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PubMed2003

Antimicrobial proficiency testing of National Nosocomial Infections Surveillance System hospital laboratories.

OBJECTIVE: The National Nosocomial Infections Surveillance (NNIS) System personnel report trends in antimicrobial-resistant pathogens. To validate select antimicrobial susceptibility testing results and to identify test methods that tend to produce errors, we conducted proficiency testing among NNIS System hospital laboratories. SETTING: NNIS System hospital laboratories in the United States. METHODS: Each laboratory received five organisms (ie, an imipenem-resistant Serratia marcescens, an oxacillin-resistant Staphylococcus aureus, a vancomycin-resistant Enterococcus faecalis, a vancomycin-intermediate Staphylococcus epidermidis, and an extended-spectrum beta-lactamase (ESbetaL)-producing Klebsiella pneumoniae). Testing results were compared with reference testing results from the Centers for Disease Control and Prevention. RESULTS: Of 138 laboratories testing imipenem against the Serratia marcescens strain, 110 (80%) correctly reported minimum inhibitory concentrations (MICs) or zone sizes in the resistant range. All 193 participating laboratories correctly reported the Staphylococcus aureus strain as oxacillin resistant Of the 193 laboratories, 169 (88%) reported correct MICs or zone sizes for the vancomycin-resistant Enterococcus faecalis. One hundred sixty-two (84%) of 193 laboratories demonstrated the ability to detect a vancomycin-intermediate strain of Staphylococcus epidermidis, however, disk diffusion performed poorly when testing both staphylococci and enterococci with vancomycin. Although laboratory personnel correctly reported nonsusceptible extended-spectrum cephalosporins and aztreonam results for K. pneumoniae, only 98 (51%) of 193 correctly reported this organism as an ESbetaL producer. CONCLUSION: Overall, NNIS System hospital laboratory personnel detected most emerging resistance patterns. Disk diffusion continues to be unreliable for vancomycin testing of staphylococci and must be used cautiously for enterococci. Further education on the processing of ESbetaL-producing organisms is warranted.

Bacteria↗

PubMed1990

Mechanisms of resistance of enterococci to beta-lactam antibiotics.

Two mechanisms are responsible for resistance of enterococci to beta-lactam antibiotics: alterations of penicillin-binding proteins and production of a beta-lactamase. The latter has been found in a few clinical isolates of Enterococcus faecalis, whereas the former appears to account for resistance in most strains. A correlation has been established between the amount of a particular penicillin-binding protein which has a low affinity for penicillin and the level of resistance. The higher activity of some penicillins, as compared to cephalosporins, has been related to the relatively higher affinity for these penicillins of the penicillin-binding protein involved in the mechanism of resistance. Alterations in the autolytic enzyme pattern have been associated with the paradoxical response to bactericidal activity of penicillin often exhibited by Enterococcus faecalis clinical isolates.

Anti-Bacterial Agents↗

PubMed1996

Incidence of high-level gentamicin resistance in enterococci at Johannesburg Hospital.

OBJECTIVE: To document the incidence of high-level gentamicin resistance (HLGR) in enterococcal isolates at Johannesburg Hospital. DESIGN: Survey of laboratory isolates. SETTING: Academic hospitals. BACTERIAL STRAINS: Consecutive samples of enterococcal isolates. MAIN OUTCOME MEASURE: The incidence of HLGR in enterococcal isolates. RESULTS: The incidence of HLGR was 26.5% of Enterococcus faecalis isolates and 20% of E. faecium isolates grown during the study period. CONCLUSIONS: High-level gentamicin resistance is common among enterococci isolated at Johannesburg Hospital, and this observation must be considered in defining strategies for the management of invasive enterococcal infections in the future.

Anti-Bacterial Agents↗

PubMed2001

Bacteriocin production in vancomycin-resistant and vancomycin-susceptible Enterococcus isolates of different origins.

Bacteriocin production was determined for 218 Enterococcus isolates (Enterococcus faecalis [93] and E. faecium [125]) obtained from different origins (human clinical samples [87], human fecal samples [78], sewage [28], and chicken samples [25]) and showing different vancomycin susceptibility patterns (vancomycin resistant, all of them vanA positive [56], and vancomycin susceptible [162]). All enterococcal isolates were randomly selected except for the vancomycin-resistant ones. A total of 33 isolates of eight different bacterial genera were used as indicators for bacteriocin production. Forty-seven percent of the analyzed enterococcal isolates were bacteriocin producers (80.6% of E. faecalis and 21.6% of E. faecium isolates). The percentage of bacteriocin producers was higher among human clinical isolates (63.2%, 81.8% of vancomycin-resistant isolates and 60.5% of vancomycin-susceptible ones) than among isolates from the other origins (28 to 39.3%). Only one out of the 15 vancomycin-resistant isolates from human fecal samples was a bacteriocin producer, while 44.4% of fecal vancomycin-susceptible isolates were. The bacteriocin produced by the vanA-containing E. faecium strain RC714, named bacteriocin RC714, was further characterized. This bacteriocin activity was cotransferred together with the vanA genetic determinant to E. faecalis strain JH2-2. Bacteriocin RC714 was purified to homogeneity and its primary structure was determined by amino acid sequencing, showing an identity of 88% and a similarity of 92% with the previously described bacteriocin 31 from E. faecalis YI717. The presence of five different amino acids in bacteriocin RC714 suggest that this could be a new bacteriocin. The results obtained suggest that the epidemiology of vancomycin resistance may be influenced by different factors, including bacteriocin production.

Amino Acid Sequence↗

PubMed1997

Light microscopic, immunohistochemical, and electron microscopic features of amyloid arthropathy in chickens.

Amyloid arthropathy has been recently recognized as a spontaneous syndrome in chickens. Predominantly, femorotibial and tarsometatarsal joints were affected, showing (peri) articular orange amyloid deposits. Immunohistochemical evaluation revealed the amyloid to be of the reactive type. Induction of amyloid arthropathy in chickens was carried out using a single intravenous injection of Enterococcus faecalis cultures. In the naturally occurring and the induced cases, amyloid deposits were found in the hypertrophic synovial villi and in the articular cartilage, particularly in the superficial layer and in the nutritional blood vessel walls. Highly sulfated glycosaminoglycans (GAGs) were found in the amyloid deposits. Ultrastructurally, bundles of amyloid fibrils were seen in invaginations of synoviocytes and chondrocytes. Immunogold electron microscopy failed to reveal signs of intracellular amyloid formation. The predilection site for amyloid deposition in the major leg joints of the chickens with reactive amyloid could be explained by the arthritic condition caused by Enterococcus faecalis bacteriaemia. The polyarthritis triggers hepatic acute phase protein synthesis and increases the vascular serum amyloid A (SAA) supply to the joint. Inflammatory and degenerative changes in the articular cartilage and adjoining tissues result in an increase of highly sulphated GAGs, which are considered to enhance deposition of SAA as amyloid.

Amyloid↗

PubMed2002

Laboratory contamination of specimens with quality control strains of vancomycin-resistant enterococci in Ontario.

Cross-contamination with laboratory control strains of vancomycin-resistant Enterococcus faecalis was documented in 15 clinical specimens from nine clinical microbiology laboratories in Ontario, Canada. Laboratories should be alert to the possibility of contamination of specimens with vancomycin-resistant enterococci from the laboratory environment. Molecular typing of strains may assist in elucidating the source of such contamination.

DNA, Bacterial↗

PubMed1992

Concomitant high-level vancomycin and penicillin resistance in clinical isolates of enterococci.

Enterococci are important nosocomial pathogens among which resistance to multiple antibiotics is being recognized with increasing frequency. We characterized three clinical isolates from three New York City hospitals that demonstrated concomitant resistance to vancomycin (one VanA, two VanB phenotypes) and high-level resistance to penicillin. Two Enterococcus faecium strains were intrinsically highly resistant to penicillin and showed very low affinity for penicillin of penicillin-binding protein 5. Unlike previously described glycopeptide-resistant enterococci, these strains were not hypersusceptible to beta-lactam agents after vancomycin induction, and combinations of penicillin and vancomycin were not synergistic against them. A third isolate, Enterococcus faecalis, produced beta-lactamase. Two of the three strains were also highly resistant to all aminoglycosides. Emergence of concomitant high-level resistance to multiple antibiotic classes among enterococci considerably narrows the therapeutic options for treatment of infections due to these opportunistic pathogens.

Aged↗

PubMedDate not supplied

In vitro susceptibility to everninomycin of gram-positive nosocomial pathogens isolated from intensive care units in Germany.

BACKGROUND: Gram-positive bacteria are increasingly found to be causative pathogens in nosocomial infections, and the occurrence of vancomycin resistance in enterococci as well as staphylococci has prompted the investigation of alternative antimicrobial agents active against these strains. Everninomycin, a new oligosaccharide antibiotic, has excellent in vitro activity against gram-positive bacteria, including those resistant to vancomycin. However, avilamycin, a related compound, has been used in Europe as a growth promoter in animal food for years and concern has been raised that cross-resistance in clinical isolates may arise. METHODS: We studied a collection of 268 nosocomial gram-positive isolates from intensive care unit patients with nosocomial pneumonia, urinary tract infection and sepsis, using standard in vitro susceptibility testing. RESULTS: It could be shown that all species tested were exquisitely sensitive to everninomycin (MIC(90) of 0.38 microg/ml for Staphylococcus aureus, 0.5 microg/ml for enterococci and 0.75 microg/ml for coagulase-negative staphylococci). Furthermore, no difference could be observed between methicillin-resistant and methicillin-sensitive S. aureus or between Enterococcus faecium and Enterococcus faecalis. CONCLUSIONS: These results suggest that everninomycin is a promising antibiotic for the treatment of nosocomial infections in intensive care unit patients and that the use of a related substance as an additive in animal food has not yet promoted resistance in clinical isolates.

Aminoglycosides↗

PubMed2006

Use of competitive DNA hybridization to identify differences in the genomes of bacteria.

Although recent technological advances in DNA sequencing and computational biology now allow scientists to compare entire microbial genomes, comparisons of closely related bacterial species and individual isolates by whole-genome sequencing approaches remains prohibitively expensive for most laboratories. Here we report the development and testing of a biochemical approach for targeted sequencing of only those chromosomal regions that differ between two DNA preparations. The method, designated GFE (genome fragment enrichment) uses competitive solution hybridization and positive selection to obtain genomic DNA fragments that are present in one pool of fragments but not another. Repeated comparisons of the genomes of Enterococcus faecalis and E. faecium led to the identification of 225 putative genome-specific DNA fragments. Species and strain variations within these fragments were confirmed by both experimental and bioinformatic analyses. The E. faecalis genome-specific sequences identified included both a preponderance of those predicted to encode surface-exposed proteins, as well as several previously described unique marker regions embedded within highly conserved rrn operons. The GFE strategy we describe efficiently identified genomic differences between two enterococcal genomes, and will be widely applicable for studying genetic variation among closely related bacterial species.

DNA, Bacterial↗

PubMed2004

Characterization of faecal enterococci from rabbits for the selection of probiotic strains.

AIMS: To characterize the facultative anaerobic intestinal microbiota of healthy rabbits, especially enterococci, for the selection of potential probiotic strains. METHODS AND RESULTS: Phenotypic and molecular methods were used to identify enterococcal isolates. Results obtained indicated that enterococcal microbiota widely varied among individuals both in size and in composition. Antibacterial and haemolytic activities, and resistance to acid and bile salts were determined. A small group of strains produced bacteriocins active against listeriae and indigenous clostridia and therefore they were selected as potential probiotics. One such strain, 8G, was assayed for colonization capacity. Results obtained suggested that the fate of the introduced strain depended on the composition of the enterococcal indigenous microbiota. CONCLUSIONS: Enterococcus faecalis and Ent. faecium are the predominant enterococcal species in the gut of rabbits. Other species of lactic acid bacteria were not recovered. SIGNIFICANCE AND IMPACT OF THE STUDY: The enterococcal intestinal microbiota of healthy rabbits has been characterized in detail. Monitoring the fate of an introduced probiotic in vivo is required in order to evaluate potential probiotic strains.

Animals↗

PubMed1992

The effect of selective decontamination of the digestive tract on gastrointestinal enterococcal colonization in ITU patients.

OBJECTIVE: The effect of selective decontamination of the digestive tract (SDD) on Intensive Therapy Unit (ITU)-acquired enterococcal infection and colonization was studied. Changes in the predominant species isolated and resistance patterns to antimicrobial agents were also studied. DESIGN: Three groups were investigated: historical control (HC), contemporaneous control (CC) and patients receiving SDD (topical polymyxin, amphotericin B and tobramycin throughout ITU stay with intravenous ceftazidime for the first 3 days only). SETTING: Adult general ITU with 7 beds. PATIENTS: Patients with a nasogastric tube in situ and who were likely to remain in ITU for 48 h or longer were recruited. RESULTS: Enterococcal infections occurred in 3 of 84 HC patients and 2 of 91 CC patients. There were no unit-acquired enterococcal infections in the SDD group. There were 140 episodes of enterococcal colonization occurring in 112 patients, with significantly more in the SDD and CC groups (p < 0.05. There were no significant differences in antibiotic sensitivities between the three groups. Enterococcus faecalis was the most frequently isolated species. CONCLUSION: SDD does not predispose to enterococcal infection but does encourage colonization in patients receiving the regimen and other patients in ITU at the same. There is a complex interaction of factors which influence faecal flora and the likelihood of patients becoming colonized or infected with enterococci.

Administration, Buccal↗

PubMed1992

Characterization of enterococcal isolates by restriction enzyme analysis of genomic DNA.

A restriction enzyme analysis (REA) of chromosomal DNA for the intra-species characterization of enterococci is reported. The DNA was extracted by a rapid method and digested with the restriction enzyme Sal I to provide a characteristic 'fingerprint' consisting of 10-20 bands in the 1.6-5.0 kb range. One hundred and eighty enterococcal isolates were examined; 5 were type strains, 15 from an out-patient clinic and 160 from a geographically isolated British Antarctic Survey Base. The epidemiologically unrelated out-patient clinic isolates gave readily distinguishable patterns, whereas isolates from the geographically isolated community showed evidence of colonization. This technique provided a highly discriminatory method of isolate characterization for Enterococcus faecalis, E. faecium and E. durans suitable for epidemiological studies. A sample of isolates were probed with 16 + 23 S ribosomal RNA from Escherichia coli. Discrimination between isolates was poorer than with REA, although good correlation was observed between the results of the two techniques.

Antarctic Regions↗

PubMed2003

Anomalies in species identification of enterococci from veterinary sources using a commercial biochemical identification system.

AIMS: A commercial biochemical panel ID kit was used to identify presumptive enterococci isolates of veterinary or agricultural origin obtained during different steps of culture. METHODS AND RESULTS: Fifty isolates identified as enterococci using a genus PCR assay were tested for genus and species identification using the BBL Crystal Identification Gram-Positive ID kit (Becton Dickinson, Sparks, MD, USA). Following sub-culture of the isolates three times, 59% agreement with the original panel ID was obtained. After four and six sub-cultures, percentage agreement increased to 61 and 64%, respectively. Nineteen of the 50 cultures were identified as both Enterococcus faecalis and E. faecium. CONCLUSIONS: As a result of the variability between speciation of isolates following re-culture, additional methods for speciation are warranted. SIGNIFICANCE AND IMPACT OF THE STUDY: This study suggests that the identification of the genus and species of non-human enterococcal isolates can vary greatly during successive passages when using this kit.

Animals↗

PubMed2004

In vitro effect of levofloxacin and vancomycin combination against high level aminoglycoside-resistant enterococci.

The in vitro effects of levofloxacin and vancomycin in combination were evaluated against high level aminoglycoside-resistant (HLAR) enterococci using chequerboard and time-kill curve techniques. We examined 28 strains of enterococci comprising 17 Enterococcus faecalis, 10 E. faecium and one E. durans. The combination of vancomycin and levofloxacin had indifferent activity against all isolates according to chequerboard microdilution method, but was synergistic for two isolates, one E. faecium and one E. faecalis, using the time-kill curve method. Both strains were levofloxacin resistant and had high level aminoglycoside resistance to gentamicin and streptomycin. Antagonism was not detected in any strain. The results of this study suggested that the combination of vancomycin with levofloxacin does not often show synergistic effect against high level aminoglycoside-resistant enterococci.

Aminoglycosides↗

PubMedDate not supplied

[Study of probiotic properties of bacteriocinogenic Lactobacillus plantarum G3/3(13) strain].

Bacteriocinogenic culture Lactobacillus plantarum G3/3(13) was tested according to the basic requirements to probiotic strains. The reidentification of the strain was carried out using a set of methods including PCR. It was confirmed that the strain belongs to the species L. plantarum. The studied culture expressed antagonistic activity with respect to Enterococcus faecalis and E. faecium. The susceptibility test to 57 antibiotics of different groups showed that the strain was resistant to oxacillin, cefepyme, lomefloxacin, polymyxin, nalidixic acid and the antifungal antibiotics. High adhesive properties of the microorganism to the human epithelial cells (medium adhesion index was 4.6-5.6; microorganism adhesion index was 4.9-5.9) were shown. Obtained results allow to recommend L. plantarum G3/3(13) for the construction of probiotics.

Anti-Bacterial Agents↗

PubMed1994

Revised estimates of enterococcal chromosomal sizes.

We previously reported that the chromosomal sizes of four strains of enterococci ranged from 2,045 to 2,761 kb. Extensive analysis and mapping subsequently confirmed the size of Enterococcus faecalis strain OG1 as 2,825 kb (prior size estimate range, 2,750-2,761 kb) (Murray et al., J. Bacteriol. 175, 5216, 1993). However, using variable conditions of electrophoresis and additional digestions, revised size estimates for the other strains are 2,852-3,093 kb for E. faecalis strain JH2-2 (prior range, 2,008-2,135 kb), 2,910-3,065 kb for E. faecalis strain HH67 (prior range, 2,170-2,288 kb), and 2,334-2,558 for E. faecium strain GE-1 (prior range, 2,045-2,155 kb). The earlier underestimations of the chromosomal sizes were due to the inconsistent presence of a large fragment, likely caused by shearing of the DNA during handling, causing it to be considered a partial digestion product, and failure to resolve multiple fragments of the same approximate size.

Chromosomes, Bacterial↗

PubMed2006

[Genotypes of aminoglycoside-modifying enzyme and clinical study of high-level gentamycin resistant enterococcus].

OBJECTIVE: To determine the antibiotics resistance, aminoglycoside-modifying enzymes and homology of high-level gentamycin resistant enterococcus in clinical specimens. METHODS: The high-level gentamicin resistant (HLGR) isolates were screened by the agar method and the resistance of 14 antimicrobial agents was determined by K-B method. The aminoglycoside-modifying enzyme genes were detected by polymerase chain reaction (PCR). Pulsed-field gel electrophoresis (PFGE) was used to analyze the homology of HLGR isolates. RESULTS: The ratio of HLGR was 64.2% (68/106). Among the HLGR,there were no isolates resistant to linezolid, vancomycin and tecoplanin, and Enterococcus faecium was more resistant to beta-lactam antibiotics and quinolone than Enterococcus faecalis. The positive rate of aac(6')-Ie-aph(2')-Ia was 92.6% and 3 isolates had the resistance gene mostly similar to aph(2')-Id. And among 51 HLGR isolates from the hospitalized patients, PFGE grouped 17 E. faecalis isolates into 4 clusters (A-D), and 33 E. faecium isolates into 8 clusters (A-H) with A cluster as predominant. CONCLUSION: HLGR has become the important antibiotic resistance bacteria which results in nosocomial infection; and aac(6')-Ie-aph(2')-Ia is the main aminoglycoside-modifying enzyme gene which causes HLGR.

Drug Resistance, Bacterial↗

PubMed2005

Survivability of vancomycin resistant enterococci and fitness cost of vancomycin resistance acquisition.

AIMS: To investigate the survivability of vancomycin resistant enterococci (VRE) under dry starvation conditions and the fitness cost of vancomycin resistance. METHODS: VRE colonies on cotton swabs were incubated at room temperature in a sterile box and cultured weekly until cultures no longer showed growth. Negative swabs inoculated into brain heart infusion (BHI) broth were subcultured to blood agar after 24, 48, and 72 hours of incubation to resuscitate viable but non-culturable cells. Stability of the vancomycin resistance determinant and of the DNA fingerprint pattern was determined by multiplex polymerase chain reaction (PCR) and repetitive PCR, respectively. Tests for fitness cost were carried out on the same VRE isolates and 28 hospital vancomycin sensitive enterococci (VSE) isolates by incubation and measurement of optical density using a microplate reader and comparing maximum growth rate and lag phase duration between VRE and VSE, using independent samples t tests. RESULTS: Mean maximum time of recovery by primary culture was 8.5 weeks for Enterococcus faecalis VRE and 21.8 weeks for E. faecium VRE. Two of two E. faecalis isolates were resuscitated after 24 hours in BHI broth, and two of five E. faecium isolates after 72 hours. No fitness cost of vancomycin resistance was demonstrated. CONCLUSIONS: VRE can survive for prolonged periods in a dry starvation state, retaining their genetic complement, including vancomycin resistance determinants, and show little or no fitness cost of vancomycin resistance. Thus, the rate of entry required for VRE to become, and remain, endemic in the community is relatively small.

Cross Infection↗