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PubMed1990

High efficiency introduction of plasmid DNA into glycine treated Enterococcus faecalis by electroporation.

A highly efficient electroporation system for Enterococcus faecalis was developed by systematically optimizing different parameters. One parameter found to be particularly critical for electroporation was cultivation of E. faecalis in medium containing a high glycine concentration, prior to electroporation. Osmotic stabilization of cells with 0.5 M sucrose was also found to be critical during glycine treatment. 10(6) transformants per microgram of plasmid DNA were consistently obtained within 48 h. Electrocompetent preparations of E.-faecalis could be stored at - 70 degrees C without loss of competence.

DNA, Bacterial↗
PubMed1991

The conjugative transposon Tn925: enhancement of conjugal transfer by tetracycline in Enterococcus faecalis and mobilization of chromosomal genes in Bacillus subtilis and E. faecalis.

The effects of tetracycline on transfer of the conjugative, tetracycline-resistance transposon. Tn925, as well as the ability of the transposon to promote the transfer of chromosomal genes was examined in Enterococcus faecalis and Bacillus subtilis. To test for chromosomal transfer, multiply-marked strains of each organism, each carrying a single chromosomal copy of Tn925, were mated on filters with suitable recipient strains, under conditions where transformation and transduction were precluded. In both cases, transfer of a variety of chromosomal genes, at frequencies comparable to the frequency of Tn925 transfer, was detected readily. The presence of Tn925 in one of the members of the mating pair was absolutely required for chromosomal transfer, but transfer of Tn925 did not accompany every chromosomal transfer event. The results were consistent with a mating event resembling a type of cell fusion, allowing for extensive recombination between the genomes of the mating partners. Growth of Tn925-containing donor cells in the presence of tetracycline increased the transfer frequency of Tn925 by about tenfold in E. faecalis, but not in B. subtilis.

Bacillus subtilis↗
PubMed1989

Study on pathogenesis of Enterococcus faecalis in urinary tract.

The rate of isolation of Enterococcus faecalis as the causative bacterium of complicated urinary tract infections has been increasing. However, the pathogenicity of this bacterium in the urinary tract has not been clarified. Thus, the authors investigated how E. faecalis is pathogenic to the urinary tract, using mice with experimental urinary tract infection. The investigation revealed that this bacterium when sufficiently numerous can be directly pathogenic. The bacterium can be pathogenic indirectly when present with other typical urinary bacteria such as E. coli.

Animals↗
PubMed1994

Activities and regulation of the enzymes involved in the first and the third steps of the aspartate biosynthetic pathway in Enterococcus faecium.

The enzymes aspartokinase and homoserine dehydrogenase catalyze the reaction at key branching points in the aspartate pathway of amino acid biosynthesis. Enterococcus faecium has been found to contain two distinct aspartokinases and a single homoserine dehydrogenase. Aspartokinase isozymes eluted on gel filtration chromatography at molecular weights greater than 250,000 and about 125,000. The molecular weight of homoserine dehydrogenase was determined to be 220,000. One aspartokinase isozyme was slightly inhibited by meso-diaminopimelic acid. Another aspartokinase was repressed and inhibited by lysine. Although the level of diaminopimelate-sensitive (DAPs) enzyme was not much affected by growth conditions, the activity of lysine-sensitive (Lyss) aspartokinase disappeared rapidly during the stationary phase and was depressed in rich media. The synthesis of homoserine dehydrogenase was controlled by threonine and methionine. Threonine also inhibited the specific activity of this enzyme. The regulatory properties of aspartokinase isozymes and homoserine dehydrogenase from E. faecium are discussed and compared with those from Bacillus subtilis.

Amino Acids↗
PubMed1996

Enterococcus faecalis cytolysin and lactocin S of Lactobacillus sake.

Strains of Enterococcus faecalis and Lactobacillus sake have been found to express lantibiotics with unusual properties. The enterococcal lantibiotic is unusual in that it consists of two dissimilar subunits, both putatively containing modifications consistent with those found in other lantibiotics. The enterococcal lantibiotic is also unusual in the number of proteolytic steps involved in secretion signal removal and activation. Moreover, it has been observed to contribute to enterococcal disease in humans and in animal models. Structural studies of lactocin S, expressed by a strain of L. sake highlight unique properties including the presence of D-alanine within its structure, and a protease putatively responsible for lactocin S secretion signal peptide removal which, itself, lacks a signal or propeptide sequence. Despite the unusual properties of each of these lantibiotics, the operons encoding each, and accompanying auxiliary functions, are collinear suggesting a common ancestry. The accretion of interdigitating DNA sequences between genes encoded within the lactocin S determinant are unique to that determinant, however, and are of unknown function.

Amino Acid Sequence↗
PubMed1989

Identification of aggregation substances of Enterococcus faecalis cells after induction by sex pheromones. An immunological and ultrastructural investigation.

The sex pheromone system of Enterococcus faecalis is responsible for the clumping response of a plasmid carrying donor strain with a corresponding plasmid free recipient strain due to the production of sex pheromones by the recipient strain. The clumping response is mediated by a surface material (called aggregation substance) which is synthesized upon addition of sex pheromones to the cultures. Here we show that after induction a dense layer of "hairlike" structures is formed on the cell wall of the bacteria. These hairlike structures are responsible for the cell-cell contact which leads to the aggregation of cells. Formation of these structures was specific, only occurring after the addition of homologous sex pheromone.

Biological Factors↗
PubMed1989

Localization of aggregation substances of Enterococcus faecalis after induction by sex pheromones. An ultrastructural comparison using immuno labelling, transmission and high resolution scanning electron microscopic techniques.

The distribution of sex pheromone induced aggregation substance was studied on the cell surface of various Enterococcus faecalis strains. In the accompanying paper we have shown that the aggregation substance appears as a layer of hairlike structures. Using direct and indirect immunogold technique, transmission electron microscopy and high resolution scanning electron microscopy we investigated the appearance and distribution of the aggregation substance. The "hairs" increase in number with increasing exposure to sex pheromones (maximum density: 1300/microns2). We show that these structures are unequally distributed over the cell surface, even if the cells were induced by sex pheromones for a long period of time. Statistical analysis of the unequal distribution indicates that aggregation substance is incorporated into pre-existing "old" cell-walls and that this incorporation shows a saturation ca. 40 min after addition of sex pheromones.

Biological Factors↗
PubMed1992

Identification of new sex pheromone plasmids in Enterococcus faecalis.

We describe the identification of the following new sex pheromone plasmids in Enterococcus faecalis: a haemolysin-bacteriocin plasmid, pIP964; three R plasmids, pIP1017, pIP1438 and pIP1440; and two cryptic conjugative plasmids, pIP1141 and pMV120. The identification was based on the formation of cell aggregates on filter membranes during conjugation, on efficient transfer in broth matings, and on a positive clumping reaction of cells carrying these plasmids. In addition these plasmids hybridized with DNA probes specific for sex pheromone-induced structural genes encoding surface proteins required for conjugative transfer of the plasmids.

DNA Probes↗
PubMed1992

Sex pheromone plasmid pAD1-encoded surface exclusion protein of Enterococcus faecalis.

During conjugative transfer of sex pheromone plasmids of Enterococcus faecalis a so-called surface exclusion protein reduces the frequency with which these plasmids are transferred to cells already possessing the same plasmid. We report here the DNA sequence of a 3.8 kb fragment of the sex pheromone plasmid pAD1 containing the structural gene sea1 for surface exclusion protein and a small open reading frame (ORF) upstream of sea1. Surface exclusion protein Sea1 was found to be highly homologous to the surface exclusion protein Sec10 encoded by the sex pheromone plasmid pCF10. Hybridization studies with DNA probes derived from the structural gene sea1 demonstrated that, with the exception of pAM373, all known sex pheromone plasmids carry a homologous gene. These studies also indicated that the genetic organization is similar in these plasmids, with the structural gene for surface exclusion protein being located 5' to that for aggregation substance.

Amino Acid Sequence↗
PubMed1993

Characterization of the heat shock response in Enterococcus faecalis.

We have characterized the general properties of the heat shock response of the Gram-positive hardy bacterium Enterococcus faecalis. The heat resistance (60 degrees C or 62.5 degrees C, 30 min) of log phase cells of E. faecalis grown at 37 degrees C was enhanced by exposing cells to a prior heat shock at 45 degrees C or 50 degrees C for 30 min. These conditioning temperatures also induced ethanol (22%, v/v) tolerance. The onset of thermotolerance was accompanied by the synthesis of a number of heat shock proteins. The most prominent bands had molecular weights in the range of 48 to 94kDa. By Western blot analysis two of them were found to be immunologically related to the well known DnaK (72kDa) and GroEL (63kDa) heat shock proteins of Escherichia coli. Four other proteins showing little or no variations after exposure to heat are related to DnaJ, GrpE and Lon (La) E. coli proteins and to the Bacillus subtilis sigma 43 factor. Ethanol (2% or 4%, v/v) treatments elicited a similar response although there was a weaker induction of heat shock proteins than with heat shock.

Antigens, Bacterial↗
PubMedDate not supplied

Bactericidal effect of penicillin, ampicillin, and amoxicillin alone and in combination with tobramycin against Enterococcus faecalis as determined by kill-kinetic studies.

Fifteen strains of Enterococcus faecalis, all clinical blood culture isolates from patients with endocarditis, were studied by kill-kinetic experiments using penicillin G, ampicillin and amoxicillin alone and in combination with tobramycin. The median minimal inhibitory concentrations (MIC), were penicillin 4 mg/l, ampicillin 2 mg/l, amoxicillin 2 mg/l and tobramycin 32 mg/l. Equipotent doses of the antibiotics (1/2 x MIC, 1 x MIC and 4 x MIC) were used in the kill-kinetic studies. Synergism was studied using a combination of 1/2 x MIC of the beta-lactam antibiotic and 8 mg/l of tobramycin. The bactericidal activity did not exceed 10(+3) cfu/ml at 5 hours for any single compound. After 5 h all three beta-lactam antibiotics in combination with tobramycin resulted in synergism, i.e. more than one hundredfold reduction of colony forming units (cfu) as compared to the most active single agent. Amoxicillin had a significantly higher bactericidal potential than ampicillin or penicillin both alone and in combination with tobramycin. The clinical significance of these findings warrants further studies in vivo.

Amoxicillin↗
PubMed1995

Enterococcus faecalis colonisation and endocarditis in five intensive care patients as late sequelae of selective decontamination.

OBJECTIVE: To describe Enterococcus faecalis colonisation and endocarditis in 5 intensive care patients after treatment with selective decontamination (SDD). SETTING: Intensive care unit (ICU) in a general hospital. PATIENTS: The patients were admitted to the ICU because of adult respiratory distress syndrome, polytrauma (2 patients), abdominal aortic surgery and gastrointestinal surgery. Because these patients needed mechanical ventilation they received systemic cefotaxime and SDD (polymyxin E, amphotericin B and norfloxacin). RESULTS: Colonisation with E. faecalis was documented in all patients. Intravascular catheter-related infection with E. faecalis occurred in 4 patients. None of the patients received antibiotics active against, E. faecalis, because body temperature normalised after catheter removal. In the course of his ICU stay one patient died. Autopsy showed E. faecalis endocarditis. The other 4 patients recovered from their primary illness, but had to be readmitted after several months because of E. faecalis endocarditis. One of these patients died. One patient recovered of endocarditis, but one year later valve surgery was necessary. The other 2 patients needed acute valve replacement. The latter 3 patients survived. CONCLUSION: We observed 5 patients with E. faecalis endocarditis as a late and severe sequela of SDD during their ICU stay.

Aged↗
PubMedDate not supplied

The influence of Enterococcus faecalis on the morphology and the antibody-binding capacity of the intestinal bacteria of ten healthy human volunteers.

The influence of Enterococcus faecalis on the morphology of the bacterial cells which make up the gut microflora and on the levels of circulating IgG bound to the gut microflora was assessed. After 29 days of pretreatment monitoring, ten healthy human volunteers ingested 10(7) viable cells of E. faecalis three times daily, for 21 days. After this treatment another 21 days of follow-up completed the study. Each volunteer delivered eleven faecal samples during the entire study period of 71 days with a 7 day interval. Before and after the faeces sampling period, blood samples were collected from all volunteers. The influence of the ingestion of E. faecalis on the morphology of the gut microflora was measured by image analysis. In addition, the binding of circulating IgG to intestinal bacteria in all intermediate faecal samples was measured by means of quantitative immunofluorescence. The oral administration of E. faecalis resulted in a significant change of the morphological composition of the gut microflora and in a significant decrease in IgG-binding capacity of the gut microflora.

Adult↗
PubMed1992

New high-content disks for determination of high-level aminoglycoside resistance in clinical isolates of Enterococcus faecalis.

Disks impregnated with 500 and 1000 micrograms of streptomycin, 1000 micrograms of kanamycin and 250 and 500 micrograms of gentamicin were used for detection of high-level resistance to aminoglycosides in 120 clinical isolates of Enterococcus faecalis. Fifty-seven strains were highly resistant to streptomycin, 80 to kanamycin and 41 to gentamicin. Using disks containing 500 micrograms of streptomycin, 1000 micrograms of kanamycin and 500 micrograms of gentamicin strains resistant to high levels of these drugs (97.9%, 100% and 100%, respectively) were accurately detected. Better discrimination between high-level and low-level resistance was achieved with a 500 micrograms streptomycin or gentamicin disk. Zone-size breakpoints are proposed for detection of high-level resistance by disk diffusion.

Drug Resistance, Microbial↗
PubMed1988

Aortic endocarditis caused by gentamicin-resistant Enterococcus faecalis.

A case is reported of a patient who developed aortic endocarditis after protracted hospitalization during which time the patient received multiple courses of aminoglycoside and cephalosporin therapy. The infecting organism, an Enterococcus faecalis strain, was resistant to gentamicin and to the combination of ampicillin with gentamicin. Valve replacement was performed after four weeks of therapy with ampicillin, and enterococci were isolated from cardiac vegetations. Valve replacement may be necessary to achieve cure of endocarditis due to enterococci highly resistant to gentamicin.

Aged↗
PubMed1990

Recent taxonomic changes in the genus Enterococcus.

Recent taxonomic changes among the catalase-negative gram-positive cocci have led to the recognition of a separate genus for bacteria previously considered to be members of the genus Streptococcus. Presently, a total of 12 species of the genus Enterococcus have been described. The characteristics, taxonomy and nomenclature of these species, along with the possible clinical implications of revised enterococcal taxonomy, are discussed.

Enterococcus faecalis↗
PubMed1988

Turbidimetric response of Staphylococcus aureus and Enterococcus faecalis to daptomycin.

The response to daptomycin of three strains of Staphylococcus aureus (one of which was methicillin-resistant) and a strain of Enterococcus faecalis was investigated by continuous turbidimetric monitoring. Daptomycin caused partial inhibition of bacterial growth at concentrations well below those suppressing growth overnight. The activity of the drug was more than 200-fold greater in the presence of a physiological concentration of calcium (2.5 mmol/l) than in its absence. Variants present in cultures inhibited by daptomycin were shown to exhibit decreased susceptibility to the drug and the susceptibility declined further on repeat exposure; however, there was a slow but incomplete reversion to susceptibility on sequential subculture in drug-free broth. Resistance was difficult to induce when the concentration of calcium in the broth exceeded 2.5 mmol/l.

Anti-Bacterial Agents↗
PubMedDate not supplied

Recurrent septicemias with Enterococcus faecium.

A 17-year-old male patient with extrahepatic biliary atresia underwent an orthotopic liver transplantation in September 1994. In blood cultures drawn in November and (6 weeks later) December 1994, from bile secretions in May 1995, stool in June 1995 an wound abscess in August 1995, ampicillin-resistant Enterococcus faecium was isolated. Pulsed-field gel electrophoresis demonstrated the clonal identity of the isolates. To our knowledge, repeated infections with the same E. faecium strain over a period of 9 months have not been described before. As multiple-resistant enterococci may colonize and reinfect liver transplant recipients for such a long time, preoperative antibiotic therapy should be administered cautiously in order not to select these organisms.

Adolescent↗