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Colonization and microbiology of the motile enterococci in a patient population.

The motile enterococci with the vanC gene have intrinsic low-level resistance to vancomycin, but have not been implicated in a nosocomial outbreak. We determined the colonization rate of motile enterococci in hospitalized and nonhospitalized patients. Perianal or stool specimens were cultured in Enterococcosel broth supplemented with 6 micrograms of vancomycin per mL. Rapid motility and pigment tests were performed on all enterococci isolated. A total of 82 motile and/or pigmented enterococci were isolated from 679 patients for a colonization rate of 12.1%. There were 43 Enterococcus gallinarum, 32 Enterococcus casseliflavus, 4 Enterococcus flavescens, and 3 Enterococcus mundtii identified. The E. gallinarum vancomycin MIC90 was 32 micrograms/mL and the E. casseliflavus vancomycin MIC90 was 8 micrograms/mL.

Anal Canal↗

Comparison of phenotypic and genotypic taxonomic methods for the identification of dairy enterococci.

The purpose of the study was to assess the phenotypic and genotypic taxonomic congruence in order to allow species allocation of dairy enterococci. A total of 364 enterococci isolated from ewes'milk and cheese from four Portuguese Registered Designation of Origin areas and 25 type and reference strains of Enterococcus spp. were characterized by a polyphasic taxonomical approach involving 40 physiological and biochemical tests, whole-cell protein profiles, amplification of 16S-23S intergenic spacer regions (ITS-PCR) and subsequent restriction analysis (ARDRA). Ribotyping was also performed with reference strains and a subset of 146 isolates. Numerical hierarchic data analysis showed that single-technique identification levels increase from the physiological and biochemical tests to the protein approach, being lower with ITS/ARDRA and ribotyping. Cross-analysis confirmed a higher unmatching level in all pairwise combinations involving physiological and biochemical data. Whole-cell protein profiles followed by ITS/ARDRA identified 89% of the enterococci. Reliable identification of enterococci from milk and cheese could be obtained by analysis of whole-cell protein profiles. ITS-PCR can be used to confirm E. durans and E. faecium and ARDRA further confirms E. faecalis. Results revealed E. faecalis, E. durans, E. hirae and E. faecium as the prevalent species, although species prevalence showed some degree of variation among the areas.

Animals↗

Mode of binding of fibrinogen, fibronectin and iron-binding proteins by animal enterococci.

Sixty-two animal enterococci were examined for their binding of bovine fibrinogen, porcine fibronectin, bovine lactoferrin, bovine apotransferrin and human holotransferrin in the particle agglutination assay (PAA). Individual strains expressed binding of selected glycoproteins to various degrees (0, 1, 2, 3), whereas bovine fibrinogen binding of enterococci from goats, rabbits and rodents was the strongest (3) in general. Porcine fibronectin was bound weakly (1 or 2) by enterococci from horses, dogs, poultry, rabbits and rodents, while most of the goat isolates and half of the dog feed isolates did not bind fibronectin (0). Bovine lactoferrin was bound especially by the isolates from rodents and rabbits. Bovine apotransferrin was bound very weakly (1) by only a few isolates. Human holotransferrin was bound to a greater extent than apotransferrin by some isolates from rabbits and rodents. Since multiresistant strains are preferred in our binding studies, enterococci were also examined for their antibiotic resistance pattern. Almost all investigated isolates were resistant at least to one antibiotic. However, some strains displayed resistance to five or six antibiotics of 10 antibiotics tested. In a study of the inhibitory effect of heparin, porcine mucin and hyaluronic acid, the greatest effect was observed after heparin treatment of bacterial cells. These observations, as well as the expression of heparin binding by most strains, may suggest that at least one mode of enterococcal attachment utilizes glycosaminoglycan chains present on the surface of adherent cells.

Animals↗

Bactericidal activity and synergy studies of BAL9141, a novel pyrrolidinone-3-ylidenemethyl cephem, tested against streptococci, enterococci and methicillin-resistant staphylococci.

BAL9141 has been reported to have inhibitory activity against methicillin-resistant Staphylococcus aureus (MRSA), many enterococci, and streptococci with various resistant patterns. BAL9141 potency was assessed by time-kill curves alone or with subinhibitory concentrations of gentamicin (MIC/4). BAL9141 exhibited bactericidal activity alone against all the streptococci and staphylococci. Among ampicillin-susceptible enterococci, BAL9141 was bactericidal against some strains, but no BAL9141 inhibition was observed of ampicillin-resistant Enterococcus faecium. The activity of BAL9141 with gentamicin was slightly enhanced (not synergy) or indifferent against staphylococci. BAL9141 demonstrated bactericidal action alone against Enterococcus faecalis and some E. faecium strains (-4.8 to -6.0 log10 CFU/mL), but static effects were also noted. Drug interactions with gentamicin showed early synergy (4-8 h) for all enterococci, and indifference or synergy at 24 h (no antagonism). BAL9141 (< or = 8 mg/L) showed promising bactericidal activity alone and synergy with gentamicin against some of the vancomycin-resistant enterococci tested.

Anti-Bacterial Agents↗

Clonal diversity of vancomycin-resistant enterococci from an outbreak in a tertiary care university hospital.

BACKGROUND: Enterococci have become important nosocomial pathogens and now account for approximately 12% of nosocomial infections. Enterococci can be transferred from patient to patient and from health care personnel to patient. We investigated the clonal diversity of vancomycinresistant enterococci (VRE) causing an outbreak of infections and attempted to determine the patterns of spread of these bacteria in a university hospital. METHODS: Ribotyping was used to examine the clonal diversity of 50 VRE isolates, including 23 from wounds, 14 from urine, 8 from blood, 3 from the rectum, 1 from drainage, and 1 from the cornea. RESULTS: Nine patients were infected with Enterococcus faecalis, 10 with Enterococcus faecium, 3 with both E faecalis and E faecium, and 1 with Enterococcus avium. The results suggest that the sources of the VRE infections included endogenous strains and strains acquired by transmission from attending staff or from the environment. Three patients were infected by both nosocomial and endogenous strains. CONCLUSIONS: These data suggest that the collection and analysis of several isolates from repeated specimens is necessary to obtain a fuller understanding of the epidemiology and population structure of antibiotic-resistant enterococci.

Anti-Bacterial Agents↗

Survival of enterococci during hospital laundry processing.

Much remains to be elucidated about the epidemiology of nosocomial enterococcal infections. Enterococci are, however, known to be relatively thermotolerant, and several studies have shown that under laboratory conditions many strains are able to survive the time/temperature combinations of the UK Department of Health recommendations for the decontamination of used linen (HSG(95)18). We therefore wished to investigate the efficacy of decontamination of enterococci from hospital linen in working hospital laundries. The thermotolerance of 40 strains of Enterococcus faecalis and Enterococcus faecium was first determined. Reduction by a factor of greater than 10(5) was achieved in only two of 40 strains after 3 min at 71 degrees C or 10 min at 65 degrees C, the time/temperature combinations specified by the Department of Health for the disinfection of used linen. During experimental challenge of 10 working hospital laundries, however, we demonstrated successful decontamination of laundry artificially contaminated with enterococci. This was shown to take place during the washing stage. Our study suggests that, despite the relative thermotolerance of enterococci, the time/temperature combinations specified in HSG(95)18 should be adequate for their decontamination in hospital laundries.

Decontamination↗

Aminoglycoside resistance in enterococci.

High-level aminoglycoside resistance in enterococci is mediated generally by aminoglycoside-modifying enzymes, which eliminate the synergistic bactericidal effect usually seen when a cell wall-active agent is combined with an aminoglycoside. Clinical microbiology laboratories currently screen for aminoglycoside resistance in enterococci by testing gentamicin and streptomycin susceptibility. If the recently detected aminoglycoside resistance genes, aph(2")-Ib, aph(2")-Ic, and aph(2")-Id, become more prevalent among clinical isolates, the approach for detecting susceptibility to aminoglycoside synergism in enterococci will require modification. More potent aminoglycosides need to be developed that will be resistant to modification by a broad spectrum of aminoglycoside-modifying enzymes present in enterococci.

Anti-Bacterial Agents↗

Clinical prevalence, antimicrobial susceptibility, and geographic resistance patterns of enterococci: results from the SENTRY Antimicrobial Surveillance Program, 1997-1999.

As part of the SENTRY Antimicrobial Resistance Surveillance Program, a total of 4998 strains of enterococci isolated from 1997 to 1999 were processed. The occurrence of enterococcal infections by species and site of infection was analyzed, as were the occurrence of vancomycin-resistant enterococci (VRE) and their resistance phenotypes and genotypes. Trends in antimicrobial susceptibility to a variety of agents (including experimental compounds) were also reported. Enterococci accounted for >9% of isolates from all bloodstream infections (BSIs) in North America. Ampicillin was active against strains from Latin America and Europe but not against those from the United States and Canada. US isolates were considerably more resistant to vancomycin (17% resistant strains in 1999) than were those from patients in the rest of the world. The highest proportion of VRE was observed among BSI isolates (81.7%). Quinupristin-dalfopristin, chloramphenicol, and doxycycline were the most active agents tested against VRE. The results of this study confirm the worldwide trend in increasing occurrence of enterococci and the emerging pattern of antimicrobial resistance among such isolates.

Anti-Bacterial Agents↗

Enterococci and vancomycin resistance.

The frequency of infections with multiply antibiotic-resistant gram-positive bacteria is increasing, and in some cases these organisms remain susceptible only to the glycopeptides vancomycin and teicoplanin. The appearance of transferable high-level glycopeptide resistance in enterococci--producing some strains that are now resistant to all available antibiotics--is thus a cause for concern. The enterococci readily colonize the bowel, spread rapidly among hospital patients, and transfer their antibiotic resistances widely among themselves and other gram-positive species. Glycopeptide resistance has not yet transferred in vivo to other significant pathogens, but experimental transfer to Staphylococcus aureus has been achieved in vitro. The emergence of glycopeptide-resistant enterococci has been encouraged by the increasing use of aminoglycosides, cephalosporins, and quinolones for the treatment of infections due to gram-negative bacteria and glycopeptides for infections due to staphylococci and clostridium difficile. In Europe this antibiotic pressure has been aggravated by the use of the glycopeptide avoparcin in animal feeds. The enterococci may now be poised to disseminate glycopeptide resistance among other more pathogenic gram-positive bacteria.

Animals↗

Epidemiology of the genetic elements responsible for acquired glycopeptide resistance in enterococci.

Five genotypes of acquired glycopeptide resistance have been documented in enterococci, with vanA and vanB being the most globally widespread and prevalent. Resistance results from the production of peptidoglycan precursors with reduced binding affinity for glycopeptides and is encoded by complex clusters of van genes. The prototype VanA element is Tn1546, a 10.8-kb transposon that carries the vanRSHAXYZ genes. Diverse VanA elements exist, but all share the vanRSHAX cluster and are believed to be derived from a Tn1546 progenitor. The sequences of these genes are remarkably conserved, with only a few point mutations identified. VanA elements do however vary by the presence of deletions and insertion sequences (IS) in nonessential genes (orf1, orf2, vanY, and vanZ) and intergenic regions. IS transposition probably plays a key role in VanA element evolution. By contrast, vanB gene clusters show greater sequence divergence. Three vanB alleles have been reported, of which vanB2 appears to be the most widespread, generally as part of Tn5382 and related elements. To date, only four Enterococcus faecium strains with VanD resistance have been reported, and each contained a distinct vanD allele. The VanE and VanG types have each been identified in single strains of Enterococcus faecalis. The existence of distinct genotypes, together with the allelic nature of vanB and vanD, suggests that van clusters have transferred to enterococci on multiple occasions from undefined donor species, with subsequent horizontal dissemination, particularly of VanA and VanB elements, among enterococci. Characterization of glycopeptide resistance elements yields information on their evolution and broadens our insights into the epidemiology of resistant enterococci.

Anti-Bacterial Agents↗

Association between antecedent intravenous antimicrobial exposure and isolation of vancomycin-resistant enterococci.

Vancomycin-resistant enterococci (VRE) have become important causes of nosocomial infections. This study evaluated the association between a variety of intravenous antimicrobial exposures and the isolation of VRE using two control groups: (1) a vancomycin-susceptible enterococci (VSE) group, to assess factors associated with development of VRE, and (2) a nonenterococci control group, to assess factors associated with positive cultures for enterococci without regard to vancomycin resistance. After adjusting for the effect of other antimicrobials, time at risk, and patient morbidity, compared to vancomycin-susceptible enterococci controls, exposures to imipenem (OR = 4.9, 95% CI = 1.6-14.1) and ceftazidime (OR = 2.6, 95% CI = 1.1-6.1) were significant predictors of VRE. When compared to nonenterococci controls, exposures to ampicillin (OR = 20.1, 95% CI = 1.5-263.1) and imipenem (OR = 5.1, 95% CI = 1.5-17.1) were significantly associated with VRE. Neither piperacillin nor vancomycin was associated with VRE compared to either control group. This study offers further evidence that the replacement of broad-spectrum cephalosporins by extended-spectrum penicillins, specifically piperacillin, may be effective in reducing VRE.

Adult↗

Effect of tylosin used as a growth promoter on the occurrence of macrolide-resistant enterococci and staphylococci in pigs.

This study was conducted to investigate the effect of tylosin used as a growth promoter on the occurrence of macrolide-resistant enterococci and staphylococci in pigs. Two identical feeding experiments with 10 pigs each were conducted to evaluate the effect of tylosin on the occurrence of erythromycin resistance among enterococci sampled from feces and Staphylococcus hyicus sampled by skin swabs. Half of the pigs were fed antibiotic-free feed and the other half feed with tylosin (30 microg/g) added. For each pig, weekly fecal samples and skin swabs were collected and examined for erythromycin-susceptible and erythromycin-resistant enterococci and S. hyicus, respectively. There was an immediate effect of tylosin on the fraction of resistant enterococci recovered, increasing by a factor 2.4; the effect on S. hyicus was more gradual, increasing at a rate of about 8% per day and totaling a five-fold increase over 20 days. Thus, a clear effect on resistance occurrence was demonstrated, not only in intestinal bacteria, but also on bacteria sampled from the skin.

Analysis of Variance↗

Distribution of the erm (B) gene, tetracycline resistance genes, and Tn1545-like transposons in macrolide- and lincosamide-resistant enterococci from pigs and humans.

The distribution of the erm (B) and the tetracycline resistance genes tet(K), tet(L), tet(M), tet(O), and tet(S) was investigated among macrolide- and lincosamide-resistant enterococci originating from humans, pigs, and pork carcasses. The presence of transposons of the Tn916/Tn1545 family was also traced in these isolates. Furthermore, the porcine strains were tested for the presence of glycopeptide resistance genes vanA and vanB. The erm(B) gene was found in 85% of the porcine and in all human isolates. Ninety-eight percent of the porcine and 89% of the human erm(B)-positive enterococci carried the tet(M) gene. Seventy-seven percent and 70%, respectively, of these strains harbored a Tn1545-like element. Tet(L) was observed in 68% of the porcine and in 65% of the human enterococci. The other tetracycline resistance genes were very rare and the glycopeptide resistance genes vanA and vanB were not detected among the porcine isolates. The similar frequencies of resistance genes and the highly mobile Tn1545-like transposon among porcine and human enterococci might indicate exchange of resistant strains or their resistance genes between humans and pigs or the existence of a common reservoir.

Animals↗

The prevalence of fecal colonization of enterococci, the resistance of the isolates to ampicillin, vancomycin, and high-level aminoglycosides, and the clonal relationship among isolates.

The gastrointestinal tract carriage of enterococci was searched in 150 hospitalized patients and 100 outpatients, and clonal relatedness of the isolates and their resistance to ampicillin, vancomycin, and high-level streptomycin and gentamicin were investigated. A stool sample or rectal swab collected from each patient was inoculated into appropriate media within an hour. Enterococcus species were identified by using conventional biochemical tests, API-20 Strep assay, and BBL crystal kit. Antibiotic susceptibility tests were performed using Kirby-Bauer disk diffusion method. A polymerase chain reaction (PCR) was used to detect vanA and vanB genes. Pulsed-field gel electrophoresis (PFGE) and arbitrarily primed-polymerase chain reaction (AP-PCR) methods were used for molecular typing of the strains. Enterococci were isolated from 90 (60%) of the specimens collected from 150 inpatients. Of these 90 isolates, 37 (41%) had high-level gentamicin resistance, 36 (40%) had high-level streptomycin resistance, and 50 (55.6%) had ampicillin resistance. Fecal colonization was found in 30% of the outpatients. Resistances to ampicillin, high-level streptomycin, and gentamicin were 13%, 10%, and 3%, in these patients' isolates, respectively. No vancomycin-resistant enterococci were detected by both agar diffusion and PCR assays in our study. Both typing procedures were applied on 78 Enterococcus strains isolated from inpatients. AP-PCR typing showed that 30 (50.8%) of the 59 E. faecium and 5 (50%) of the 10 E. faecalis strains were clonally related. These values were found to be 12 (20.3%) and two (20%) by PFGE, respectively. The typing procedures did not find any clustered strains in the six E. durans and three E. avium isolates. Neither PFGE nor AP-PCR result was significantly different among the sensitive and resistant strains. Our results indicate that the high prevalence of colonization with ampicillin and highlevel aminoglycoside-resistant enterococci is an important problem in our medical center. The high clonal diversity among the isolates indicates limited spread of antibiotic-resistant strains between patients.

Aminoglycosides↗

Synergistic bactericidal activity of rat serum with vancomycin against enterococci.

Animal models of infectious diseases may not predict clinical efficacy when species-related factors come into play. Recently, unexpected bactericidal activity of vancomycin alone against enterococci was observed in a rat model of endocarditis. A factor or factors in rat serum, but not rabbit or human serum, enhanced in vitro killing by vancomycin in four of five clinical isolates of enterococci. Bactericidal activity was maintained on dilution of rat serum to 5.0% and after exposure of serum to 56 degrees C for 30 min. Activity was lost by heating at 60 degrees C for 2 h, ultrafiltration, or absorption with bentonite or heat-killed bacteria. Rat serum appears to contain a factor or factors that contribute bactericidal activity to vancomycin, a drug normally bacteriostatic for these enterococci. The mechanism by which this factor enhances killing of enterococci by vancomycin is unknown.

Animals↗

In-vitro activity of azlocillin and other beta-lactam antibiotics against enterococci.

The susceptibility of enterococci to 16 beta-lactam antibiotics (5 penicillins, 11 cephalosporins) was evaluated by determining MICs. All cephalosporins are ineffective against these bacteria. Penicillin G should not be used to treat infections with enterococci. For less serious infections due to enterococci, ampicillin is certainly the drug of first choice. Azlocillin, mezlocillin and piperacillin are the most effective drugs to prevent superinfections with enterococci in clinically and bacteriologically defined situations.

Anti-Bacterial Agents↗

Penicillin-aminoglycoside synergy and post-antibiotic effect for enterococci.

The post-antibiotic effect (PAE) on seven strains of enterococci was studied using a novel ATP method and conventional viable counting. There was a good correlation between the results of the two techniques. In strains exhibiting low-level resistance to penicillin and streptomycin, PAEs of 1 to 2.4 h (mean 1.8 h) resulted from exposure to penicillin; shorter PAEs were induced by streptomycin (0.2-0.3 h; mean 0.2 h). Addition of streptomycin to pencillin increased the duration of penicillin-induced PAEs by two-fold to three-fold. Enterococci exhibiting high-level streptomycin resistance but low-level penicillin resistance did not exhibit a PAE with streptomycin but exhibited a short PAE with gentamicin (0.3-0.6 h). For these strains, the addition of gentamicin, but not streptomycin, to penicillin increased the penicillin-induced PAEs. Penicillin alone or in combination with streptomycin or gentamicin did not induce PAEs for a single strain of Enterococcus faecium which exhibited high-level resistance to both penicillin and streptomycin. During penicillin-induced PAEs, extracellular ATP was detectable and only during this time period were enterococci susceptible to the action of gentamicin. The addition of aminoglycosides to penicillin not only extended the PAE for enterococci but also the periods when organisms leaked ATP and were susceptible to growth inhibition by gentamicin.

Adenosine Triphosphate↗

The activity in vitro of trospectomycin sulphate (U-63366F) against aminoglycoside-resistant enterococci.

Trospectomycin (U-63366F), a 6'-propyl analogue of spectinomycin, was tested against aminoglycoside-resistant enterococci. The MIC90 for Enterococcus faecalis was 4 mg/l and that for E. faecium was 8 mg/l. Trospectomycin alone was not bactericidal for enterococci, with MBC90 4096 mg/l for both E. faecalis and E. faecium. The addition of commercially available polyvalent immunoglobulin decreased significantly both the MICs and the MBCs and the rendered trospectomycin bactericidal for enterococci. Chequerboard titration of a combination of trospectomycin with ampicillin revealed an FIC index of 1.0 for all the isolates tested. Time-kill curves also did not show any enhancement of bactericidal activity of ampicillin when combined with trospectomycin. A combination of ampicillin and gentamicin was synergistic for enterococci under similar experimental conditions. Trospectomycin can be used as a safe alternative to aminoglycosides or beta-lactam antibiotics in enterococcal infection where bactericidal activity is not required, or in the event of serious side effects from these two classes of antibiotics.

Aminoglycosides↗