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Scientific Reports Dec 2020The aim of this study was to investigate how carbohydrates (glucose or sucrose) affect the characteristics of Enterococcus faecalis (E. faecalis) planktonic and biofilm...
The aim of this study was to investigate how carbohydrates (glucose or sucrose) affect the characteristics of Enterococcus faecalis (E. faecalis) planktonic and biofilm in vitro. For this study, E. faecalis was cultured in tryptone-yeast extract broth with 0% glucose + 0% sucrose, 0.5% glucose, 1% glucose, 0.5% sucrose, or 1% sucrose. Viability of E. faecalis was examined by colony forming unit counting assays. Biofilm formation was assessed by measuring extracellular DNA (eDNA), a component of the biofilm matrix. Quantitative real-time PCR (qRT-PCR) was performed to investigate the expression of virulence-associated genes. Field emission scanning electron microscopy analysis, confocal laser scanning microscopy analysis, and crystal violet colorimetric assay were conducted to study E. faecalis biofilms. E. faecalis showed the highest viability and eDNA levels in 1% sucrose medium in biofilms. The result of qRT-PCR showed that the virulence-associated genes expressed highest in 1% sucrose-grown biofilms and in 1% glucose-grown planktonic cultures. E. faecalis showed highly aggregated biofilms and higher bacteria and exopolysaccharide (EPS) bio-volume in sucrose than in 0% glucose + 0% sucrose or glucose. The results indicate that the production of eDNA and EPS and expression of virulence-associated genes in E. faecalis are affected by the concentration of carbohydrates in biofilm or planktonic culture.
Topics: Culture Media; Enterococcus faecalis; Extracellular Polymeric Substance Matrix; Microbial Viability; Virulence
PubMed: 33318537
DOI: 10.1038/s41598-020-78998-5 -
Mikrobiyoloji Bulteni Jan 2020Enterococci, which are commonly found in the environment, cause serious infections despite the absence of well-defined virulence factors and toxins. Knowing the...
Enterococci, which are commonly found in the environment, cause serious infections despite the absence of well-defined virulence factors and toxins. Knowing the virulence properties of enterococci is important to understand the complex pathogenic structures. In this study, we aimed to investigate the virulence factors (asa1, hyl, cylA, efa, ebp, ace, esp, gelE, sprE, fsrA, fsrB, fsrC genes, gelatinase activity, hemolysin, hydrogen peroxide and biofilm production) and antibiotic resistance of Enterococcus faecium and Enterococcus faecalis strains isolated from clinical specimens. A total of 110 enterococcus isolates which were accepted as infectious agents were included in the study. The polymerase chain reaction method was used to identify the isolates and to detect virulence genes. Characteristics of hemolysis, biofilm formation, hydrogen peroxide production and gelatinase activity were investigated by phenotypic methods. The antibiotic susceptibility test was performed with VITEK 2 automated system. E.faecalis ATCC 29212 standard strain was used as a quality control in all tests. Of the 110 enterococci isolates included in the study, 61 were identified as E.faecium and 49 as E.faecalis. The efa gene was the most frequently detected virulence gene (92.7%), followed by ace (83.6%), esp (66.4%), ebp (60.0%), cylA (50.9%), hyl (46.4%), asa1 (45.5%), gelE, sprE, fsrC (33.6%), fsrA (12.7%) and fsrB (11.8%). All genes except hyl were higher in E.faecalis isolates and the difference was statistically significant (p<0.05). Twenty-five (51%) E.faecalis and 1 (1.6%) E.faecium isolates had beta-hemolysis and the difference was statistically significant (p= 0.000). Seven (11.5%) E.faecium and 4 (8.2%) E.faecalis isolates formed biofilm, but the difference was not statistically significant (p> 0.05). Two (3.3%) E.faecium and 14 (28.6%) E.faecalis isolates exhibited gelatinase activity and the difference between the two species was statistically significant (p= 0.000). Hydrogen peroxide production was not detected in any of the isolates. The highest resistance rate was determined against ciprofloxacin (70.9%). The resistance to ampicillin was 69.1%, high level streptomycin 65.1%, high level gentamicin 39.4%, vancomycin and teicoplanin 4.5%, and linezolid 1.8%. In conclusion, our data indicated that virulence factors except hyl gene and biofilm production were higher in E.faecalis isolates but E.faecium isolates were more resistant to antibiotics. In order to prevent infection of such virulent or resistant isolates in the hospital setting, infection control measures must be followed. In vivo studies are needed for the better understanding of the virulence of enterococci.
Topics: Anti-Bacterial Agents; Drug Resistance, Bacterial; Enterococcus faecalis; Enterococcus faecium; Gram-Positive Bacterial Infections; Humans; Microbial Sensitivity Tests; Virulence Factors
PubMed: 32050876
DOI: 10.5578/mb.68810 -
Toxins Apr 2013Enterococcus faecalis is a Gram-positive commensal member of the gut microbiota of a wide range of organisms. With the advent of antibiotic therapy, it has emerged as a... (Review)
Review
Enterococcus faecalis is a Gram-positive commensal member of the gut microbiota of a wide range of organisms. With the advent of antibiotic therapy, it has emerged as a multidrug resistant, hospital-acquired pathogen. Highly virulent strains of E. faecalis express a pore-forming exotoxin, called cytolysin, which lyses both bacterial and eukaryotic cells in response to quorum signals. Originally described in the 1930s, the cytolysin is a member of a large class of lanthionine-containing bacteriocins produced by Gram-positive bacteria. While the cytolysin shares some core features with other lantibiotics, it possesses unique characteristics as well. The current understanding of cytolysin biosynthesis, structure/function relationships, and contribution to the biology of E. faecalis are reviewed, and opportunities for using emerging technologies to advance this understanding are discussed.
Topics: Animals; Bacteriocins; Cross Infection; Enterococcus faecalis; Gram-Positive Bacterial Infections; Humans; Molecular Structure
PubMed: 23628786
DOI: 10.3390/toxins5050895 -
Virulence Nov 2017Enterococcus faecalis and Enterococcus faecium are common inhabitants of the human gastrointestinal tract, as well as frequent opportunistic pathogens. Enterococci cause... (Review)
Review
Enterococcus faecalis and Enterococcus faecium are common inhabitants of the human gastrointestinal tract, as well as frequent opportunistic pathogens. Enterococci cause a range of infections including, most frequently, infections of the urinary tract, catheterized urinary tract, bloodstream, wounds and surgical sites, and heart valves in endocarditis. Enterococcal infections are often biofilm-associated, polymicrobial in nature, and resistant to antibiotics of last resort. Understanding Enterococcal mechanisms of colonization and pathogenesis are important for identifying new ways to manage and intervene with these infections. We review vertebrate and invertebrate model systems applied to study the most common E. faecalis and E. faecium infections, with emphasis on recent findings examining Enterococcal-host interactions using these models. We discuss strengths and shortcomings of each model, propose future animal models not yet applied to study mono- and polymicrobial infections involving E. faecalis and E. faecium, and comment on the significance of anti-virulence strategies derived from a fundamental understanding of host-pathogen interactions in model systems.
Topics: Animals; Anti-Bacterial Agents; Biofilms; Disease Models, Animal; Enterococcus faecalis; Enterococcus faecium; Gram-Positive Bacterial Infections; Humans
PubMed: 28102784
DOI: 10.1080/21505594.2017.1279766 -
Microbiology and Immunology 2000An oxalate-degrading Enterococcus faecalis was isolated from human stools under anaerobic conditions. The bacteria required a poor nutritional environment and repeated...
An oxalate-degrading Enterococcus faecalis was isolated from human stools under anaerobic conditions. The bacteria required a poor nutritional environment and repeated subculturing to maintain their oxalate-degrading ability. The E. faecalis produced 3 proteins (65, 48, and 40 kDa) that were not produced by non-oxalate-degrading E. faecalis as examined by SDS-PAGE. Antibodies against oxalyl-coenzyme A decarboxylase (65 kDa) and formyl-coenzyme A transferase (48 kDa) obtained from Oxalobacter formigenes (an oxalate-degrading anaerobic bacterium in the human intestine) reacted with 2 of the proteins (65 and 48 kDa) from the E. faecalis as examined by Western blottings. This is the first report on the isolation of oxalate-degrading facultative anaerobic bacteria from humans.
Topics: Adult; Animals; Bacterial Proteins; Colony Count, Microbial; Culture Media; Electrophoresis, Polyacrylamide Gel; Enterococcus faecalis; Feces; Female; Humans; Male; Oxalates; Rabbits
PubMed: 10832966
DOI: 10.1111/j.1348-0421.2000.tb02489.x -
Journal of Bacteriology Nov 2017Transmembrane Ser/Thr kinases containing extracellular PASTA domains are ubiquitous among and Such PASTA kinases regulate critical processes, including antibiotic...
Transmembrane Ser/Thr kinases containing extracellular PASTA domains are ubiquitous among and Such PASTA kinases regulate critical processes, including antibiotic resistance, cell division, toxin production, and virulence, and are essential for viability in certain organisms. Based on studies with purified extracellular and intracellular fragments of PASTA kinases, a model for signaling has been proposed, in which the extracellular PASTA domains bind currently undefined ligands (typically thought to be peptidoglycan, or fragments thereof) to drive kinase dimerization, which leads to enhanced kinase autophosphorylation and enhanced phosphorylation of substrates. However, this model has not been rigorously tested is a Gram-positive intestinal commensal and major antibiotic-resistant opportunistic pathogen. In , the PASTA kinase IreK drives intrinsic resistance to cell wall-active antimicrobials, suggesting that such antimicrobials may trigger IreK signaling. Here we show that IreK responds to cell wall stress by enhancing its phosphorylation and that of a downstream substrate. This response requires both the extracellular PASTA domains and specific phosphorylatable residues in the kinase domain. Thus, our results provide evidence, with an intact full-length PASTA kinase in its native physiological environment, that supports the prevailing model of PASTA kinase signaling. In addition, we show that IreK responds to a signal associated with growth and/or cell division, in the absence of cell wall-active antimicrobials. Surprisingly, the ability of IreK to respond to growth and/or division does not require the extracellular PASTA domains, suggesting that IreK monitors multiple parameters for sensory input Transmembrane Ser/Thr kinases containing extracellular PASTA domains are ubiquitous among and and regulate critical processes. The prevailing model for signaling by PASTA kinases proposes that the extracellular PASTA domains bind ligands to drive kinase dimerization, enhanced autophosphorylation, and enhanced phosphorylation of substrates. However, this model has not been rigorously tested We show that the PASTA kinase IreK of responds to cell wall stress by enhancing its phosphorylation and that of a downstream substrate. This response requires the PASTA domains and phosphorylatable residues in the kinase domain. Thus, our results provide evidence, with an intact full-length PASTA kinase in its native physiological environment, that supports the prevailing model of PASTA kinase signaling.
Topics: Cell Membrane; Enterococcus faecalis; Gene Expression Regulation, Bacterial; Models, Biological; Phosphorylation; Protein Processing, Post-Translational; Protein Serine-Threonine Kinases; Signal Transduction; Stress, Physiological
PubMed: 28808126
DOI: 10.1128/JB.00363-17 -
The Journal of Infectious Diseases Dec 2017Enterococcus faecalis is one of the most frequently isolated bacterial species in wounds yet little is known about its pathogenic mechanisms in this setting. Here, we...
Enterococcus faecalis is one of the most frequently isolated bacterial species in wounds yet little is known about its pathogenic mechanisms in this setting. Here, we used a mouse wound excisional model to characterize the infection dynamics of E faecalis and show that infected wounds result in 2 different states depending on the initial inoculum. Low-dose inocula were associated with short-term, low-titer colonization whereas high-dose inocula were associated with acute bacterial replication and long-term persistence. High-dose infection and persistence were also associated with immune cell infiltration, despite suppression of some inflammatory cytokines and delayed wound healing. During high-dose infection, the multiple peptide resistance factor, which is involved in resisting immune clearance, contributes to E faecalis fitness. These results comprehensively describe a mouse model for investigating E faecalis wound infection determinants, and suggest that both immune modulation and resistance contribute to persistent, nonhealing wounds.
Topics: Animals; Disease Models, Animal; Enterococcus faecalis; Gram-Positive Bacterial Infections; Immune Evasion; Male; Mice, Inbred C57BL; Wound Infection
PubMed: 29045678
DOI: 10.1093/infdis/jix541 -
Microbiology (Reading, England) Nov 2011Enterococcus faecalis is a ubiquitous bacterium that is capable of surviving in a broad range of natural environments, including the human host, as either a natural... (Review)
Review
Enterococcus faecalis is a ubiquitous bacterium that is capable of surviving in a broad range of natural environments, including the human host, as either a natural commensal or an opportunistic pathogen involved in severe hospital-acquired infections. How such opportunistic pathogens cause fatal infections is largely unknown but it is likely that they are equipped with sophisticated systems to perceive external signals and interact with eukaryotic cells. Accordingly, being partially exposed at the cell exterior, some surface-associated proteins are involved in several steps of the infection process. Among them are lipoproteins, representing about 25 % of the surface-associated proteins, which could play a major role in bacterial virulence processes. This review focuses on the identification of 90 lipoprotein-encoding genes in the genome of the E. faecalis V583 clinical strain and their putative roles, and provides a transcriptional comparison of microarray data performed in environmental conditions including blood and urine. Taken together, these data suggest a potential involvement of lipoproteins in E. faecalis virulence, making them serious candidates for vaccine production.
Topics: Bacterial Proteins; Computational Biology; Enterococcus faecalis; Gene Expression Regulation, Bacterial; Genome, Bacterial; Lipoproteins; Membrane Proteins; Oligonucleotide Array Sequence Analysis; Proteome; Virulence
PubMed: 21903750
DOI: 10.1099/mic.0.053314-0 -
Comparison of Biofilm Removal Efficiency among Bacteriophage PBEF129, Its Endolysin, and Cefotaxime.Viruses Mar 2021is a Gram-positive pathogen which colonizes human intestinal surfaces, forming biofilms, and demonstrates a high resistance to many antibiotics. Especially, antibiotics...
is a Gram-positive pathogen which colonizes human intestinal surfaces, forming biofilms, and demonstrates a high resistance to many antibiotics. Especially, antibiotics are less effective for eradicating biofilms and better alternatives are needed. In this study, we have isolated and characterized a bacteriophage, PBEF129, infecting . PBEF129 infected a variety of strains of , including those exhibiting antibiotic resistance. Its genome is a linear double-stranded DNA, 144,230 base pairs in length. Its GC content is 35.9%. The closest genomic DNA sequence was found in phage vB_EfaM_Ef2.3, with a sequence identity of 99.06% over 95% query coverage. Furthermore, 75 open reading frames (ORFs) were functionally annotated and five tRNA-encoding genes were found. ORF 6 was annotated as a phage endolysin having an L-acetylmuramoyl-l-alanine amidase activity. We purified the enzyme as a recombinant protein and confirmed its enzymatic activity. The endolysin's host range was observed to be wider than its parent phage PBEF129. When applied to bacterial biofilm on the surface of in vitro cultured human intestinal cells, it demonstrated a removal efficacy of the same degree as cefotaxime, but much lower than its parent bacteriophage.
Topics: Bacteriophages; Biofilms; Endopeptidases; Enterococcus faecalis; Gram-Positive Bacterial Infections; Host Specificity
PubMed: 33800040
DOI: 10.3390/v13030426 -
The Analyst Nov 2012The heptapeptide ARHPHPH was identified from biofilms and planktonic cultures of two different strains of Enterococcus faecalis, V583 and ATCC 29212, using matrix...
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.
Topics: Amino Acid Sequence; Bacterial Proteins; Biofilms; Enterococcus faecalis; Escherichia coli; Molecular Imaging; Oligopeptides; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
PubMed: 22962657
DOI: 10.1039/c2an35922g