associated genes in E. faecalis from clinical as well as food origins of isolation. These results help to avoid preconditioning enterococci in food production processes and to understand the complex mechanisms in E. faecalis' switch to pathogenicity. PMID:20227595

  9. Inhibition of initial adhesion of uropathogenic Enterococcus faecalis by biosurfactants from Lactobacillus isolates.

    PubMed Central

    Velraeds, M M; van der Mei, H C; Reid, G; Busscher, H J

    1996-01-01

    In this study, 15 Lactobacillus isolates were found to produce biosurfactants in the mid-exponential and stationary growth phases. The stationary-phase biosurfactants from lactobacillus casei subsp. rhamnosus 36 and ATCC 7469, Lactobacillus fermentum B54, and Lactobacillus acidophilus RC14 were investigated further to determine their capacity to inhibit the initial adhesion of uropathogenic Enterococcus faecalis 1131 to glass in a parallel-plate flow chamber. The initial deposition rate of E. faecalis to glass with an adsorbed biosurfactant layer from L. acidophilus RC14 or L. fermentum B54 was significantly decreased by approximately 70%, while the number of adhering enterococci after 4 h of adhesion was reduced by an average of 77%. The surface activity of the biosurfactants and their activity inhibiting the initial adhesion of E. faecalis 1131 were retained after dialysis (molecular weight cutoff, 6,000 to 8,000) and freeze-drying. Fourier transform infrared spectroscopy and X-ray photoelectron spectroscopy revealed that the freeze-dried biosurfactants from L. acidophilus RC14 and L. fermentum B54 were richest in protein, while those from L. casei subsp. rhamnosus 36 and ATCC 7469 had relatively high polysaccharide and phosphate contents. PMID:8787394

  10. Enterococcus faecalis reconfigures its gene regulatory network activation under copper exposure

    PubMed Central

    Latorre, Mauricio; Galloway-Peña, Jessica; Roh, Jung Hyeob; Budinich, Marko; Reyes-Jara, Angélica; Murray, Barbara E.; Maass, Alejandro; González, Mauricio

    2014-01-01

    A gene regulatory network was generated in the bacterium Enterococcus faecalis in order to understand how this organism can activate its expression under different copper concentrations. The topological evaluation of the network showed common patterns described in other organisms. Integrating microarray experiments allowed the identification of sub-networks activated under low (0.05 mM CuSO4) and high (0.5 mM CuSO4) copper concentrations. The analysis indicates the presence of specific functionally activated modules induced by copper, highlighting the regulons LysR, ArgR as global regulators and CopY, Fur and LexA as local regulators. Taking advantage of the fact that E. faecalis presented a homeostatic module isolated, we produced an in vivo intervention removing this system from the cell without affecting the connectivity of the global transcriptional network. This strategy led us to find that this bacterium can reconfigure its gene expression to maintain cellular homeostasis, activating new modules principally related to glucose metabolism and transcriptional processes. Finally, these results position E. faecalis as the organism having the most complete and controllable systemic model of copper homeostasis available to date. PMID:24382465

  11. Antimicrobial activity of tetraacetylethylenediamine-sodium perborate versus sodium hypochlorite against Enterococcus faecalis

    PubMed Central

    Shakouie, Sahar; Salem Milani, Amin; Eskandarnejad, Mahsa; Rahimi, Saeed; Froughreyhani, Mohammad; Galedar, Saeede; Ranjbar, Ehsan

    2016-01-01

    Background. This study evaluated the antimicrobial activity of Tetraacetylethylenediamine-sodium perborate (TAED-SP) in comparison to 2.5% and 5% sodium hypochlorite (NaOCl) against Enterococcus faecalis. Methods. A standard suspension of E. faecalis was inoculated on 60 plates containing Mueller-Hinton agar culture medium. Four sterile disks of Beckman filtration paper were placed on each plate. TAED-SP, 5% and 2.5% NaOCl were placed on three disks. Sterile physiologic saline was placed on the fourth disk as negative control. After 24-hour incubation, the diameter of the inhibition zone around the disks was measured using a transparent ruler. One-way Analysis of Variance (ANOVA) was used to compare the mean zone of microbial growth in the groups. P-values less than 0.05 were considered statistically significant. Results. There was a significant difference in the diameter of the inhibition zones between groups (P < 0.05). The Tukey post hoc test showed a higher diameter of the inhibitory zone with TAED-SP than that of 2.5% NaOCl. However, there were no significant differences between the inhibitory zones of TAED-SP and 5% NaOCl. Conclusion. TAED-SP and 5% NaOCl have similar antibacterial activity against E. faecalis; however, TAED-SP has a greater antibacterial effect compared to 2.5% NaOCl. PMID:27092214

  12. In Vitro Antimicrobial Effect of a Cold Plasma Jet against Enterococcus faecalis Biofilms

    PubMed Central

    Jiang, Chunqi; Schaudinn, Christoph; Jaramillo, David E.; Webster, Paul; Costerton, J. William

    2012-01-01

    The hypothesis that a cold plasma jet has the antimicrobial effect against Enterococcus faecalis biofilms was tested in vitro. 27 hydroxyapatite discs were incubated with E. faecalis for six days to form a monoculture biofilm on the disc surface. The prepared substrata were divided into three groups: the negative control, the positive control (5.25% NaOCl solution), and the plasma treatment group. Resultant colony-forming unit counts were associated with observations of bacterial cell morphology changes using scanning electron microscopy (SEM). Treatment of E. faecalis biofilm with the plasma and 5.25% NaOCl for 5 min resulted in 93.1% and 90.0% kill (P < 0.0001), respectively. SEM detected that nearly no intact bacteria were discernible for the plasma-exposed HA disc surfaces. The demonstrated bactericidal effect of the plasma with direct surface contact may be due to the enhanced oxidation by the locally produced reactive plasma species. PMID:22461988

  13. Enterococcus faecalis Ebp pili are important for cell-cell aggregation and intraspecies gene transfer.

    PubMed

    La Rosa, Sabina Leanti; Montealegre, Maria Camila; Singh, Kavindra V; Murray, Barbara E

    2016-05-01

    Enterococcus faecalis is an opportunistic pathogen that ranks among the leading causes of biofilm-associated infections. We previously demonstrated that the endocarditis- and biofilm-associated pili (Ebp) of E. faecalis play a major role in biofilm formation, adherence to abiotic surfaces and experimental infections. In this study, derivatives of E. faecalis strain OG1 were engineered to further characterize functions of Ebp pili. Loss of pili resulted in a 36-fold decrease in the number of closely associated cells when OG1RFΔebpABC was mixed with OG1SSpΔebpABC, compared with mixing the Ebp+ parental strains. In addition, using the Ebp+ parental strains as donor and recipient, we found a statistically significant increase (280-360 %, P < 0.05) in the frequency of plasmid transfer versus using Ebp-  mutants in the conjugation experiments. These results demonstrate a previously unrecognized role of Ebp pili, namely, as important contributors to microscale cell aggregation and horizontal spread of genetic material. PMID:26967674

  14. Genome-based characterization of hospital-adapted Enterococcus faecalis lineages.

    PubMed

    Raven, Kathy E; Reuter, Sandra; Gouliouris, Theodore; Reynolds, Rosy; Russell, Julie E; Brown, Nicholas M; Török, M Estée; Parkhill, Julian; Peacock, Sharon J

    2016-01-01

    Vancomycin-resistant Enterococcus faecalis (VREfs) is an important nosocomial pathogen(1,2). We undertook whole genome sequencing of E. faecalis associated with bloodstream infection in the UK and Ireland over more than a decade to determine the population structure and genetic associations with hospital adaptation. Three lineages predominated in the population, two of which (L1 and L2) were nationally distributed, and one (L3) geographically restricted. Genome comparison with a global collection identified that L1 and L3 were also present in the USA, but were genetically distinct. Over 90% of VREfs belonged to L1-L3, with resistance acquired and lost multiple times in L1 and L2, but only once followed by clonal expansion in L3. Putative virulence and antibiotic resistance genes were over-represented in L1, L2 and L3 isolates combined, versus the remainder. Each of the three main lineages contained a mixture of vancomycin-resistant and -susceptible E. faecalis (VSEfs), which has important implications for infection control and antibiotic stewardship. PMID:27572164

  15. Characterization of Small-Colony Variants of Enterococcus faecalis Isolated from Chickens with Amyloid Arthropathy▿

    PubMed Central

    Petersen, Andreas; Chadfield, Mark S.; Christensen, Jens P.; Christensen, Henrik; Bisgaard, Magne

    2008-01-01

    In this study we report the isolation and characterization of normal-sized and small-colony variants of Enterococcus faecalis from outbreaks of amyloid arthropathy in chickens. Postmortem examinations of 59 chickens revealed orange deposits in the knee joints, typical for amyloid arthropathy. Bacterial cultures from 102 joints and 43 spleens exhibited pure (n = 88) and mixed (n = 11) cultures of normal (n = 60) and pinpoint (n = 28) colonies of E. faecalis. Pulsed-field gel electrophoresis of 62 isolates demonstrated seven different band patterns with at most two band size variations, and multilocus sequence typing demonstrated two different sequence types, sharing six out of seven alleles, suggesting a close evolutionary relationship between isolates obtained from four outbreaks. In addition, all isolates were clonally related to an amyloid arthropathy reference strain from The Netherlands, previously shown to be globally dispersed. Initial investigation of the isolated small-colony variant phenotype revealed no difference in whole-cell protein profiling between normal and pinpoint colonies. However, the pinpoint colony isolates appeared to be more virulent in an in vivo challenge model in chickens than their normal-sized-colony counterparts. In addition, pinpoint morphology and associated slow growth were expressed without reversion after in vitro and in vivo passage, suggesting a genuine altered phenotype, and in some instances normal colonies converted to pinpoint morphology postinfection. In conclusion, small-colony variants of E. faecalis are described for the first time from veterinary clinical sources and in relation to amyloid arthropathy in chickens. PMID:18579713

  16. Involvement of PhoP-PhoS homologs in Enterococcus faecalis virulence.

    PubMed

    Teng, Fang; Wang, Ling; Singh, Kavindra V; Murray, Barbara E; Weinstock, George M

    2002-04-01

    Eleven PhoP-PhoS homolog pairs were identified by searching the Enterococcus faecalis V583 genome sequence database at The Institute for Genomic Research with the Bacillus subtilis PhoP-PhoS sequences. Each pair appears to be a potential two-component system composed of a response regulator and a sensor kinase. Seven of the homologs were disrupted in E. faecalis strain OG1RF. TX10293, a mutant disrupted in one of these genes (etaR, the first gene of the gene pair designated etaRS), showed delayed killing and a higher 50% lethal dose in a mouse peritonitis model. The predicted EtaR protein sequence showed greatest similarity to LisR of Listeria monocytogenes (77%) and CsrR of Streptococcus pyogenes (70%); EtaS is 53% similar to LisK and 54% similar to CsrS. When grown in vitro, the TX10293 mutant was more sensitive to low pH (pH 3.4) and more resistant to high temperature (55 degrees C) than wild-type OG1RF. In conclusion, many potential two-component systems are identified for E. faecalis, one of which, EtaRS, was shown to be involved in stress response and virulence. PMID:11895963

  17. Transcriptome profiling of TDC cluster deletion mutant of Enterococcus faecalis V583.

    PubMed

    Perez, Marta; Ladero, Victor; Del Rio, Beatriz; Redruello, Begoña; de Jong, Anne; Kuipers, Oscar P; Kok, Jan; Martin, M Cruz; Fernandez, Maria; Alvarez, Miguel A

    2016-09-01

    The species Enterococcus faecalis is able to catabolise the amino acid tyrosine into the biogenic amine tyramine by the tyrosine decarboxilase (TDC) pathway Ladero et al. (2012) [1]. The TDC cluster comprises four genes: tyrS, an aminoacyl-tRNA synthetase-like gene; tdcA, which encodes the tyrosine decarboxylase; tyrP, a tyrosine/tyramine exchanger gene and nhaC-2, which encodes an Na(+)/H(+) antiporter and whose role in the tyramine biosynthesis remains unknown [2]. In E. faecalis V583 the last three genes are co-transcribed as a single polycistronic mRNA forming the catabolic operon, while tyrS is transcribed independently of the catabolic genes as a monocistronic mRNA [2]. The catabolic operon is transcriptionally induced by tyrosine and acidic pH. On the opposite, the tyrS expression is repressed by tyrosine concentrations [2]. In this work we report the transcriptional profiling of the TDC cluster deletion mutant (E. faecalis V583 ΔTDC) [2] compared to the wild-type strain, both grown in M17 medium supplemented with tyrosine. The transcriptional profile data of TDC cluster-regulated genes were deposited in the Gene Expression Omnibus (GEO) database under accession no. GSE77864. PMID:27408815

  18. A comparison of monomicrobial versus polymicrobial Enterococcus faecalis bacteriuria in a French University Hospital.

    PubMed

    Fourcade, C; Canini, L; Lavigne, J-P; Sotto, A

    2015-08-01

    Enterococci are of considerable relevance in the hospital setting. Their most common location is the urinary tract, where they may be responsible for both colonization and infections. They are often associated with the presence of other microorganisms. The aim was to compare monomicrobial and polymicrobial Enterococcus faecalis bacteriuria. A retrospective study was performed on the demographic, clinical, and laboratory data of 299 patients who had presented with E. faecalis bacteriuria in 2012 at a University Hospital. The bacteriuria was polymicrobial in 46.1 % of cases and in 36.4 % of cases was responsible for a urinary tract infection. Infections appeared to be more prevalent in the polymicrobial than the monomicrobial group (42 % vs 32 %, p = 0.06). Half of the patients who presented with urinary tract colonization received antibiotic treatment (54/ out of 10). A multivariate analysis adjusted for age (adjusted odds ratio [AOR] = 1.02 per year, p = 0.006), gender (AOR = 2.2, p = 0.007), and clinical classification (colonization or infection, AOR = 1.6, p = 0.091), showed that diabetes mellitus (AOR = 2.0, p = 0.04), hospital length of stay exceeding 28 days (AOR = 2.0, p = 0.03), and presence of a urinary catheter (AOR = 2.4, p = 0.001) were all factors associated with polymicrobial E. faecalis bacteriuria. A reduction in the length of hospital stay and the use of urinary catheters would appear to be required to decrease the incidence of urinary tract colonization and infections by polymicrobial E. faecalis. Improper use of antibiotics to treat urinary tract colonization remains a major concern. PMID:25987245

  19. The Heterodimeric ABC Transporter EfrCD Mediates Multidrug Efflux in Enterococcus faecalis

    PubMed Central

    Hürlimann, Lea M.; Corradi, Valentina; Hohl, Michael; Bloemberg, Guido V.; Tieleman, D. Peter

    2016-01-01

    Nosocomial infections with Enterococcus faecalis are an emerging health problem. However, drug efflux pumps contributing to intrinsic drug resistance are poorly studied in this Gram-positive pathogen. In this study, we functionally investigated seven heterodimeric ABC transporters of E. faecalis that are annotated as drug efflux pumps. Deletion of ef0789-ef0790 on the chromosome of E. faecalis resulted in increased susceptibility to daunorubicin, doxorubicin, ethidium, and Hoechst 33342, and the corresponding transporter was named EfrCD. Unexpectedly, the previously described heterodimeric multidrug ABC transporter EfrAB contributes marginally to drug efflux in the endogenous context of E. faecalis. In contrast, heterologous expression in Lactococcus lactis revealed that EfrAB, EfrCD, and the product of ef2226-ef2227 (EfrEF) mediate the efflux of fluorescent substrates and confer resistance to multiple dyes and drugs, including fluoroquinolones. Four of seven transporters failed to exhibit drug efflux activity for the set of drugs and dyes tested, even upon overexpression in L. lactis. Since all seven transporters were purified as heterodimers after overexpression in L. lactis, a lack of drug efflux activity is not attributed to poor expression or protein aggregation. Reconstitution of the purified multidrug transporters EfrAB, EfrCD, and EfrEF in proteoliposomes revealed functional coupling between ATP hydrolysis and drug binding. Our analysis creates an experimental basis for the accurate prediction of drug efflux transporters and indicates that many annotated multidrug efflux pumps might be incapable of drug transport and thus might fulfill other physiological functions in the cell. PMID:27381387

  20. Antibacterial Activity of Diode Laser and Sodium Hypochlorite in Enterococcus Faecalis-Contaminated Root Canals

    PubMed Central

    Sohrabi, Khosrow; Sooratgar, Aidin; Zolfagharnasab, Kaveh; Kharazifard, Mohammad Javad; Afkhami, Farzaneh

    2016-01-01

    Introduction: The aim of the present in vitro study was to evaluate the disinfection ability of 980-nm diode laser in comparison with sodium hypochlorite (NaOCl) as a common root canal irrigant in canals infected with Enterococcus faecalis (E. faecalis). Methods and Materials: The root canals of 18 extracted single-rooted premolars were prepared by rotary system. After decoronation, the roots were autoclaved. One specimen was chosen for the negative control, and the remaining teeth were incubated with E. faecalis suspension for two weeks. Subsequently, one specimen was selected as the positive control and the remaining samples were divided into two groups (n=8). The samples of the first group were irrigated with 5.25% NaOCl and the second group were treated with a 980-nm diode laser. Microbial samples were taken from the root canals and bacterial cultivation was carried out. The average value and the standard deviation of colony-forming units (CFU) of each specimen were measured using descriptive statistics. The student’s t-test was used to compare the reduction in CFU in each group. The equality of variance of CFU was measured by the Levene’s test. Results: NaOCl resulted in 99.87% removal of the bacteria and showed significantly more antibacterial effect compared to the 980-nm diode laser which led to 96.56% bacterial reduction (P<0.05). Conclusion: Although 5.25% NaOCl seems to reduce E. faecalis more effectively, the diode laser also reduced the bacterial count. Therefore a 980-nm diode laser could be considered as a complementary disinfection method in root canal treatment. PMID:26843870

  1. Global Regulation of Gene Expression by the MafR Protein of Enterococcus faecalis.

    PubMed

    Ruiz-Cruz, Sofía; Espinosa, Manuel; Goldmann, Oliver; Bravo, Alicia

    2015-01-01

    Enterococcus faecalis is a natural inhabitant of the human gastrointestinal tract. However, as an opportunistic pathogen, it is able to colonize other host niches and cause life-threatening infections. Its adaptation to new environments involves global changes in gene expression. The EF3013 gene (here named mafR) of E. faecalis strain V583 encodes a protein (MafR, 482 residues) that has sequence similarity to global response regulators of the Mga/AtxA family. The enterococcal OG1RF genome also encodes the MafR protein (gene OG1RF_12293). In this work, we have identified the promoter of the mafR gene using several in vivo approaches. Moreover, we show that MafR influences positively the transcription of many genes on a genome-wide scale. The most significant target genes encode components of PTS-type membrane transporters, components of ABC-type membrane transporters, and proteins involved in the metabolism of carbon sources. Some of these genes were previously reported to be up-regulated during the growth of E. faecalis in blood and/or in human urine. Furthermore, we show that a mafR deletion mutant strain induces a significant lower degree of inflammation in the peritoneal cavity of mice, suggesting that enterococcal cells deficient in MafR are less virulent. Our work indicates that MafR is a global transcriptional regulator. It might facilitate the adaptation of E. faecalis to particular host niches and, therefore, contribute to its potential virulence. PMID:26793169

  2. Global Regulation of Gene Expression by the MafR Protein of Enterococcus faecalis

    PubMed Central

    Ruiz-Cruz, Sofía; Espinosa, Manuel; Goldmann, Oliver; Bravo, Alicia

    2016-01-01

    Enterococcus faecalis is a natural inhabitant of the human gastrointestinal tract. However, as an opportunistic pathogen, it is able to colonize other host niches and cause life-threatening infections. Its adaptation to new environments involves global changes in gene expression. The EF3013 gene (here named mafR) of E. faecalis strain V583 encodes a protein (MafR, 482 residues) that has sequence similarity to global response regulators of the Mga/AtxA family. The enterococcal OG1RF genome also encodes the MafR protein (gene OG1RF_12293). In this work, we have identified the promoter of the mafR gene using several in vivo approaches. Moreover, we show that MafR influences positively the transcription of many genes on a genome-wide scale. The most significant target genes encode components of PTS-type membrane transporters, components of ABC-type membrane transporters, and proteins involved in the metabolism of carbon sources. Some of these genes were previously reported to be up-regulated during the growth of E. faecalis in blood and/or in human urine. Furthermore, we show that a mafR deletion mutant strain induces a significant lower degree of inflammation in the peritoneal cavity of mice, suggesting that enterococcal cells deficient in MafR are less virulent. Our work indicates that MafR is a global transcriptional regulator. It might facilitate the adaptation of E. faecalis to particular host niches and, therefore, contribute to its potential virulence. PMID:26793169

  3. Enterococcus faecalis Gene Transfer under Natural Conditions in Municipal Sewage Water Treatment Plants†

    PubMed Central

    Marcinek, Herbert; Wirth, Reinhard; Muscholl-Silberhorn, Albrecht; Gauer, Matthias

    1998-01-01

    The ability of Enterococcus faecalis to transfer various genetic elements under natural conditions was tested in two municipal sewage water treatment plants. Experiments in activated sludge basins of the plants were performed in a microcosm which allowed us to work under sterile conditions; experiments in anoxic sludge digestors were performed in dialysis bags. We used the following naturally occurring genetic elements: pAD1 and pIP1017 (two so-called sex pheromone plasmids with restricted host ranges, which are transferred at high rates under laboratory conditions); pIP501 (a resistance plasmid possessing a broad host range for gram-positive bacteria, which is transferred at low rates under laboratory conditions); and Tn916 (a conjugative transposon which is transferred under laboratory conditions at low rates to gram-positive bacteria and at very low rates to gram-negative bacteria). The transfer rate between different strains of E. faecalis under natural conditions was, compared to that under laboratory conditions, at least 105-fold lower for the sex pheromone plasmids, at least 100-fold lower for pIP501, and at least 10-fold lower for Tn916. In no case was transfer from E. faecalis to another bacterial species detected. By determining the dependence of transfer rates for pIP1017 on bacterial concentration and extrapolating to actual concentrations in the sewage water treatment plant, we calculated that the maximum number of transfer events for the sex pheromone plasmids between different strains of E. faecalis in the municipal sewage water treatment plant of the city of Regensburg ranged from 105 to 108 events per 4 h, indicating that gene transfer should take place under natural conditions. PMID:9464401

  4. The Heterodimeric ABC Transporter EfrCD Mediates Multidrug Efflux in Enterococcus faecalis.

    PubMed

    Hürlimann, Lea M; Corradi, Valentina; Hohl, Michael; Bloemberg, Guido V; Tieleman, D Peter; Seeger, Markus A

    2016-09-01

    Nosocomial infections with Enterococcus faecalis are an emerging health problem. However, drug efflux pumps contributing to intrinsic drug resistance are poorly studied in this Gram-positive pathogen. In this study, we functionally investigated seven heterodimeric ABC transporters of E. faecalis that are annotated as drug efflux pumps. Deletion of ef0789-ef0790 on the chromosome of E. faecalis resulted in increased susceptibility to daunorubicin, doxorubicin, ethidium, and Hoechst 33342, and the corresponding transporter was named EfrCD. Unexpectedly, the previously described heterodimeric multidrug ABC transporter EfrAB contributes marginally to drug efflux in the endogenous context of E. faecalis In contrast, heterologous expression in Lactococcus lactis revealed that EfrAB, EfrCD, and the product of ef2226-ef2227 (EfrEF) mediate the efflux of fluorescent substrates and confer resistance to multiple dyes and drugs, including fluoroquinolones. Four of seven transporters failed to exhibit drug efflux activity for the set of drugs and dyes tested, even upon overexpression in L. lactis Since all seven transporters were purified as heterodimers after overexpression in L. lactis, a lack of drug efflux activity is not attributed to poor expression or protein aggregation. Reconstitution of the purified multidrug transporters EfrAB, EfrCD, and EfrEF in proteoliposomes revealed functional coupling between ATP hydrolysis and drug binding. Our analysis creates an experimental basis for the accurate prediction of drug efflux transporters and indicates that many annotated multidrug efflux pumps might be incapable of drug transport and thus might fulfill other physiological functions in the cell. PMID:27381387

  5. The effect of different root canal medicaments on the elimination of Enterococcus faecalis ex vivo

    PubMed Central

    Dammaschke, Till; Jung, Nina; Harks, Inga; Schafer, Edgar

    2013-01-01

    Objective: The aim of this study was to evaluate the antimicrobial effect of chlorhexidine gel (CHX-G) 2%, chlorhexidine powder (CHX-P) 1%, povidone-iodine (PVP-I), polyhexanide and camphorated-and-mentholated chlorophenol (ChKM) ex vivo. Materials and Methods: For every medicament group 10 root segments (15 mm long) of extracted human teeth were prepared to ISO-size 45 and sterilized (n = 50). The root segments were then inoculated with Enterococcus faecalis and aerobically incubated at 37°C. After 1 week, ten root canals were filled with one of the medicaments, respectively and aerobically incubated at 37°C for another week. Ten teeth served as positive controls and were filled with sterile saline solution. After 7 days, the medicaments were inactivated and all root canals were instrumented to ISO-size 50. The obtained dentin samples were dispersed in Ringer solution followed by the preparation of serial dilutions. 10 μl per sample were applied to an agar plate and incubated at 37°C for 48 h. The colony forming units were counted and the reduction factors (RFs) were calculated and statistically analyzed. Results: Compared with the positive controls all medicaments exhibited an antibacterial effect against E. faecalis. The RFs for CHX-G, CHX-P and ChKM were significantly higher compared to PVP-I and polyhexanide (P < 0.05). In contrast to PVP-I and polyhexanide, CHX-G, CHX-P and ChKM were able to eliminate E. faecalis from all dentin samples. Conclusions: Within the limitations of this ex vivo investigation, 2% CHX-G and CHX-P were as effective as ChKM against E. faecalis. Thus, when choosing a root canal medicament the better biocompatibility of CHX compared with ChKM should be taken in consideration. PMID:24932119

  6. Draft Genome Sequence of the Bacteriocinogenic Strain Enterococcus faecalis DBH18, Isolated from Mallard Ducks (Anas platyrhynchos)

    PubMed Central

    Arbulu, Sara; Jimenez, Juan J.; Borrero, Juan; Sánchez, Jorge; Frantzen, Cyril; Herranz, Carmen; Nes, Ingolf F.; Cintas, Luis M.; Diep, Dzung B.

    2016-01-01

    Here, we report the draft genome sequence of Enterococcus faecalis DBH18, a bacteriocinogenic lactic acid bacterium (LAB) isolated from mallard ducks (Anas platyrhynchos). The assembly contains 2,836,724 bp, with a G+C content of 37.6%. The genome is predicted to contain 2,654 coding DNA sequences (CDSs) and 50 RNAs. PMID:27417838

  7. Draft Genome Sequence of the Bacteriocinogenic Strain Enterococcus faecalis DBH18, Isolated from Mallard Ducks (Anas platyrhynchos).

    PubMed

    Arbulu, Sara; Jimenez, Juan J; Borrero, Juan; Sánchez, Jorge; Frantzen, Cyril; Herranz, Carmen; Nes, Ingolf F; Cintas, Luis M; Diep, Dzung B; Hernández, Pablo E

    2016-01-01

    Here, we report the draft genome sequence of Enterococcus faecalis DBH18, a bacteriocinogenic lactic acid bacterium (LAB) isolated from mallard ducks (Anas platyrhynchos). The assembly contains 2,836,724 bp, with a G+C content of 37.6%. The genome is predicted to contain 2,654 coding DNA sequences (CDSs) and 50 RNAs. PMID:27417838

  8. Expression of Alzheimer-Type Neurofibrillary Epitopes in Primary Rat Cortical Neurons Following Infection with Enterococcus faecalis

    PubMed Central

    Underly, Robert; Song, Mee-Sook; Dunbar, Gary L.; Weaver, Charles L.

    2016-01-01

    The neurofibrillary tau pathology and amyloid deposits seen in Alzheimer’s disease (AD) also have been seen in bacteria-infected brains. However, few studies have examined the role of these bacteria in the generation of tau pathology. One suggested link between infection and AD is edentulism, the complete loss of teeth. Edentulism can result from chronic periodontal disease due to infection by Enterococcus faecalis. The current study assessed the ability to generate early Alzheimer-like neurofibrillary epitopes in primary rat cortical neurons through bacterial infection by E. faecalis. Seven-day old cultured neurons were infected with E. faecalis for 24 and 48 h. An upward molecular weight shift in tau by Western blotting (WB) and increased appearance of tau reactivity in cell bodies and degenerating neurites was found in the 48 h infection group for the antibody CP13 (phospho-Serine 202). A substantial increase in reactivity of Alz-50 was seen at 24 and 48 h after infection. Furthermore, extensive microtubule-associated protein 2 (MAP2) reactivity also was seen at 24 and 48 h post-infection. Our preliminary findings suggest a potential link between E. faecalis infection and intracellular changes that may help facilitate early AD-like neurofibrillary pathology. HighlightsEnterococcus faecalis used in the generation of AD neurofibrillary epitopes in rat.Infection increases Alz-50, phospho-Serine 202 tau, and MAP2 expression.Infection by Enterococcus may play a role in early Alzheimer neurofibrillary changes. PMID:26834627

  9. The Capability of Tyramine Production and Correlation between Phenotypic and Genetic Characteristics of Enterococcus faecium and Enterococcus faecalis Strains

    PubMed Central

    Bargossi, Eleonora; Gardini, Fausto; Gatto, Veronica; Montanari, Chiara; Torriani, Sandra; Tabanelli, Giulia

    2015-01-01

    The aim of this study was to investigate the diversity of tyramine production capability of four Enterococcus strains in buffered systems in relation to their genetic characteristics and environmental conditions. Cells of the strains Enterococcus faecalis EF37 and ATCC 29212, and E. faecium FC12 and FC643 were re-suspended in phosphate/citrate buffers with different pH, NaCl concentration and incubation temperature. At intervals, cell viability and tyramine production were assessed by plate counting and HPLC analysis, respectively. The activity of a purified tyrosine decarboxylase (TDC) was determined under the same conditions, as a reference. Reduced loss in cell viability was observed in all the tested conditions, except for pH 4 after 24 h. The TDC activity was greatly heterogeneous within the enterococci: EF37 and FC12 produced the higher tyramine concentrations, ATCC 29212 showed a reduced decarboxylase activity, while EF643 did not accumulate detectable amounts of tyramine in all the conditions assayed. Among the considerate variables, temperature was the most influencing factor on tyramine accumulation for enterococcal cells. To further correlate the phenotypic and genetic characteristics of the enterococci, the TDC operon region carrying the genes tyrosine decarboxylase (tyrDC), tyrosine/tyramine permease (tyrP), and Na+/H+ antiporter (nhaC-2) was amplified and sequenced. The genetic organization and nucleotide sequence of this operon region were highly conserved in the enterococcal strains of the same species. The heterogeneity in tyramine production found between the two E. faecalis strains could be ascribed to different regulation mechanisms not yet elucidated. On the contrary, a codon stop was identified in the translated tyrDC sequence of E. faecium FC643, supporting its inability to accumulate tyramine in the tested conditions. In addition, the presence of an additional putative tyrosine decarboxylase with different substrate specificity and genetic

  10. Virulence Genes among Enterococcus faecalis and Enterococcus faecium Isolated from Coastal Beaches and Human and Nonhuman Sources in Southern California and Puerto Rico

    PubMed Central

    Talavera, Ginamary Negrón; Hernández, Luis A. Ríos; Ambrose, Richard F.; Jay, Jennifer A.

    2016-01-01

    Most Enterococcus faecalis and E. faecium are harmless to humans; however, strains harboring virulence genes, including esp, gelE, cylA, asa1, and hyl, have been associated with human infections. E. faecalis and E. faecium are present in beach waters worldwide, yet little is known about their virulence potential. Here, multiplex PCR was used to compare the distribution of virulence genes among E. faecalis and E. faecium isolated from beaches in Southern California and Puerto Rico to isolates from potential sources including humans, animals, birds, and plants. All five virulence genes were found in E. faecalis and E. faecium from beach water, mostly among E. faecalis. gelE was the most common among isolates from all source types. There was a lower incidence of asa1, esp, cylA, and hyl genes among isolates from beach water, sewage, septage, urban runoff, sea wrack, and eelgrass as compared to human isolates, indicating that virulent strains of E. faecalis and E. faecium may not be widely disseminated at beaches. A higher frequency of asa1 and esp among E. faecalis from dogs and of asa1 among birds (mostly seagull) suggests that further studies on the distribution and virulence potential of strains carrying these genes may be warranted. PMID:27144029

  11. Virulence Genes among Enterococcus faecalis and Enterococcus faecium Isolated from Coastal Beaches and Human and Nonhuman Sources in Southern California and Puerto Rico.

    PubMed

    Ferguson, Donna M; Talavera, Ginamary Negrón; Hernández, Luis A Ríos; Weisberg, Stephen B; Ambrose, Richard F; Jay, Jennifer A

    2016-01-01

    Most Enterococcus faecalis and E. faecium are harmless to humans; however, strains harboring virulence genes, including esp, gelE, cylA, asa1, and hyl, have been associated with human infections. E. faecalis and E. faecium are present in beach waters worldwide, yet little is known about their virulence potential. Here, multiplex PCR was used to compare the distribution of virulence genes among E. faecalis and E. faecium isolated from beaches in Southern California and Puerto Rico to isolates from potential sources including humans, animals, birds, and plants. All five virulence genes were found in E. faecalis and E. faecium from beach water, mostly among E. faecalis. gelE was the most common among isolates from all source types. There was a lower incidence of asa1, esp, cylA, and hyl genes among isolates from beach water, sewage, septage, urban runoff, sea wrack, and eelgrass as compared to human isolates, indicating that virulent strains of E. faecalis and E. faecium may not be widely disseminated at beaches. A higher frequency of asa1 and esp among E. faecalis from dogs and of asa1 among birds (mostly seagull) suggests that further studies on the distribution and virulence potential of strains carrying these genes may be warranted. PMID:27144029

  12. β-Lactam combinations with daptomycin provide synergy against vancomycin-resistant Enterococcus faecalis and Enterococcus faecium

    PubMed Central

    Smith, Jordan R.; Barber, Katie E.; Raut, Animesh; Aboutaleb, Mostafa; Sakoulas, George; Rybak, Michael J.

    2015-01-01

    Objectives Enterococcus faecalis (Efc) and Enterococcus faecium (Efm) are frequently resistant to vancomycin and β-lactams (BLs). In vitro data suggest synergy between several BLs and glycopeptides or lipopeptides against resistant pathogens. Our objective was to conduct combination MIC and time–kill experiments to evaluate BL synergy with daptomycin against enterococci. Methods Fifteen Efc and 20 Efm strains were evaluated for daptomycin enhancement via combination MICs. Daptomycin MICs were obtained by microdilution in the absence and presence of ceftaroline, ertapenem, cefepime, ceftriaxone, cefotaxime, cefazolin and ampicillin. Two Efc strains (R6981 and R7808) and one isogenic daptomycin-susceptible/daptomycin-non-susceptible Efm pair (8019/5938) were evaluated in time–kill experiments. Daptomycin at 0.5 × MIC was used in combination with BL at biological free concentration. Strain 5938 was evaluated for enhancement of daptomycin binding in fluorescently labelled daptomycin (BoDipy) experiments. Results Ceftaroline reduced daptomycin MIC values the most against all strains. In time–kill experiments, ceftaroline, ertapenem, cefepime, ceftriaxone and ampicillin demonstrated synergy with daptomycin against all strains, cefazolin demonstrated none and cefotaxime demonstrated synergy against only R7808. Bacterial reduction at 24 h was greater for daptomycin + ceftaroline, ertapenem, cefepime, ceftriaxone or ampicillin for all strains compared with any single agent or daptomycin + cefazolin or cefotaxime (P < 0.001). In BoDipy daptomycin experiments, ceftaroline enhanced daptomycin binding most compared with all other agents (P < 0.001). Conclusions The data support the potential use of daptomycin/BL combination therapy in infections caused by VRE. Combination regimens, other than those involving cefazolin and cefotaxime, provide better kill compared with daptomycin alone. Further clinical research involving daptomycin combinations is

  13. Interference in Pheromone-Responsive Conjugation of a High-Level Bacitracin Resistant Enterococcus faecalis Plasmid of Poultry Origin

    PubMed Central

    Tremblay, Cindy-Love; Archambault, Marie

    2013-01-01

    The current study reports on contact interference of a high-level bacitracin- resistant pheromone-responsive plasmid of Enterococcus faecalis strain 543 of poultry origin during conjugative transfer of bcr antimicrobial resistance genes using a polyclonal antiserum aggregation substance44–560 (AS). After induction with pheromones produced by the recipient strain E. faecalis JH2-2, clumping of the donor E. faecalis strain 543 was observed as well as high transfer frequencies of bcr in short time broth mating. Filter mating assays from donor strain E. faecalis 543 to the recipient strain E. faecalis JH2-2 revealed conjugative transfer of asa1 (AS), bcrRAB and traB (negative regulator pheromone response) genes. The presence of these genes in transconjugants was confirmed by antimicrobial susceptibility testing, PCR, Southern hybridization and sequencing. A significant reduction in formation of aggregates was observed when the polyclonal anti-AS44–560 was added in the pheromone-responsive conjugation experiments as compared to the induced state. Moreover, interference of anti-AS44–560 antibodies in pheromone-responsive conjugation was demonstrated by a reduction in horizontal transfer of asa1 and bcr genes between E. faecalis strain 543 and E. faecalis JH2-2. Reducing the pheromone-responsive conjugation of E. faecalis is of interest because of its clinical importance in the horizontal transfer of antimicrobial resistance. PMID:24030654

  14. Effects of Enterococcus faecalis CECT 7121 on Cryptosporidium parvum infection in mice.

    PubMed

    Del Coco, Valeria F; Sparo, Mónica D; Sidoti, Alicia; Santín, Mónica; Basualdo, Juan Angel; Córdoba, María Alejandra

    2016-08-01

    Cryptosporidium is an opportunistic protozoan parasite of humans and animals worldwide and causes diarrheal disease that is typically self-limiting in immunocompetent hosts but often life threatening to immunocompromised individuals. However, there is a lack of completely efficient therapy available. Probiotics have attracted the attention as potential antiparasite compounds against protozoa involved in intestinal infections. This study investigated the effects of administration of probiotic Enterococcus faecalis CECT 7121 on Cryptosporidium parvum infection in immunosuppressed mice. Effects on C. parvum infection at the intestinal mucosa were studied and scored at each portion of the gut. It was demonstrated that Ef CECT 7121 interfered with C. parvum infection when both probiotic and parasite were present in the same intestinal location suggesting that Ef CECT 7121 supplementation can alleviate the negative effects of C. parvum infection. PMID:27193238

  15. Inductive effects of environmental concentration of atrazine on Escherichia coli and Enterococcus faecalis.

    PubMed

    Koutsotoli, A D; Dimou, D S; Alamanos, Y P; Maipa, V E

    2005-01-01

    Atrazine solutions (0.1, 1, 10 and 100 microg/L) inoculated with Escherichia coli and Enterococcus faecalis under natural conditions significantly increased (p < or = 0.05) the population levels of both test bacteria; it indicates the ability of bacterial cells to degrade atrazine and to use the original compound or its degradation products as nutrient(s). In some cases, alterations in the morphology of the colonies were also observed on selective solid media. Biochemical differentiation was also found and, on the other hand, a loss of culturability was recorded; this suggests that bacteria have entered in a viable but nonculturable state. A re-appearance of the colonies occurred after inoculation on tryptone-soy agar with atrazine. PMID:16408845

  16. Treatment of enterococcus faecalis bacteria by a helium atmospheric cold plasma brush with oxygen addition

    SciTech Connect

    Chen Wei; Huang Jun; Wang Xingquan; Lv Guohua; Zhang Guoping; Du Ning; Liu Xiaodi; Guo Lihong; Yang Size

    2012-07-01

    An atmospheric cold plasma brush suitable for large area and low-temperature plasma-based sterilization is designed. Results demonstrate that the He/O{sub 2} plasma more effectively kills Enterococcus faecalis than the pure He plasma. In addition, the sterilization efficiency values of the He/O{sub 2} plasma depend on the oxygen fraction in Helium gas. The atmospheric cold plasma brush using a proper ratio of He/O{sub 2} (2.5%) reaches the optimum sterilization efficiency. After plasma treatment, the cell structure and morphology changes can be observed by the scanning electron microscopy. Optical emission measurements indicate that reactive species such as O and OH play a significant role in the sterilization process.

  17. Treatment of enterococcus faecalis bacteria by a helium atmospheric cold plasma brush with oxygen addition

    NASA Astrophysics Data System (ADS)

    Chen, Wei; Huang, Jun; Du, Ning; Liu, Xiao-Di; Wang, Xing-Quan; Lv, Guo-Hua; Zhang, Guo-Ping; Guo, Li-Hong; Yang, Si-Ze

    2012-07-01

    An atmospheric cold plasma brush suitable for large area and low-temperature plasma-based sterilization is designed. Results demonstrate that the He/O2 plasma more effectively kills Enterococcus faecalis than the pure He plasma. In addition, the sterilization efficiency values of the He/O2 plasma depend on the oxygen fraction in Helium gas. The atmospheric cold plasma brush using a proper ratio of He/O2 (2.5%) reaches the optimum sterilization efficiency. After plasma treatment, the cell structure and morphology changes can be observed by the scanning electron microscopy. Optical emission measurements indicate that reactive species such as O and OH play a significant role in the sterilization process.

  18. Antimicrobial activity of herbal medicines (tulsi extract, neem extract) and chlorhexidine against Enterococcus faecalis in Endodontics: An in vitro study

    PubMed Central

    Chandrappa, Pradeep Muttagadur; Dupper, Akash; Tripathi, Pragya; Arroju, Ramakrishna; Sharma, Preeti; Sulochana, Konthoujam

    2015-01-01

    Background: Successful endodontic treatment depends on effective disinfection and complete sealing of root canal. Various medicaments are advised for disinfecting root canal, such as herbal and non-herbal medicaments. This study was done to assess the antimicrobial activity of herbal medicines (neem extract, tulsi extract) and chlorhexidine against Enterococcus faecalis in Endodontics. Materials and Methods: Agar diffusion method was used to evaluate the antimicrobial action of different medicines. Sixty samples were segregated into four groups with 15 samples in each: Group I: chlorhexidine 2%, Group II: neem extract, Group III: tulsi extract, and Group IV: distilled water. The inhibition zones against E. faecalis were recorded and statistically assessed using one-way analysis of variance (ANOVA) test (P < 0.05). Results: Significant antibacterial effect against E. faecalis was observed with chlorhexidine followed by neem extract and tulsi extract. Conclusion: Herbal medicines seemed to be effective against E. faecalis compared to 2% chlorhexidine gluconate. PMID:26942123

  19. Identification of an N-terminal formylated, two-peptide bacteriocin from Enterococcus faecalis 710C.

    PubMed

    Liu, Xiaoji; Vederas, John C; Whittal, Randy M; Zheng, Jing; Stiles, Michael E; Carlson, Denise; Franz, Charles M A P; McMullen, Lynn M; van Belkum, Marco J

    2011-05-25

    Enterococcus faecalis 710C, isolated from beef product, has a broad antimicrobial activity spectrum against foodborne pathogens. Two bacteriocins, enterocin 7A (Ent7A) and enterocin 7B (Ent7B), were purified from the culture supernatant of E. faecalis 710C and characterized using matrix-assisted laser desorption ionization-time-of-flight mass spectrometry and electrospray infusion tandem mass spectrometry analyses. These data and subsequent genetic analysis showed that Ent7A and Ent7B are produced without N-terminal leader sequences and have amino acid sequences that are identical to those of enterocins MR10A and MR10B, respectively. However, the observed masses for Ent7A and Ent7B are 5200.80 and 5206.65 Da (monoisotopic mass), respectively, which are higher than the theoretical molecular masses of MR10A and MR10B, respectively. This study provides evidence that both Ent7A and Ent7B are formylated on the N-terminal methionine residue. Purified Ent7A and Ent7B are active against spoilage microorganisms and foodborne pathogens, including Clostridium sporogenes , Listeria monocytogenes , and Staphylococcus aureus as well as Brevundimonas diminuta , which has been associated with infections among immune-suppressed cancer patients. PMID:21469734

  20. Involvement of Enterococcus faecalis Small RNAs in Stress Response and Virulence

    PubMed Central

    Michaux, Charlotte; Hartke, Axel; Martini, Cecilia; Reiss, Swantje; Albrecht, Dirk; Budin-Verneuil, Aurélie; Sanguinetti, Maurizio; Engelmann, Susanne; Hain, Torsten; Verneuil, Nicolas

    2014-01-01

    Candidate small RNAs (sRNAs) have recently been identified in Enterococcus faecalis, a Gram-positive opportunistic pathogen, and six of these candidate sRNAs with unknown functions were selected for a functional study. Deletion mutants and complemented strains were constructed, and their virulence was tested. We were unable to obtain the ef0869-0870 mutant, likely due to an essential role, and the ef0820-0821 sRNA seemed not to be involved in virulence. In contrast, the mutant lacking ef0408-0409 sRNA, homologous to the RNAII component of the toxin-antitoxin system, appeared more virulent and more able to colonize mouse organs. The three other mutants showed reduced virulence. In addition, we checked the responses of these mutant strains to several stresses encountered in the gastrointestinal tract or during the infection process. In parallel, the activities of the sRNA promoters were measured using transcriptional fusion constructions. To attempt to identify the regulons of these candidate sRNAs, proteomics profiles of the mutant strains were compared with that of the wild type. This showed that the selected sRNAs controlled the expression of proteins involved in diverse cellular processes and the stress response. The combined data highlight the roles of certain candidate sRNAs in the adaptation of E. faecalis to environmental changes and in the complex transition process from a commensal to a pathogen. PMID:24914223

  1. The Intraperitoneal Transcriptome of the Opportunistic Pathogen Enterococcus faecalis in Mice

    PubMed Central

    Muller, Cécile; Cacaci, Margherita; Sauvageot, Nicolas; Sanguinetti, Maurizio; Rattei, Thomas; Eder, Thomas; Giard, Jean-Christophe; Kalinowski, Jörn; Hain, Torsten; Hartke, Axel

    2015-01-01

    Enterococcus faecalis is a Gram-positive lactic acid intestinal opportunistic bacterium with virulence potential. For a better understanding of the adapation of this bacterium to the host conditions, we performed a transcriptome analysis of bacteria isolated from an infection site (mouse peritonitis) by RNA-sequencing. We identified a total of 211 genes with significantly higher transcript levels and 157 repressed genes. Our in vivo gene expression database reflects well the infection process since genes encoding important virulence factors like cytolysin, gelatinase or aggregation substance as well as stress response proteins, are significantly induced. Genes encoding metabolic activities are the second most abundant in vivo induced genes demonstrating that the bacteria are metabolically active and adapt to the special nutrient conditions of the host. α- and β- glucosides seem to be important substrates for E. faecalis inside the host. Compared to laboratory conditions, the flux through the upper part of glycolysis seems to be reduced and more carbon may enter the pentose phosphate pathway. This may reflect the need of the bacteria under infection conditions to produce more reducing power for biosynthesis. Another important substrate is certainly glycerol since both pathways of glycerol catabolism are strongly induced. Strongly in vivo induced genes should be important for the infection process. This assumption has been verified in a virulence test using well characterized mutants affected in glycerol metabolism. This showed indeed that mutants unable to metabolize this sugar alcohol are affected in organ colonisation in a mouse model. PMID:25978463

  2. Endophthalmitis Caused by Enterococcus faecalis: Clinical Features, Antibiotic Sensitivities, and Outcomes

    PubMed Central

    Kuriyan, Ajay E.; Sridhar, Jayanth; Flynn, Harry W.; Smiddy, William E.; Albini, Thomas A.; Berrocal, Audina M.; Forster, Richard K.; Belin, Peter J.; Miller, Darlene

    2014-01-01

    Purpose To report the clinical features, antibiotic sensitivities, and visual acuity (VA) outcomes of endophthalmitis caused by Enterococcus faecalis. Study Design Retrospective, observational case series. Methods A consecutive case series of patients with culture-positive endophthalmitis caused by E. faecalis between January 1, 2002 and December 31, 2012 at an academic referral center. Results Of 14 patients identified, clinical settings included bleb-associated (n=8), post-cataract surgery (n=4), and post-penetrating keratoplasty (n=2). All isolates were vancomycin sensitive. When comparing isolates in the current study to isolates from 1990–2001, the minimal inhibitory concentration required to inhibit 90% of isolates (MIC 90, μg/ml) increased for ciprofloxacin (4 from 1), erythromycin (256 from 4), and penicillin (8 from 4), indicating higher levels of resistance. The MIC 90 remained the same for vancomycin (2) and linezolid (2). Presenting VA ranged from hand motion to no light perception. Initial treatment strategies were vitreous tap and intravitreal antibiotic injection (n=12) and pars plana vitrectomy with intravitreal antibiotic injection (n=2). VA outcomes were ≤ 20/400 in 13 (93%) of 14 patients. Conclusions Although all isolates were sensitive to vancomycin and linezolid, higher MIC 90s for isolates in the current study, compared to isolates from 1990 to 2001, occurred with ciprofloxacin, erythromycin, and penicillin. Despite prompt treatment, most patients had poor outcomes. PMID:25089354

  3. The Presence and Origin of Enterococcus faecalis in Cabo Rojo, Puerto Rico

    NASA Astrophysics Data System (ADS)

    Zachman, A. J.; Sturm, P.; Viqueira Ríos, R.

    2015-12-01

    Currently, a watershed management plan is being developed for Cabo Rojo region in Southwest Puerto Rico. This project fills in major gaps for water quality data on the Rio Viejo, a tributary on the Guanajibio River. The Rio Viejo flows through the town of Cabo Rojo, a town of 51,245 people. The project has identified 5 sites along the river to track bacterial loads. In the tropics, Enterococcus faecalis is an important indicator for fecal contamination in surface waters as it does not reproduce as quickly soils as E. coli. A combination of EPA 1600 and 9230B from Standard Methods for the Examination of Water and Wastewater for identification of E. faecalis were utilized. The assay is a four step procedure that identifies the four criteria of bacteria in the group D Streptococcus system. The criteria require that the bacteria are Gram-positive cocci and Esculin-positive. There also must be growth in Brain Heart Infusion Broth at 35C and 45C as well as growth in Brain Heart Infusion broth + 6.5% NaCl. Further research will be conducted at North Carolina State University to ascertain the vertebrate species that is the source of the contamination through the use of qPCR.

  4. Overexpression, crystallization and preliminary X-ray crystallographic analysis of phosphopantetheine adenylyltransferase from Enterococcus faecalis

    SciTech Connect

    Kang, Ji Yong; Lee, Hyung Ho; Yoon, Hye Jin; Kim, Hyoun Sook; Suh, Se Won

    2006-11-01

    Phosphopantetheine adenylyltransferase from En. faecalis was crystallized and X-ray diffraction data were collected to 2.70 Å resolution. Phosphopantetheine adenylyltransferase, an essential enzyme in the coenzyme A biosynthetic pathway, catalyzes the reversible transfer of an adenylyl group from ATP to 4′-phosphopantetheine, yielding 3′-dephospho-CoA and pyrophosphate. Enterococcus faecalis PPAT has been overexpressed in Escherichia coli as a fusion with a C-terminal purification tag and crystallized at 297 K using a reservoir solution consisting of 0.1 M sodium HEPES pH 7.5, 0.8 M sodium dihydrogen phosphate and 0.8 M potassium dihydrogen phosphate. X-ray diffraction data were collected to 2.70 Å at 100 K. The crystals belong to the primitive tetragonal space group P4{sub 1} (or P4{sub 3}), with unit-cell parameters a = b = 160.81, c = 225.68 Å. Four copies of the hexameric molecule are likely to be present in the asymmetric unit, giving a crystal volume per protein weight (V{sub M}) of 3.08 Å{sup 3} Da{sup −1} and a solvent content of 60.1%.

  5. Bactericial effect of a non-thermal plasma needle against Enterococcus faecalis biofilms

    NASA Astrophysics Data System (ADS)

    Jiang, Chunqi; Schaudinn, C.; Jaramillo, D. E.; Sedghizadeh, P. P.; Webster, P.; Costerton, J. W.

    2011-10-01

    Up to 3 cm long submillimeter-in-scale plasma needle was generated in ambient atmosphere for root canal disinfection. Powered with 1-2 kHz, multi-kilovolt nanosecond electric pulses, this He/(1%)O2 plasma jet consists of ionization fronts propagating at speeds of the order of 107 cm/s. Plasma treatment of Enterococcus faecalis biofilms on hydroxyapatite (HA) discs for 5 min resulted in severe damage of the bacterial cells and sterilized HA surfaces of more than 3 mm in diameter, observed by the scanning electron microscopy. With a curing dielectric microtube placed 1 cm or less below the nozzle, the plasma jet entered even at a sharp angle and followed the curvature of the tube, and reached the bottom of the tube. The bactericidal effect of the plasma needle against E. faecalis biofilm grown on the inner surfaces of the tube was demonstrated. However, the bactericidal effect weakens or diminishes for the bacteria grown deeper in the tube, indicating improvement of the plasma treatment scheme is needed. Mechanisms of the plasma bactericidal effects are discussed. Supported by the National Institute of Dental and Craniofacial Research and the Air Force Office of Scientific Research.

  6. Structural Studies on Cytosolic Domain of Magnesium Transporter MgtE from Enterococcus faecalis

    SciTech Connect

    Ragumani, S.; Sauder, J; Burley, S; Swaminathan, S

    2009-01-01

    Magnesium (Mg{sup 2+}) is an essential element for growth and maintenance of living cells. It acts as a cofactor for many enzymes and is also essential for stability of the plasma membrane. There are two distinct classes of magnesium transporters identified in bacteria that convey Mg{sup 2+} from periplasm to cytoplasm [ATPase-dependent (MgtA and MgtB) and constitutively active (CorA and MgtE)]. Previously published work on Mg{sup 2+} transporters yielded structures of full length MgtE from Thermus thermophilus, determined at 3.5 {angstrom} resolution, and its cytoplasmic domain with and without bond Mg{sup 2+} determined at 2.3 and 3.9 {angstrom} resolution, respectively. Here, they report the crystal structure of the Mg{sup 2+} bound form of the cytosolic portion of MgtE (residues 6-262) from Enterococcus faecalis at 2.2 {angstrom} resolution. The present structure and magnesium bound cytosolic domain structure from T. thermophilus (PDB ID: 2YVY) are structurally similar. Three magnesium binding sites are common to both MgtE full length and the present structure. Their work revealed an additional Mg{sup 2+} binding site in the E. faecalis structure. In this report, they discuss the functional significance of Mg{sup 2+} binding sites in the cytosolic domains of MgtE transporters.

  7. [Emergence of linezolid-resistant Enterococcus faecalis strains from two inpatients in a pediatric ward].

    PubMed

    Nihonyanagi, Shin; Adachi, Yuzuru; Onuki, Tomoyo; Nakazaki, Nobuhiko; Hirata, Yasuyosi; Fujiki, Kuniko; Takayama, Yoko; Kanoh, Yuhsaku; Bandoh, Yuki; Dantsuji, Yurika; Hanaki, Hideaki; Sunakawa, Keisuke

    2012-09-01

    We report herein on the isolation of three linezolid-resistant Enterococcus faecalis strains in 2011 from two pediatric inpatients at Kitasato University Hospital, Japan. Three linezolid resistant strains were isolated from two patients who shared the same room of a pediatric inpatient ward. Two linezolid resistant strains were isolated from patient A who had been treated with a total of 17,600mg of linezolid during 60 days of hospitalization (strains 1 and 2). The linezolid resistant E. faecalis persisted through the time that the patient had been discharged from the hospital. Another linezolid resistant strain was isolated from patient B who had no history of linezolid administration. The resistant strain in patient B phased out spontaneously. The minimum inhibitory concentration of linezolid in these strains ranged from 8.0 to 16.0 microg/mL. PCR amplification of the chromosomal gene encoding domain V of the 23S rRNA and subsequent nucleotide sequencing revealed that all the strains had at least one G2576T mutation. The pulse-field-gel electrophoretograms of the DNA treated with the SmaI restriction enzyme showed an identical profile suggesting that they were derived from a single resistant strain. These results suggested that the resistant strain occurred in patient A and was transmitted to patient B within the inpatient ward. PMID:23198574

  8. Effect of high-intensity focused ultrasound on Enterococcus faecalis planktonic suspensions and biofilms.

    PubMed

    Iqbal, Kulsum; Ohl, Siew-Wan; Khoo, Boo-Cheong; Neo, Jennifer; Fawzy, Amr S

    2013-05-01

    In this study, the effect of high-intensity focused ultrasound (HIFU) on Enterococcus faecalis on both planktonic suspensions and biofilms was investigated. E. faecalis persist in secondary dental infections as biofilms. Glass-bottom Petri dishes with biofilms were centered at the focal point of the HIFU wave generated by a 250-kHz transducer. Specimens were subjected to HIFU exposure at different periods of 30, 60 and 120 s. The viable bacteria, removal effect and bacterial viability of biofilms attached to the Petri dish surface were studied by colony-forming units (CFUs), scanning electron microscopy and confocal microscopy, respectively. The removal and bactericidal effects of HIFU are dependent on the exposure time. A significant reduction in biofilm thickness and CFU was found with the increase in HIFU exposure. The removal or bactericidal effect of HIFU was more significant starting from 60 s of exposure. This study highlighted the potential application of HIFU as a novel method for root canal disinfection. PMID:23453374

  9. Enterococcus faecalis Clones in Poultry and in Humans with Urinary Tract Infections, Vietnam

    PubMed Central

    Poulsen, Louise Ladefoged; Bisgaard, Magne; Son, Nguyen Thai; Trung, Nguyen Vu; An, Hoang Manh

    2012-01-01

    Enterococcus spp. as pathogens have increased, but the sources of infection often remain unclear. To investigate whether poultry might be a reservoir for E. faecalis–associated urinary tract infections (UTIs) in humans, we characterized E. faecalis isolates from patients in Vietnam with UTIs during January 2008–January 2010 and poultry living in close contact with them by multilocus sequence typing (MLST), pulsed-field gel electrophoresis, analysis of antimicrobial drug susceptibility patterns, and sequencing of virulence genes. In 7 (23%) of 31 UTI cases, we detected identical MLST, indistinguishable or closely related pulsed-field gel electrophoresis patterns, and similar antimicrobial drug susceptibility patterns. Isolates from urine and poultry showed identical virulence gene profiles, except for 1 variation, and individual genes showed identical sequences. The homology of isolates from urine and poultry further indicates the zoonotic potential and global spread of E. faecalis sequence type 16, which recently was reported in humans with endocarditis and in pigs in Denmark. PMID:22709904

  10. Antibiotic Resistance and Biofilm Formation of Enterococcus faecalis in Patient and Environmental Samples

    PubMed Central

    Talebi, Malihe; Asghari Moghadam, Nastaran; Mamooii, Zeynab; Enayati, Mohsen; Saifi, Mahnaz; Pourshafie, Mohammad Reza

    2015-01-01

    Background: Enterococci are opportunistic pathogens and are a major factor in nosocomial infections. They may contain ebp operon, which upon expression makes them highly prone to biofilm formation on biotic and abiotic surfaces. Objectives: The aim of the current study was to detect the polymorphism of ebp genes in Enterococcus faecalis. Materials and Methods: Samples were isolated from patients (n = 58) and hospital environments (n = 32) of two hospitals in Tehran, Iran. All enterococcal species were identified by species-specific polymerase chain reaction (PCR); the antibiotic resistance pattern against nine antibiotics was determined. The ebp A, ebp B, ebp C and srt C genes were detected by PCR and the biofilm formation by the isolates was evaluated using the microtiter plate method. The genetic diversity of ebp genes was analyzed by restriction fragment length polymorphism (RFLP). Results: The results indicated that, 86% of patient and 29% of environmental isolates carried ebp genes. The ability of the isolates to strongly attach was 62% and 71% for patient and environmental samples, respectively. The RFLP of the ebp showed no genetic variations amongst the isolates. Conclusions: The results of the antibiotic resistance and other data suggest that there is a possible common clone of E. faecalis, which could rapidly disseminate in patients and the environment. PMID:26587208

  11. In vitro evaluation of the antimicrobial efficacy of chitosan and other endodontic irrigants against Enterococcus faecalis.

    PubMed

    Shenoi, Pratima R; Morey, Elakshi S; Makade, Chetana S; Gunwal, Mohit K; Khode, Rajiv T; Wanmali, Sunay S

    2016-01-01

    The success of endodontic treatment is directly enhanced by elimination of microorganisms in infected root canals. Recently, chitosan, a natural, nontoxic biopolymer, has been introduced as an irrigant that has the capacity to remove the smear layer. The antimicrobial properties of chitosan as an endodontic irrigant have not yet been explored. The purpose of this study was to compare the antimicrobial efficacy of BioPure MTAD, 0.2% chitosan, 1% chitosan, 2% chlorhexidine gluconate, and 3% sodium hypochlorite (NaOCl) against Enterococcus faecalis, which is frequently isolated from persistent root canal infections. The agar well diffusion method was used to measure the antimicrobial activities of these irrigants. Saline was used as a negative control. The order of effectiveness was determined by the measurement of inhibition zones. Data were analyzed using 1-way analysis of variance and the Duncan multiple range test. BioPure MTAD had a significantly larger mean inhibition zone against E faecalis than the other irrigants (P < 0.001). Although 0.2% chitosan did not show any inhibition zones, 1% chitosan was as effective as 3% NaOCl (P = 0.352), and both irrigants showed significantly greater effectivity than 2% chlorhexidine (P < 0.001). Thus, 1% chitosan can be an effective natural antimicrobial substitute for synthetic irrigants. PMID:27599284

  12. Identification and Characterization of a Bacitracin Resistance Network in Enterococcus faecalis

    PubMed Central

    Fang, Chong; Shaaly, Aishath; Leslie, David J.; Weimar, Marion R.; Kalamorz, Falk; Carne, Alan; Cook, Gregory M.

    2014-01-01

    Resistance of Enterococcus faecalis against antimicrobial peptides, both of host origin and produced by other bacteria of the gut microflora, is likely to be an important factor in the bacterium's success as an intestinal commensal. The aim of this study was to identify proteins with a role in resistance against the model antimicrobial peptide bacitracin. Proteome analysis of bacitracin-treated and untreated cells showed that bacitracin stress induced the expression of cell wall-biosynthetic proteins and caused metabolic rearrangements. Among the proteins with increased production, an ATP-binding cassette (ABC) transporter with similarity to known peptide antibiotic resistance systems was identified and shown to mediate resistance against bacitracin. Expression of the transporter was dependent on a two-component regulatory system and a second ABC transporter, which were identified by genome analysis. Both resistance and the regulatory pathway could be functionally transferred to Bacillus subtilis, proving the function and sufficiency of these components for bacitracin resistance. Our data therefore show that the two ABC transporters and the two-component system form a resistance network against antimicrobial peptides in E. faecalis, where one transporter acts as the sensor that activates the TCS to induce production of the second transporter, which mediates the actual resistance. PMID:24342648

  13. Biodegradation of C.I. Reactive Red 195 by Enterococcus faecalis strain YZ66.

    PubMed

    Mate, Madhuri Sahasrabudhe; Pathade, Girish

    2012-03-01

    Synthetic dyes are extensively used in textile dyeing, paper, printing, colour photography, pharmaceutics, cosmetics and other industries. Among these, azodyes represents the largest and most versatile class of synthetic dyes. As high as 50% of the dyes are released into the environment during manufacture and usage. Traditional methods of treatment are found to be expensive and have operational problems. Biological decolourization has been investigated as a method to transform, degrade or mineralize azo dyes. In the present studies bacteria from soil from dye waste area, dye waste, sewage and dung were subjected to acclimatization with C.I. Reactive Red 195 an azo dye, in the basal nutrient media. The most promising bacterial isolate was used for further dye degradation studies. The 16s rRNA gene sequencing and biochemical characteristics revealed the isolated organism as Enterococcus faecalis strain YZ66. The strain showed 99.5% decolourization of the selected dye (Reactive Red 195-50 mg/l) within one and half hour in static anoxic condition. The optimum pH and temperature for the decolourization was 5.0 and 40°C respectively. The biodegradation was monitored by UV-Vis, FTIR, TLC and HPLC. The final products were characterized by Gas chromatography and Mass Spectrophotometry. Toxicity study demonstrated no toxicity of the biodegradation product. The results suggest that the isolated organism E. faecalis strain YZ 66 can be used as a useful tool to treat waste water containing reactive dyes. PMID:22805800

  14. Antibacterial Efficacy of Aqueous Ozone in Root Canals Infected by Enterococcus faecalis

    PubMed Central

    Hubbezoglu, Ihsan; Zan, Recai; Tunc, Tutku; Sumer, Zeynep

    2014-01-01

    Background: In endodontics, the elimination of resistant bacteria such as Enterococcus faecalis plays an important role for treatment success in root canals. Therefore, new alternative irrigants (instead of sodium hypochlorite) have been researched to achieve ideal endodontic treatment. Objectives: The aim of the present study was to evaluate and to compare the antibacterial effect of aqueous ozone with different concentrations and techniques of application (manual and ultrasonic) against E. faecalis in human root canals. Patients and Methods: Eighty single-root mandibular premolar teeth were selected, prepared and sterilized. E. faecalis was incubated in the root canals and kept at 37°C for 24 h. The teeth were divided into four main groups each has 20 members: NaOCl (positive control) group; 8 ppm aqueous ozone group; 12 ppm aqueous ozone group; and 16 ppm aqueous ozone group. While half of the specimens were disinfected with aqueous ozone by manual technique, the other half was disinfected with the aqueous ozone by ultrasonic technique. Conventional irrigation technique was simultaneously applied with ultrasonic vibration that was produced by VDW.ULTRA device. The disinfection procedures were performed for 180 s to ensure standardization of all the working groups. Paper points (placed in the root canals before and after the disinfection procedures) were transferred to Eppendorf tubes containing 0.5 mL of brain heart infusion broth. Then, 50 μL of the suspension was inoculated onto broth agar media. Microbial colonies were counted, and the data were evaluated statistically using 2-way analysis of variance (ANOVA) and Tukey tests. Results: Although the antibacterial effect of 16 ppm aqueous ozone using a manual technique had an insufficient effect, its ultrasonic application technique resulted in complete disinfection in the root canals. Conclusions: The bactericidal activity of high concentration of aqueous ozone combined with ultrasonic application technique

  15. Estradiol protects female rats against sepsis induced by Enterococcus faecalis improving leukocyte bactericidal activity.

    PubMed

    Saia, Rafael Simone; Garcia, Fabíola Morales; Cárnio, Evelin Capellari

    2015-10-01

    Enterococcus faecalis is a Gram-positive bacteria described as an important causative agent of sepsis. The contact between host leukocytes and bacteria activates the innate immunity, participating as the first defense mechanism against infection. Pro-inflammatory cytokines [including tumor necrosis factor (TNF)-α and interleukin-1β] and nitric oxide (NO) are essential to recruitment of leukocytes into the infectious focus as well as their activation for phagocytosis. Beyond the bacteria species, gender has been considered another factor to predict outcome in septic patients. Studies suggest that females exhibit a protective advantage during sepsis models, being gonadal hormones possible modulators of functions of immune cells. Nevertheless, the role of estradiol during Gram-positive infection remains a literature gap. Our aims were to investigate whether estradiol protects rats against bacterial dissemination during E. faecalis-induced sepsis. We determined whether estradiol modulates the local and systemic inflammatory response, as well as the cell migration into the infectious focus and the bactericidal capacity of leukocytes. Our findings demonstrated that estradiol pre-treated rats showed a dose-dependent reduction in bacterial counts in peritoneal lavage fluid (PLF) and in liver. Moreover, TNF-α and nitrate levels were increased in plasma, while only TNF-α was increased in the PLF in estradiol-treated rats. The prevention of bacterial dissemination may be related to the enhanced neutrophil and macrophage migration into the peritoneal cavity. Furthermore, estradiol improved the phagocytic and bactericidal ability of these both inflammatory cells. Taken together, the present study clearly demonstrates an important protective role of estradiol against sepsis induced by E. faecalis in female rats. PMID:26143494

  16. Pilin and Sortase Residues Critical for Endocarditis- and Biofilm-Associated Pilus Biogenesis in Enterococcus faecalis

    PubMed Central

    Nielsen, Hailyn V.; Flores-Mireles, Ana L.; Kau, Andrew L.; Kline, Kimberly A.; Pinkner, Jerome S.; Neiers, Fabrice; Normark, Staffan; Henriques-Normark, Birgitta

    2013-01-01

    Enterococci commonly cause hospital-acquired infections, such as infective endocarditis and catheter-associated urinary tract infections. In animal models of these infections, a long hairlike extracellular protein fiber known as the endocarditis- and biofilm-associated (Ebp) pilus is an important virulence factor for Enterococcus faecalis. For Ebp and other sortase-assembled pili, the pilus-associated sortases are essential for fiber formation as they create covalent isopeptide bonds between the sortase recognition motif and the pilin-like motif of the pilus subunits. However, the molecular requirements governing the incorporation of the three pilus subunits (EbpA, EbpB, and EbpC) have not been investigated in E. faecalis. Here, we show that a Lys residue within the pilin-like motif of the EbpC subunit was necessary for EbpC polymerization. However, incorporation of EbpA into the pilus fiber only required its sortase recognition motif (LPXTG), while incorporation of EbpB only required its pilin-like motif. Only the sortase recognition motif would be required for incorporation of the pilus tip subunit, while incorporation of the base subunit would only require the pilin recognition motif. Thus, these data support a model with EbpA at the tip and EbpB at the base of an EbpC polymer. In addition, the housekeeping sortase, SrtA, was found to process EbpB and its predicted catalytic Cys residue was required for efficient cell wall anchoring of mature Ebp pili. Thus, we have defined molecular interactions involved in fiber polymerization, minor subunit organization, and pilus subcellular compartmentalization in the E. faecalis Ebp pilus system. These studies advance our understanding of unique molecular mechanisms of sortase-assembled pilus biogenesis. PMID:23913319

  17. New trends in dentistry: plant extracts against Enterococcus faecalis. The efficacy compared to chlorhexidine.

    PubMed

    Castilho, Adriana Lígia de; Saraceni, Cintia Helena Coury; Díaz, Ingrit Elida Collantes; Paciencia, Mateus Luís Barradas; Suffredini, Ivana Barbosa

    2013-01-01

    Enterococcus faecalis is an important pathogen associated with endodontic diseases, and its elimination and control are of paramount importance, as it represents one of the major causes of failure in the treatment of endodontic disease. Twenty-five plant extracts obtained from Brazilian forests were found to be effective against planktonic E. faecalis and were subjected to two traditional antibacterial assays, the microdilution broth assay (MDBA) and the disk diffusion assay (DDA), using chlorhexidine (CHX) as a control. Seven out of 25 extracts showed significant antibacterial activity and were tested in a biofilm assay, and three of these extracts were subjected to chemical fractionation. Residues were tested for their antibacterial activity, and the first chemical findings were described based on thin layer chromatography (TLC). Extracts obtained from Ipomoea alba, Symphonia globulifera and Moronobea coccinea showed significant bactericidal activity in the MDBA. The same I. alba and S. globulifera extracts, as well as the extract obtained from Connarus ruber var. ruber, showed significant activity in the DDA. RH2O obtained from Psidium densicomum and Stryphnodendron pulcherrimum showed better antibacterial activity compared to the respective crude extracts and CHX. TLC analysis showed that phenolic compounds and triterpenes represent the first findings of chemical groups that may occur in all species. The results of the present study include the discovery of six active extracts against planktonic E. faecalis and support further testing via assays involving biofilm formation, as well as the determination of the compounds' chemical profiles, as their activity was significantly better than that observed for CHX. PMID:23538423

  18. Human salivary proteins with affinity to lipoteichoic acid of Enterococcus faecalis.

    PubMed

    Baik, Jung Eun; Choe, Hyuk-Il; Hong, Sun Woong; Kang, Seok-Seong; Ahn, Ki Bum; Cho, Kun; Yun, Cheol-Heui; Han, Seung Hyun

    2016-09-01

    Enterococcus faecalis is associated with refractory apical periodontitis and its lipoteichoic acid (Ef.LTA) is considered as a major virulence factor. Although the binding proteins of Ef.LTA may play an important role for mediating infection and immunity in the oral cavity, little is known about Ef.LTA-binding proteins (Ef.LTA-BPs) in saliva. In this study, we identified salivary Ef.LTA-BPs with biotinylated Ef.LTA (Ef.LTA-biotin) through mass spectrometry. The biotinylation of Ef.LTA was confirmed by binding capacity with streptavidin-FITC on CHO/CD14/TLR2 cells. The biological activity of Ef.LTA-biotin was determined based on the induction of nitric oxide and macrophage inflammatory protein-1α in a macrophage cell-line, RAW 264.7. To identify salivary Ef.LTA-BPs, the Ef.LTA-biotin was mixed with a pool of human saliva obtained from nine healthy subjects followed by precipitation with a streptavidin-coated bead. Ef.LTA-BPs were then separated with 12% SDS-PAGE and subjected to the mass spectrometry. Six human salivary Ef.LTA-BPs including short palate lung and nasal epithelium carcinoma-associated protein 2, zymogen granule protein 16 homolog B, hemoglobin subunit α and β, apolipoprotein A-I, and lipocalin-1 were identified with statistical significance (P<0.05). Ef.LTA-BPs were validated with lipocalin-1 using pull-down assay. Hemoglobin inhibited the biofilm formation of E. faecalis whereas lipocalin-1 did not show such effect. Collectively, the identified Ef.LTA-BPs could provide clues for our understanding of the pathogenesis of E. faecalis and host immunity in oral cavity. PMID:27474971

  19. Photodynamic therapy in root canals contaminated with Enterococcus faecalis using curcumin as photosensitizer.

    PubMed

    da Frota, Matheus Franco; Guerreiro-Tanomaru, Juliane Maria; Tanomaru-Filho, Mario; Bagnato, Vanderlei Salvador; Espir, Camila Galetti; Berbert, Fabio Luis Camargo Villela

    2015-09-01

    The aim of the study was to evaluate the photodynamic therapy (PDT) effect on root canals contaminated with Enterococcus faecalis using a light emitting diode (LED) light and a curcumin solution (CUR) as photosensitizer (PS). Eighty root canals from uniradicular human teeth were prepared with Protaper Universal rotary system and contaminated with E. faecalis for 21 days. They were divided as: GIa-PDT (CUR, pre-irradiation for 5 + 5 min of irradiation); GIb-PDT (CUR, pre-irradiation for 5 + 10 min of irradiation); GIIa-(CUR, pre-irradiation for 5 + 5 min without irradiation); GIIb-(CUR pre-irradiation for 5 + 10 min of irradiation); GIIIa-(physiological solution and irradiation for 5 min); and GIIIb-(physiological solution and irradiation for 10 min); positive and negative control groups. Collections from root canals were made at time intervals of 21 days after contamination, immediately after treatment, and 7 days after treatment, and submitted to colony forming units per milliter (CFU mL(-1)) counts. The data were submitted to ANOVA and Tukey multiple comparison tests, at a level of significance of 5 %. In the immediate post-treatment collection, group GIa showed greater bacterial reduction in comparison with GIIa, GIIb, GIIIa, GIIIb, and positive control (P < 0.05). At 7 days post-treatment, GIa showed significant bacterial reduction only in comparison with GIIIa (P < 0.05). Curcumin as sensitizer was effective by 5 min LED irradiation but not by 10 min irradiation PDT using LED light, and curcumin as PS was not effective in eliminating E. faecalis. No difference was observed for periods of irradiation. PMID:25502831

  20. Erythromycin resistance and virulence genes in Enterococcus faecalis from swine in China.

    PubMed

    Zou, Li-Kou; Wang, Hong-Ning; Zeng, Bo; Li, Jin-Niang; Li, Xu-Ting; Zhang, An-Yun; Zhou, Ying-Shun; Yang, Xin; Xu, Chang-Wen; Xia, Qing-Qing

    2011-01-01

    This study aims to describe the erythromycin resistance phenotypes and genotypes, and the prevalence of virulence genes of Enterococcus faecalis isolated from swine in China. A total of 117 nonreplicate E. faecalis isolates, obtained from 502 clinical samples taken from different pig farms between 2007 and 2009 were included in the study. Minimum inhibitory concentrations were determined using the broth microdilution method. All of the isolates were screened for the presence of seven virulence genes (ace, asa1, cylA, efaA, esp, gelE, and hyl). In addition, the DNA from rythromycin-resistant isolates were amplified with primers specific for erythromycin resistance erm(A), erm(B), erm(C), mef(A/E), and msr(C) genes. Results show that erythromycin, tylosin, and ciprofloxacin resistance rates in E. faecalis were 66.67% (n=78), 66.67% (n=78), and 64.10% (n=75), respectively. About 69.23% of isolates (n=81) were positive for gelE, 48.72% (n=57) for ace, 15.38% (n=18) for efa, 7.69% (n=9) for asa1, and 6.84% (n=8) for esp. Among the erythromycin-resistant isolates, erm(B) (n=54) was the most prevalent resistance gene, followed by erm(A) (n=37). A significant correlation was found between the presence of the gelE virulence gene and erythromycin resistance (P<0.05). These findings suggest that enterococci from swine should be regarded with caution because they can be reservoirs for antimicrobial resistance and virulence genes. PMID:21344149

  1. Biofilm forming capacity of Enterococcus faecalis on Gutta-percha points treated with four disinfectants using confocal scanning laser microscope: An in vitro study

    PubMed Central

    Ravi Chandra, Polavarapu Venkata; Kumar, Vemisetty Hari; Reddy, Surakanti Jayaprada; Kiran, Dandolu Ram; Krishna, Muppala Nagendra; Kumar, Golla Vinay

    2015-01-01

    Background: The aim of this study was to evaluate and compare the in vitro biofilm forming capacity of Enterococcus faecalis on Gutta-percha points disinfected with four disinfectants. Materials and Methods: A total of 50 Gutta-percha points used in this study were divided into four test groups based on disinfectant (5.25% sodium hypochlorite, 2% chlorhexidine gluconate, 20% neem, 13% benzalkonium chloride [BAK]), and one control group. The Gutta-percha points were initially treated with corresponding disinfectants followed by anaerobic incubation in Brain Heart Infusion broth suspended with human serum and E. faecalis strain for 14 days. After incubation, these Gutta-percha points were stained with Acridine Orange (Sigma – Aldrich Co., St. Louis, MO, USA) and 0.5 mm thick cross section samples were prepared. The biofilm thickness of E. faecalis was analyzed quantitatively using a confocal scanning laser microscope. Results statistically analyzed using analysis of variance. P < 0.05 was considered to be significant. Results: Confocal scanning laser microscope showed reduced amount of E. faecalis biofilm on Gutta-percha points treated with BAK and sodium hypochlorite. Post-hoc (least square differences) test revealed that there is no statistically significant difference between BAK and sodium hypochlorite groups (P > 0.05). Conclusion: This study illustrates that the Gutta-percha points disinfected with sodium hypochlorite and BAK showed minimal biofilm growth on its surface. PMID:26288622

  2. Antibacterial Efficacy of Calcium Hypochlorite with Vibringe Sonic Irrigation System on Enterococcus faecalis: An In Vitro Study.

    PubMed

    Dumani, Aysin; Guvenmez, Hatice Korkmaz; Yilmaz, Sehnaz; Yoldas, Oguz; Kurklu, Zeliha Gonca Bek

    2016-01-01

    Aim. The purpose of this study was to compare the in vitro efficacy of calcium hypochlorite (Ca[OCl]2) and sodium hypochlorite (NaOCl) associated with sonic (Vibringe) irrigation system in root canals which were contaminated with Enterococcus faecalis. Material and Methods. The root canals of 84 single-rooted premolars were enlarged up to a file 40, autoclaved, inoculated with Enterococcus faecalis, and incubated for 21 days. The samples were divided into 7 groups according to the irrigation protocol: G0: no treatment; G1: distilled water; G2: 2.5% NaOCl; G3: 2.5% Ca(OCl)2; G4: distilled water with sonic activation; G5: 2.5% NaOCl with sonic activation; and G6: 2.5% Ca(OCl)2 with sonic activation. Before and after decontamination procedures microbiological samples were collected and the colony-forming units were counted and the percentages of reduction were calculated. Results. Distilled water with syringe irrigation and sonic activation groups demonstrated poor antibacterial effect on Enterococcus faecalis compared to other experimental groups (p < 0.05). There was no statistically significant difference between syringe and sonic irrigation systems with Ca(OCl)2 and NaOCl. Conclusion. The antimicrobial property of Ca(OCl)2 has been investigated and compared with that of NaOCl. Both conventional syringe irrigation and sonic irrigation were found effective at removing E. faecalis from the root canal of extracted human teeth. PMID:27218106

  3. Antibacterial Efficacy of Calcium Hypochlorite with Vibringe Sonic Irrigation System on Enterococcus faecalis: An In Vitro Study

    PubMed Central

    Dumani, Aysin; Guvenmez, Hatice Korkmaz; Yilmaz, Sehnaz; Yoldas, Oguz; Kurklu, Zeliha Gonca Bek

    2016-01-01

    Aim. The purpose of this study was to compare the in vitro efficacy of calcium hypochlorite (Ca[OCl]2) and sodium hypochlorite (NaOCl) associated with sonic (Vibringe) irrigation system in root canals which were contaminated with Enterococcus faecalis. Material and Methods. The root canals of 84 single-rooted premolars were enlarged up to a file 40, autoclaved, inoculated with Enterococcus faecalis, and incubated for 21 days. The samples were divided into 7 groups according to the irrigation protocol: G0: no treatment; G1: distilled water; G2: 2.5% NaOCl; G3: 2.5% Ca(OCl)2; G4: distilled water with sonic activation; G5: 2.5% NaOCl with sonic activation; and G6: 2.5% Ca(OCl)2 with sonic activation. Before and after decontamination procedures microbiological samples were collected and the colony-forming units were counted and the percentages of reduction were calculated. Results. Distilled water with syringe irrigation and sonic activation groups demonstrated poor antibacterial effect on Enterococcus faecalis compared to other experimental groups (p < 0.05). There was no statistically significant difference between syringe and sonic irrigation systems with Ca(OCl)2 and NaOCl. Conclusion. The antimicrobial property of Ca(OCl)2 has been investigated and compared with that of NaOCl. Both conventional syringe irrigation and sonic irrigation were found effective at removing E. faecalis from the root canal of extracted human teeth. PMID:27218106

  4. Bacteriocin Protein BacL1 of Enterococcus faecalis Is a Peptidoglycan d-Isoglutamyl-l-lysine Endopeptidase*

    PubMed Central

    Kurushima, Jun; Hayashi, Ikue; Sugai, Motoyuki; Tomita, Haruyoshi

    2013-01-01

    Enterococcus faecalis strains are commensal bacteria in humans and other animals, and they are also the causative agent of opportunistic infectious diseases. Bacteriocin 41 (Bac41) is produced by certain E. faecalis clinical isolates, and it is active against other E. faecalis strains. Our genetic analyses demonstrated that the extracellular products of the bacL1 and bacA genes, which are encoded in the Bac41 operon, coordinately express the bacteriocin activity against E. faecalis. In this study, we investigated the molecular functions of the BacL1 and BacA proteins. Immunoblotting and N-terminal amino acid sequence analysis revealed that BacL1 and BacA are secreted without any processing. The coincidental treatment with the recombinant BacL1 and BacA showed complete bacteriocin activity against E. faecalis, but neither BacL1 nor BacA protein alone showed the bacteriocin activity. Interestingly, BacL1 alone demonstrated substantial degrading activity against the cell wall fraction of E. faecalis in the absence of BacA. Furthermore, MALDI-TOF MS analysis revealed that BacL1 has a peptidoglycan d-isoglutamyl-l-lysine endopeptidase activity via a NlpC/P60 homology domain. These results collectively suggest that BacL1 serves as a peptidoglycan hydrolase and, when BacA is present, results in the lysis of viable E. faecalis cells. PMID:24235140

  5. Prevalence of putative virulence factors and antimicrobial susceptibility of Enterococcus faecalis isolates from patients with dental Diseases

    PubMed Central

    Salah, Randa; Dar-Odeh, Najla; Abu Hammad, Osama; Shehabi, Asem A

    2008-01-01

    Background This study investigated the prevalence of Enterococcus faecalis, its putative virulence factors and antimicrobial susceptibility in individuals with and without dental diseases. A total of 159 oral rinse specimens were collected from patients (n = 109) suffering from dental diseases and healthy controls (n = 50). Results E. faecalis was detected using only culture in 8/109 (7.3%) of the patients with various types of dental diseases, whereas no E. faecalis was found in the healthy controls weather using both culture and PCR. Phenotype characterizations of the 8 E. faecalis isolates indicated that 25% of the isolates produced haemolysin and 37.5% produced gelatinase. Most important virulence genes; collagen binding protein (ace) and endocarditis antigen (efaA) were present in all 8 E. faecalis isolates, while haemolysin activator gene (cylA) was detected only in 25% of isolates, and all isolates were negative for esp gene. All E. faecalis isolates were 100% susceptible to ampicillin, chloramphenicol, ciprofloxacin, vancomycin, and teicoplanin, and to less extent to erythromycin (62.5%). Conclusion This study shows that all E. faecalis isolates were recovered only from patients with dental diseases especially necrotic pulps, and all isolates carried both collagen binding protein and endocarditis antigen genes and highly susceptible to frequently used antimicrobial drugs in Jordan. PMID:18513445

  6. The mazEF toxin–antitoxin system as an attractive target in clinical isolates of Enterococcus faecium and Enterococcus faecalis

    PubMed Central

    Soheili, Sara; Ghafourian, Sobhan; Sekawi, Zamberi; Neela, Vasantha Kumari; Sadeghifard, Nourkhoda; Taherikalani, Morovat; Khosravi, Afra; Ramli, Ramliza; Hamat, Rukman Awang

    2015-01-01

    The toxin–antitoxin (TA) system is a regulatory system where two sets of genes encode the toxin and its corresponding antitoxin. In this study, the prevalence of TA systems in independently isolated clinical isolates of Enterococcus faecium and Enterococcus faecalis was determined, the dominant TA system was identified, different virulence genes in E. faecium and E. faecalis were surveyed, the level of expression of the virulence and TA genes in normal and stress conditions was determined, and finally their associations with the TA genes were defined. Remarkably, the analysis demonstrated higBA and mazEF in all clinical isolates, and their locations were on chromosomes and plasmids, respectively. On the other hand, a quantitative analysis of TA and virulence genes revealed that the expression level in both genes is different under normal and stress conditions. The results obtained by anti-mazF peptide nucleic acids demonstrated that the expression level of virulence genes had decreased. These findings demonstrate an association between TA systems and virulence factors. The mazEF on the plasmids and the higBA TA genes on the chromosomes of all E. faecium and E. faecalis strains were dominant. Additionally, there was a decrease in the expression of virulence genes in the presence of anti-mazF peptide nucleic acids. Therefore, it is suggested that mazEF TA systems are potent and sensitive targets in all E. faecium and E. faecalis strains. PMID:26005332

  7. Biochemical characterization of an anti-Candida factor produced by Enterococcus faecalis

    PubMed Central

    2012-01-01

    Background Because Candida albicans is resistant to several antifungal antibiotics, there is a need to identify other less toxic natural products, particularly antimicrobial proteins, peptides or bacteriocin like inhibitory substances. An attempt has been made to purify and characterise an anti-Candida compound produced by Enterococcus faecalis. Results An anti-Candida protein (ACP) produced by E. faecalis active against 8 C. albicans strains was characterised and partially purified. The ACP showed a broad-spectrum activity against multidrug resistant C. albicans MTCC 183, MTCC 7315, MTCC 3958, NCIM 3557, NCIM 3471 and DI. It was completely inactivated by treatment with proteinase K and partially by pronase E. The ACP retained biological stability after heat-treatment at 90°C for 20 min, maintained activity over a pH range 6–10, and remained active after treatment with α-amylase, lipase, organic solvents, and detergents. The antimicrobial activity of the E. faecalis strain was found exclusively in the extracellular filtrate produced in the late logarithmic growth phase. The highest activity (1600 AU mL-1) against C. albicans MTCC 183 was recorded at 48 h of incubation, and activity decreased thereafter. The peptide showed very low haemagglutination and haemolytic activities against human red blood cells. The antimicrobial substance was purified by salt-fractionation and chromatography. Partially purified ACP had a molecular weight of approximately 43 KDa in Tricine-PAGE analysis. The 12 amino acid N terminal sequence was obtained by Edman degradation. The peptide was de novo sequenced by ESI-MS, and the deduced combined sequence when compared to other bacteriocins and antimicrobial peptide had no significant sequence similarity. Conclusions The inhibitory activity of the test strain is due to the synthesis of an antimicrobial protein. To our knowledge, this is the first report on the isolation of a promising non-haemolytic anti-Candida protein from E

  8. In Vitro Activities of Telavancin and Vancomycin against Biofilm-Producing Staphylococcus aureus, S. epidermidis, and Enterococcus faecalis Strains▿

    PubMed Central

    LaPlante, Kerry L.; Mermel, Leonard A.

    2009-01-01

    We investigated the activities of telavancin and vancomycin against biofilm-producing Staphylococcus and Enterococcus strains. At clinically attainable concentrations, telavancin was active against bacteria embedded in biofilm (minimal biofilm eradication concentration [MBEC], 0.125 to 2 μg/ml) and inhibited biofilm formation at concentrations below the MIC. Vancomycin did not demonstrate the same activity (MBEC, ≥512 μg/ml) against Staphylococcus aureus and Enterococcus faecalis. Telavancin may have a unique role in biofilm-associated infections. PMID:19451302

  9. Genome sequences of copper resistant and sensitive Enterococcus faecalis strains isolated from copper-fed pigs in Denmark

    PubMed Central

    2015-01-01

    Six strains of Enterococcus faecalis (S1, S12, S17, S18, S19 and S32) were isolated from copper fed pigs in Denmark. These Gram-positive bacteria within the genus Enterococcus are able to survive a variety of physical and chemical challenges by the acquisition of diverse genetic elements. The genome of strains S1, S12, S17, S18, S19 and S32 contained 2,615, 2,769, 2,625, 2,804, 2,853 and 2,935 protein-coding genes, with 41, 42, 27, 42, 32 and 44 genes encoding antibiotic and metal resistance, respectively. Differences between Cu resistant and sensitive E. faecalis strains, and possible co-transfer of Cu and antibiotic resistance determinants were detected through comparative genome analysis. PMID:26203344

  10. Complete Genome Sequence of Enterococcus faecalis Strain P8-1 Isolated from Wild Magellanic Penguin (Spheniscus magellanicus) Feces on the South Coast of Brazil.

    PubMed

    Prichula, Janira; Campos, Fabricio Souza; Pereira, Rebeca Inhoque; Cardoso, Leonardo Almansa; Wachholz, Guilherme Raffo; Pieta, Luiza; Mariot, Roberta Fogliatto; de Moura, Tiane Martin; Tavares, Maurício; d'Azevedo, Pedro Alves; Frazzon, Jeverson; Frazzon, Ana Paula Guedes

    2016-01-01

    Enterococcus faecalis strains have a ubiquitous nature that allows them to survive in different niches. Studies involving enterococci isolated from marine animals are scarce. Therefore, in this study, we report the complete genome sequence of E. faecalis strain P8-1 isolated from feces of a Magellanic penguin on the south coast of Brazil. PMID:26769928

  11. Complete Genome Sequence of Enterococcus faecalis Strain P8-1 Isolated from Wild Magellanic Penguin (Spheniscus magellanicus) Feces on the South Coast of Brazil

    PubMed Central

    Prichula, Janira; Campos, Fabricio Souza; Pereira, Rebeca Inhoque; Cardoso, Leonardo Almansa; Wachholz, Guilherme Raffo; Pieta, Luiza; Mariot, Roberta Fogliatto; de Moura, Tiane Martin; Tavares, Maurício; d’Azevedo, Pedro Alves; Frazzon, Ana Paula Guedes

    2016-01-01

    Enterococcus faecalis strains have a ubiquitous nature that allows them to survive in different niches. Studies involving enterococci isolated from marine animals are scarce. Therefore, in this study, we report the complete genome sequence of E. faecalis strain P8-1 isolated from feces of a Magellanic penguin on the south coast of Brazil. PMID:26769928

  12. High-level plasmid-mediated gentamicin resistance and pheromone response of plasmids present in clinical isolates of Enterococcus faecalis.

    PubMed Central

    Shiojima, M; Tomita, H; Tanimoto, K; Fujimoto, S; Ike, Y

    1997-01-01

    Eleven pheromone-responding plasmids encoding erythromycin or gentamicin resistance were isolated from multiresistant clinical Enterococcus faecalis isolates. The plasmids were classified into six types with respect to their pheromone responses. The three erythromycin resistance plasmids responded to different pheromones. Of the eight gentamicin resistance plasmids, four plasmids responded to same pheromone. Southern hybridization studies showed that the genes involved in regulation of the pheromone response were conserved in the drug resistance plasmids. PMID:9056018

  13. Enterococcus faecalis 3-hydroxy-3-methylglutaryl coenzyme A synthase, an enzyme of isopentenyl diphosphate biosynthesis.

    PubMed

    Sutherlin, Autumn; Hedl, Matija; Sanchez-Neri, Barbara; Burgner, John W; Stauffacher, Cynthia V; Rodwell, Victor W

    2002-08-01

    Biosynthesis of the isoprenoid precursor isopentenyl diphosphate (IPP) proceeds via two distinct pathways. Sequence comparisons and microbiological data suggest that multidrug-resistant strains of gram-positive cocci employ exclusively the mevalonate pathway for IPP biosynthesis. Bacterial mevalonate pathway enzymes therefore offer potential targets for development of active site-directed inhibitors for use as antibiotics. We used the PCR and Enterococcus faecalis genomic DNA to isolate the mvaS gene that encodes 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) synthase, the second enzyme of the mevalonate pathway. mvaS was expressed in Escherichia coli from a pET28 vector with an attached N-terminal histidine tag. The expressed enzyme was purified by affinity chromatography on Ni(2+)-agarose to apparent homogeneity and a specific activity of 10 micromol/min/mg. Analytical ultracentrifugation showed that the enzyme is a dimer (mass, 83.9 kDa; s(20,w), 5.3). Optimal activity occurred in 2.0 mM MgCl(2) at 37(o)C. The DeltaH(a) was 6,000 cal. The pH activity profile, optimum activity at pH 9.8, yielded a pK(a) of 8.8 for a dissociating group, presumably Glu78. The stoichiometry per monomer of acetyl-CoA binding was 1.2 +/- 0.2 and that of covalent acetylation was 0.60 +/- 0.02. The K(m) for the hydrolysis of acetyl-CoA was 10 microM. Coupled conversion of acetyl-CoA to mevalonate was demonstrated by using HMG-CoA synthase and acetoacetyl-CoA thiolase/HMG-CoA reductase from E. faecalis. PMID:12107122

  14. Atomic force microscopy visualization of injuries in Enterococcus faecalis surface caused by Er,Cr:YSGG and diode lasers

    PubMed Central

    López-Jiménez, Lidia; Viñas, Miguel; Vinuesa, Teresa

    2015-01-01

    Aim: To visualize by Atomic Force Microscopy the alterations induced on Enterococcus. faecalis surface after treatment with 2 types of laser: Erbium chromium:yttrium-scandium-gallium-garnet (Er,Cr:YSGG) laser and Diode laser. Material and Methods: Bacterial suspensions from overnight cultures of E. faecalis were irradiated during 30 seconds with the laser-lights at 1 W and 2 W of power, leaving one untreated sample as control. Surface alterations on treated E. faecalis were visualized by Atomic Force Microscopy (AFM) and its surface roughness determined. Results: AFM imaging showed that at high potency of laser both cell morphology and surface roughness resulted altered, and that several cell lysis signs were easily visualized. Surface roughness clearly increase after the treatment with Er,Cr:YSGG at 2W of power, while the other treatments gave similar values of surface roughness. The effect of lasers on bacterial surfaces visualized by AFM revealed drastic alterations. Conclusions: AFM is a good tool to evaluate surface injuries after laser treatment; and could constitute a measure of antimicrobial effect that can complete data obtained by determination of microbial viability. Key words:Atomic force microscopy, Er,Cr:YSGG laser, diode laser, Enterococcus faecalis, surface roughness. PMID:25475770

  15. Systems biology approach for mapping the response of human urothelial cells to infection by Enterococcus faecalis

    PubMed Central

    Dozmorov, Mikhail G; Kyker, Kimberly D; Saban, Ricardo; Shankar, Nathan; Baghdayan, Arto S; Centola, Michael B; Hurst, Robert E

    2007-01-01

    Background To better understand the response of urinary epithelial (urothelial) cells to Enterococcus faecalis, a uropathogen that exhibits resistance to multiple antibiotics, a genome-wide scan of gene expression was obtained as a time series from urothelial cells growing as a layered 3-dimensional culture similar to normal urothelium. We herein describe a novel means of analysis that is based on deconvolution of gene variability into technical and biological components. Results Analysis of the expression of 21,521 genes from 30 minutes to 10 hours post infection, showed 9553 genes were expressed 3 standard deviations (SD) above the system zero-point noise in at least 1 time point. The asymmetric distribution of relative variances of the expressed genes was deconvoluted into technical variation (with a 6.5% relative SD) and biological variation components (>3 SD above the mode technical variability). These 1409 hypervariable (HV) genes encapsulated the effect of infection on gene expression. Pathway analysis of the HV genes revealed an orchestrated response to infection in which early events included initiation of immune response, cytoskeletal rearrangement and cell signaling followed at the end by apoptosis and shutting down cell metabolism. The number of poorly annotated genes in the earliest time points suggests heretofore unknown processes likely also are involved. Conclusion Enterococcus infection produced an orchestrated response by the host cells involving several pathways and transcription factors that potentially drive these pathways. The early time points potentially identify novel targets for enhancing the host response. These approaches combine rigorous statistical principles with a biological context and are readily applied by biologists. PMID:18047719

  16. The Transcriptome of the Nosocomial Pathogen Enterococcus faecalis V583 Reveals Adaptive Responses to Growth in Blood

    PubMed Central

    Vebø, Heidi C.; Snipen, Lars; Nes, Ingolf F.; Brede, Dag A.

    2009-01-01

    Background Enterococcus faecalis plays a dual role in human ecology, predominantly existing as a commensal in the alimentary canal, but also as an opportunistic pathogen that frequently causes nosocomial infections like bacteremia. A number of virulence factors that contribute to the pathogenic potential of E. faecalis have been established. However, the process in which E. faecalis gains access to the bloodstream and establishes a persistent infection is not well understood. Methodology/Principal Findings To enhance our understanding of how this commensal bacterium adapts during a bloodstream infection and to examine the interplay between genes we designed an in vitro experiment using genome-wide microarrays to investigate what effects the presence of and growth in blood have on the transcriptome of E. faecalis strain V583. We showed that growth in both 2xYT supplemented with 10% blood and in 100% blood had a great impact on the transcription of many genes in the V583 genome. We identified several immediate changes signifying cellular processes that might contribute to adaptation and growth in blood. These include modulation of membrane fatty acid composition, oxidative and lytic stress protection, acquisition of new available substrates, transport functions including heme/iron transporters and genes associated with virulence in E. faecalis. Conclusions/Significance The results presented here reveal that cultivation of E. faecalis in blood in vitro has a profound impact on its transcriptome, which includes a number of virulence traits. Observed regulation of genes and pathways revealed new insight into physiological features and metabolic capacities which enable E. faecalis to adapt and grow in blood. A number of the regulated genes might potentially be useful candidates for development of new therapeutic approaches for treatment of E. faecalis infections. PMID:19888459

  17. Molecular Epidemiologic Analysis of Enterococcus faecalis Isolates in Cuba by Multilocus Sequence Typing

    PubMed Central

    Kobayashi, Nobumichi; Nagashima, Shigeo

    2009-01-01

    We carried out the first study of Enterococcus faecalis clinical isolates in Cuba by multilocus sequence typing linking the molecular typing data with the presence of virulence determinants and the antibiotic resistance genes. A total of 23 E. faecalis isolates recovered from several clinic sources and geographic areas of Cuba during a period between 2000 and 2005 were typed by multilocus sequence typing. Thirteen sequence types (STs) including five novel STs were identified, and the ST 64 (clonal complex [CC] 8), ST 6 (CC2), ST 21(CC21), and ST 16 (CC58) were found in more than one strain. Sixty-seven percent of STs corresponded to STs reported previously in Spain, Poland, and The Netherlands, and other STs (ST115, ST64, ST6, and ST40) were genetically close to those detected in the United States. Prevalence of both antimicrobial resistance genes [aac(6′)-aph(2″), aph(3′), ant(6), ant(3″)(9), aph(2″)-Id, aph(2″)-Ic, erm(B), erm(A), erm(C), mef(A), tet(M), and tet(L)] and virulence genes (agg, gelE, cylA, esp, ccf, and efaAfs) were examined by polymerase chain reaction. Aminoglycoside resistance genes aac(6′)-Ie-aph(2″)-Ia, aph(3′), ant(6), ant(3″)(9) were more frequently detected in ST6, ST16, ST23, ST64, and ST115. The multidrug resistance was distributed to all STs detected, except for ST117 and singleton ST225. The presence of cyl gene was specifically linked to the ST64 and ST16. Presence of the esp, gel, and agg genes was not specific to any particular ST. This research provided the first insight into the population structure of E. faecalis in Cuba, that is, most Cuban strains were related to European strains, whereas others to U.S. strains. The CC2, CC21, and CC8, three of the biggest CCs in the world, were evidently circulating in Cuba, associated with multidrug resistance and virulence traits. PMID:19857135

  18. Partial purification and characterization of bacteriocin produced by Enterococcus faecalis DU10 and its probiotic attributes.

    PubMed

    Perumal, Venkatesh; Repally, Ayyanna; Dasari, Ankaiah; Venkatesan, Arul

    2016-10-01

    A novel bacteriocin produced by avian duck isolated lactic acid bacterium Enterococcus faecalis DU10 was isolated. This bacteriocin showed a broad spectrum of antibacterial activity against important food-borne pathogens and was purified by size exclusion chromatography followed by reverse-phase high-performance liquid chromatography in a C-18 column. Tricine-SDS PAGE revealed the presence of a band with an estimated molecular mass of 6.3 kDa. The zymogram clearly linked the antimicrobial activity with this band. This result was further confirmed by mass-assisted laser desorption ionization time-of-flight mass spectrometry, since a sharp peak corresponding to 6.313 kDa was detected and the functional groups were revealed by Fourier transform infrared spectroscopy. Bacteriocin DU10 activity was found sensitive to proteinase-K and pepsin and partially affected by trypsin and α-chymotrypsin. The activity of bacteriocin DU10 was partially resistant to heat treatments ranging from 30 to 90°C for 30 min. It also withstood a treatment at 121°C for 10 min. Cytotoxicity of bacteriocin DU10 by methyl-thiazolyl-diphenyl-tetrazolium bromide assay showed that the viability of HT-29 and HeLa cells decreased 60 ± 0.7% and 43 ± 4.8%, respectively, in the presence of 3,200 AU/mL of bacteriocin. The strain withstood 0.3% w/v of bile oxgall and pH 2 affected the bacterial growth between 2 and 4 hr of incubation. Adhesion properties examined with HT-29 cell line showed 69.85% initial population of strain E. faecalis DU10, which was found to be strongly adhered to this cell line. These results conclude bacteriocin DU10 may be used as a potential biopreservative and E. faecalis DU10 may be used as a potential probiont to control Salmonella infections. PMID:26786752

  19. Biofilms of Enterococcus faecalis and Enterococcus faecium isolated from the processing of ricotta and the control of these pathogens through cleaning and sanitization procedures.

    PubMed

    da Silva Fernandes, Meg; Kabuki, Dirce Yorika; Kuaye, Arnaldo Yoshiteru

    2015-05-01

    The biofilm formation of Enterococcus faecalis and Enterococcus faecium isolated from the processing of ricotta on stainless steel coupons was evaluated, and the effect of cleaning and sanitization procedures in the control of these biofilms was determined. The formation of biofilms was observed while varying the incubation temperature (7, 25 and 39°C) and time (0, 1, 2, 4, 6 and 8 days). At 7°C, the counts of E. faecalis and E. faecium were below 2 log10 CFU/cm(2). For the temperatures of 25 and 39°C, after 1 day, the counts of E. faecalis and E. faecium were 5.75 and 6.07 log10 CFU/cm(2), respectively, which is characteristic of biofilm formation. The tested sanitation procedures a) acid-anionic tensioactive cleaning, b) anionic tensioactive cleaning+sanitizer and c) acid-anionic tensioactive cleaning+sanitizer were effective in removing the biofilms, reducing the counts to levels below 0.4 log10 CFU/cm(2). The sanitizer biguanide was the least effective, and peracetic acid was the most effective. These studies revealed the ability of enterococci to form biofilms and the importance of the cleaning step and the type of sanitizer used in sanitation processes for the effective removal of biofilms. PMID:25702883

  20. CRISPR-Cas and Restriction-Modification Act Additively against Conjugative Antibiotic Resistance Plasmid Transfer in Enterococcus faecalis

    PubMed Central

    Price, Valerie J.; Huo, Wenwen; Sharifi, Ardalan

    2016-01-01

    ABSTRACT Enterococcus faecalis is an opportunistic pathogen and a leading cause of nosocomial infections. Conjugative pheromone-responsive plasmids are narrow-host-range mobile genetic elements (MGEs) that are rapid disseminators of antibiotic resistance in the faecalis species. Clustered regularly interspaced short palindromic repeat (CRISPR)-Cas and restriction-modification confer acquired and innate immunity, respectively, against MGE acquisition in bacteria. Most multidrug-resistant E. faecalis isolates lack CRISPR-Cas and possess an orphan locus lacking cas genes, CRISPR2, that is of unknown function. Little is known about restriction-modification defense in E. faecalis. Here, we explore the hypothesis that multidrug-resistant E. faecalis strains are immunocompromised. We assessed MGE acquisition by E. faecalis T11, a strain closely related to the multidrug-resistant hospital isolate V583 but which lacks the ~620 kb of horizontally acquired genome content that characterizes V583. T11 possesses the E. faecalis CRISPR3-cas locus and a predicted restriction-modification system, neither of which occurs in V583. We demonstrate that CRISPR-Cas and restriction-modification together confer a 4-log reduction in acquisition of the pheromone-responsive plasmid pAM714 in biofilm matings. Additionally, we show that the orphan CRISPR2 locus is functional for genome defense against another pheromone-responsive plasmid, pCF10, only in the presence of cas9 derived from the E. faecalis CRISPR1-cas locus, which most multidrug-resistant E. faecalis isolates lack. Overall, our work demonstrated that the loss of only two loci led to a dramatic reduction in genome defense against a clinically relevant MGE, highlighting the critical importance of the E. faecalis accessory genome in modulating horizontal gene transfer. Our results rationalize the development of antimicrobial strategies that capitalize upon the immunocompromised status of multidrug-resistant E. faecalis

  1. CRISPR-Cas and Restriction-Modification Act Additively against Conjugative Antibiotic Resistance Plasmid Transfer in Enterococcus faecalis.

    PubMed

    Price, Valerie J; Huo, Wenwen; Sharifi, Ardalan; Palmer, Kelli L

    2016-01-01

    Enterococcus faecalis is an opportunistic pathogen and a leading cause of nosocomial infections. Conjugative pheromone-responsive plasmids are narrow-host-range mobile genetic elements (MGEs) that are rapid disseminators of antibiotic resistance in the faecalis species. Clustered regularly interspaced short palindromic repeat (CRISPR)-Cas and restriction-modification confer acquired and innate immunity, respectively, against MGE acquisition in bacteria. Most multidrug-resistant E. faecalis isolates lack CRISPR-Cas and possess an orphan locus lacking cas genes, CRISPR2, that is of unknown function. Little is known about restriction-modification defense in E. faecalis. Here, we explore the hypothesis that multidrug-resistant E. faecalis strains are immunocompromised. We assessed MGE acquisition by E. faecalis T11, a strain closely related to the multidrug-resistant hospital isolate V583 but which lacks the ~620 kb of horizontally acquired genome content that characterizes V583. T11 possesses the E. faecalis CRISPR3-cas locus and a predicted restriction-modification system, neither of which occurs in V583. We demonstrate that CRISPR-Cas and restriction-modification together confer a 4-log reduction in acquisition of the pheromone-responsive plasmid pAM714 in biofilm matings. Additionally, we show that the orphan CRISPR2 locus is functional for genome defense against another pheromone-responsive plasmid, pCF10, only in the presence of cas9 derived from the E. faecalis CRISPR1-cas locus, which most multidrug-resistant E. faecalis isolates lack. Overall, our work demonstrated that the loss of only two loci led to a dramatic reduction in genome defense against a clinically relevant MGE, highlighting the critical importance of the E. faecalis accessory genome in modulating horizontal gene transfer. Our results rationalize the development of antimicrobial strategies that capitalize upon the immunocompromised status of multidrug-resistant E. faecalis. IMPORTANCE

  2. Spread of multidrug-resistant Enterococcus faecalis within the household setting.

    PubMed

    Leite-Martins, Liliana; Meireles, Diana; Bessa, Lucinda J; Mendes, Ângelo; de Matos, Augusto J; da Costa, Paulo Martins

    2014-10-01

    Advances in veterinary medicine have resulted in the survival of many animals with severe illness or infectious diseases. In addition, increased usage of antimicrobial agents for veterinary purposes has contributed to the worldwide problem of increasing antimicrobial resistance. The objective of this study was to contribute to better understand the potential and implications for the spread of antimicrobial-resistant enterococci between pets receiving antimicrobial treatments and their owners. Three household aggregates (HA A, B, and C) were selected for this study. Information was collected on individual and clinical parameters of both humans and animals that cohabit. For this study, samples of feces, oral secretions, skin and fur of pets, as well as owners' feces and hands and exposed household surfaces and objects were also collected. All enterococci isolates were analyzed for antimicrobial susceptibility. Based on the antimicrobial resistance patterns and origin of isolates, ERIC-PCR analysis was performed on selected isolates to evaluate phylogenetic relationships. In all three HA, Enterococcus faecalis clonal spread was detected between pets and the respective owners, confirming the in-home interanimal species dissemination. Additionally, fecal enterococci colonization of other body parts of the same animal and dissemination of those same enterococci to household surfaces and objects were also observed. Our results demonstrate that enterococcal clones were found in pets in multiple body sites, their human cohabitants, and shared domestic objects. PMID:24617521

  3. Structural analysis and proteolytic activation of Enterococcus faecalis cytolysin, a novel lantibiotic.

    PubMed

    Booth, M C; Bogie, C P; Sahl, H G; Siezen, R J; Hatter, K L; Gilmore, M S

    1996-09-01

    Clinical isolates of Enterococcus faecalis more commonly produce a cytolysin than do commensal isolates. Epidemiologic evidence and animal-model studies have established a role for the cytolysin in the pathogenesis of enterococcal disease. The cytolysin consists of two structural subunits, CylLL and CylLS, that are activated by a third component, CylA. Genetic and biochemical characterization of CylA indicate that it is a serine protease, and that activation putatively results from cleavage of one or both cytolysin subunits. Genetic evidence also suggests that the cytolysin subunits are related to the rapidly growing class of bacteriocins termed lantibiotics. However, unlike lantibiotics, the cytolysin is lytic for eukaryotic as well as prokaryotic cells, and it consists of two structural subunits. This report describes the purification and characterization of the cytolysin subunits and detection of lanthionine-type post-translational modifications within their structures. Furthermore, the cleavage specificity of the CylA activator is reported and it is shown that proteolytic activation of both subunits is essential for activity. PMID:8898386

  4. In Vitro Comparison of the Effectiveness of Chlorhexidine and Two Calcium Hydroxide Formulations on Enterococcus Faecalis

    PubMed Central

    Sharifian, Mohammad Reza; Shokouhinejad, Noushin; Aligholi, Marzieh; Emaneini, Mohammad; Katebi, Arash; Assadian, Hadi

    2008-01-01

    INTRODUCTION: The aim of this in vitro study was to compare the effectiveness of three intracanal medicaments in disinfecting the root canal and dentin of experimentally infected human teeth with Enterococcus faecalis (EF). MATERIALS AND METHODS: One hundred extracted human single-rooted teeth were used. After root canal preparation, teeth were mounted in epoxy resin. Following sterilization, the teeth were infected for 28 days with EF. Then root canals were filled with one of three different disinfectants: viscous 2% Chlorhexidine (CHX), calcium hydroxide paste (CH) or a mixture of CH and CHX (n=30 in each group). Antimicrobial assessments were performed at 1, 3 and 7 days (n=10 in each time period). Microbial samples were obtained from root canals before and after the experiment. Also dentin samples were examined. The data was analyzed using Two- Way ANOVA test. RESULTS: The findings showed that there was no difference between experimental groups at different time periods. The mixture of CH/CHX in 7 days was able to eliminate EF completely from root canal system. The most elimination of EF was from dentinal tubules. CONCLUSION: According to the results of this in vitro study, viscous 2% CHX, mixture of CH with distilled water and 2% CHX are all effective disinfectants. PMID:24146671

  5. Heat-Killed Enterococcus faecalis EF-2001 Ameliorates Atopic Dermatitis in a Murine Model.

    PubMed

    Choi, Eun-Ju; Iwasa, Masahiro; Han, Kwon-Il; Kim, Wan-Jae; Tang, Yujiao; Hwang, Young Joung; Chae, Jeong Ryong; Han, Weon Cheol; Shin, Yu-Su; Kim, Eun-Kyung

    2016-03-01

    Recent reports have shown the immunomodulatory effect of heat-killed lactic acid bacteria. Atopic dermatitis (AD) is an allergic skin disease, caused by immune dysregulation among other factors. The aim of this study was to assess the effect of heat-killed Enterococcus faecalis EF-2001 (EF-2001) on AD. We established an in vivo AD model by repeated local exposure of Dermatophagoides farinae extract (DFE; house dust mite extract) and 2,4-dinitrochlorobenzene (DNCB) to the ears of mice. After oral administration of EF-2001 for four weeks, the epidermal and dermal ear thickness, mast cell infiltration, and serum immunoglobulin levels were measured. In addition, the gene expression levels of pathogenic cytokines in the ears, lymph nodes, and splenocytes were assayed. EF-2001 attenuated AD symptoms based on the ear thickness, histopathological analysis, and serum immunoglobulin levels. Moreover, EF-2001 decreased the DFE/DNCB-induced expression of various pathogenic cytokines in the ears, lymph nodes, and splenocytes. These results suggest that EF-2001 has therapeutic potential in the treatment of AD owing to its immunomodulatory effects. PMID:26959058

  6. Alterations in peptidoglycan precursors and vancomycin susceptibility in Tn917 insertion mutants of Enterococcus faecalis 221.

    PubMed Central

    Handwerger, S

    1994-01-01

    Derivatives of the highly vancomycin-resistant Enterococcus faecalis strain 221 (MIC, 1,024 micrograms/ml) harboring Tn917 insertions in vanR, vanH, and vanA were compared with the parent strain and the susceptible plasmid-free strain JH2-2 (MIC, 2 micrograms/ml). Cytoplasmic pools of UDP-N-acetyl-muramyl-peptide precursors of strain 221 contained the depsipeptide-terminating precursor as well as elevated levels of both the tripeptide and tetrapeptide precursors. Insertional inactivation of vanR resulted in the loss of carboxypeptidase activity, full susceptibility to vancomycin, and precursor pools similar to those of JH2-2. For the vanA insertional mutant the MBC of vancomycin was fourfold higher than that for JH2-2, and the mutant had increased levels of tripeptide and tetrapeptide precursors compared with those for JH2-2. The vanH insertional mutant showed elevated levels of these precursors, as well as a small amount of depsipeptide, and both the MIC and the MBC of vancomycin were increased compared with those for JH2-2. These findings suggest that DD-carboxypeptidase activity, under the control of vanR, results in increased pools of both tripeptide and tetrapeptide precursors, which may contribute to survival in the presence of vancomycin. PMID:8203839

  7. Molecular cloning, expression, and characterization of a novel endo-alpha-N-acetylgalactosaminidase from Enterococcus faecalis.

    PubMed

    Goda, Hatsumi M; Ushigusa, Kota; Ito, Hiromi; Okino, Nozomu; Narimatsu, Hisashi; Ito, Makoto

    2008-10-31

    We report here the molecular cloning, expression and characterization of a novel endo-alpha-N-acetylgalactosaminidase, classified into the GH101 family, from Enterococcus faecalis (endo-EF). The recombinant endo-EF was found to catalyze the liberation of core1-disaccharides (Galbeta1-3GalNAc) from core1-pNP (Galbeta1-3GalNAcalpha-pNP) like other GH101 family enzymes. However, endo-EF seems to differ in specificity from the GH101 enzymes reported to date, because it was able to release trisaccharides from core2-pNP (Galbeta1-3[GlcNAcbeta1-6]GalNAcalpha-pNP) and tetrasaccharides from Gal-core2-pNP (Galbeta1-3[Galbeta1-3GlcNAcbeta1-6]GalNAcalpha-pNP). Interestingly, the enzyme could transfer not only core1-disaccharides but also core2-trisaccharides to alkanols generating alkyl-oligosaccharides. Endo-EF should facilitate O-glycoprotein research. PMID:18725192

  8. Heat-Killed Enterococcus faecalis EF-2001 Ameliorates Atopic Dermatitis in a Murine Model

    PubMed Central

    Choi, Eun-Ju; Iwasa, Masahiro; Han, Kwon-Il; Kim, Wan-Jae; Tang, Yujiao; Hwang, Young Joung; Chae, Jeong Ryong; Han, Weon Cheol; Shin, Yu-Su; Kim, Eun-Kyung

    2016-01-01

    Recent reports have shown the immunomodulatory effect of heat-killed lactic acid bacteria. Atopic dermatitis (AD) is an allergic skin disease, caused by immune dysregulation among other factors. The aim of this study was to assess the effect of heat-killed Enterococcus faecalis EF-2001 (EF-2001) on AD. We established an in vivo AD model by repeated local exposure of Dermatophagoides farinae extract (DFE; house dust mite extract) and 2,4-dinitrochlorobenzene (DNCB) to the ears of mice. After oral administration of EF-2001 for four weeks, the epidermal and dermal ear thickness, mast cell infiltration, and serum immunoglobulin levels were measured. In addition, the gene expression levels of pathogenic cytokines in the ears, lymph nodes, and splenocytes were assayed. EF-2001 attenuated AD symptoms based on the ear thickness, histopathological analysis, and serum immunoglobulin levels. Moreover, EF-2001 decreased the DFE/DNCB-induced expression of various pathogenic cytokines in the ears, lymph nodes, and splenocytes. These results suggest that EF-2001 has therapeutic potential in the treatment of AD owing to its immunomodulatory effects. PMID:26959058

  9. Crystal structure of enterococcus faecalis sly A-like transcriptional factor.

    SciTech Connect

    Wu, R.; Zhang, R.; Zagnitko, O.; Dementieva, I.; Maltsev, N.; Watson, J. D.; Laskowski, R.; Gornicki, P.; Joachimiak, A.; Univ. of Chicago; European Bioinformatics Inst.

    2003-05-30

    The crystal structure of a SlyA transcriptional regulator at 1.6 {angstrom} resolution is presented, and structural relationships between members of the MarR/SlyA family are discussed. The SlyA family, which includes SlyA, Rap, Hor, and RovA proteins, is widely distributed in bacterial and archaeal genomes. Current evidence suggests that SlyA-like factors act as repressors, activators, and modulators of gene transcription. These proteins have been shown to up-regulate the expression of molecular chaperones, acid-resistance proteins, and cytolysin, and down-regulate several biosynthetic enzymes. The structure of SlyA from Enterococcus faecalis, determined as a part of an ongoing structural genomics initiative (www.mcsg.anl.gov), revealed the same winged helix DNA-binding motif that was recently found in the MarR repressor from Escherichia coli and the MexR repressor from Pseudomonas aeruginosa, a sequence homologue of MarR. Phylogenetic analysis of the MarR/SlyA family suggests that Sly is placed between the SlyA and MarR subfamilies and shows significant sequence similarity to members of both subfamilies.

  10. Insidious Onset of Tetraparesis due to Cervical Epidural Abscess from Enterococcus faecalis.

    PubMed

    Soultanis, Konstantinos Chr; Sakellariou, Vasileios I; Starantzis, Konstantinos A; Stavropoulos, Nikolaos A; Papagelopoulos, Panayiotis J

    2013-01-01

    We report a case of cervical epidural abscess from Enterococcus faecalis, which caused an insidious onset of tetraparesis. This 53-year-old female with a history of diabetes mellitus and chronic renal failure under hemodialysis presented with pain and progressive weakness of upper and lower extremities without fever. Although a recent MRI she did at the beginning of symptoms showed no significant pathologies, except for a cervical disc herniation and adjacent spinal degeneration, and stenosis that confused the diagnostic procedure, newer imaging with CT and MRI, which was performed due to progression of tetraparesis, revealed the formation of a cervical epidural abscess. Surgical drainage was done after a complete infection workup. The patient showed immediate neurological improvement after surgery. She received antibiotics intravenously for 3 weeks and orally for another 6 weeks. The patient was free from complications 24 months after surgery. A high index of suspicion is most important in making a rapid and correct diagnosis of spinal epidural abscess. The classic clinical triad (fever, local pain, and neurologic deficits) is not sensitive enough for early detection. Continuous clinical, laboratory, and imaging monitoring are of paramount importance. Early diagnosis and surgical intervention could optimize the final functional outcome. PMID:23573096

  11. Adaptation of Enterococcus faecalis to Daptomycin Reveals an Ordered Progression to Resistance

    PubMed Central

    Miller, Corwin; Kong, Jiayi; Tran, Truc T.; Arias, Cesar A.; Saxer, Gerda

    2013-01-01

    With increasing numbers of hospital-acquired antibiotic resistant infections each year and staggering health care costs, there is a clear need for new antimicrobial agents, as well as novel strategies to extend their clinical efficacy. While genomic studies have provided a wealth of information about the alleles associated with adaptation to antibiotics, they do not provide essential information about the relative importance of genomic changes, their order of appearance, or potential epistatic relationships between adaptive changes. Here we used quantitative experimental evolution of a single polymorphic population in continuous culture with whole-genome sequencing and allelic frequency measurements to study daptomycin (DAP) resistance in the vancomycin-resistant clinical pathogen Enterococcus faecalis S613. Importantly, we sustained both planktonic and nonplanktonic (i.e., biofilm) populations in coculture as the concentration of antibiotic was raised, facilitating the development of more ecological complexity than is typically observed in laboratory evolution. Quantitative experimental evolution revealed a clear order and hierarchy of genetic changes leading to resistance, the signaling and metabolic pathways responsible, and the relative importance of these mutations to the evolution of DAP resistance. Despite the relative simplicity of this ex vivo approach compared to the ecological complexity of the human body, we showed that experimental evolution allows for rapid identification of clinically relevant adaptive molecular pathways and new targets for drug design in pathogens. PMID:23959318

  12. Characterization of lead-resistant river isolate Enterococcus faecalis and assessment of its multiple metal and antibiotic resistance.

    PubMed

    Aktan, Yasin; Tan, Sema; Icgen, Bulent

    2013-06-01

    Contamination of surface waters has a direct impact on the public health of entire communities. Microorganisms inhabiting contaminated surface waters have developed mechanisms of coping with a variety of toxic metals and drugs. Investigations were carried out to isolate and identify lead-resistant bacteria from the river Kızılırmak along the city of Kırıkkale, Turkey. Of the 33 lead-resistant isolates, one isolate with a minimal inhibitory concentration of 1,200 mg L(-1) was isolated and identified as Enterococcus faecalis by using biochemical tests and 16S rRNA sequencing. Lead-resistant E. faecalis isolate was found out to be resistant to other heavy metals like aluminum, lithium, barium, chromium, iron, silver, tin, nickel, zinc, and strontium and to drugs like amikacin, aztreonam, and gentamicin. E. faecalis harbored four plasmids with the molecular sizes of 1.58, 3.06, 22.76, and 28.95 kb. Plasmid profile analyses of cured derivatives revealed that the lead resistance ability of E. faecalis was still existing despite the elimination of all the plasmids. Moreover, the antibiotic resistance pattern of the cured derivatives did not demonstrate any change from the parental strain. Our findings indicated that the lead resistance genes of E. faecalis were located on the chromosomal DNA rather than the plasmid. PMID:23079796

  13. Antimicrobial activity of some essential oils against oral multidrug-resistant Enterococcus faecalis in both planktonic and biofilm state

    PubMed Central

    Benbelaïd, Fethi; Khadir, Abdelmounaïm; Abdoune, Mohamed Amine; Bendahou, Mourad; Muselli, Alain; Costa, Jean

    2014-01-01

    Objective To evaluate some essential oils in treatment of intractable oral infections, principally caused by biofilm of multidrug-resistant Enterococcus faecalis (E. faecalis), such as persistent endodontic infections in which their treatment exhibits a real challenge for dentists. Methods Ten chemically analyzed essential oils by gas chromatography-mass spectrometry were evaluated for antimicrobial activity against sensitive and resistant clinical strains of E. faecalis in both planktonic and biofilm state using two methods, disk diffusion and broth micro-dilution. Results Studied essential oils showed a good antimicrobial activity and high ability in E. faecalis biofilm eradication, whether for sensitive or multidrug-resistant strains, especially those of Origanum glandulosum and Thymbra capitata with interesting minimum inhibitory concentration, biofilm inhibitory concentration, and biofilm eradication concentration values which doesn't exceed 0.063%, 0.75%, and 1.5%, respectively. Conclusions Findings of this study indicate that essential oils extracted from aromatic plants can be used in treatment of intractable oral infections, especially caused by biofilm of multidrug-resistant E. faecalis. PMID:25182948

  14. The polyamine N-acetyltransferase-like enzyme PmvE plays a role in the virulence of Enterococcus faecalis.

    PubMed

    Martini, Cecilia; Michaux, Charlotte; Bugli, Francesca; Arcovito, Alessandro; Iavarone, Federica; Cacaci, Margherita; Paroni Sterbini, Francesco; Hartke, Axel; Sauvageot, Nicolas; Sanguinetti, Maurizio; Posteraro, Brunella; Giard, Jean-Christophe

    2015-01-01

    We previously showed that the mutant strain of Enterococcus faecalis lacking the transcriptional regulator SlyA is more virulent than the parental strain. We hypothesized that this phenotype was due to overexpression of the second gene of the slyA operon, ef_3001, renamed pmvE (for polyamine metabolism and virulence of E. faecalis). PmvE shares strong homologies with N(1)-spermidine/spermine acetyltransferase enzymes involved in the metabolism of polyamines. In this study, we used an E. faecalis strain carrying the recombinant plasmid pMSP3535-pmvE (V19/p3535-pmvE), which allows the induction of pmvE by addition of nisin. Thereby, we showed that the overexpression of PmvE increased the virulence of E. faecalis in the Galleria mellonella infection model, as well as the persistence within peritoneal macrophages. We were also able to show a direct interaction between the His-tagged recombinant PmvE (rPmvE) protein and putrescine by the surface plasmon resonance (SPR) technique on a Biacore instrument. Moreover, biochemical assays showed that PmvE possesses an N-acetyltransferase activity toward polyamine substrates. Our results suggest that PmvE contributes to the virulence of E. faecalis, likely through its involvement in the polyamine metabolism. PMID:25385793

  15. Antibacterial and residual antimicrobial activities against Enterococcus faecalis biofilm: A comparison between EDTA, chlorhexidine, cetrimide, MTAD and QMix

    PubMed Central

    Zhang, Rui; Chen, Min; Lu, Yan; Guo, Xiangjun; Qiao, Feng; Wu, Ligeng

    2015-01-01

    We compared the antibacterial and residual antimicrobial activities of five root canal irrigants (17% EDTA,2% chlorhexidine,0.2% cetrimide, MTAD, and QMix) in a model of Enterococcus faecalis biofilm formation. Sixty dentin blocks with 3-week E. faecalis biofilm were divided into six equal groups and flushed with irrigant for 2 min or left untreated. A blank control group was also established. Antibacterial activities of the irrigants were evaluated by counting colony forming units. To test residual antimicrobial activities, 280 dentin blocks were divided into seven equal groups and flushed with irrigant for 2 min or left untreated and then incubated with E. faecalis suspension for 48 h, or used as a blank. No bacteria were observed in the blank control group. The number of viable E. faecalis was significantly fewer in the irrigant-treated groups compared with the untreated control (P < 0.05). Among the five irrigants, QMix had the strongest antibacterial activity. Residual antimicrobial activities of CHX were significantly higher at 12 h, 24 h and 36 h compared to untreated control (P < 0.05). All five root canal irrigants were effective to some extent against E. faecalis, but QMix and CHX had the strongest, and CHX the longest (up to 36 h), antimicrobial activity. PMID:26245711

  16. Antibacterial and residual antimicrobial activities against Enterococcus faecalis biofilm: A comparison between EDTA, chlorhexidine, cetrimide, MTAD and QMix.

    PubMed

    Zhang, Rui; Chen, Min; Lu, Yan; Guo, Xiangjun; Qiao, Feng; Wu, Ligeng

    2015-01-01

    We compared the antibacterial and residual antimicrobial activities of five root canal irrigants (17% EDTA,2% chlorhexidine,0.2% cetrimide, MTAD, and QMix) in a model of Enterococcus faecalis biofilm formation. Sixty dentin blocks with 3-week E. faecalis biofilm were divided into six equal groups and flushed with irrigant for 2 min or left untreated. A blank control group was also established. Antibacterial activities of the irrigants were evaluated by counting colony forming units. To test residual antimicrobial activities, 280 dentin blocks were divided into seven equal groups and flushed with irrigant for 2 min or left untreated and then incubated with E. faecalis suspension for 48 h, or used as a blank. No bacteria were observed in the blank control group. The number of viable E. faecalis was significantly fewer in the irrigant-treated groups compared with the untreated control (P < 0.05). Among the five irrigants, QMix had the strongest antibacterial activity. Residual antimicrobial activities of CHX were significantly higher at 12 h, 24 h and 36 h compared to untreated control (P < 0.05). All five root canal irrigants were effective to some extent against E. faecalis, but QMix and CHX had the strongest, and CHX the longest (up to 36 h), antimicrobial activity. PMID:26245711

  17. Isolation and identification of Enterococcus faecalis membrane proteins using membrane shaving, 1D SDS/PAGE, and mass spectrometry.

    PubMed

    Cathro, Peter; McCarthy, Peter; Hoffmann, Peter; Zilm, Peter

    2016-06-01

    Enterococcus faecalis is a significant nosocomial pathogen, which is able to survive in diverse environments and resist killing with antimicrobial therapies. The expression of cell membrane proteins play an important role in how bacteria respond to environmental stress. As such, the capacity to identify and study membrane protein expression is critical to our understanding of how specific proteins influence bacterial survival. Here, we describe a combined approach to identify membrane proteins of E. faecalis ATCC V583 using membranes fractionated by either 1D SDS/PAGE or membrane shaving, coupled with LC-ESI mass spectrometry. We identified 222 membrane-associated proteins, which represent approximately 24% of the predicted membrane-associated proteome: 170 were isolated using 1D SDS/PAGE and 68 with membrane shaving, with 36 proteins being common to both the techniques. Of the proteins identified by membrane shaving, 97% were membrane-associated with the majority being integral membrane proteins (89%). Most of the proteins identified with known physiology are involved with transportation across the membrane. The combined 1D SDS/PAGE and membrane shaving approach has produced the greatest number of membrane proteins identified from E. faecalis to date. These protocols will aid future researchers investigating changes in the membrane proteome of E. faecalis by improving our understanding of how E. faecalis adapts and responds to its environment. PMID:27419061

  18. Combined effect of a mixture of tetracycline, acid, and detergent, and Nisin against Enterococcus faecalis and Actinomyces viscosus biofilms

    PubMed Central

    Balto, Hanan A.; Shakoor, Zahid A.; Kanfar, Maha A.

    2015-01-01

    Objectives: To evaluate the combined effect of a mixture of tetracycline, acid, and detergent (MTAD) and Nisin against Enterococcus faecalis (E. faecalis) and Actinomyces viscosus (A. viscosus) biofilms. Methods: This study was conducted between June and December 2013 in collaboration with Dental Caries Research Chair, College of Dentistry, King Saud University, Riyadh, Saudi Arabia. Single-species biofilms (n=9/species/observation period) were generated on membrane filter discs and subjected to 5, 10, or 15 minute incubation with MTADN (MTAD with 3% Nisin), 5.25% sodium hypochlorite (NaOCl), or normal saline. The colony forming units were counted using the Dark field colony counter. Results: A 100% bactericidal effect of 5.25% NaOCl was noted during the 3 observation periods; a significant reduction (p=0.000) in mean survival rates of E. faecalis (77.3+13.6) and A. viscosus (39.6+12.6) was noted after 5 minutes exposure to MTADN compared with normal saline (78000000+5291503) declining to almost no growth after 10 and 15 minutes. The survival rates of the E. faecalis and A. viscosus biofilm were no different after treatment with MTADN and 5.25% NaOCl at the 3 observation periods (p=1.000). Conclusion: A combination of MTAD and Nisin was as effective as NaOCl against E. faecalis and A. viscosus biofilms. PMID:25719587

  19. Antibacterial and residual antimicrobial activities against Enterococcus faecalis biofilm: A comparison between EDTA, chlorhexidine, cetrimide, MTAD and QMix

    NASA Astrophysics Data System (ADS)

    Zhang, Rui; Chen, Min; Lu, Yan; Guo, Xiangjun; Qiao, Feng; Wu, Ligeng

    2015-08-01

    We compared the antibacterial and residual antimicrobial activities of five root canal irrigants (17% EDTA,2% chlorhexidine,0.2% cetrimide, MTAD, and QMix) in a model of Enterococcus faecalis biofilm formation. Sixty dentin blocks with 3-week E. faecalis biofilm were divided into six equal groups and flushed with irrigant for 2 min or left untreated. A blank control group was also established. Antibacterial activities of the irrigants were evaluated by counting colony forming units. To test residual antimicrobial activities, 280 dentin blocks were divided into seven equal groups and flushed with irrigant for 2 min or left untreated and then incubated with E. faecalis suspension for 48 h, or used as a blank. No bacteria were observed in the blank control group. The number of viable E. faecalis was significantly fewer in the irrigant-treated groups compared with the untreated control (P < 0.05). Among the five irrigants, QMix had the strongest antibacterial activity. Residual antimicrobial activities of CHX were significantly higher at 12 h, 24 h and 36 h compared to untreated control (P < 0.05). All five root canal irrigants were effective to some extent against E. faecalis, but QMix and CHX had the strongest, and CHX the longest (up to 36 h), antimicrobial activity.

  20. An in vitro study on the effects of nisin on the antibacterial activities of 18 antibiotics against Enterococcus faecalis.

    PubMed

    Tong, Zhongchun; Zhang, Yuejiao; Ling, Junqi; Ma, Jinglei; Huang, Lijia; Zhang, Luodan

    2014-01-01

    Enterococcus faecalis rank among the leading causes of nosocomial infections worldwide and possesses both intrinsic and acquired resistance to a variety of antibiotics. Development of new antibiotics is limited, and pathogens continually generate new antibiotic resistance. Many researchers aim to identify strategies to effectively kill this drug-resistant pathogen. Here, we evaluated the effect of the antimicrobial peptide nisin on the antibacterial activities of 18 antibiotics against E. faecalis. The MIC and MBC results showed that the antibacterial activities of 18 antibiotics against E. faecalis OG1RF, ATCC 29212, and strain E were significantly improved in the presence of 200 U/ml nisin. Statistically significant differences were observed between the results with and without 200 U/ml nisin at the same concentrations of penicillin or chloramphenicol (p<0.05). The checkerboard assay showed that the combination of nisin and penicillin or chloramphenicol had a synergetic effect against the three tested E. faecalis strains. The transmission electron microscope images showed that E. faecalis was not obviously destroyed by penicillin or chloramphenicol alone but was severely disrupted by either antibiotic in combination with nisin. Furthermore, assessing biofilms by a confocal laser scanning microscope showed that penicillin, ciprofloxacin, and chloramphenicol all showed stronger antibiofilm actions in combination with nisin than when these antibiotics were administered alone. Therefore, nisin can significantly improve the antibacterial and antibiofilm activities of many antibiotics, and certain antibiotics in combination with nisin have considerable potential for use as inhibitors of this drug-resistant pathogen. PMID:24586598

  1. Transcriptome Analysis of Enterococcus faecalis during Mammalian Infection Shows Cells Undergo Adaptation and Exist in a Stringent Response State

    PubMed Central

    Frank, Kristi L.; Colomer-Winter, Cristina; Grindle, Suzanne M.; Lemos, José A.; Schlievert, Patrick M.; Dunny, Gary M.

    2014-01-01

    As both a commensal and a major cause of healthcare-associated infections in humans, Enterococcus faecalis is a remarkably adaptable organism. We investigated how E. faecalis adapts in a mammalian host as a pathogen by characterizing changes in the transcriptome during infection in a rabbit model of subdermal abscess formation using transcriptional microarrays. The microarray experiments detected 222 and 291 differentially regulated genes in E. faecalis OG1RF at two and eight hours after subdermal chamber inoculation, respectively. The profile of significantly regulated genes at two hours post-inoculation included genes involved in stress response, metabolism, nutrient acquisition, and cell surface components, suggesting genome-wide adaptation to growth in an altered environment. At eight hours post-inoculation, 88% of the differentially expressed genes were down-regulated and matched a transcriptional profile consistent with a (p)ppGpp-mediated stringent response. Subsequent subdermal abscess infections with E. faecalis mutants lacking the (p)ppGpp synthetase/hydrolase RSH, the small synthetase RelQ, or both enzymes, suggest that intracellular (p)ppGpp levels, but not stringent response activation, influence persistence in the model. The ability of cells to synthesize (p)ppGpp was also found to be important for growth in human serum and whole blood. The data presented in this report provide the first genome-wide insights on E. faecalis in vivo gene expression and regulation measured by transcriptional profiling during infection in a mammalian host and show that (p)ppGpp levels affect viability of E. faecalis in multiple conditions relevant to mammalian infection. The subdermal abscess model can serve as a novel experimental system for studying the E. faecalis stringent response in the context of the mammalian immune system. PMID:25545155

  2. Native Microbial Colonization of Drosophila melanogaster and Its Use as a Model of Enterococcus faecalis Pathogenesis▿ †

    PubMed Central

    Cox, Christopher R.; Gilmore, Michael S.

    2007-01-01

    Enterococci are commensal organisms of the gastrointestinal (GI) tracts of a broad range of mammalian and insect hosts, but they are also leading causes of nosocomial infection. Little is known about the ecological role of enterococci in the GI tract consortia. To develop a tractable model for studying the roles of these organisms as commensals and pathogens, we characterized the Drosophila melanogaster microflora and examined the occurrence of enterococci in the gastrointestinal consortium of Drosophila. In a survey of laboratory-reared Drosophila and wild-captured flies, we found that Drosophila was naturally colonized by representatives of five bacterial phyla. Among these organisms were several species of enterococci, including Enterococcus faecalis, Enterococcus faecium, Enterococcus gallinaraum, and Enterococcus durans, as well as a previously detected but uncultured Enterococcus species. Drosophila could be cured of enterococcal carriage by antibiotic treatment and could be reassociated with laboratory strains. High-level colonization by a well-characterized strain expressing the enterococcal cytolysin was found to be detrimental to Drosophila compared to the effect of an isogenic, noncytolytic control. The anatomical distribution of enterococci in the Drosophila GI tract was determined by immunohistochemical staining of thin sections of naturally colonized and reassociated flies. PMID:17220307

  3. Biofilm formation on polystyrene under different temperatures by antibiotic resistant Enterococcus faecalis and Enterococcus faecium isolated from food

    PubMed Central

    Marinho, A.R.; Martins, P.D.; Ditmer, E.M.; d’Azevedo, P.A.; Frazzon, J.; Van Der Sand, S.T.; Frazzon, A.P.G.

    2013-01-01

    The ability of antibiotic resistant E. faecalis and E. faecium isolated from food to form biofilm at different temperatures in the absence or presence of 0.75% glucose was evaluated. A synergistic effect on biofilm at 10 °C, 28 °C, 37 °C and 45 °C and glucose was observed for E. faecalis and E. faecium. PMID:24294231

  4. Defining Daptomycin Resistance Prevention Exposures in Vancomycin-Resistant Enterococcus faecium and E. faecalis

    PubMed Central

    Werth, B. J.; Steed, M. E.; Ireland, C. E.; Tran, T. T.; Nonejuie, P.; Murray, B. E.; Rose, W. E.; Sakoulas, G.; Pogliano, J.; Arias, C. A.

    2014-01-01

    Daptomycin is used off-label for enterococcal infections; however, dosing targets for resistance prevention remain undefined. Doses of 4 to 6 mg/kg of body weight/day approved for staphylococci are likely inadequate against enterococci due to reduced susceptibility. We modeled daptomycin regimens in vitro to determine the minimum exposure to prevent daptomycin resistance (Dapr) in enterococci. Daptomycin simulations of 4 to 12 mg/kg/day (maximum concentration of drug in serum [Cmax] of 57.8, 93.9, 123.3, 141.1, and 183.7 mg/liter; half-life [t1/2] of 8 h) were tested against one Enterococcus faecium strain (S447) and one Enterococcus faecalis strain (S613) in a simulated endocardial vegetation pharmacokinetic/pharmacodynamic model over 14 days. Samples were plated on media containing 3× the MIC of daptomycin to detect Dapr. Mutations in genes encoding proteins associated with cell envelope homeostasis (yycFG and liaFSR) and phospholipid metabolism (cardiolipin synthase [cls] and cyclopropane fatty acid synthetase [cfa]) were investigated in Dapr derivatives. Dapr derivatives were assessed for changes in susceptibility, surface charge, membrane depolarization, cell wall thickness (CWT), and growth rate. Strains S447 and S613 developed Dapr after simulations of 4 to 8 mg/kg/day but not 10 to 12 mg/kg/day. MICs for Dapr strains ranged from 8 to 256 mg/liter. Some S613 derivatives developed mutations in liaF or cls. S447 derivatives lacked mutations in these genes. Dapr derivatives from both strains exhibited lowered growth rates, up to a 72% reduction in daptomycin-induced depolarization and up to 6-nm increases in CWT (P < 0.01). Peak/MIC and AUC0–24/MIC ratios (AUC0–24 is the area under the concentration-time curve from 0 to 24 h) associated with Dapr prevention were 72.1 and 780 for S447 and 144 and 1561 for S613, respectively. Daptomycin doses of 10 mg/kg/day may be required to prevent Dapr in serious enterococcal infections. PMID:24957825

  5. Cyclooxygenase-2 Generates the Endogenous Mutagen trans-4-Hydroxy-2-nonenal in Enterococcus faecalis-infected Macrophages

    PubMed Central

    Wang, Xingmin; Allen, Toby D.; Yang, Yonghong; Moore, Danny R.; Huycke, Mark M.

    2013-01-01

    Infection of macrophages by the human intestinal commensal Enterococcus faecalis generates DNA damage and chromosomal instability in mammalian cells through bystander effects. These effects are characterized by clastogenesis and damage to mitotic spindles in target cells and are mediated, in part, by trans-4-hydroxy-2-nonenal (4-HNE). In this study we investigated the role of cyclooxygenase (COX) and lipoxygenase (LOX) in producing this reactive aldehyde using E. faecalis-infected macrophages and interleukin-10 knockout mice colonized with this commensal. 4-HNE production by E. faecalis-infected macrophages was significantly reduced by COX and LOX inhibitors. The infection of macrophages led to decreased Cox1 and Alox5 expression while COX-2 and 4-HNE increased. Silencing Alox5 and Cox1 with gene-specific siRNAs had no effect on 4-HNE production. In contrast, silencing Cox2 significantly decreased 4-HNE production by E. faecalis-infected macrophages. Depleting intracellular glutathione increased 4-HNE production by these cells. Next, to confirm COX-2 as a source for 4-HNE, we assayed the products generated by recombinant human COX-2 and found 4-HNE in a concentration-dependent manner using arachidonic acid as a substrate. Finally, tissue macrophages in colon biopsies from interleukin-10 knockout mice colonized with E. faecalis were positive for COX-2 by immunohistochemical staining. This was associated with increased staining for 4-HNE-protein adducts in surrounding stroma. These data show that E. faecalis, a human intestinal commensal, can trigger macrophages to produce 4-HNE through COX-2. Importantly, it reinforces the concept of COX-2 as a procarcinogenic enzyme capable of damaging DNA in target cells through bystander effects that contribute to colorectal carcinogenesis. PMID:23321929

  6. Enterococcus faecalis inhibits superantigen toxic shock syndrome toxin-1-induced interleukin-8 from human vaginal epithelial cells through tetramic acids.

    PubMed

    Brosnahan, Amanda J; Merriman, Joseph A; Salgado-Pabón, Wilmara; Ford, Bradley; Schlievert, Patrick M

    2013-01-01

    The vaginal mucosa can be colonized by many bacteria including commensal organisms and potential pathogens, such as Staphylococcus aureus. Some strains of S. aureus produce the superantigen toxic shock syndrome toxin-1, which can penetrate the vaginal epithelium to cause toxic shock syndrome. We have observed that a female was mono-colonized with Enterococcus faecalis vaginally as tested in aerobic culture, even upon repeated culture for six months, suggesting this organism was negatively influencing colonization by other bacteria. In recent studies, we demonstrated an "outside-in" mechanism of cytokine signaling and consequent inflammation that facilitates the ability of potential pathogens to initiate infection from mucosal surfaces. Thus, we hypothesized that this strain of E. faecalis may make anti-inflammatory factors which block disease progression of more pathogenic organisms. E. faecalis MN1 inhibited interleukin-8 production from human vaginal epithelial cells in response to the vaginal pathogens Candida albicans, Gardnerella vaginalis, and Neisseria gonorrhoeae, as well as to toxic shock syndrome toxin-1. We further demonstrated that this organism secretes two tetramic acid compounds which appear responsible for inhibition of interleukin-8 production, as well as inhibition of T cell proliferation due to toxic shock syndrome toxin-1. Microbicides that include anti-inflammatory molecules, such as these tetramic acid compounds naturally produced by E. faecalis MN1, may be useful in prevention of diseases that develop from vaginal infections. PMID:23613823

  7. Antimicrobial Effect of Lippia sidoides and Thymol on Enterococcus faecalis Biofilm of the Bacterium Isolated from Root Canals

    PubMed Central

    Veras, H. N. H.; Rodrigues, F. F. G.; Botelho, M. A.; Menezes, I. R. A.; Coutinho, H. D. M.; da Costa, J. G. M.

    2014-01-01

    The species Lippia sidoides Cham. (Verbenaceae) is utilized in popular medicine as a local antiseptic on the skin and mucosal tissues. Enterococcus faecalis is the bacterium isolated from root canals of teeth with persistent periapical lesions and has the ability to form biofilm, where it is responsible for the failure of endodontic treatments. Essential oil of L. sidoides (EOLS) and its major component, thymol, were evaluated for reducing the CFU in biofilms of E. faecalis in vitro. The essential oil was obtained by hydrodistillation and examined with respect to the chemical composition, by gas chromatography-mass spectrometry (GC-MS). The GC-MS analysis has led to the identification of thymol (84.9%) and p-cymene (5.33%). EOLS and thymol reduced CFU in biofilms of E. faecalis in vitro (time of maturation, 72 h), with an exposure time of 30 and 60 min at concentrations of 2.5 and 10%. There was no statistical difference in effect between EOLS and thymol, demonstrating that this phenolic monoterpene was the possible compound responsible for the antimicrobial activity of EOLS. This study provides a basis for the possible utilization of EOLS as an adjuvant in the treatment of root canals that show colonization by E. faecalis. PMID:24683344

  8. Sunlight mediated inactivation mechanisms of Enterococcus faecalis and Escherichia coli in clear water versus waste stabilization pond water.

    PubMed

    Kadir, Khalid; Nelson, Kara L

    2014-03-01

    Escherichia coli and enterococci have been previously reported to differ in the mechanisms and conditions that affect their sunlight-mediated inactivation in waste stabilization ponds. This study was undertaken to further characterize these mechanisms, using simulated sunlight and single strains of laboratory-grown E. coli and Enterococcus faecalis, with a focus on characterizing the contribution of exogenous reactive oxygen species to the inactivation process. We found that direct damage by UVB light (280-320 nm) was not a significant inactivation mechanism for either organism. E. coli inactivation was strongly dependent on dissolved oxygen concentrations and the presence of UVB wavelengths but E. coli were not susceptible to inactivation by exogenous sensitizers present in waste stabilization pond water. In contrast, E. faecalis inactivation in pond water occurred primarily through exogenous mechanisms, with strong evidence that singlet oxygen is an important transient reactive species. The exogenous mechanism could utilize wavelengths into the visible spectrum and sensitizers were mainly colloidal, distributed between 0.2 and ∼1 μm in size. Singlet oxygen is likely an important endogenous species in both E. faecalis and E. coli inactivation due to sunlight. Although the two organisms had similar inactivation rates in buffered, clear water, the inactivation rate of E. faecalis was 7 times greater than that of E. coli in air-saturated pond water at circumneutral pH due to its susceptibility to exogenous sensitizers and longer wavelengths. PMID:24188579

  9. Immunostimulatory Effects Triggered by Enterococcus faecalis CECT7121 Probiotic Strain Involve Activation of Dendritic Cells and Interferon-Gamma Production.

    PubMed

    Molina, Matías Alejandro; Díaz, Ailén Magalí; Hesse, Christina; Ginter, Wiebke; Gentilini, María Virginia; Nuñez, Guillermo Gabriel; Canellada, Andrea Mercedes; Sparwasser, Tim; Berod, Luciana; Castro, Marisa Silvia; Manghi, Marcela Alejandra

    2015-01-01

    Probiotics can modulate the immune system, conferring beneficial effects on the host. Understanding how these microorganisms contribute to improve the health status is still a challenge. Previously, we have demonstrated that Enterococcus faecalis CECT7121 implants itself and persists in the murine gastrointestinal tract, and enhances and skews the profile of cytokines towards the Th1 phenotype in several biological models. Given the importance of dendritic cells (DCs) in the orchestration of immunity, the aim of this work was to elucidate the influence of E. faecalis CECT7121 on DCs and the outcome of the immune responses. In this work we show that E. faecalis CECT7121 induces a strong dose-dependent activation of DCs and secretion of high levels of IL-12, IL-6, TNFα, and IL-10. This stimulation is dependent on TLR signaling, and skews the activation of T cells towards the production of IFNγ. The influence of this activation in the establishment of Th responses in vivo shows the accumulation of specific IFNγ-producing cells. Our findings indicate that the activation exerted by E. faecalis CECT7121 on DCs and its consequence on the cellular adaptive immune response may have broad therapeutic implications in immunomodulation. PMID:25978357

  10. Tyramine biosynthesis is transcriptionally induced at low pH and improves the fitness of Enterococcus faecalis in acidic environments.

    PubMed

    Perez, Marta; Calles-Enríquez, Marina; Nes, Ingolf; Martin, Maria Cruz; Fernandez, Maria; Ladero, Victor; Alvarez, Miguel A

    2015-04-01

    Enterococcus faecalis is a commensal bacterium of the human gut that requires the ability to pass through the stomach and therefore cope with low pH. E. faecalis has also been identified as one of the major tyramine producers in fermented food products, where they also encounter acidic environments. In the present work, we have constructed a non-tyramine-producing mutant to study the role of the tyramine biosynthetic pathway, which converts tyrosine to tyramine via amino acid decarboxylation. Wild-type strain showed higher survival in a system that mimics gastrointestinal stress, indicating that the tyramine biosynthetic pathway has a role in acid resistance. Transcriptional analyses of the E. faecalis V583 tyrosine decarboxylase cluster showed that an acidic pH, together with substrate availability, induces its expression and therefore the production of tyramine. The protective role of the tyramine pathway under acidic conditions appears to be exerted through the maintenance of the cytosolic pH. Tyramine production should be considered important in the adaptability of E. faecalis to acidic environments, such as fermented dairy foods, and to survive passage through the human gastrointestinal tract. PMID:25529314

  11. IS256 abolishes gelatinase activity and biofilm formation in a mutant of the nosocomial pathogen Enterococcus faecalis V583.

    PubMed

    Perez, Marta; Calles-Enríquez, Marina; del Rio, Beatriz; Ladero, Victor; Martín, María Cruz; Fernández, María; Alvarez, Miguel A

    2015-07-01

    Enterococcus faecalis is one of the most controversial species of lactic acid bacteria. Some strains are used as probiotics, while others are associated with severe and life-threatening nosocomial infections. Their pathogenicity depends on the acquisition of multidrug resistance and virulence factors. Gelatinase, which is required in the first steps of biofilm formation, is an important virulence determinant involved in E. faecalis pathogenesis, including endocarditis and peritonitis. The gene that codes for gelatinase (gelE) is controlled by the Fsr quorum-sensing system, whose encoding genes (fsrA, fsrB, fsrC, and fsrD) are located immediately upstream of gelE. The integration of a DNA fragment into the fsr locus of a derived mutant of E. faecalis V583 suppressed the gelatinase activity and prevented biofilm formation. Sequence analysis indicated the presence of IS256 integrated into the fsrC gene at nucleotide position 321. Interestingly, IS256 is also associated with biofilm formation in Staphylococcus epidermidis and Staphylococcus aureus. This is the first description of an insertion sequence that prevents biofilm formation in E. faecalis. PMID:25966618

  12. Comparative evaluation of antimicrobial efficacy of QMix™ 2 in 1, sodium hypochlorite, and chlorhexidine against Enterococcus faecalis and Candida albicans

    PubMed Central

    Elakanti, Soujanya; Cherukuri, Gayathri; Rao, Venkateswara G; Chandrasekhar, Veeramachaneni; Rao, Anitha S; Tummala, Muralidhar

    2015-01-01

    Aim/Objective: The aim of this study is to compare the antimicrobial efficacy of QMix™ 2 in 1, sodium hypochlorite (NaOCl), and chlorhexidine (CHX) against Enterococcus faecalis and Candida albicans. Materials and Methods: Eighty freshly extracted, single-rooted human mandibular premolar teeth were instrumented and autoclaved. Samples were divided into two groups of 40 teeth each based on the type of microorganism used. Group I was inoculated with E. faecalis and Group II with C. albicans and incubated for 3 days. Each group was subdivided into four subgroups based on the type of irrigant used. Group IA, IIA, 5.25% NaOCl; Group IB, IIB, 2% CHX; Group IC, IIC, QMix™ 2 in 1; and Group ID, IID, 0.9% saline (the control group). Ten microliters of the sample from each canal was taken and was placed on Brain Heart Infusion agar and Sabouraud dextrose agar. The plates were incubated at 37°C for 24 h and colony forming units (CFUs) that were grown were counted. Data was analyzed with analysis of variance (ANOVA) followed by post-hoc Games-Howell test. Results: The greatest antimicrobial effects were observed in samples treated with QMix™ 2 in 1 (P < 0.001). No statistical significant difference was found between 5.25% NaOCl and 2% CHX (P > 0.001) against E. faecalis and C. albicans. Conclusion: QMix™ 2 in 1 demonstrated significant antimicrobial efficacy against E. faecalis and C. albicans. PMID:25829691

  13. Immunostimulatory Effects Triggered by Enterococcus faecalis CECT7121 Probiotic Strain Involve Activation of Dendritic Cells and Interferon-Gamma Production

    PubMed Central

    Molina, Matías Alejandro; Díaz, Ailén Magalí; Hesse, Christina; Ginter, Wiebke; Gentilini, María Virginia; Nuñez, Guillermo Gabriel; Canellada, Andrea Mercedes; Sparwasser, Tim; Berod, Luciana; Castro, Marisa Silvia; Manghi, Marcela Alejandra

    2015-01-01

    Probiotics can modulate the immune system, conferring beneficial effects on the host. Understanding how these microorganisms contribute to improve the health status is still a challenge. Previously, we have demonstrated that Enterococcus faecalis CECT7121 implants itself and persists in the murine gastrointestinal tract, and enhances and skews the profile of cytokines towards the Th1 phenotype in several biological models. Given the importance of dendritic cells (DCs) in the orchestration of immunity, the aim of this work was to elucidate the influence of E. faecalis CECT7121 on DCs and the outcome of the immune responses. In this work we show that E. faecalis CECT7121 induces a strong dose-dependent activation of DCs and secretion of high levels of IL-12, IL-6, TNFα, and IL-10. This stimulation is dependent on TLR signaling, and skews the activation of T cells towards the production of IFNγ. The influence of this activation in the establishment of Th responses in vivo shows the accumulation of specific IFNγ-producing cells. Our findings indicate that the activation exerted by E. faecalis CECT7121 on DCs and its consequence on the cellular adaptive immune response may have broad therapeutic implications in immunomodulation. PMID:25978357

  14. Transferable plasmid-mediated resistance to linezolid due to cfr in a human clinical isolate of Enterococcus faecalis.

    PubMed

    Diaz, Lorena; Kiratisin, Pattarachai; Mendes, Rodrigo E; Panesso, Diana; Singh, Kavindra V; Arias, Cesar A

    2012-07-01

    Nonmutational resistance to linezolid is due to the presence of cfr, which encodes a methyltransferase responsible for methylation of A2503 in the 23S rRNA. The cfr gene was first described in animal isolates of staphylococci, and more recently, it has been identified in Staphylococcus aureus from human clinical infections, including in an outbreak of methicillin-resistant S. aureus. In enterococci, cfr has been described in an animal isolate of Enterococcus faecalis from China. Here, we report an isolate of linezolid-resistant E. faecalis (603-50427X) recovered from a patient in Thailand who received prolonged therapy with the antibiotic for the treatment of atypical mycobacterial disease. The isolate lacked mutations in the genes coding for 23S rRNA and L3 and L4 ribosomal proteins and belonged to the multilocus sequence type (MLST) 16 (ST16), which is commonly found in enterococcal isolates from animal sources. Resistance to linezolid was associated with the presence of cfr on an ~97-kb transferable plasmid. The cfr gene environment exhibited DNA sequences similar to those of other cfr-carrying plasmids previously identified in staphylococci (nucleotide identity, 99 to 100%). The cfr-carrying plasmid was transferable by conjugation to a laboratory strain of E. faecalis (OG1RF) but not to Enterococcus faecium or S. aureus. The cfr gene was flanked by IS256-like sequences both upstream and downstream. This is the first characterization of the potential horizontal transferability of the cfr gene from a human linezolid-resistant isolate of E. faecalis. PMID:22491691

  15. Ex situ study of Enterococcus faecalis survival in the recreational waters of the southern coast of the Caspian Sea

    PubMed Central

    Irankhah, Sahar; Soudi, Mohammad Reza; Gharavi, Sara

    2016-01-01

    Background and Objectives: The US Environmental Protection Agency has suggested faecal enterococci as the primary bacterial indicators. Of more importance is their direct correlation with swimmer-associated gastroenteritis in recreation water quality monitoring. In contrast to other seawater bodies with 3.5% salinity, the recreational waters in the southern coast of the Caspian Sea possess its own salinity (about 1% w/v) and thus require further investigations to determine the capacity of Enterococcus faecalis as the sole primary microbial index in this unique aquatic environment. Materials and Methods: The survey of the presence and survival of E. faecalis as a microbial index in the recreational waters of the southern Caspian Sea was carried out using a microcosm as an experimental model. The concentration of E. faecalis cells in samples of seawater were estimated by a standard membrane filtration method using m-Enterococcus agar as the selective culture medium. As the current standard culture-based methods are not reliable enough for the detection of non-growing, damaged and under-tension bacteria, PCR was used to identify the possible VBNC form of the bacterium after disappearance of the culturable cells. Results and Conclusion: A continuous decline in the number of culturable E. faecalis cells resulted in apparent elimination of the bacteria from seawater in a defined period. Detection of intact DNA was possible in the following 60 days. The salinity of about 1% and the self-purification properties of the Caspian Sea make the conditions feasible for the use of this microorganism as a measure of water quality throughout the region. The results confirmed the presence of damaged bacterial cells, namely VBNC forms, indicating the necessity of examining of the sea water samples by using molecular approaches or repair procedures. PMID:27307975

  16. Escherichia coli and Enterococcus faecalis are able to incorporate and enhance a pre-formed Gardnerella vaginalis biofilm.

    PubMed

    Castro, Joana; Machado, Daniela; Cerca, Nuno

    2016-04-01

    Gardnerella vaginalis is the most frequent microorganism found in bacterial vaginosis (BV), while Escherichia coli and Enterococcus faecalis are amongst the most frequent pathogens found in urinary tract infections (UTIs). This study aimed to evaluate possible interactions between UTIs pathogens and G. vaginalis using an in vitro dual-species biofilm model. Our results showed that dual-species biofilms reached significantly higher bacterial concentration than monospecies biofilms. Moreover, visualization of dual-populations species in the biofilms, using the epifluorescence microscopy, revealed that all of the urogenital pathogens coexisted with G. vaginalis. In conclusion, our work demonstrates that uropathogens can incorporate into mature BV biofilms. PMID:26782142

  17. Heterologous expression and characterization of tyrosine decarboxylase from Enterococcus faecalis R612Z1 and Enterococcus faecium R615Z1.

    PubMed

    Liu, Fang; Xu, Wenjuan; Du, Lihui; Wang, Daoying; Zhu, Yongzhi; Geng, Zhiming; Zhang, Muhan; Xu, Weimin

    2014-04-01

    Tyrosine decarboxylase (TDC) is responsible for tyramine production and can catalyze phenylalanine to produce β-phenylethylamine. Enterococcus strains are a group of bacteria predominantly producing tyramine and β-phenylethylamine in water-boiled salted duck. In this study, the heterologous expression and characterization of two TDCs from Enterococcus faecalis R612Z1 (612TDC) and Enterococcus faecium R615Z1 (615TDC) were studied. The recombinant putative proteins of 612TDC and 615TDC were heterologously expressed in Escherichia coli. 612TDC is a 620-amino-acid protein with a molecular mass of 70.0 kDa, whereas 615TDC is a 625-amino-acid protein with a molecular mass of 70.3 kDa. Both 612TDC and 615TDC showed an optimum temperature of 25 °C for the tyrosine and phenylalanine substrates. However, 612TDC revealed maximal activity at pH 5.5, whereas 615TDC revealed maximal activity at pH 6.0. Kinetic studies showed that 612TDC and 615TDC exhibited higher specificity for tyrosine than for phenylalanine. The catalysis abilities of both 612TDC and 615TDC for phenylalanine were restrained significantly with the increase in NaCl concentration, but this was not the case for tyrosine. This study revealed that the enzyme properties of the purified recombinant 612TDC and 615TDC were similar, although their amino acid sequences had 84% identity. PMID:24680070

  18. RNA of Enterococcus faecalis Strain EC-12 Is a Major Component Inducing Interleukin-12 Production from Human Monocytic Cells.

    PubMed

    Nishibayashi, Ryoichiro; Inoue, Ryo; Harada, Yuri; Watanabe, Takumi; Makioka, Yuko; Ushida, Kazunari

    2015-01-01

    Interleukin-12 (IL-12) is an important cytokine for the immunomodulatory effects of lactic acid bacteria (LAB). Using murine immune cells, we previously reported that the RNA of Enterococcus faecalis EC-12, a LAB strain exerting probiotic-like beneficial effects, is the major IL-12-inducing immunogenic component. However, it was recently revealed that bacterial RNA can be a ligand for Toll-like receptor (TLR) 13, which is only expressed in mice. Because TLR13 is not expressed in humans, the immuno-stimulatory and -modulatory effects of LAB RNA in human cells should be augmented excluding TLR13 contribution. In experiment 1 of this study, the role of LAB RNA in IL-12 induction in human immune cells was studied using three LAB strains, E.faecalis EC-12, Lactobacillus gasseri JCM5344, and Bifidobacterium breve JCM1192. RNase A treatment of heat-killed LAB significantly decreased the IL-12 production of human peripheral blood mononuclear cells on stimulation, while RNase III treatment revealed virtually no effects. Further, IL-12 production against heat-killed E. faecalis EC-12 was abolished by depleting monocytes. These results demonstrated that single stranded RNA (ssRNA) of LAB is a strong inducer of IL-12 production from human monocytes. In experiment 2, major receptor for ssRNA of E. faecalis EC-12 was identified using THP-1 cells, a human monocytic cell line. The type of RNA molecules of E. faecalis EC-12 responsible for IL-12 induction was also identified. IL-12 production induced by the total RNA of E. faecalis EC-12 was significantly reduced by the treatment of siRNA for TLR8 but not for TLR7. Furthermore, both 23S and 16S rRNA, but not mRNA, of E. faecalis EC-12 markedly induced IL-12 production from THP-1 cells. These results suggested that the recognition of ssRNA of E. faecalis EC-12 was mediated by TLR8 and that rRNA was the RNA molecule that exhibited IL-12-inducing ability in human cells. PMID:26083838

  19. RNA of Enterococcus faecalis Strain EC-12 Is a Major Component Inducing Interleukin-12 Production from Human Monocytic Cells

    PubMed Central

    Nishibayashi, Ryoichiro; Inoue, Ryo; Harada, Yuri; Watanabe, Takumi; Makioka, Yuko; Ushida, Kazunari

    2015-01-01

    Interleukin-12 (IL-12) is an important cytokine for the immunomodulatory effects of lactic acid bacteria (LAB). Using murine immune cells, we previously reported that the RNA of Enterococcus faecalis EC-12, a LAB strain exerting probiotic-like beneficial effects, is the major IL-12-inducing immunogenic component. However, it was recently revealed that bacterial RNA can be a ligand for Toll-like receptor (TLR) 13, which is only expressed in mice. Because TLR13 is not expressed in humans, the immuno-stimulatory and -modulatory effects of LAB RNA in human cells should be augmented excluding TLR13 contribution. In experiment 1 of this study, the role of LAB RNA in IL-12 induction in human immune cells was studied using three LAB strains, E.faecalis EC-12, Lactobacillus gasseri JCM5344, and Bifidobacterium breve JCM1192. RNase A treatment of heat-killed LAB significantly decreased the IL-12 production of human peripheral blood mononuclear cells on stimulation, while RNase III treatment revealed virtually no effects. Further, IL-12 production against heat-killed E. faecalis EC-12 was abolished by depleting monocytes. These results demonstrated that single stranded RNA (ssRNA) of LAB is a strong inducer of IL-12 production from human monocytes. In experiment 2, major receptor for ssRNA of E. faecalis EC-12 was identified using THP-1 cells, a human monocytic cell line. The type of RNA molecules of E. faecalis EC-12 responsible for IL-12 induction was also identified. IL-12 production induced by the total RNA of E. faecalis EC-12 was significantly reduced by the treatment of siRNA for TLR8 but not for TLR7. Furthermore, both 23S and 16S rRNA, but not mRNA, of E. faecalis EC-12 markedly induced IL-12 production from THP-1 cells. These results suggested that the recognition of ssRNA of E. faecalis EC-12 was mediated by TLR8 and that rRNA was the RNA molecule that exhibited IL-12-inducing ability in human cells. PMID:26083838

  20. Incongruence between the cps type 2 genotype and host-related phenotypes of an Enterococcus faecalis food isolate.

    PubMed

    Gaspar, Frédéric Bustos; Montero, Natalia; Akary, Elodie; Teixeira, Neuza; Matos, Renata; Gonzalez-Zorn, Bruno; Barreto Crespo, Maria Teresa; Serror, Pascale; Silva Lopes, Maria de Fátima

    2012-08-17

    Enterococcus faecalis is a nosocomial opportunistic pathogen, but is also found in fermented food products where it plays a fundamental role in the fermentation process. Previously, we have described the non-starter E. faecalis cheese isolate QA29b as harboring virulence genes and proven to be virulent in Galleria mellonella virulence model. In this study, we further characterized this food strain concerning traits relevant for the host-pathogen relationship. QA29b was found to belong to sequence type (ST) 72, a common ST among food isolates, and thus we consider it as a good representative of food E. faecalis strains. It demonstrated high ability to form biofilms, to adhere to epithelial cells and was readily eliminated by J774.A1 macrophage cells. Despite carrying the cps locus associated with the capsular polysaccharide CPS 2 type, cps genes were not expressed, likely due to an IS6770 inserted in the cpsC-cpsK promoter region. This work constitutes the first study of traits important for interaction, colonization and infection in the host performed on a good representative of E. faecalis food isolates. Reported results stress the need for a reliable serotyping assay of E. faecalis, as cps genotyping may not be reliable. Overall, QA29b characterization shows that despite its virulence potential in an insect model, this food strain is readily eliminated by mammalian macrophages. Thus, fine tuned approaches combining cellular and mammalian models are needed to address and elucidate the multifactorial aspect of virulence potential associated with food isolates. PMID:22831818

  1. Aggregation and Binding Substances Enhance Pathogenicity in Rabbit Models of Enterococcus faecalis Endocarditis

    PubMed Central

    Schlievert, Patrick M.; Gahr, Pamala J.; Assimacopoulos, Aris P.; Dinges, Martin M.; Stoehr, Jennifer A.; Harmala, John W.; Hirt, Helmut; Dunny, Gary M.

    1998-01-01

    We investigated the importance of enterococcal aggregation substance (AS) and enterococcal binding substance (EBS) in rabbit models of Enterococcus faecalis cardiac infections. First, American Dutch belted rabbits were injected intraventricularly with 108 CFU and observed for 2 days. No clinical signs of illness developed in animals given AS− EBS− organisms, and all survived. All rabbits given AS− EBS+ organisms developed signs of illness, including significant pericardial inflammation, but only one of six died. All animals given AS+ EBS− organisms developed signs of illness, including pericardial inflammation, and survived. All rabbits given AS+ EBS+ organisms developed signs of illness and died. None of the rabbits receiving AS+ EBS+ organisms showed gross pericardial inflammation. The lethality and lack of inflammation are consistent with the presence of a superantigen. Rabbit and human lymphocytes were highly stimulated in vitro by cell extracts, but not cell-free culture fluids, of AS+ EBS+ organisms. In contrast, cell extracts from AS− EBS− organisms weakly stimulated lymphocyte proliferation. Culture fluids from human lymphocytes stimulated with AS+/EBS+ enterococci contained high levels of gamma interferon and tumor necrosis factor alpha (TNF-α) and TNF-β, which is consistent with functional stimulation of T-lymphocyte proliferation and macrophage activation. Subsequent experiments examined the abilities of the same strains to cause endocarditis in a catheterization model. New Zealand White rabbits underwent transaortic catheterization for 2 h, at which time catheters were removed and animals were injected with 2 × 109 CFU of test organisms. None of the animals given AS− EBS− organisms developed vegetations or showed autopsy evidence of tissue damage. Rabbits given AS− EBS+ or AS+ EBS− organisms developed small vegetations and had splenomegaly at autopsy. All rabbits given AS+ EBS+ organisms developed large vegetations and had

  2. Role of the mannose receptor in phagocytosis of Enterococcus faecalis strain EC-12 by antigen-presenting cells

    PubMed Central

    Tsuruta, Takeshi; Inoue, Ryo; Nagino, Takayuki; Nishibayashi, Ryoichiro; Makioka, Yuko; Ushida, Kazunari

    2013-01-01

    Abstract The aim of this study was to clarify the phagocytic mechanisms of a heat-killed cell preparation of Enterococcus faecalis strain EC-12 (EC-12) by antigen-presenting cells (APCs). Fluorescein isothiocyanate (FITC)-labeled EC-12 was cocultured with peritoneal macrophage and the amount of EC-12 phagocytosed by peritoneal macrophages was measured using a microplate fluorometer. Peritoneal macrophages from toll-like receptor (TLR)2-, TLR7-, and MyD88-deficient knockout (KO) mice exhibited similar levels of EC-12 phagocytosis to those from wild-type mice. Similarly, dectin-1 neutralization of peritoneal macrophages had no effect on EC-12 phagocytosis. However, blockade of the mannose receptor (MR) significantly decreased the amount of EC-12 phagocytosed by peritoneal macrophages; the same effect was observed in bone marrow-derived macrophages and dendritic cells. Our findings suggest that MR plays a major role in EC-12 phagocytosis by the APCs. This aim of this study was to clarify the phagocytic mechanisms of a heat-killed cell preparation of Enterococcus faecalis strain EC-12 (EC-12) by antigen-presenting cells (APCs). Our findings suggest that mannose receptor (MR) plays a major role in EC-12 phagocytosis by the APCs. PMID:23801521

  3. Protein translation machinery holds a key for transition of planktonic cells to biofilm state in Enterococcus faecalis: A proteomic approach.

    PubMed

    Qayyum, Shariq; Sharma, Divakar; Bisht, Deepa; Khan, Asad U

    2016-06-10

    Enterococcus faecalis is a member of human gut microflora causing nosocomial infection involving biofilm formation. Ethyl methyl sulfonate induced mutants were analysed using crystal violet assay, SEM and CLSM microscopy which confirmed AK-E12 as biofilm efficient and AK-F6 as biofilm deficient mutants. Growth curve pattern revealed AK-E12 was fast growing whereas, AK-F6 was found slow growing mutant. 2D-Electrophorosis and MALDI-TOF analysis revealed over and underexpression of many translation-elongation associated proteins in mutants compared to wild type. Protein translation elongation factor G, translation elongation factor Tu and ribosomal subunit interface proteins were underexpressed and UTP-glucose-1-phosphate uridylyl transferase and cell division protein divIVA were overexpressed in AK-E12 as compared to wild type. In AK-F6, except 10 kDa chaperonin which was over-expressed other selected proteins were found to be suppressed. RT-PCR confirmed proteomic data except for the translation elongation factor G which showed contradictory data of proteome expression in AK-E12. Protein-protein interaction networks were constructed using STRING 10.0 which demonstrated strong connection of translation-elongation proteins with other proteins. Hence, it concludes from the data that translation elongation factors are important in transition of planktonic cells to biofilm cells in Enterococcus faecalis. PMID:27144316

  4. The Enterococcus faecalis cytolysin determinant and its relationship to those encoding lantibiotics.

    PubMed

    Bogie, C P; Hancock, L E; Gilmore, M S

    1995-01-01

    The E. faecalis cytolysin represents a new class of cytolytic agents which are related to a family of antibacterial peptides termed lantibiotics. Despite considerable similarity at the genetic level, the E. faecalis cytolysin differs from the lantibiotics in several respects. First, the E. faecalis cytolysin consists of two dissimilar precursors, both of which are required to effect target cell lysis. A second important difference is that the E. faecalis cytolysin is active against eukaryotic as well as Gram-positive prokaryotic cells. Originally identified as a haemolysin [32], the E. faecalis cytolysin has been shown to make a contribution to bacterial virulence in endocarditis [8, 33] and endophthalmitis [7] models, and the cytolytic phenotype is enriched among clinical isolates of the organism [2, 3]. Similarities between the E. faecalis cytolysin and lantibiotics such as nisin (which is used as a food preservative in several countries [34] and is the subject of continuing attempts at rational design of lantibiotic-based food preservatives [25]), and the observation of an association between the E. faecalis cytolysin and bacterial virulence indicate that it may be possible to engineer lantibiotics to a point where undesired toxic or cytolytic activities will arise. Further comparison of the chemical, structural and biological properties of the E. faecalis cytolysin and classical lantibiotics will define these limits. PMID:8586241

  5. Focal targeting by human β-defensin 2 disrupts localized virulence factor assembly sites in Enterococcus faecalis.

    PubMed

    Kandaswamy, Kumaravel; Liew, Tze Horng; Wang, Charles Y; Huston-Warren, Emily; Meyer-Hoffert, Ulf; Hultenby, Kjell; Schröder, Jens M; Caparon, Michael G; Normark, Staffan; Henriques-Normark, Birgitta; Hultgren, Scott J; Kline, Kimberly A

    2013-12-10

    Virulence factor secretion and assembly occurs at spatially restricted foci in some Gram-positive bacteria. Given the essentiality of the general secretion pathway in bacteria and the contribution of virulence factors to disease progression, the foci that coordinate these processes are attractive antimicrobial targets. In this study, we show in Enterococcus faecalis that SecA and Sortase A, required for the attachment of virulence factors to the cell wall, localize to discrete domains near the septum or nascent septal site as the bacteria proceed through the cell cycle. We also demonstrate that cationic human β-defensins interact with E. faecalis at discrete septal foci, and this exposure disrupts sites of localized secretion and sorting. Modification of anionic lipids by multiple peptide resistance factor, a protein that confers antimicrobial peptide resistance by electrostatic repulsion, renders E. faecalis more resistant to killing by defensins and less susceptible to focal targeting by the cationic antimicrobial peptides. These data suggest a paradigm in which focal targeting by antimicrobial peptides is linked to their killing efficiency and to disruption of virulence factor assembly. PMID:24191013

  6. Inhibitory effect of gels loaded with a low concentration of antibiotics against biofilm formation by Enterococcus faecalis and Porphyromonas gingivalis.

    PubMed

    A Algarni, Amnah; H Yassen, Ghaeth; L Gregory, Richard

    2015-09-01

    We explored longitudinally the inhibitory effect of gels loaded with 1 mg/mL modified triple antibiotic paste (MTAP) or double antibiotic paste (DAP) against biofilm formation by Enterococcus faecalis and Porphyromonas gingivalis. Methylcellulose-based antibiotic gels of MTAP (ciprofloxacin, metronidazole and clindamycin) and DAP (ciprofloxacin and metronidazole) were prepared at a concentration of 1 mg/mL. Individually cultured E. faecalis and P. gingivalis bacterial suspensions were treated with MTAP, DAP, or placebo (vehicle only) gels at different dilutions and allowed to grow in 96-well microtiter plates. Untreated bacterial suspensions served as a negative control. Crystal violet assays were used to evaluate biofilm formation after 48 h. The ability of the gels to inhibit biofilm formation was determined immediately, and at 1 month and 3 months after the gels had been prepared. Data were analyzed using a mixed-model ANOVA. The MTAP and DAP gels significantly reduced biofilm formation by both bacterial species at all time points, regardless of the tested dilution. No-significant differences in biofilm-inhibitory effects between MTAP and DAP gels were observed at the majority of the tested dilutions through various time points. Gels loaded with 1 mg/mL MTAP and DAP demonstrated a significant antibiofilm effect against E.faecalis and P. gingivalis. PMID:26369485

  7. The Fibronectin-Binding Protein EfbA Contributes to Pathogenesis and Protects against Infective Endocarditis Caused by Enterococcus faecalis

    PubMed Central

    Singh, Kavindra V.; La Rosa, Sabina Leanti; Somarajan, Sudha R.; Roh, Jung Hyeob

    2015-01-01

    EfbA is a PavA-like fibronectin adhesin of Enterococcus faecalis previously shown to be important in experimental urinary tract infection. Here, we expressed and purified the E. faecalis OG1RF EfbA and confirmed that this protein binds with high affinity to immobilized fibronectin, collagen I, and collagen V. We constructed an efbA deletion mutant and demonstrated that its virulence was significantly attenuated (P < 0.0006) versus the wild type in a mixed inoculum rat endocarditis model. Furthermore, efbA deletion resulted in diminished ability to bind fibronectin (P < 0.0001) and reduced biofilm (P < 0.001). Reintroduction of efbA into the original chromosomal location restored virulence, adherence to fibronectin, and biofilm formation to wild-type levels. Finally, vaccination of rats with purified recombinant EfbA protein protected against OG1RF endocarditis (P = 0.008 versus control). Taken together, our results demonstrate that EfbA is an important factor involved in E. faecalis endocarditis and that rEfbA immunization is effective in preventing such infection, likely by interfering with bacterial adherence. PMID:26351286

  8. Biochemical and Structural Basis for Inhibition of Enterococcus faecalis Hydroxymethylglutaryl-CoA Synthase, mvaS, by Hymeglusin

    SciTech Connect

    Skaff, D. Andrew; Ramyar, Kasra X.; McWhorter, William J.; Barta, Michael L.; Geisbrecht, Brian V.; Miziorko, Henry M.

    2012-07-25

    Hymeglusin (1233A, F244, L-659-699) is established as a specific {beta}-lactone inhibitor of eukaryotic hydroxymethylglutaryl-CoA synthase (HMGCS). Inhibition results from formation of a thioester adduct to the active site cysteine. In contrast, the effects of hymeglusin on bacterial HMG-CoA synthase, mvaS, have been minimally characterized. Hymeglusin blocks growth of Enterococcus faecalis. After removal of the inhibitor from culture media, a growth curve inflection point at 3.1 h is observed (vs 0.7 h for the uninhibited control). Upon hymeglusin inactivation of purified E. faecalis mvaS, the thioester adduct is more stable than that measured for human HMGCS. Hydroxylamine cleaves the thioester adduct; substantial enzyme activity is restored at a rate that is 8-fold faster for human HMGCS than for mvaS. Structural results explain these differences in enzyme-inhibitor thioester adduct stability and solvent accessibility. The E. faecalis mvaS-hymeglusin cocrystal structure (1.95 {angstrom}) reveals virtually complete occlusion of the bound inhibitor in a narrow tunnel that is largely sequestered from bulk solvent. In contrast, eukaryotic (Brassica juncea) HMGCS binds hymeglusin in a more solvent-exposed cavity.

  9. Effects Of Myrcia Ovata Cambess. Essential Oil On Planktonic Growth Of Gastrointestinal Microorganisms and Biofilm Formation Of Enterococcus Faecalis

    PubMed Central

    Cândido, Cinthya S.; Portella, Cadmo Silton A.; Laranjeira, Bruno J.; da Silva, Sérgio S.; Arriaga, Angela M.C.; Santiago, Gilvandete M. P.; Gomes, Geovany A.; Almeida, Paulo César; Carvalho, Cibele B. M.

    2010-01-01

    The essential oil from the leaves of Myrcia ovata Cambess., commonly used in Brazil for the treatment of gastric illnesses, was screened for antimicrobial activity and action in the formation of microbial biofilms by Enterococcus faecalis. The oil was obtained by hydrodistillation using a clevenger-type system. Its chemical composition was analyzed using GC and GC-MS. Both MIC and MBC of the essential oil were determined by broth microdilution techniques and agar dilution method. The essential oil showed antimicrobial activity against E. faecalis, Escherichia coli, Pseudomonas aeruginosa, Salmonella choleraesuis, Staphylococcus aureus, Streptococcus pneumoniae and Candida parapsilosis. The results showed that the essential oil of M. ovata Cambess. was effective against the formation of biofilm by E. faecalis when compared with the control. Four volatile compounds, representing 92.1 % of the oil, were identified and geranial was the major component (50.4 %). At the best of our knowledge, this is the first report of the chemical composition and antimicrobial activity of the essential oil from leaves of M. ovata. PMID:24031537

  10. β-Lactams Enhance Daptomycin Activity against Vancomycin-Resistant Enterococcus faecalis and Enterococcus faecium in In Vitro Pharmacokinetic/Pharmacodynamic Models

    PubMed Central

    Smith, Jordan R.; Barber, Katie E.; Raut, Animesh

    2015-01-01

    Enterococcus faecalis and Enterococcus faecium are frequently resistant to vancomycin and β-lactams. In enterococcal infections with reduced glycopeptide susceptibility, combination therapy is often administered. Our objective was to conduct pharmacokinetic/pharmacodynamic (PK/PD) models to evaluate β-lactam synergy with daptomycin (DAP) against resistant enterococci. One E. faecalis strain (R6981) and two E. faecium strains (R6370 and 8019) were evaluated. DAP MICs were obtained. All strains were evaluated for response to LL37, an antimicrobial peptide, in the presence and absence of ceftaroline (CPT), ertapenem (ERT), and ampicillin (AMP). After 96 h, in vitro models were run simulating 10 mg DAP/kg body weight/day, 600 mg CPT every 8 h (q8h), 2 g AMP q4h, and 1 g ERT q24h, both alone and in combination against all strains. DAP MICs were 2, 4, and 4 μg/ml for strains R6981, R6370, and 8019, respectively. PK/PD models demonstrated bactericidal activity with DAP-CPT, DAP-AMP, and DAP-ERT combinations against strain 8019 (P < 0.001 and log10 CFU/ml reduction of >2 compared to any single agent). Against strains R6981 and R6370, the DAP-AMP combination demonstrated enhancement against R6370 but not R6981, while the combinations of DAP-CPT and DAP-ERT were bactericidal, demonstrated enhancement, and were statistically superior to all other regimens at 96 h (P < 0.001) against both strains. CPT, ERT, and AMP similarly augmented LL37 killing against strain 8019. In strains R6981 and R6370, CPT and ERT aided LL37 more than AMP (P < 0.001). Compared to DAP alone, combination regimens provide better killing and prevent resistance. Clinical research involving DAP combinations is warranted. PMID:25753639

  11. Mortality in kittens is associated with a shift in ileum mucosa-associated enterococci from Enterococcus hirae to biofilm-forming Enterococcus faecalis and adherent Escherichia coli.

    PubMed

    Ghosh, Anuradha; Borst, Luke; Stauffer, Stephen H; Suyemoto, Mitsu; Moisan, Peter; Zurek, Ludek; Gookin, Jody L

    2013-11-01

    Approximately 15% of foster kittens die before 8 weeks of age, with most of these kittens demonstrating clinical signs or postmortem evidence of enteritis. While a specific cause of enteritis is not determined in most cases, these kittens are often empirically administered probiotics that contain enterococci. The enterococci are members of the commensal intestinal microbiota but also can function as opportunistic pathogens. Given the complicated role of enterococci in health and disease, it would be valuable to better understand what constitutes a "healthy" enterococcal community in these kittens and how this microbiota is impacted by severe illness. In this study, we characterized the ileum mucosa-associated enterococcal community of 50 apparently healthy and 50 terminally ill foster kittens. In healthy kittens, Enterococcus hirae was the most common species of ileum mucosa-associated enterococci and was often observed to adhere extensively to the small intestinal epithelium. These E. hirae isolates generally lacked virulence traits. In contrast, non-E. hirae enterococci, notably Enterococcus faecalis, were more commonly isolated from the ileum mucosa of kittens with terminal illness. Isolates of E. faecalis had numerous virulence traits and multiple antimicrobial resistances. Moreover, the attachment of Escherichia coli to the intestinal epithelium was significantly associated with terminal illness and was not observed in any kitten with adherent E. hirae. These findings identify a significant difference in the species of enterococci cultured from the ileum mucosa of kittens with terminal illness compared to the species cultured from healthy kittens. In contrast to prior case studies that associated enteroadherent E. hirae with diarrhea in young animals, these controlled studies identified E. hirae as more often isolated from healthy kittens and adherence of E. hirae as more common and extensive in healthy kittens than in sick kittens. PMID:23966487

  12. Mortality in Kittens Is Associated with a Shift in Ileum Mucosa-Associated Enterococci from Enterococcus hirae to Biofilm-Forming Enterococcus faecalis and Adherent Escherichia coli

    PubMed Central

    Ghosh, Anuradha; Borst, Luke; Stauffer, Stephen H.; Suyemoto, Mitsu; Moisan, Peter; Zurek, Ludek

    2013-01-01

    Approximately 15% of foster kittens die before 8 weeks of age, with most of these kittens demonstrating clinical signs or postmortem evidence of enteritis. While a specific cause of enteritis is not determined in most cases, these kittens are often empirically administered probiotics that contain enterococci. The enterococci are members of the commensal intestinal microbiota but also can function as opportunistic pathogens. Given the complicated role of enterococci in health and disease, it would be valuable to better understand what constitutes a “healthy” enterococcal community in these kittens and how this microbiota is impacted by severe illness. In this study, we characterized the ileum mucosa-associated enterococcal community of 50 apparently healthy and 50 terminally ill foster kittens. In healthy kittens, Enterococcus hirae was the most common species of ileum mucosa-associated enterococci and was often observed to adhere extensively to the small intestinal epithelium. These E. hirae isolates generally lacked virulence traits. In contrast, non-E. hirae enterococci, notably Enterococcus faecalis, were more commonly isolated from the ileum mucosa of kittens with terminal illness. Isolates of E. faecalis had numerous virulence traits and multiple antimicrobial resistances. Moreover, the attachment of Escherichia coli to the intestinal epithelium was significantly associated with terminal illness and was not observed in any kitten with adherent E. hirae. These findings identify a significant difference in the species of enterococci cultured from the ileum mucosa of kittens with terminal illness compared to the species cultured from healthy kittens. In contrast to prior case studies that associated enteroadherent E. hirae with diarrhea in young animals, these controlled studies identified E. hirae as more often isolated from healthy kittens and adherence of E. hirae as more common and extensive in healthy kittens than in sick kittens. PMID:23966487

  13. Effects of microgravity on the virulence of Listeria monocytogenes, Enterococcus faecalis, Candida albicans, and methicillin-resistant Staphylococcus aureus.

    PubMed

    Hammond, Timothy G; Stodieck, Louis; Birdsall, Holly H; Becker, Jeanne L; Koenig, Paul; Hammond, Jeffrey S; Gunter, Margaret A; Allen, Patricia L

    2013-11-01

    To evaluate effects of microgravity on virulence, we studied the ability of four common clinical pathogens--Listeria monocytogenes, methicillin-resistant Staphylococcus aureus (MRSA), Enterococcus faecalis, and Candida albicans--to kill wild type Caenorhabditis elegans (C. elegans) nematodes at the larval and adult stages. Simultaneous studies were performed utilizing spaceflight, clinorotation in a 2-D clinorotation device, and static ground controls. The feeding rate of worms for killed E. coli was unaffected by spaceflight or clinorotation. Nematodes, microbes, and growth media were separated until exposed to true or modeled microgravity, then mixed and grown for 48 h. Experiments were terminated by paraformaldehyde fixation, and optical density measurements were used to assay residual microorganisms. Spaceflight was associated with reduced virulence for Listeria, Enterococcus, MRSA, and Candida for both larval and adult C. elegans. These are the first data acquired with a direct in vivo assay system in space to demonstrate virulence. Clinorotation reproduced the effects of spaceflight in some, but not all, virulence assays: Candida and Enterococcus were less virulent for larval worms but not adult worms, whereas virulence of MRSA and Listeria were unaffected by clinorotation in tests with both adult and larval worms. We conclude that four common clinical microorganisms are all less virulent in space. PMID:24283929

  14. Dietary Enterococcus faecalis LAB31 Improves Growth Performance, Reduces Diarrhea, and Increases Fecal Lactobacillus Number of Weaned Piglets

    PubMed Central

    Hu, Yuanliang; Dun, Yaohao; Li, Shenao; Zhang, Dongxiao; Peng, Nan; Zhao, Shumiao; Liang, Yunxiang

    2015-01-01

    Lactic acid bacteria (LAB) have been shown to enhance performance of weaned piglets. However, few studies have reported the addition of LAB Enterococcus faecalis as alternatives to growth promoting antibiotics for weaned piglets. This study evaluated the effects of dietary E. faecalis LAB31 on the growth performance, diarrhea incidence, blood parameters, fecal bacterial and Lactobacillus communities in weaned piglets. A total of 360 piglets weaned at 26 ± 2 days of age were randomly allotted to 5 groups (20 pens, with 4 pens for each group) for a trial of 28 days: group N (negative control, without antibiotics or probiotics); group P (Neomycin sulfate, 100 mg/kg feed); groups L, M and H (supplemented with E. faecalis LAB31 0.5×109, 1.0×109, and 2.5×109 CFU/kg feed, respectively). Average daily gain and feed conversion efficiency were found to be higher in group H than in group N, and showed significant differences between group H and group P (P0 < 0.05). Furthermore, groups H and P had a lower diarrhea index than the other three groups (P0 < 0.05). Denaturing gradient gel electrophoresis (DGGE) showed that the application of probiotics to the diet changed the bacterial community, with a higher bacterial diversity in group M than in the other four groups. Real-time PCR revealed that the relative number of Lactobacillus increased by addition of probiotics, and was higher in group H than in group N (P0 < 0.05). However, group-specific PCR-DGGE showed no obvious difference among the five groups in Lactobacillus composition and diversity. Therefore, the dietary addition of E. faecalis LAB31 can improve growth performance, reduce diarrhea, and increase the relative number of Lactobacillus in feces of weaned piglets. PMID:25617897

  15. The Lysozyme-Induced Peptidoglycan N-Acetylglucosamine Deacetylase PgdA (EF1843) Is Required for Enterococcus faecalis Virulence

    PubMed Central

    Ladjouzi, Rabia; Le Jeune, André; Hébert, Laurent; Thorpe, Simon; Courtin, Pascal; Chapot-Chartier, Marie-Pierre; Prajsnar, Tomasz K.; Foster, Simon J.

    2012-01-01

    Lysozyme is a key component of the innate immune response in humans that provides a first line of defense against microbes. The bactericidal effect of lysozyme relies both on the cell wall lytic activity of this enzyme and on a cationic antimicrobial peptide activity that leads to membrane permeabilization. Among Gram-positive bacteria, the opportunistic pathogen Enterococcus faecalis has been shown to be extremely resistant to lysozyme. This unusual resistance is explained partly by peptidoglycan O-acetylation, which inhibits the enzymatic activity of lysozyme, and partly by d-alanylation of teichoic acids, which is likely to inhibit binding of lysozyme to the bacterial cell wall. Surprisingly, combined mutations abolishing both peptidoglycan O-acetylation and teichoic acid alanylation are not sufficient to confer lysozyme susceptibility. In this work, we identify another mechanism involved in E. faecalis lysozyme resistance. We show that exposure to lysozyme triggers the expression of EF1843, a protein that is not detected under normal growth conditions. Analysis of peptidoglycan structure from strains with EF1843 loss- and gain-of-function mutations, together with in vitro assays using recombinant protein, showed that EF1843 is a peptidoglycan N-acetylglucosamine deacetylase. EF1843-mediated peptidoglycan deacetylation was shown to contribute to lysozyme resistance by inhibiting both lysozyme enzymatic activity and, to a lesser extent, lysozyme cationic antimicrobial activity. Finally, EF1843 mutation was shown to reduce the ability of E. faecalis to cause lethality in the Galleria mellonella infection model. Taken together, our results reveal that peptidoglycan deacetylation is a component of the arsenal that enables E. faecalis to thrive inside mammalian hosts, as both a commensal and a pathogen. PMID:22961856

  16. Evaluation of Antimicrobial Effects of Different Concentrations of Triple Antibiotic Paste on Mature Biofilm of Enterococcus faecalis

    PubMed Central

    Frough Reyhani, Mohammad; Rahimi, Saeed; Fathi, Zahra; Shakouie, Sahar; Salem Milani, Amin; Soroush Barhaghi, Mohammad Hossein; Shokri, Javad

    2015-01-01

    Background and aims. Triple antibiotic paste (TAP) is widely used in endodontics for root canal disinfection, particularly in regenerative procedures. The aim of this in vitro study was to evaluate the antimicrobial effects of different concentrations of TAP at 1-, 2-, 3-, and 4-week intervals on mature Enterococcus faecalis biofilm. Materials and methods. A total of 287 extracted one-rooted human central incisors were infected with E. faecalis ATCC 29212 after removing the crown and preparation. The root canal space was filled with one of the 0.01-, 0.1-, 1-, 10-, 100-, and 1000-mg/mL concentrations of TAP or normal saline (control). The root canal dentin was sampled after 1, 2, 3, and 4 weeks. The dentinal shavings were cultured on Mueller-Hinton agar plates after serial dilutions. The classic colony-forming unit (CFU) counting technique was used to determine remaining bacterial counts. Data were analyzed by using the two-way ANOVA, post hoc Tukey tests and one-way ANOVA (P<0.05). Results. TAP completely eliminated E. faecalis biofilms at all the intervals at concentrations of 1000, 100, and 10 mg/mL, whereas 1-, 0.1-, and 0.01-mg/mL TAP resulted in significant reduction of CFU means compared with the control group. There were no statistically significant differences between the four time intervals. Conclusion. Use of lower concentrations of TAP at short term could eradicate E. faecalis biofilm and decrease high-concentration side effects. PMID:26697145

  17. Evaluation of Antimicrobial Effects of Different Concentrations of Triple Antibiotic Paste on Mature Biofilm of Enterococcus faecalis.

    PubMed

    Frough Reyhani, Mohammad; Rahimi, Saeed; Fathi, Zahra; Shakouie, Sahar; Salem Milani, Amin; Soroush Barhaghi, Mohammad Hossein; Shokri, Javad

    2015-01-01

    Background and aims. Triple antibiotic paste (TAP) is widely used in endodontics for root canal disinfection, particularly in regenerative procedures. The aim of this in vitro study was to evaluate the antimicrobial effects of different concentrations of TAP at 1-, 2-, 3-, and 4-week intervals on mature Enterococcus faecalis biofilm. Materials and methods. A total of 287 extracted one-rooted human central incisors were infected with E. faecalis ATCC 29212 after removing the crown and preparation. The root canal space was filled with one of the 0.01-, 0.1-, 1-, 10-, 100-, and 1000-mg/mL concentrations of TAP or normal saline (control). The root canal dentin was sampled after 1, 2, 3, and 4 weeks. The dentinal shavings were cultured on Mueller-Hinton agar plates after serial dilutions. The classic colony-forming unit (CFU) counting technique was used to determine remaining bacterial counts. Data were analyzed by using the two-way ANOVA, post hoc Tukey tests and one-way ANOVA (P<0.05). Results. TAP completely eliminated E. faecalis biofilms at all the intervals at concentrations of 1000, 100, and 10 mg/mL, whereas 1-, 0.1-, and 0.01-mg/mL TAP resulted in significant reduction of CFU means compared with the control group. There were no statistically significant differences between the four time intervals. Conclusion. Use of lower concentrations of TAP at short term could eradicate E. faecalis biofilm and decrease high-concentration side effects. PMID:26697145

  18. Influence of the length of remaining root canal filling and post space preparation on the coronal leakage of Enterococcus faecalis

    PubMed Central

    Mozini, Alexandra Conca Alves; Vansan, Luis P.; Sousa Neto, Manoel D.; Pietro, Rosimeire

    2009-01-01

    This study evaluated the sealing ability of different lengths of remaining root canal filling and post space preparation against coronal leakage of Enterococcus faecalis. Forty-one roots of maxillary incisors were biomechanically prepared, maintaining standardized canal diameter at the middle and coronal thirds. The roots were autoclaved and all subsequent steps were undertaken in a laminar flow chamber. The canals of 33 roots were obturated with AH Plus sealer and gutta-percha. The root canal fillings were reduced to 3 predetermined lengths (n=11): G1=6 mm, G2=4 mm and G3=2 mm. The remaining roots served as positive and negative controls. Bacterial leakage test apparatuses were fabricated with the roots attached to Eppendorf tubes keeping 2 mm of apex submerged in BHI in glass flasks. The specimens received an E. faecalis inoculum of 1 x 107 cfu/mL every 3 days and were observed for bacterial leakage daily during 60 days. Data were submitted to ANOVA, Tukey’s test and Fisher’s test. At 60 days, G1 (6 mm) and G2 (4 mm) presented statistically similar results (p>0.05) (54.4% of specimens with bacterial leakage) and both groups differed significantly (p<0.01) from G3 (2 mm), which presented 100% of specimens with E. faecalis leakage. It may be concluded that the shortest endodontic obturation remnant leaked considerably more than the other lengths, although none of the tested conditions avoids coronal leakage of E. faecalis. PMID:24031339

  19. Antibacterial effect of calcium hydroxide combined with chlorhexidine on Enterococcus faecalis: a systematic review and meta-analysis

    PubMed Central

    SAATCHI, Masoud; SHOKRANEH, Ali; NAVAEI, Hooman; MARACY, Mohammad Reza; SHOJAEI, Hasan

    2014-01-01

    Objective Enterococcus faecalis (E. faecalis) is the most frequently isolated strain in failed endodontic therapy cases since it is resistant to calcium hydroxide (CH). Whether a combination of CH and chlorhexidine (CHX) is more effective than CH alone against E. faecalis is a matter of controversy. Thus, the aim of this study was to conduct a systematic review and meta-analysis of the literature. Material and Methods A comprehensive search in PubMed, EMbase, EBSCOhost, The Cochrane Library, SciELO, and BBO databases, Clinical trials registers, Open Grey, and conference proceedings from the earliest available date to February 1, 2013 was carried out and the relevant articles were identified by two independent reviewers. Backward and forward search was performed and then inclusion and exclusion criteria were applied. The included studies were divided into "comparisons" according to the depth of sampling and dressing period of each medicament. Meta-analysis was performed using Stata software 10.0. The level of significance was set at 0.05. Results Eighty-five studies were retrieved from databases and backward/forward searches. Fortyfive studies were considered as relevant (5 in vivo, 18 in vitro, 18 ex vivo, and 4 review articles). Nine studies were included for meta-analysis. Inter-observer agreement (Cohen kappa) was 0.93. The included studies were divided into 21 comparisons for meta-analysis. Chi-square test showed the comparisons were heterogeneous (p<0.001). Random effect model demonstrated no significant difference between CH/CHX mixture and CH alone in their effect on E. faecalis (p=0.115). Conclusions According to the evidence available now, mixing CH with CHX does not significantly increase the antimicrobial activity of CH against E. faecalis. It appears that mixing CH with CHX does not improve its ex vivo antibacterial property as an intracanal medicament against E. faecalis. Further in vivo studies are necessary to confirm and correlate the findings of

  20. The influence of different peritoneal dialysis fluids on the in vitro activity of ampicillin, daptomycin, and linezolid against Enterococcus faecalis.

    PubMed

    Kussmann, M; Schuster, L; Zeitlinger, M; Pichler, P; Reznicek, G; Wiesholzer, M; Burgmann, H; Poeppl, W

    2015-11-01

    Intraperitoneal administration of antibiotics is recommended for the treatment of peritoneal dialysis-related peritonitis. However, little data are available on a possible interference between peritoneal dialysis fluids and the activity of antimicrobial agents. Thus, the present in vitro study set out to investigate the influence of different peritoneal dialysis fluids on the antimicrobial activity of ampicillin, linezolid, and daptomycin against Enterococcus faecalis. Time-kill curves in four different peritoneal dialysis fluids were performed over 24 h with four different concentrations (1 × MIC, 4 × MIC, 8 × MIC, 30 × MIC) of each antibiotic evaluated. Cation-adjusted Mueller-Hinton broth was used as the comparator solution. All four peritoneal dialysis fluids evaluated had a bacteriostatic effect on the growth of Enterococcus faecalis. Compared to the cation-adjusted Mueller-Hinton broth comparator solution, the antimicrobial activity of all antibiotics tested was reduced. For ampicillin and linezolid, no activity was found in any peritoneal dialysis fluid, regardless of the concentration. Daptomycin demonstrated dose-dependent activity in all peritoneal dialysis fluids. Bactericidal activity was observed at the highest concentrations evaluated in Dianeal® PDG4 and Extraneal®, but not in concentrations lower than 30 × MIC and not in Nutrineal® PD4 and Physioneal® 40. The antimicrobial activity of ampicillin and linezolid is limited in peritoneal dialysis fluids in vitro. Daptomycin is highly effective in peritoneal dialysis fluids and might, thus, serve as an important treatment option in peritoneal dialysis-related peritonitis. Further studies are needed to evaluate the clinical impact of the present findings. PMID:26337433

  1. Behavior of Listeria monocytogenes in a multi-species biofilm with Enterococcus faecalis and Enterococcus faecium and control through sanitation procedures.

    PubMed

    da Silva Fernandes, Meg; Kabuki, Dirce Yorika; Kuaye, Arnaldo Yoshiteru

    2015-05-01

    The formation of mono-species biofilm (Listeria monocytogenes) and multi-species biofilms (Enterococcus faecium, Enterococcus faecalis, and L. monocytogenes) was evaluated. In addition, the effectiveness of sanitation procedures for the control of the multi-species biofilm also was evaluated. The biofilms were grown on stainless steel coupons at various incubation temperatures (7, 25 and 39°C) and contact times (0, 1, 2, 4, 6 and 8 days). In all tests, at 7°C, the microbial counts were below 0.4 log CFU/cm(2) and not characteristic of biofilms. In mono-species biofilm, the counts of L. monocytogenes after 8 days of contact were 4.1 and 2.8 log CFU/cm(2) at 25 and 39°C, respectively. In the multi-species biofilms, Enterococcus spp. were present at counts of 8 log CFU/cm(2) at 25 and 39°C after 8 days of contact. However, the L. monocytogenes in multi-species biofilms was significantly affected by the presence of Enterococcus spp. and by temperature. At 25°C, the growth of L. monocytogenes biofilms was favored in multi-species cultures, with counts above 6 log CFU/cm(2) after 8 days of contact. In contrast, at 39°C, a negative effect was observed for L. monocytogenes biofilm growth in mixed cultures, with a significant reduction in counts over time and values below 0.4 log CFU/cm(2) starting at day 4. Anionic tensioactive cleaning complemented with another procedure (acid cleaning, disinfection or acid cleaning+disinfection) eliminated the multi-species biofilms under all conditions tested (counts of all micro-organisms<0.4 log CFU/cm(2)). Peracetic acid was the most effective disinfectant, eliminating the multi-species biofilms under all tested conditions (counts of the all microorganisms <0.4 log CFU/cm(2)). In contrast, biguanide was the least effective disinfectant, failing to eliminate biofilms under all the test conditions. PMID:25655573

  2. The Two-Component System GrvRS (EtaRS) Regulates ace Expression in Enterococcus faecalis OG1RF

    PubMed Central

    Singh, Kavindra V.; La Rosa, Sabina Leanti; Cohen, Ana Luisa V.; Murray, Barbara E.

    2014-01-01

    Expression of ace (adhesin to collagen of Enterococcus faecalis), encoding a virulence factor in endocarditis and urinary tract infection models, has been shown to increase under certain conditions, such as in the presence of serum, bile salts, urine, and collagen and at 46°C. However, the mechanism of ace/Ace regulation under different conditions is still unknown. In this study, we identified a two-component regulatory system GrvRS as the main regulator of ace expression under these stress conditions. Using Northern hybridization and β-galactosidase assays of an ace promoter-lacZ fusion, we found transcription of ace to be virtually absent in a grvR deletion mutant under the conditions that increase ace expression in wild-type OG1RF and in the complemented strain. Moreover, a grvR mutant revealed decreased collagen binding and biofilm formation as well as attenuation in a murine urinary tract infection model. Here we show that GrvR plays a major role in control of ace expression and E. faecalis virulence. PMID:25385790

  3. Antibacterial Activity of Synthetic Peptides Derived from Lactoferricin against Escherichia coli ATCC 25922 and Enterococcus faecalis ATCC 29212

    PubMed Central

    León-Calvijo, María A.; Leal-Castro, Aura L.; Almanzar-Reina, Giovanni A.; Rosas-Pérez, Jaiver E.; García-Castañeda, Javier E.; Rivera-Monroy, Zuly J.

    2015-01-01

    Peptides derived from human and bovine lactoferricin were designed, synthesized, purified, and characterized using RP-HPLC and MALDI-TOF-MS. Specific changes in the sequences were designed as (i) the incorporation of unnatural amino acids in the sequence, the (ii) reduction or (iii) elongation of the peptide chain length, and (iv) synthesis of molecules with different number of branches containing the same sequence. For each peptide, the antibacterial activity against Escherichia coli ATCC 25922 and Enterococcus faecalis ATCC 29212 was evaluated. Our results showed that Peptides I.2 (RWQWRWQWR) and I.4 ((RRWQWR)4K2Ahx2C2) exhibit bigger or similar activity against E. coli (MIC 4–33 μM) and E. faecalis (MIC 10–33 μM) when they were compared with lactoferricin protein (LF) and some of its derivate peptides as II.1 (FKCRRWQWRMKKLGA) and IV.1 (FKCRRWQWRMKKLGAPSITCVRRAE). It should be pointed out that Peptides I.2 and I.4, containing the RWQWR motif, are short and easy to synthesize; our results demonstrate that it is possible to design and obtain synthetic peptides that exhibit enhanced antibacterial activity using a methodology that is fast and low-cost and that allows obtaining products with a high degree of purity and high yield. PMID:25815317

  4. The effects of solution chemistry on the sticking efficiencies of viable Enterococcus faecalis: An atomic force microscopy and modeling study

    NASA Astrophysics Data System (ADS)

    Cail, Tracy L.; Hochella, Michael F.

    2005-06-01

    Atomic force microscopy (AFM) and Derjaguin-Landau-Verwey-Overbeek (DLVO) theory in combination with the interaction force boundary layer (IFBL) model have been used to empirically and theoretically calculate sticking efficiencies (α) of Enterococcus faecalis cells against a silica glass surface. Sticking efficiencies were calculated in solutions of varying pH and ionic strength and related to maximum distances of transport through a hypothetical soil block using colloid filtration theory. AFM measurements show that the repulsive and attractive forces between E. faecalis cells and a glass surface are a function of ionic strength but are less sensitive to changes in solution pH. Zeta (ζ)-potential measurements of the cells and glass surfaces correlate with these trends. Calculated DLVO energy profiles predict much greater sensitivity to changing solution chemistry. Sticking efficiencies derived from AFM measurements range from 9.6 × 10 -17 to 1 in solutions of low ionic strength (IS) and from 2.6 × 10 -33 to 1 at higher IS. Corresponding α values determined from DLVO theory are essentially zero in all tested solutions. Sticking efficiencies calculated in this study are smaller than values determined from column and field studies in similar systems; however, α derived from AFM data and the IFBL model more closely represent field data than do values calculated from DLVO energy values. A comparison with different methods of calculating α suggests that reversible adhesion may be significant in column-scale transport studies.

  5. Identification of vancomycin interaction with Enterococcus faecalis within 30 min of interaction time using Raman spectroscopy.

    PubMed

    Assmann, Cora; Kirchhoff, Johanna; Beleites, Claudia; Hey, Jessica; Kostudis, Sophia; Pfister, Wolfgang; Schlattmann, Peter; Popp, Jürgen; Neugebauer, Ute

    2015-11-01

    Vancomycin is an important glycopeptide antibiotic which is used to treat serious infections caused by Gram-positive bacteria. However, during the last years, a tremendous rise in vancomycin resistances, especially among Enterococci, was reported, making fast diagnostic methods inevitable. In this contribution, we apply Raman spectroscopy to systematically characterize vancomycin-enterococci interactions over a time span of 90 min using a sensitive Enterococcus faecalis strain and two different vancomycin concentrations above the minimal inhibitory concentration (MIC). Successful action of the drug on the pathogen could be observed already after 30 min of interaction time. Characteristic spectral changes are visualized with the help of multivariate statistical analysis (linear discriminant analysis and partial least squares regressions). Those changes were employed to train a statistical model to predict vancomycin treatment based on the Raman spectra. The robustness of the model was tested using data recorded by an independent operator. Classification accuracies of >90 % were obtained for vancomycin concentrations in the lower range of a typical trough serum concentration recommended for most patients during appropriate vancomycin therapy. Characterization of drug-pathogen interactions by means of label-free spectroscopic methods, such as Raman spectroscopy, can provide the knowledge base for innovative and fast susceptibility tests which could speed up microbiological analysis as well as finding applications in novel antibiotic screenings assays. Graphical Abstract E. faecalis is incubated with vancomycin and characterized by means of Raman spectroscopy after different time points. Characteristic spectral changes reveal efficient vancomycin-enterococci-interaction. PMID:26231687

  6. Decolorization and detoxification of sulfonated toxic diazo dye C.I. Direct Red 81 by Enterococcus faecalis YZ 66.

    PubMed

    Sahasrabudhe, Madhuri M; Saratale, Rijuta G; Saratale, Ganesh D; Pathade, Girish R

    2014-01-01

    Isolated Enterococcus faecalis YZ 66 strain shows ability to decolorize various industrial dyes among which, it showed complete decolorization and degradation of toxic, sulfonated recalcitrant diazo dye Direct Red 81 (50 mg/L) within 1.5 h of incubation under static anoxic condition. The optimum pH and temperature for decolorization was 7.0 and 40°C, respectively. Significant induction in the activity of intracellular oxidoreductive enzymes suggested its involvement in the decolorization of Direct Red 81. The biodegradation of Direct Red 81 was monitored by UV-Visible, FT-IR spectroscopy and HPLC. The final products were characterized by GC-MS and possible pathway of the degradation of the dye was proposed. The phytotoxicity assay (with respect to plants Sorghum vulgare and Phaseolus mungo) revealed that the degradation of Direct Red 81 produced nontoxic metabolites. Finally E. faecalis was employed to decolorize actual industrial effluent showing decolorization (in terms of ADMI value) with moderate COD and BOD reduction. Moreover the result increases the applicability of the strain for the treatment of industrial wastewaters containing dye pollutants. PMID:25649265

  7. Inactivation of Staphylococcus aureus and Enterococcus faecalis by a direct-current, cold atmospheric-pressure air plasma microjet☆

    PubMed Central

    Tian, Ye; Sun, Peng; Wu, Haiyan; Bai, Na; Wang, Ruixue; Zhu, Weidong; Zhang, Jue; Liu, Fuxiang

    2010-01-01

    Objective A direct-current, cold atmospheric-pressure air plasma microjet (PMJ) was performed to inactivate Staphylococcus aureus (S. aureus) and Enterococcus faecalis (E. faecalis) in air. The process of sterilization and morphology of bacteria was observed. We wish to know the possible inactivation mechanisms of PMJ and explore a potential application in dental and other temperature sensitive treatment. Methods In this study, we employed a direct current, atmospheric pressure, cold air PMJ to inactivate bacterias. Scanning electron microscopy was employed to evaluate the morphology of S. aureus and showed rupture of cell walls after the plasma treatment and Optical emission spectrum (OES) were used to understand the possible inactivation mechanisms of PMJ. Results The inactivation rates could reach 100% in 5 min. When the distance between the exit nozzle of the PMJ device and Petri dish was extended from 1 cm to 3 cm, effective inactivation was also observed with a similar inactivation curve. Conclusion The inactivation of bacteria is attributed to the abundant reactive oxygen and nitrogen species, as well as ultroviolet radiation in the plasma. Different life spans and defensibilities of these killing agents may hold the key to understanding the different inactivation curves at different treatment distances. PMID:23554639

  8. Antibacterial activity of synthetic peptides derived from lactoferricin against Escherichia coli ATCC 25922 and Enterococcus faecalis ATCC 29212.

    PubMed

    León-Calvijo, María A; Leal-Castro, Aura L; Almanzar-Reina, Giovanni A; Rosas-Pérez, Jaiver E; García-Castañeda, Javier E; Rivera-Monroy, Zuly J

    2015-01-01

    Peptides derived from human and bovine lactoferricin were designed, synthesized, purified, and characterized using RP-HPLC and MALDI-TOF-MS. Specific changes in the sequences were designed as (i) the incorporation of unnatural amino acids in the sequence, the (ii) reduction or (iii) elongation of the peptide chain length, and (iv) synthesis of molecules with different number of branches containing the same sequence. For each peptide, the antibacterial activity against Escherichia coli ATCC 25922 and Enterococcus faecalis ATCC 29212 was evaluated. Our results showed that Peptides I.2 (RWQWRWQWR) and I.4 ((RRWQWR)4K2Ahx2C2) exhibit bigger or similar activity against E. coli (MIC 4-33 μM) and E. faecalis (MIC 10-33 μM) when they were compared with lactoferricin protein (LF) and some of its derivate peptides as II.1 (FKCRRWQWRMKKLGA) and IV.1 (FKCRRWQWRMKKLGAPSITCVRRAE). It should be pointed out that Peptides I.2 and I.4, containing the RWQWR motif, are short and easy to synthesize; our results demonstrate that it is possible to design and obtain synthetic peptides that exhibit enhanced antibacterial activity using a methodology that is fast and low-cost and that allows obtaining products with a high degree of purity and high yield. PMID:25815317

  9. Development of an intracanal mature Enterococcus faecalis biofilm and its susceptibility to some antimicrobial intracanal medications; an in vitro study

    PubMed Central

    Saber, Shehab El-Din Mohamed; El-Hady, Soha A.

    2012-01-01

    Objectives: To develop a mature biofilm of Enterococcus faecalis inside the root canal system and to test its susceptibility to some antimicrobial medications in vitro. Methods: Single rooted premolars were mechanically enlarged, sterilized, and then infected with a clinical isolate of E. faecalis. Biofilm formation and maturation was monitored using SEM. Biofilm bacteria were exposed to Amoxicillin+clavulanate, Ciprofloxacin, Clindamycin, Doxycycline, and calcium hydroxide as intracanal medications for 1 week. Finally bacterial samples were collected, and colony-forming units were enumerated. Results: SEM examination confirmed the formation of a mature biofilm at the end of the incubation period. All the chemotherapeutic agents used were significantly better than Calcium hydroxide in elimination of biofilm bacteria. The antimicrobial effect of Amoxicillin + clavulanate, Ciprofloxacin and Clindamycin was significantly better than Doxycycline (P=.05). However the difference in the antimicrobial effectiveness among them was statistically non-significant (P=.05). Conclusions: The method used for bacterial biofilm development and maturation is reliable and can be used to assess the anti bacterial potential of endodontic materials. Also, the local application of antibacterial agents can be beneficial in resistant cases of apical periodontitis but only after careful culture and sensitivity testing to choose the appropriate agent for the existing flora. PMID:22229006

  10. Eep confers lysozyme resistance to enterococcus faecalis via the activation of the extracytoplasmic function sigma factor SigV.

    PubMed

    Varahan, Sriram; Iyer, Vijayalakshmi S; Moore, William T; Hancock, Lynn E

    2013-07-01

    Enterococcus faecalis is a commensal bacterium found in the gastrointestinal tract of most mammals, including humans, and is one of the leading causes of nosocomial infections. One of the hallmarks of E. faecalis pathogenesis is its unusual ability to tolerate high concentrations of lysozyme, which is an important innate immune component of the host. Previous studies have shown that the presence of lysozyme leads to the activation of SigV, an extracytoplasmic function (ECF) sigma factor in E. faecalis, and that the deletion of sigV increases the susceptibility of the bacterium toward lysozyme. Here, we describe the contribution of Eep, a membrane-bound zinc metalloprotease, to the activation of SigV under lysozyme stress by its effects on the stability of the anti-sigma factor RsiV. We demonstrate that the Δeep mutant phenocopies the ΔsigV mutant in lysozyme, heat, ethanol, and acid stress susceptibility. We also show, using an immunoblot analysis, that in an eep deletion mutant, the anti-sigma factor RsiV is only partially degraded after lysozyme exposure, suggesting that RsiV is processed by unknown protease(s) prior to the action of Eep. An additional observation is that the deletion of rsiV, which results in constitutive SigV expression, leads to chaining of cells, suggesting that SigV might be involved in regulating cell wall-modifying enzymes important in cell wall turnover. We also demonstrate that, in the absence of eep or sigV, enterococci bind significantly more lysozyme, providing a plausible explanation for the increased sensitivity of these mutants toward lysozyme. PMID:23645601

  11. The Antibacterial Efficacy of Biopure MTAD in Root Canal Contaminated with Enterococcus faecalis

    PubMed Central

    Kamberi, Blerim; Bajrami, Donika; Stavileci, Miranda; Omeragiq, Shuhreta; Dragidella, Fatmir; Koçani, Ferit

    2012-01-01

    Aim. The purpose of this in vitro study was to assess the antimicrobial efficacy of Biopure MTAD against E. faecalis in contaminated root canals. Materials and Methods. Forty-two single rooted extracted human teeth were inoculated with E. faecalis and incubated for four weeks. The samples were divided in two control and five experimental groups irrigated with 1.5% sodium hypochlorite solution (NaOCl); 3% NaOCl; BioPure MTAD; 1.5% NaOCl/17% EDTA; or 3% NaOCl/17% EDTA. After a one-week incubation, complete disinfection was confirmed by the absence of turbidity in the incubation media. Dentin shavings were taken from samples with no turbidity to verify whether E. faecalis was present in dentin tubules. Results were analyzed statistically using Fisher's exact test, with the level of significance set at P < 0.05. Results. Statistical analysis of the data obtained at Day 7 and after dentin shaving analysis showed that BioPure MTAD had significantly greater antibacterial activity than 1.5% NaOCl, 1.5% NaOCl/17% EDTA and 3% NaOCl/17% EDTA. No significant difference was detected between MTAD and 3% NaOCl. Conclusions. These findings suggest that BioPure MTAD possesses superior bactericidal activity compared with NaOCl and EDTA against E. faecalis. PMID:22991671

  12. The Antibacterial Efficacy of Biopure MTAD in Root Canal Contaminated with Enterococcus faecalis.

    PubMed

    Kamberi, Blerim; Bajrami, Donika; Stavileci, Miranda; Omeragiq, Shuhreta; Dragidella, Fatmir; Koçani, Ferit

    2012-01-01

    Aim. The purpose of this in vitro study was to assess the antimicrobial efficacy of Biopure MTAD against E. faecalis in contaminated root canals. Materials and Methods. Forty-two single rooted extracted human teeth were inoculated with E. faecalis and incubated for four weeks. The samples were divided in two control and five experimental groups irrigated with 1.5% sodium hypochlorite solution (NaOCl); 3% NaOCl; BioPure MTAD; 1.5% NaOCl/17% EDTA; or 3% NaOCl/17% EDTA. After a one-week incubation, complete disinfection was confirmed by the absence of turbidity in the incubation media. Dentin shavings were taken from samples with no turbidity to verify whether E. faecalis was present in dentin tubules. Results were analyzed statistically using Fisher's exact test, with the level of significance set at P < 0.05. Results. Statistical analysis of the data obtained at Day 7 and after dentin shaving analysis showed that BioPure MTAD had significantly greater antibacterial activity than 1.5% NaOCl, 1.5% NaOCl/17% EDTA and 3% NaOCl/17% EDTA. No significant difference was detected between MTAD and 3% NaOCl. Conclusions. These findings suggest that BioPure MTAD possesses superior bactericidal activity compared with NaOCl and EDTA against E. faecalis. PMID:22991671

  13. Enterococcus faecalis Glycolipids Modulate Lipoprotein-Content of the Bacterial Cell Membrane and Host Immune Response

    PubMed Central

    Otto, Andreas; Sava, Irina G.; Wobser, Dominique; Bao, Yinyin; Hese, Katrin; Broszat, Melanie; Henneke, Philipp; Becher, Dörte; Huebner, Johannes

    2015-01-01

    In this study, we investigated the impact of the cell membrane composition of E. faecalis on its recognition by the host immune system. To this end, we employed an E. faecalis deletion mutant (ΔbgsA) that does not synthesize the major cell membrane glycolipid diglycosyl-diacylglycerol (DGlcDAG). Proteomic analysis revealed that 13 of a total of 21 upregulated surface-associated proteins of E. faecalis ΔbgsA were lipoproteins. This led to a total lipoprotein content in the cell membrane of 35.8% in ΔbgsA compared to only 9.4% in wild-type bacteria. Increased lipoprotein content strongly affected the recognition of ΔbgsA by mouse macrophages in vitro with an increased stimulation of TNF-α production by heat-fixed bacteria and secreted antigens. Inactivation of the prolipoprotein diacylglycerol transferase (lgt) in ΔbgsA abrogated TNF-α induction by a ΔbgsA_lgt double mutant indicating that lipoproteins mediate increased activation of mouse macrophages by ΔbgsA. Heat-fixed ΔbgsA bacteria, culture supernatant, or cell membrane lipid extract activated transfected HEK cells in a TLR2-dependent fashion; the same was not true of wild-type bacteria. In mice infected intraperitoneally with a sublethal dose of E. faecalis we observed a 70% greater mortality in mice infected with ΔbgsA compared with wild-type-infected mice. Increased mortality due to ΔbgsA infection was associated with elevated plasma levels of the inflammatory cytokines TNF-α, IL-6 and MIP-2. In summary, our results provide evidence that an E. faecalis mutant lacking its major bilayer forming glycolipid DGlcDAG upregulates lipoprotein expression leading to increased activation of the host innate immune system and virulence in vivo. PMID:26172831

  14. Surface-Associated Lipoproteins Link Enterococcus faecalis Virulence to Colitogenic Activity in IL-10-Deficient Mice Independent of Their Expression Levels

    PubMed Central

    Lengfelder, Isabella; Lagkouvardos, Ilias; Steck, Natalie; Roh, Jung H.; Tchaptchet, Sandrine; Bao, Yinyin; Hansen, Jonathan J.; Huebner, Johannes; Carroll, Ian M.; Murray, Barbara E.; Sartor, R. Balfour; Haller, Dirk

    2015-01-01

    The commensal Enterococcus faecalis is among the most common causes of nosocomial infections. Recent findings regarding increased abundance of enterococci in the intestinal microbiota of patients with inflammatory bowel diseases and induction of colitis in IL-10-deficient (IL-10-/-) mice put a new perspective on the contribution of E. faecalis to chronic intestinal inflammation. Based on the expression of virulence-related genes in the inflammatory milieu of IL-10-/- mice using RNA-sequencing analysis, we characterized the colitogenic role of two bacterial structures that substantially impact on E. faecalis virulence by different mechanisms: the enterococcal polysaccharide antigen and cell surface-associated lipoproteins. Germ-free wild type and IL-10-/- mice were monoassociated with E. faecalis wild type OG1RF or the respective isogenic mutants for 16 weeks. Intestinal tissue and mesenteric lymph nodes (MLN) were collected to characterize tissue pathology, loss of intestinal barrier function, bacterial adhesion to intestinal epithelium and immune cell activation. Bone marrow-derived dendritic cells (BMDC) were stimulated with bacterial lysates and E. faecalis virulence was additionally investigated in three invertebrate models. Colitogenic activity of wild type E. faecalis (OG1RF score: 7.2±1.2) in monoassociated IL-10-/- mice was partially impaired in E. faecalis lacking enterococcal polysaccharide antigen (ΔepaB score: 4.7±2.3; p<0.05) and was almost completely abrogated in E. faecalis deficient for lipoproteins (Δlgt score: 2.3±2.3; p<0.0001). Consistently both E. faecalis mutants showed significantly impaired virulence in Galleria mellonella and Caenorhabditis elegans. Loss of E-cadherin in the epithelium was shown for all bacterial strains in inflamed IL-10-/- but not wild type mice. Inactivation of epaB in E. faecalis reduced microcolony and biofilm formation in vitro, altered bacterial adhesion to intestinal epithelium of germ-free Manduca sexta larvae

  15. Methicillin-resistant Staphylococcus aureus, Vancomycin-resistant Enterococcus faecalis and Enterococcus faecium active Dimeric Isobutyrylphloroglucinol from Ivesia gordonii

    PubMed Central

    Ahmed, Marwa H.; Ibrahim, Mohamed Ali; Zhang, Jin; Melek, Farouk R.; El-Hawary, Seham S.; Jacob, Melissa R.; Muhammad, Ilias

    2014-01-01

    Bioassay-guided fractionation of the chloroform soluble fraction of stem, leave, and flower extracts of the American plant Ivesia gordonii led to the isolation of a new dimeric acylphloroglucinol 3,3′-diisobutyryl-2,6′-dimethoxy-4,6,2′,4′-tetrahydroxy-5,5′dimethyldiphenyl methane (1), to which we have assigned the trivial name of ivesinol (1), together with a known monomeric acylphloroglucinol, 1,5-dihydroxy-2-(2′-methylpropionyl)-3-methoxy-6-methylbenzene (2). The structures of the isolated compounds were characterized using 1D- and 2D NMR spectroscopy, including COSY, HMQC, HMBC, and ROESY experiments as well as mass spectrometery. Ivesinol (1) showed potent activity against Staphylococcus aureus (SA) and methicillin-resistant S. aureus (MRSA) with IC50/MIC/MBC values of 0.10/1.25/>20 µg/mL and 0.05/0.31/>20 µg/mL, respectively (vs. IC50/MIC/MBC 0.133/0.5/1.0 µg/mL and 0.128/0.5/1.0 µg/mL of ciprofloxacin), while the corresponding monomer 2 was found to be less active. Compound 1 also demonstrated strong activity against vancomycin-resistant Enterococcus faecium (VRE) with IC50/MIC/MBC values 0.22/1.25/>20 µg/mL, whereas the reference standard ciprofloxacin was found to be inactive against this strain. In addition, compound 2 showed moderate activity against two species of Candida and Cryptococcus neoformans, while 1 was inactive against these fungi. In order to evaluate the influence of acyl group(s) in phloroglucinol (3) as a ligand, the mono- (4) and diacetylphloroglucinol (5) were prepared from 3, and evaluated for their in vitro SA, MRSA, and VRE activities, where 2,4-diacetylphloroglucinol (5) showed potent activity, like 1, against SA, MRSA, and VRE (ATCC 700221) with IC50/ MIC values of 0.27/2.5 µg/mL, 0.23/2.5 µg/mL, and 0.86/2.5 µg/mL, respectively, while 4 was inactive. PMID:24689296

  16. ANALYSIS OF ENTEROCOCCUS FAECALIS IN SAMPLES FROM TURKISH PATIENTS WITH PRIMARY ENDODONTIC INFECTIONS AND FAILED ENDODONTIC TREATMENT BY REAL-TIME PCR SYBR GREEN METHOD

    PubMed Central

    Ozbek, Selcuk M.; Ozbek, Ahmet; Erdogan, Aziz S.

    2009-01-01

    Objective: The aims of this study were to investigate the presence of Enterococcus faecalis in primary endodontic infections and failed endodontic treatments using real-time PCR and to determine the statistical importance of the presence of E. faecalis in a Turkish population with endodontic infections. Material and Methods: E. faecalis was investigated from 79 microbial samples collected from patients who were treated at the Endodontic Clinic of the Dental School of Atatürk University (Erzurum, Turkey). Microbial samples were taken from 43 patients (Group 1) with failed endodontic treatments and 36 patients (Group 2) with chronic apical periodontitis (primary endodontic infections). DNA was extracted from the samples by using a QIAamp® DNA mini-kit and analyzed with real-time PCR SYBR Green. Results: E. faecalis was detected in 41 out of 79 patients, suggesting that it exists in not less than 61% of all endodontic infections when the proportion test (z= -1.645, faecalis in 32 out of 43 (74.4%) in Group 1, and in 9 out of 36 (25%) in Group 2. Conclusions: These results suggest that E. faecalis is a frequent isolate for endodontic infections in Turkish patients, and is more often associated with failed endodontic treatments than primary endodontic infections. PMID:19936510

  17. Molecular detection of virulence factors among food and clinical Enterococcus faecalis strains in South Brazil

    PubMed Central

    Medeiros, A.W.; Pereira, R.I.; Oliveira, D.V.; Martins, P.D.; d’Azevedo, P.A.; Van der Sand, S.; Frazzon, J.; Frazzon, A.P.G

    2014-01-01

    The present report aimed to perform a molecular epidemiological survey by investigating the presence of virulence factors in E. faecalis isolated from different human clinical (n = 57) and food samples (n = 55) in Porto Alegre, Brazil, collected from 2006 to 2009. In addition, the ability to form biofilm in vitro on polystyrene and the β-haemolytic and gelatinase activities were determined. Clinical strains presented a higher prevalence of aggregation substance (agg), enterococcal surface protein (esp) and cytolysin (cylA) genes when compared with food isolates. The esp gene was found only in clinical strains. On the other hand, the gelatinase (gelE) and adherence factor (ace) genes had similar prevalence among the strains, showing the widespread occurrence of these virulence factors among food and clinical E. faecalis strains in South Brazil. More than three virulence factor genes were detected in 77.2% and 18.2% of clinical and food strains, respectively. Gelatinase and β-haemolysin activities were not associated with the presence of gelE and cylA genes. The ability to produce biofilm was detected in 100% of clinical and 94.6% of food isolates, and clinical strains were more able to form biofilm than the food isolates (Student’s t-test, p < 0.01). Results from the statistical analysis showed significant associations between strong biofilm formation and ace (p = 0.015) and gelE (p = 0.007) genes in clinical strains. In conclusion, our data indicate that E. faecalis strains isolated from clinical and food samples possess distinctive patterns of virulence factors, with a larger number of genes that encode virulence factors detected in clinical strains. PMID:24948952

  18. Enterococcus faecalis lipoteichoic acid suppresses Aggregatibacter actinomycetemcomitans lipopolysaccharide-induced IL-8 expression in human periodontal ligament cells.

    PubMed

    Im, Jintaek; Baik, Jung Eun; Kim, Kyoung Whun; Kang, Seok-Seong; Jeon, Jun Ho; Park, Ok-Jin; Kim, Hyun Young; Kum, Kee-Yeon; Yun, Cheol-Heui; Han, Seung Hyun

    2015-08-01

    Periodontitis is caused by multi-bacterial infection and Aggregatibacter actinomycetemcomitans and Enterococcus faecalis are closely associated with inflammatory periodontal diseases. Although lipopolysaccharide (LPS) of A. actinomycetemcomitans (Aa.LPS) and lipoteichoic acid of E. faecalis (Ef.LTA) are considered to be major virulence factors evoking inflammatory responses, their combinatorial effect on the induction of chemokines has not been investigated. In this study, we investigated the interaction between Aa.LPS and Ef.LTA on IL-8 expression in human periodontal ligament (PDL) cells. Aa.LPS, but not Ef.LTA, substantially induced IL-8 expression at the protein and mRNA levels. Interestingly, Ef.LTA suppressed Aa.LPS-induced IL-8 expression without affecting the binding of Aa.LPS to Toll-like receptor (TLR) 4. Ef.LTA reduced Aa.LPS-induced phosphorylation of mitogen-activated protein kinases, including ERK, JNK and p38 kinase. Furthermore, Ef.LTA inhibited the Aa.LPS-induced transcriptional activities of the activating protein 1, CCAAT/enhancer-binding protein and nuclear factor-kappa B transcription factors, all of which are known to regulate IL-8 gene expression. Ef.LTA augmented the expression of IL-1 receptor-associated kinase-M (IRAK-M), a negative regulator of TLR intracellular signaling pathways, in the presence of Aa.LPS at both the mRNA and protein levels. Small interfering RNA silencing IRAK-M reversed the attenuation of Aa.LPS-induced IL-8 expression by Ef.LTA. Collectively, these results suggest that Ef.LTA down-regulates Aa.LPS-induced IL-8 expression in human PDL cells through up-regulation of the negative regulator IRAK-M. PMID:25840438

  19. Unraveling antimicrobial resistance genes and phenotype patterns among Enterococcus faecalis isolated from retail chicken products in Japan.

    PubMed

    Hidano, Arata; Yamamoto, Takehisa; Hayama, Yoko; Muroga, Norihiko; Kobayashi, Sota; Nishida, Takeshi; Tsutsui, Toshiyuki

    2015-01-01

    Multidrug-resistant enterococci are considered crucial drivers for the dissemination of antimicrobial resistance determinants within and beyond a genus. These organisms may pass numerous resistance determinants to other harmful pathogens, whose multiple resistances would cause adverse consequences. Therefore, an understanding of the coexistence epidemiology of resistance genes is critical, but such information remains limited. In this study, our first objective was to determine the prevalence of principal resistance phenotypes and genes among Enterococcus faecalis isolated from retail chicken domestic products collected throughout Japan. Subsequent analysis of these data by using an additive Bayesian network (ABN) model revealed the co-appearance patterns of resistance genes and identified the associations between resistance genes and phenotypes. The common phenotypes observed among E. faecalis isolated from the domestic products were the resistances to oxytetracycline (58.4%), dihydrostreptomycin (50.4%), and erythromycin (37.2%), and the gene tet(L) was detected in 46.0% of the isolates. The ABN model identified statistically significant associations between tet(L) and erm(B), tet(L) and ant(6)-Ia, ant(6)-Ia and aph(3')-IIIa, and aph(3')-IIIa and erm(B), which indicated that a multiple-resistance profile of tetracycline, erythromycin, streptomycin, and kanamycin is systematic rather than random. Conversely, the presence of tet(O) was only negatively associated with that of erm(B) and tet(M), which suggested that in the presence of tet(O), the aforementioned multiple resistance is unlikely to be observed. Such heterogeneity in linkages among genes that confer the same phenotypic resistance highlights the importance of incorporating genetic information when investigating the risk factors for the spread of resistance. The epidemiological factors that underlie the persistence of systematic multiple-resistance patterns warrant further investigations with appropriate

  20. Pyruvate formate-lyase is essential for fumarate-independent anaerobic glycerol utilization in the Enterococcus faecalis strain W11.

    PubMed

    Doi, Yuki; Ikegami, Yuki

    2014-07-01

    Although anaerobic glycerol metabolism in Enterococcus faecalis requires exogenous fumarate for NADH oxidation, E. faecalis strain W11 can metabolize glycerol in the absence of oxygen without exogenous fumarate. In this study, metabolic end product analyses and reporter assays probing the expression of enzymes involved in pyruvate metabolism were performed to investigate this fumarate-independent anaerobic metabolism of glycerol in W11. Under aerobic conditions, the metabolic end products of W11 cultured with glycerol were similar to those of W11 cultured with glucose. However, when W11 was cultured anaerobically, most of the glucose was converted to l-lactate, but glycerol was converted to ethanol and formate. During anaerobic culture with glycerol, the expression of the l-lactate dehydrogenase and pyruvate dehydrogenase E1αβ genes in W11 was downregulated, whereas the expression of the pyruvate formate-lyase (Pfl) and aldehyde/alcohol dehydrogenase genes was upregulated. These changes in the expression levels caused the change in the composition of end products. A pflB gene disruptant (Δpfl mutant) of W11 could barely utilize glycerol under anaerobic conditions, but the growth of the Δpfl mutant cultured with either glucose or dihydroxyacetone (DHA) under anaerobic conditions was the same as that of W11. Glucose metabolism and DHA generates one NADH molecule per pyruvate molecule, whereas glycerol metabolism in the dehydrogenation pathway generates two NADH molecules per pyruvate molecule. These findings demonstrate that NADH generated from anaerobic glycerol metabolism in the absence of fumarate is oxidized through the Pfl-ethanol fermentation pathway. Thus, Pfl is essential to avoid the accumulation of excess NADH during fumarate-independent anaerobic glycerol metabolism. PMID:24769696

  1. Pyruvate Formate-Lyase Is Essential for Fumarate-Independent Anaerobic Glycerol Utilization in the Enterococcus faecalis Strain W11

    PubMed Central

    Ikegami, Yuki

    2014-01-01

    Although anaerobic glycerol metabolism in Enterococcus faecalis requires exogenous fumarate for NADH oxidation, E. faecalis strain W11 can metabolize glycerol in the absence of oxygen without exogenous fumarate. In this study, metabolic end product analyses and reporter assays probing the expression of enzymes involved in pyruvate metabolism were performed to investigate this fumarate-independent anaerobic metabolism of glycerol in W11. Under aerobic conditions, the metabolic end products of W11 cultured with glycerol were similar to those of W11 cultured with glucose. However, when W11 was cultured anaerobically, most of the glucose was converted to l-lactate, but glycerol was converted to ethanol and formate. During anaerobic culture with glycerol, the expression of the l-lactate dehydrogenase and pyruvate dehydrogenase E1αβ genes in W11 was downregulated, whereas the expression of the pyruvate formate-lyase (Pfl) and aldehyde/alcohol dehydrogenase genes was upregulated. These changes in the expression levels caused the change in the composition of end products. A pflB gene disruptant (Δpfl mutant) of W11 could barely utilize glycerol under anaerobic conditions, but the growth of the Δpfl mutant cultured with either glucose or dihydroxyacetone (DHA) under anaerobic conditions was the same as that of W11. Glucose metabolism and DHA generates one NADH molecule per pyruvate molecule, whereas glycerol metabolism in the dehydrogenation pathway generates two NADH molecules per pyruvate molecule. These findings demonstrate that NADH generated from anaerobic glycerol metabolism in the absence of fumarate is oxidized through the Pfl-ethanol fermentation pathway. Thus, Pfl is essential to avoid the accumulation of excess NADH during fumarate-independent anaerobic glycerol metabolism. PMID:24769696

  2. Evaluation of a PCR Assay to Detect Enterococcus faecalis in Blood and Determine Glycopeptides Resistance Genes: Van A and Van B

    PubMed Central

    Honarm, Hamidreza; Falah Ghavidel, Mahsome; Nikokar, Iraj; Rahbar Taromsari, Morteza

    2012-01-01

    Background: Bacteremia due to Enterococcus faecalis is usually caused by strains resistant to most antibiotics. Effective management of the disease is dependent on rapid detection and characterization of the bacteria, and determination its sensitivity pattern to antimicrobial drugs. The aim of this study was to investigate a more rapid and reliable assay for simultaneous diagnosis of enterococcal bacteremia and its sensitivity pattern to antimicrobial drugs. Methods: Several bacterial suspensions with different content of two standard strains of Enterococcus faecalis resistant to vancomycin were used for inoculation to defibrinated sheep blood samples. PCR and routine assay was performed on all blood samples with different bacterial content. Results: Routine assay and PCR for all inoculated blood samples with ≥5 cfu/ml was positive. Mean time for PCR and routine assays was 10 hours and 5 days, respectively. Conclusion: PCR is a more rapid and sensitive assay for simultaneous detection and characterization for Enterococcus faecalis, and determination of its sensitivity pattern to vancomycin. PMID:23115452

  3. Comparison of the Antimicrobial Efficacy of Two Antibiotics Sparfloxacin and Augmentin as Experimental Root Canal Irrigating Solutions against Enterococcus faecalis - An Invitro Study

    PubMed Central

    Venigalla, Bhuvan Shome; Surakanti, Jayaprada Reddy; Thumu, Jayaprakash; Chennamaneni, Krishna Chaitanya; Kalluru, Rama S.

    2016-01-01

    Introduction One of the main goals of endodontic treatment is root canal disinfection and to prevent subsequent chances of reinfection. Adjuvant to instrumentation, root canal irrigants are required to eliminate the bacteria found on the root canal walls and lateral canals within the dentinal tubules. Aim To measure and compare the antibacterial efficacy of two antibiotics as experimental root canal irrigating solutions against Enterococcus faecalis (E. faecalis). Materials and Methods Fifteen Brain Heart Infusion agar plates were inoculated with Enterococcus faecalis-American Type Culture Collection (ATCC) 29212. 5 micrograms (mcg) Sparfloxacin discs, 30mcg Augmentin discs, and sterile paper test discs saturated with 2% Chlorhexidine (CHX), 3% Sodium Hypochlorite (NaOCl) and 5% NaOCl solutions were placed on agar plates. Sodium Chloride 0.9% (NaCl) paper discs were used as controls. Fifteen plates were incubated aerobically at 37°C. Results were expressed as per the terms of the diameter of the inhibition zone. Results Results suggested a statistically significant difference in the zones of inhibition between five irrigating solutions (p < 0.001). Conclusion Although, zones of inhibition were found in all the groups, 5mcg Sparfloxacin and 30mcg Augmentin showed maximum antimicrobial activity against E.faecalis. PMID:27135003

  4. The Effect of Addition of an EPS Degrading Enzyme with and without Detergent to 2% Chlorhexidine on Disruption of Enterococcus faecalis Biofilm: A Confocal Laser Scanning Microscopic Study

    PubMed Central

    Nagendrababu, Venkateshbabu; John, Aby; Deivanayagam, Kandaswamy

    2015-01-01

    Background Enterococcus faecalis is one of the most commonly occurring organisms retrieved from root canal treated teeth that show refractory apical periodontitis. Though it is well known that the ability of E. faecalis to form a matrix-encased biofilm contributes to its pathogenicity, the role of extracellular dextran and DNA in biofilm formation and its effect on the susceptibility of the biofilm to chlorhexidine remains poorly understood. It was hypothesized that the addition of an Extracellular Polymeric Substance (EPS) degrading enzyme along with a detergent to chlorhexidine may increase the susceptibility of the E. faecalis biofilm. Aim To evaluate the sensitivity of Enterococcus faecalis biofilms treated with DNase enzyme and their susceptibility to 2% chlorhexidine used alone or in conjunction with a detergent in a dentin disinfection model and examine under confocal laser scanning microscopy (CLSM). Materials and Methods Semi cylindrical shaped dentin specimens were infected with E. faecalis and incubated for 24 hours. Following incubation, the infected dentin specimens were exposed for 3 minutes to the four disinfecting solutions and grouped accordingly. {Group I- Sterile saline, Group II- 2% Chlorhexidine (CHX), Group III– Dnase1 Enzyme + 2% CHX, Group IV- DNase1 Enzyme + 2% CHX & Tween 80. Bacterial viability was then assessed by staining the specimens and examining under CLSM to analyse the proportion of dead and live bacteria within the dentinal tubules. Results The Groups II, III and IV showed statistically significant (p<0.05) percentage of dead bacteria compared to the control (Group I). However there was no significant difference in the killing effectiveness within the experimental groups (II-IV) at (p<0.05). Conclusion EPS degrading enzyme (DNase I) disrupts the biofilm and increases the susceptibility of E.faecalis when exposed to 2% Chlorhexidine and the use of a surfactant with this combination significantly contributes to improving the

  5. Effects of Intracanal Irrigant MTAD Combined with Nisin at Sub-Minimum Inhibitory Concentration Levels on Enterococcus faecalis Growth and the Expression of Pathogenic Genes

    PubMed Central

    Ling, Junqi; Mao, Xueli; Ning, Yang; Deng, Dongmei

    2014-01-01

    Exposure to antibiotics is considered to be the major driver in the selection of antibiotic-resistant bacteria and may induce diverse biological responses in bacteria. MTAD is a common intracanal irrigant, but its bactericidal activity remains to be improved. Previous studies have indicated that the antimicrobial peptide nisin can significantly improve the bactericidal activity of MTAD against Enterococcus faecalis. However, the effects of MTAD and its modification at sub-minimum inhibitory concentration (sub-MIC) levels on Enterococcus faecalis growth and the expression of pathogenic genes still need to be explored. In this study, the results of post-antibiotic effects (PAE) and post-antibiotic sub-MIC effects (PASME) showed that MTADN (nisin in combination with MTAD) had the best post-antibiotic effect. E. faecalis after challenge with MTAD was less sensitive to alkaline solutions compared with MTAN (nisin in place of doxycycline in MTAD) and MTADN. E. faecalis induced with sub-MIC of MTAD generated resistance to the higher concentration, but induction of E. faecalis with MTAN did not cause resistance to higher concentrations. Furthermore, real-time polymerase chain reaction (RT-PCR) showed that the stress caused by sub-MIC exposure to MTAD, MTAN, or MTADN resulted in up- or down-regulation of nine stress genes and four virulence-associated genes in E. faecalis and resulted in different stress states. These findings suggested that nisin improved the post-antibacterial effect of MTAD at sub-MIC levels and has considerable potential for use as a modification of MTAD. PMID:24603760

  6. Effects of intracanal irrigant MTAD Combined with nisin at sub-minimum inhibitory concentration levels on Enterococcus faecalis growth and the expression of pathogenic genes.

    PubMed

    Tong, Zhongchun; Huang, Lijia; Ling, Junqi; Mao, Xueli; Ning, Yang; Deng, Dongmei

    2014-01-01

    Exposure to antibiotics is considered to be the major driver in the selection of antibiotic-resistant bacteria and may induce diverse biological responses in bacteria. MTAD is a common intracanal irrigant, but its bactericidal activity remains to be improved. Previous studies have indicated that the antimicrobial peptide nisin can significantly improve the bactericidal activity of MTAD against Enterococcus faecalis. However, the effects of MTAD and its modification at sub-minimum inhibitory concentration (sub-MIC) levels on Enterococcus faecalis growth and the expression of pathogenic genes still need to be explored. In this study, the results of post-antibiotic effects (PAE) and post-antibiotic sub-MIC effects (PASME) showed that MTADN (nisin in combination with MTAD) had the best post-antibiotic effect. E. faecalis after challenge with MTAD was less sensitive to alkaline solutions compared with MTAN (nisin in place of doxycycline in MTAD) and MTADN. E. faecalis induced with sub-MIC of MTAD generated resistance to the higher concentration, but induction of E. faecalis with MTAN did not cause resistance to higher concentrations. Furthermore, real-time polymerase chain reaction (RT-PCR) showed that the stress caused by sub-MIC exposure to MTAD, MTAN, or MTADN resulted in up- or down-regulation of nine stress genes and four virulence-associated genes in E. faecalis and resulted in different stress states. These findings suggested that nisin improved the post-antibacterial effect of MTAD at sub-MIC levels and has considerable potential for use as a modification of MTAD. PMID:24603760

  7. SalB inactivation modulates culture supernatant exoproteins and affects autolysis and viability in Enterococcus faecalis OG1RF.

    PubMed

    Shankar, Jayendra; Walker, Rachel G; Wilkinson, Mark C; Ward, Deborah; Horsburgh, Malcolm J

    2012-07-01

    The culture supernatant fraction of an Enterococcus faecalis gelE mutant of strain OG1RF contained elevated levels of the secreted antigen SalB. Using differential fluorescence gel electrophoresis (DIGE) the salB mutant was shown to possess a unique complement of exoproteins. Differentially abundant exoproteins were identified using matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry. Stress-related proteins including DnaK, Dps family protein, SOD, and NADH peroxidase were present in greater quantity in the OG1RF salB mutant culture supernatant. Moreover, several proteins involved in cell wall synthesis and cell division, including d-Ala-d-Lac ligase and EzrA, were present in reduced quantity in OG1RF salB relative to the parent strain. The salB mutant displayed reduced viability and anomalous cell division, and these phenotypes were exacerbated in a gelE salB double mutant. An epistatic relationship between gelE and salB was not identified with respect to increased autolysis and cell morphological changes observed in the salB mutant. SalB was purified as a six-histidine-tagged protein to investigate peptidoglycan hydrolytic activity; however, activity was not evident. High-pressure liquid chromatography (HPLC) analysis of reduced muropeptides from peptidoglycan digested with mutanolysin revealed that the salB mutant and OG1RF were indistinguishable. PMID:22563054

  8. Immune evasion of Enterococcus faecalis by an extracellular gelatinase that cleaves C3 and iC3b.

    PubMed

    Park, Shin Yong; Shin, Yong Pyo; Kim, Chong Han; Park, Ho Jin; Seong, Yeon Sun; Kim, Byung Sam; Seo, Sook Jae; Lee, In Hee

    2008-11-01

    Enterococcus faecalis (Ef) accounts for most cases of enterococcal bacteremia, which is one of the principal causes of nosocomial bloodstream infections (BSI). Among several virulence factors associated with the pathogenesis of Ef, an extracellular gelatinase (GelE) has been known to be the most common factor, although its virulence mechanisms, especially in association with human BSI, have yet to be demonstrated. In this study, we describe the complement resistance mechanism of Ef mediated by GelE. Using purified GelE, we determined that it cleaved the C3 occurring in human serum into a C3b-like molecule, which was inactivated rapidly via reaction with water. This C3 convertase-like activity of GelE was shown to result in a consumption of C3 and thus inhibited the activation of the complement system. Also, GelE was confirmed to degrade an iC3b that was deposited on the Ag surfaces without affecting the bound C3b. This proteolytic effect of GelE against the major complement opsonin resulted in a substantial reduction in Ef phagocytosis by human polymorphonuclear leukocytes. In addition, we verified that the action of GelE against C3, which is a central component of the complement cascade, was human specific. Taken together, it was suggested that GelE may represent a promising molecule for targeting human BSI associated with Ef. PMID:18941224

  9. Comparative evaluation of effect of different irrigation solutions against Enterococcus faecalis: A polymerase chain reaction-based study

    PubMed Central

    Seelan, R. Gnana; Kumar, Arvind; Jonathan, R.; Maheswari, Uma; Raja, Jacob; Chelliah, P.

    2015-01-01

    Enterococcus faecalis is the most isolated or detected species from oral infections including marginal periodontitis, infected root canals, periradicular abscesses and also detected in cases of failed endodontic therapy. To prevent endodontic treatment failure irrigation is mandatory for the effective removal of smear layer, pulp tissue, and microorganisms. Cultivation and other traditional identification methods have been demonstrated to have several limitations when it comes to microbiological identification. Polymerase chain reaction was selected because it has an added advantage over traditional microbiological methods. Materials and Methods: Twenty single rooted premolars were taken were taken stored in 0.1% thymol solution at 4°C decoronated to obtain 12 mm length, teeth were autoclaved at 121°C, canals were instrumented up to 35k file (International Organization for Standardization). The samples were randomly divided into three groups Group I - 5.25% sodium hypochlorite (NaOCL) and 17% ethylenediaminetetraacetic acid (EDTA), Group - II 5.25% NaOCL and 2% chlorhexidine (CHX), Group III - 5.25% NaOCL and 17% EDTA and 2% CHX. Results: The results showed that Group III which is 5% NaOCl followed by 17% EDTA and followed by 2% CHX showed maximum antimicrobial activity in all the three different time intervals. PMID:26538921

  10. VirB8-like protein TraH is crucial for DNA transfer in Enterococcus faecalis

    PubMed Central

    Fercher, Christian; Probst, Ines; Kohler, Verena; Goessweiner-Mohr, Nikolaus; Arends, Karsten; Grohmann, Elisabeth; Zangger, Klaus; Meyer, N. Helge; Keller, Walter

    2016-01-01

    Untreatable bacterial infections caused by a perpetual increase of antibiotic resistant strains represent a serious threat to human healthcare in the 21st century. Conjugative DNA transfer is the most important mechanism for antibiotic resistance and virulence gene dissemination among bacteria and is mediated by a protein complex, known as type IV secretion system (T4SS). The core of the T4SS is a multiprotein complex that spans the bacterial envelope as a channel for macromolecular secretion. We report the NMR structure and functional characterization of the transfer protein TraH encoded by the conjugative Gram-positive broad-host range plasmid pIP501. The structure exhibits a striking similarity to VirB8 proteins of Gram-negative secretion systems where they play an essential role in the scaffold of the secretion machinery. Considering TraM as the first VirB8-like protein discovered in pIP501, TraH represents the second protein affiliated with this family in the respective transfer operon. A markerless traH deletion in pIP501 resulted in a total loss of transfer in Enterococcus faecalis as compared with the pIP501 wild type (wt) plasmid, demonstrating that TraH is essential for pIP501 mediated conjugation. Moreover, oligomerization state and topology of TraH in the native membrane were determined providing insights in molecular organization of a Gram-positive T4SS. PMID:27103580