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PubMed1989

Prevalence and susceptibility of highly gentamicin resistant Enterococcus faecalis in a south London teaching hospital.

Combinations of aminoglycosides plus ampicillin usually are necessary for the satisfactory management of serious Enterococcus faecalis sepsis. A study in a South London hospital demonstrated that 7% of all E. faecalis, and 30% of those from blood cultures, were highly gentamicin resistant. Addition of gentamicin confers no benefit to the treatment of these organisms with ampicillin. We looked at the susceptibility pattern of these organisms in vitro and concluded that ampicillin plus ciprofloxacin offered the best available combination.

Anti-Bacterial Agents↗

PubMed2003

Manganese-dependent regulation of the endocarditis-associated virulence factor EfaA of Enterococcus faecalis.

There is increasing recognition of the emerging role of manganese regulation and acquisition in some pathogenic bacteria. Expression of the Enterococcus faecalis endocarditis-associated virulence factor EfaA is induced by growth in serum. It is demonstrated here that expression of the efaCBA operon encoding a putative ABC-type transporter is regulated by Mn(2+). Transcription of efaCBA and EfaA production were repressed in Mn(2+)-supplemented medium. A Mn(2+)-responsive transcriptional regulator, EfaR, sharing 27 % identity with the Corynebacterium diphtheriae diphtheria toxin repressor (DtxR), was identified. In the presence of Mn(2+), EfaR protein bound in vitro to the efaC promoter region. Analysis of the E. faecalis V583 genome revealed ten additional putative EfaR-binding sites, suggesting that manganese availability could have a broader regulatory role in infection. The results identify a new Mn(2+)-sensing regulator in enterococci that regulates the expression of a virulence factor implicated in enterococcal endocarditis.

Amino Acid Sequence↗

PubMed1994

Urinary tract infection with an Enterococcus faecalis isolate that requires vancomycin for growth.

OBJECTIVE: To characterize the nutritional requirements and potential origin of a fastidious urinary tract Enterococcus faecalis isolate that apparently requires the antimicrobial agent vancomycin to grow. DESIGN: Case report and detailed microbiologic and molecular epidemiologic analysis. SETTING: University teaching hospital. MEASUREMENTS: Growth of the vancomycin-dependent strain was monitored using various standard laboratory media with and without supplementation with vancomycin and other substrates. This strain was compared with other vancomycin-resistant but nondependent E. faecalis strains by examining plasmid profiles and pulsed-field gel electrophoresis patterns of genomic DNA and by analyzing vancomycin-resistance genes identified by the polymerase chain reaction. RESULTS: An E. faecalis isolate, strain TJ310, was isolated repeatedly from the urine of a patient receiving long-term vancomycin therapy. This strain grew in primary culture but not on subculture, suggesting an unusual growth requirement, and ultimately was found to require the glycopeptide antibiotic vancomycin to grow. Strain TJ310 appeared to be closely related to other vancomycin-resistant but nondependent E. faecalis isolates with the vanB genotype previously isolated from the same patient, suggesting that vancomycin dependence may have evolved in vivo in a vancomycin-resistant enterococcal strain during continuous exposure to high concentrations of vancomycin in the urine. CONCLUSIONS: This is the first reported example of a clinical bacterial isolate that requires an antimicrobial agent to grow.

Drug Resistance, Microbial↗

PubMed1998

Induction of ermAMR from a clinical strain of Enterococcus faecalis by 16-membered-ring macrolide antibiotics.

We cloned the MLSB resistance determinant by PCR from a clinical isolate of Enterococcus faecalis 373, which is induced more strongly by a 16-membered-ring macrolide, tylosin, than by erythromycin. To elucidate the molecular basis of resistance of E. faecalis 373, we analyzed the cloned gene, designated ermAMR, by site-directed mutagenesis and reporter gene assay. Our results showed that an arginine-to-cysteine change in the seventh codon of the putative leader peptide endowed tylosin with resistance inducibility and that TAAA duplication enabled the control region to express the downstream methylase gene at a drastically increased level.

Amino Acid Sequence↗

PubMed2005

Implication of hypR in the virulence and oxidative stress response of Enterococcus faecalis.

HypR has recently been described as the first transcriptional regulator involved in the oxidative stress response and in the intracellular survival of Enterococcus faecalis within macrophages. In order to characterize the HypR regulon, real-time quantitative RT-PCR experiments were performed. The expression of four genes involved in the oxidative stress response encoding catalase, glutathione reductase, and the two subunits of alkyl hydroperoxide reductase were down regulated in the hypR background under H(2)O(2) condition. These findings show that HypR acts as a transcriptional activator, especially during oxidative stress. In addition, DNAse I footprinting assays allowed us to identify the HypR-protected DNA regions corresponding to the "HypR box" in the hypR promoter. Moreover, the effect of the hypR mutation on the virulence of E. faecalis was evaluated in comparison with the wild-type JH2-2 strain using a mouse peritonitis model. Our results revealed that HypR appears to be an important virulence factor in E. faecalis.

Animals↗

PubMed1991

Relationship of the chemical structure and immunobiological activities of lipoteichoic acid from Streptococcus faecalis (Enterococcus hirae) ATCC 9790.

Two molecular species of lipoteichoic acid (LTA 1 and LTA 2) were isolated from whole cells of Streptococcus faecalis (Enterococcus hirae) ATCC 9790 by hydrophobic chromatography on Octyl Sepharose CL-4B. Chemical analysis revealed that LTA 1 and LTA 2 contained two and four acyl lipid anchors respectively. LTA 1 was less active than LTA 2 in inducing cytokines, except interleukin-1 (IL-1), but their in vivo antitumour effects were similar. LTA 2 was a potent inducer of tumour necrosis factor (TNF) and interferon (IFN) production and had excellent antitumour activity against Meth A fibrosarcoma established in mice. Deacylation of LTA 2 by alkaline hydrolysis abolished these biological activities. The phosphatidylglycolipid fraction derived from LTA 2 after acid hydrolysis could also induce TNF, IFN, and IL-1 production, as well as having antitumour activity against Meth A fibrosarcoma. Therefore, the lipid anchor portion of S. faecalis LTA may play an important role in the manifestation of these various biological activities.

Animals↗

PubMed2002

Amplification of the tetracycline resistance determinant of pAMalpha1 in Enterococcus faecalis requires a site-specific recombination event involving relaxase.

The small multicopy plasmid pAMalpha1 (9.75 kb) encoding tetracycline resistance in Enterococcus faecalis is known to generate tandem repeats of a 4.1-kb segment carrying tet(L) when cells are grown extensively in the presence of tetracycline. Here we show that the initial (rate-limiting) step involves a site-specific recombination event involving plasmid-encoded relaxase activity acting at two recombination sequences (RS1 and RS2) that flank the tet determinant. We also present the complete nucleotide sequence of pAMalpha1.

Anti-Bacterial Agents↗

PubMed2002

Actinomyces species, streptococci, and Enterococcus faecalis in primary root canal infections.

The purpose of this study was to evaluate the prevalence of Actinomyces species, streptococci, and Enterococcus faecalis in primary root canal infections by using a molecular genetic method. Samples were obtained from 53 infected teeth, of which 27 cases were diagnosed as acute periradicular abscesses. DNA was extracted to evaluate the occurrence of 13 bacterial species by using whole genomic DNA probes and checkerboard DNA-DNA hybridization. Polymerase chain reaction using an ubiquitous bacterial primer was undertaken to check the presence of bacterial DNA in clinical samples. All root canal samples contained bacteria as demonstrated by polymerase chain reaction. The checkerboard DNA-DNA hybridization assay allowed the detection of streptococci in 22.6% of the samples, Actinomyces species in 9.4%, and E. faecalis in 7.5%. The most prevalent species were members of the Streptococcus anginosus group. With regard to the asymptomatic lesions, the most prevalent species were S. intermedius (11.5% of the cases), E. faecalis (11.5%), and S. anginosus (7.7%). S. constellatus was the most prevalent species in pus samples (25.9% of the cases). The other most prevalent species in abscessed teeth were A. gerencseriae (14.8%), S. gordonii (11.1%), S. intermedius (11.1%), A. israelii (7.4%), S. anginosus (7.4%), and S. sanguis (7.4%). S. constellatus was the only species positively associated with acute periradicular abscess (p < 0.01).

Actinomyces↗

PubMed1991

Enterococcus faecalis: specific and non-specific interactions with human polymorphonuclear leukocytes.

In previous studies we have demonstrated that the ability of Enterococcus faecalis to adhere to and to be internalized in human urinary tract epithelial cells, Girardi Heart cells and human polymorphonuclear leukocytes (PMNs), was dependent on whether the strain had been isolated from urinary tract infections (UTI) or endocarditis (EN) respectively. These properties were further modified by growth of the organism in human serum. In the present report, using competition assays we show that adhesins containing a D-glucose moiety play a role in mediating the interactions between human PMNs and E. faecalis strains isolated from UTI and grown in brain-heart infusion broth (BHIB). On the other hand, adhesins containing both D-glucose and D-galactose moieties were involved in the interactions between PMNs and serum grown UTI isolates or EN isolates grown in either BHIB or human serum. Moreover, the impairment in the association between both UTI and EN strains after growth in serum appears to be at least partially related to a decrease in enterococcal surface hydrophobicity.

Adult↗

PubMed1994

In-vitro postantibiotic effect of sparfloxacin and ciprofloxacin against Pseudomonas aeruginosa and Enterococcus faecalis.

The effect of concentration and exposure time on the postantibiotic effect (PAE) of sparfloxacin and ciprofloxacin against Pseudomonas aeruginosa Psa-1 and Enterococcus faecalis SD-63 was evaluated by the colony counting method using centrifugation to remove the antibiotic. The microorganisms were exposed for 15, 30 and 60 min to drug concentrations ranging from 1 x MIC to 31 x MIC. The PAE was calculated as the difference in time required by test and control cultures to increase 1 log10 after the drug was removed. The MICs of sparfloxacin for P. aeruginosa and E. faecalis were 0.50 mg/L and 0.25 mg/L, respectively, whereas the MICs of ciprofloxacin were 0.25 mg/L for P. aeruginosa and 0.50 mg/L for E. faecalis. PAEs increased as a function of concentration and exposure time. Against E. faecalis no point of maximal response was reached. On the contrary, against P. aeruginosa this point was reached at 8 x MIC of ciprofloxacin and 16 x MIC of sparfloxacin PAE values of sparfloxacin for P. aeruginosa and E. faecalis ranged from 7 to 96 min and from 0 to 112 min, respectively, and those of ciprofloxacin from 8.5 to 82.33 min and from 0 to > 136 min, respectively. Significant PAEs for P. aeruginosa were obtained after 60 min exposure at 4 x MIC (62.6 min) and 2 x MIC (52.6 min) of ciprofloxacin and sparfloxacin, respectively, and at 8 x MIC (81.5 min) and 16 x MIC (81.6 min) for E. faecalis.(ABSTRACT TRUNCATED AT 250 WORDS)

Anti-Infective Agents↗

PubMed1994

Effect of orally administered heat-killed Enterococcus faecalis FK-23 preparation on leukopenia in mice treated with cyclophosphamide.

Cyclophosphamide (CY) treated mice were orally administered heat-killed Enterococcus faecalis FK-23 preparation (FK-23) to define the effect of FK-23 on leukopenia associated with chemotherapy. FK-23 had no inhibiting effect on CY induced leukopenia, whereas it augmented leukocyte reconstitution in CY-treated mice. The percentage of neutrophils increased in the mice orally given FK-23. Furthermore, an increase in a myeloid/erythroid ratio and neutrophilic lineages was found in bone marrow of FK-23 treated mice. These findings suggest that FK-23 may have a potent effect on the augmentation of leukocyte reconstituting capacity in patients receiving chemotherapy.

Animals↗

PubMed2002

Molecular analysis of vancomycin-resistant Enterococcus faecalis from Michigan hospitals during a 10 year period.

We evaluated the molecular relatedness of 47 clinical isolates of vancomycin-resistant Enterococcus faecalis collected from 15 Michigan hospitals from 1991 to 2000. There were 17 PFGE strain types for the 47 isolates. Ten of 15 hospitals demonstrated interhospital, and three of 15 intrahospital, dissemination of some isolates. Forty-two isolates (89.4%) demonstrated vanB resistance. All vanA isolates comprised unique PFGE groups, suggesting transposon dissemination or the presence of a similar plasmid. The results of this study suggest inter- and intrahospital dissemination of strains of vancomycin-resistant E. faecalis during a 10 year period in Michigan.

Anti-Bacterial Agents↗

PubMed2025

Genome-wide analysis of Enterococcus faecalis genes that facilitate interspecies competition with Lactobacillus crispatus.

Enterococci are opportunistic pathogens notorious for causing a variety of infections. While both Enterococcus faecalis and Lactobacillus crispatus are commensal residents of the vaginal tract, the molecular mechanisms that enable E. faecalis to take advantage of a vaginal biome with lower counts of lactobacilli to colonize the vaginal tract and induce aerobic vaginitis remain unknown. Here, we show that L. crispatus eradicates E. faecalis in a contact-independent manner. Using transposon sequencing to identify E. faecalis OG1RF transposon (Tn) mutants that are either under-represented or over-represented when co-cultured with L. crispatus, we found that Tn mutants with disruption in the dltABCD operon, that encodes the proteins responsible for the D-alanylation of teichoic acids, and OG1RF_11697 encoding for an uncharacterized hypothetical protein are more susceptible to killing by L. crispatus. Inversely, Tn mutants with disruption in ldh1, which encodes for L-lactate dehydrogenase, are more resistant to L. crispatus killing. Using the Galleria mellonella infection model, we show that co-injection of L. crispatus with E. faecalis OG1RF enhances larvae survival while this L. crispatus-mediated protection was lost in larvae co-infected with either L. crispatus and E. faecalisΔldh1 or Δldh1Δldh2 strains. Last, using RNA sequencing to identify E. faecalis genes that are differently expressed in the presence of L. crispatus, we found major changes in the expression of genes associated with glycerophospholipid metabolism, central metabolism, and general stress responses. The findings in this study provide insights into how E. faecalis mitigate assaults by L. crispatus.IMPORTANCEEnterococcus faecalis is an opportunistic pathogen notorious for causing a multitude of infections. As vaginal commensals, E. faecalis must interact with Lactobacillus crispatus, but how E. faecalis overcomes or mitigate assaults by L. crispatus killing remains unknown. We show that L. crispatus eradicates E. faecalis temporally in a contact-independent manner. Using high-throughput molecular approaches, we identified genetic determinants that enable E. faecalis to compete with L. crispatus. This study represents an important first step for the identification of adaptive genetic traits required for enterococci to tolerate assaults by lactobacilli.

Enterococcus faecalis↗

PubMed2005

The role of environmental changes on monospecies biofilm formation on root canal wall by Enterococcus faecalis.

Biofilm mode of growth is a strategy in microorganisms to survive harsh growth conditions. Although previous studies have established the ability of Enterococcus faecalis to survive postendodontic environmental conditions, the effect of such conditions on the ultrastructural and physiochemical features of E. faecalis biofilm has received less attention. This study aims to evaluate the effect of different growth conditions on the characteristics of E. faecalis biofilm on root canal, and the penetration of E. faecalis into dentinal tubules. Forty-five intact noncarious human maxillary molars were experimented under nutrient-rich, nutrient-deprived, aerobic, and anaerobic conditions for a period of 21 days. Scanning Electron Microscopy with Energy Dispersive X-ray microanalysis, Laser Confocal Scanning Microscopy and Light microscopic examinations were carried out. The microscopic analysis highlighted a distinct variation in the ultrastructure of the biofilms formed under different experimental conditions. The EDX microanalysis showed a significant increase in the levels of Calcium (Ca) in the biofilm structures formed under anaerobic nutrient-deprived condition (p < 0.001). The depth of bacterial penetration was significantly greater in nutrient-rich condition (p < 0.001). This study demonstrated distinct ultrastructural and physiochemical properties of the biofilms formed and dentinal tubular penetration of E. faecalis under different conditions.

Aerobiosis↗

PubMed2003

Persistence of a clone of glycopeptide-resistant Enterococcus faecalis among patients in an intensive care unit of a Greek hospital.

OBJECTIVES: To investigate an outbreak of glycopeptide-resistant Enterococcus faecalis (GREF) in the intensive care unit (ICU) of 'G. Gennimatas' General Hospital, Athens, Greece. MATERIALS AND METHODS: Between August 2000 and November 2001, 20 highly GREF isolates were recovered from severe infections of separate patients in the ICU. The isolates were tested by PCR, PFGE, mating experiments and plasmid analysis. RESULTS: All isolates carried the vanA gene. Nineteen isolates fitted to one clone by macrorestriction analysis with four subclones being consecutively detected. Each subclone seemed to predominate for a specific time period. Additionally, four GREF isolates related to the ICU clone were recovered from other wards of the hospital. CONCLUSIONS: Our findings indicate that a monoclonal GREF outbreak persisted for more than 1 year in a large Greek hospital. The rate of GREF isolation declined after the application of infection control measures.

Anti-Bacterial Agents↗

PubMed2006

Pheromone-responsive conjugative vancomycin resistance plasmids in Enterococcus faecalis isolates from humans and chicken feces.

The drug resistances and plasmid contents of a total of 85 vancomycin-resistant enterococcus (VRE) strains that had been isolated in Korea were examined. Fifty-four of the strains originated from samples of chicken feces, and 31 were isolated from hospital patients in Korea. Enterococcus faecalis KV1 and KV2, which had been isolated from a patient and a sample of chicken feces, respectively, were found to carry the plasmids pSL1 and pSL2, respectively. The plasmids transferred resistances to vancomycin, gentamicin, kanamycin, streptomycin, and erythromycin to E. faecalis strains at a high frequency of about 10(-3) per donor cell during 4 hours of broth mating. E. faecalis strains containing each of the pSL plasmids formed clumps after 2 hours of incubation in broth containing E. faecalis FA2-2 culture filtrate (i.e., the E. faecalis sex pheromone), and the plasmid subsequently transferred to the recipient strain in a 10-min short mating in broth, indicating that the plasmids are responsive to E. faecalis pheromones. The pSL plasmids did not respond to any of synthetic pheromones for the previously characterized plasmids. The pheromone specific for pSL plasmids has been designated cSL1. Southern hybridization analysis showed that specific FspI fragments from each of the pSL plasmids hybridized with the aggregation substance gene (asa1) of the pheromone-responsive plasmid pAD1, indicating that the plasmids had a gene homologous to asa1. The restriction maps of the plasmids were identical, and the size of the plasmids was estimated to be 128.1 kb. The plasmids carried five drug resistance determinants for vanA, ermB, aph(3'), aph(6'), and aac(6')/aph(2'), which encode resistance to vancomycin, erythromycin, kanamycin, streptomycin, and gentamicin/kanamycin, respectively. Nucleotide sequence analyses of the drug resistance determinants and their flanking regions are described in this report. The results described provide evidence for the exchange of genetic information between human and animal (chicken) VRE reservoirs and suggest the potential for horizontal transmission of multiple drug resistance, including vancomycin resistance, between farm animals and humans via a pheromone-responsive conjugative plasmid.

Animals↗

PubMed1997

Pheromone-inducible expression of an aggregation protein in Enterococcus faecalis requires interaction of a plasmid-encoded RNA with components of the ribosome.

Transfer of the conjugative plasmid pCF10 from Enterococcus faecalis donor strains is induced by a peptide pheromone (cCF10) secreted by recipient cells. High-efficiency transfer requires expression of an aggregation protein (Asc10) encoded by the prgB gene and positively regulated by genes in a region 3-5 kb upstream, containing prgQ-R-S. Transcriptional fusion data reported here support the results of recent molecular analysis of the 5' ends of prgB transcripts which indicated that prgB transcription occurs by readthrough from the prgQ promoter. A 530-nucleotide prgQ-encoded RNA molecule (Q(L)) with rRNA-like domains is required for Asc10 production. Q(L) and cCF10 were found to interact with the L6 and S5 ribosomal proteins, respectively. Mutational analysis of Q(L) indicates that this RNA may also directly interact with 16S RNA. Q(L) is present in ribosomes translating the prgB message, and pheromone cCF10 may affect the association of this RNA with translation complexes. Results suggest that the positive regulatory molecules act post-transcriptionally on the polycistronic message and modify a ribosome population to enhance pheromone-induced translation of prgB.

Amino Acid Sequence↗

PubMed2004

Construction of an Enterococcus faecalis Tn917-mediated-gene-disruption library offers insight into Tn917 insertion patterns.

Sequencing the insertion sites of 8,865 Tn917 insertions in Enterococcus faecalis strain OG1RF identified a hot spot in the replication terminus region corresponding to 6% of the genome where 65% of the transposons had inserted. In E. faecalis, Tn917 preferentially inserted at a 29-bp consensus sequence centered on TATAA, a 5-bp sequence that is duplicated during insertion. The regional insertion site preference at the chromosome terminus was not observed in another low-G+C gram-positive bacterium, Listeria monocytogenes, although the consensus insertion sequence was the same. The 8,865 Tn917 insertion sites sequenced in E. faecalis corresponded to only approximately 610 different open reading frames, far fewer than the predicted number of 2,400, assuming random insertion. There was no significant preference in orientation of the Tn917 insertions with either transcription or replication. Even though OG1RF has a smaller genome than strain V583 (2.8 Mb versus 3.2 Mb), the only E. faecalis strain whose sequence is in the public domain, over 10% of the Tn917 insertions appear to be in a OG1RF-specific sequence, suggesting that there are significant genomic differences among E. faecalis strains.

Bacterial Proteins↗