vanC Gene Research Articles

Detection of VanA, VanB and VanC Genes in Vancomycin Resistant Enterococci in Zagazig University Hospitals

Background: Enterococci have emerged recently as a nosocomial pathogen especially in immunocompromized individuals and in intensive care units. Objectives: This study was carried out to detect incidence of VRE to detect genotypes of vancomycin resistance Enteroccci by PCR and to do a comparison between the results MIC of VRE and PCR results. Methodology: This study was conducted from May 2013 to January 2014. It was conducted on 560 hospitalized patients in Zagazig University Hospitals including 270 males and 290 female. The specimens were collected from patients suffering from any infections. Examination of isolates by conventional methods and API 20 Strep system. Susceptibility patterns of VRE isolates to antibiotics were determined by both disc diffusion method and E test. The genotypic detection of vancomycin resistance among enterococcal isolates was done by using polymerase chain reaction (PCR). Results: This study showed that the overall incidence of vancomycin resistance among Enterococci was 35.2% (57 isolates). The study showed that 38 of isolates (66.7%) had VanA gene and their MIC were more than 64 µg/ml. Twelve (21.05%) of isolates had VanB gene and their MIC also were more than 64 µg/ml. Five of isolates (8.8%) had VanC gene. MIC of these 5 isolates were between 4 and 16 µg/ml. Our study showed that VanA – PCR had the highest sensitivity to detect resistant isolates in comparison to VanB – PCR and VanC – PCR. Conclusion: The incidece of VRE in Zagazig University Hospitals was significantly high. The commonest genotype of vancomycin resistance in Enterococci was the VanA genotype. Other genotypes seen were VanB and VanC (low level resistance to vancomycin). VanA – PCR had the highest sensitivity to detect resistant isolates in comparison to VanB – PCR and VanC – PCR.

Drug resistance in Enterococcus species in a tertiary level hospital in Assam, India

Drug resistance in Enterococcus species in a tertiary level hospital in Assam, India

Emergence of antimicrobial resistance and virulence factors among the unusual species of enterococci, from North India.

Several enterococcal species are increasingly being reported from clinical infections, besides the major species. This study was undertaken to determine the prevalence of unusual enterococcal species and their antimicrobial susceptibility patterns, virulence factors, and molecular characterization. The study was conducted in Department of Microbiology and associated Tertiary Care University Hospital in North India. Enterococcal isolates were collected for a period of 2 years from clinical specimens. Identification and elaborate phenotypic characterization was done biochemically. All the isolates were tested by Kirby-Bauer disc diffusion method and breakpoint minimum inhibitory concentration for susceptibility against standard antibiotics. Screening for vancomycin-resistant enterococci (VRE), high-level aminoglycoside resistance was done on brain heart infusion agar incorporated with 6 μg/ml vancomycin, 500 μg/ml gentamicin, and 2000 μg/ml streptomycin, respectively. VRE isolates were tested for the presence of vanA, vanB, and vanC genes and high-level gentamicin resistant (HLGR) isolates for aac-6'- aph-2' gene by polymerase chain reaction (PCR). Hemolysin and gelatinase production, hemagglutination and biofilm formation were detected along with asa1, gelE, esp, hyl, and cylA genes by multiplex PCR. Of 403 enterococci, 93 (23.07%) isolates were identified as unusual species and atypical variants. Resistance of 52.68%, 46.23%, 44.08%, and 6.45% for ampicillin, ciprofloxacin, high strength gentamicin, and vancomycin, respectively were noted. Presence of vanC gene in Enterococcus gallinarum and Enterococcus casseliflavus isolates and vanA gene in Enterococcus durans and Enterococcus hirae and aac-6'- aph-2'' gene was found in 33.14% (14/41) of the HLGR isolates. The most frequent virulence factor was biofilm production. Only a few isolates harbored asa1 (2), gelE (9), and hyl (3) genes. Considerable prevalence of pathogenic unusual species of enterococci was seen along with their emerging drug resistance and virulence. Complete identification and routine speciation is essential to limit their emergence as major species in near future.

Gastrointestinal Colonization with Vancomycin-Resistant Enterococci In Hospitalized and Outpatients.

BACKGROUND:The incidence of infection and intestinal colonization with vancomycin resistant enterococci (VRE) is increasing in many countries in the last decade. Concerning the difficult antimicrobial treatment of infections caused by VRE, decreasing the incidence and prevalence of these infections is an important factor in VRE-induced morbidity and mortality control.AIM:To determine the prevalence of gastrointestinal colonization with vancomycin resistant enterococci in hospitalized and outpatients, and to determine the genetic base of the vancomycin resistance in VRE isolates.MATERIAL AND METHODS:Seven hundred and eighty stool specimens were investigated for the gastrointestinal carriage of vancomycin-resistant enterococci (VRE). Susceptibility to vancomycin was tested in all isolates by disk-diffusion test and E-test (AB Biodisk, Sweden). Determined vancomycin resistant enterococci were than tested for detection of vanA, vanB and vanC genes by PCR.RESULTS:Vancomycin resistant strains of enterococci were isolated from 46 (16.1 %) of the 285 hospitalized patients and 5 (7.7 %) of the 65 patients living in the community (p < 0.05). The most of the highly resistant enterococci strains to vancomycin (95.2 %), were identified as E. faecium. Minimal inhibitory concentrations (MICs) to vancomycin in all 39 vanA genotypes of E. faecium and two vanA genotypes of E. fecalis were > 256 μg/ml. Three vanB genotypes of E. faecium and one vanB genotype of E. faecalis had MICs of 32 μg/ml. All six vanC genotypes of E. gallinarum had MICs of 8 μg/ml. All vanA genotypes of VRE were highly resistant to vancomycin, with MICs above 256 μg/ml. Three vanB genotypes of VR E. faecium and one VR E. fecalis were resistant, with MICs 32 μg/ml. vanC genotypes of VR E. gallinarum were intermediate resistant to vancomycin with MICs of 8 μg/ml.CONCLUSIONS:The prevalence of vancomycin resistant enterococci in Republic of Macedonia was 2-fold higher in hospitalized than in outpatients. VanA genotype was dominant in isolates of E. faecium and it was highly associated with the MIC values above the 256 μg/ml. Since most of the enterococcal infections are endogenous, there is a need for screening the colonization of patient’s intestinal flora with VRE at the hospital entry. Identification and genotyping of faecal enterococci, together with their susceptibility testing to vancomycin, could be useful marker for the infection control.

Detection of vanC1 and vanC2 Genes in an Enterococcal Isolate and vanC Genes in non-Motile Enterococcus spp.

Background: In recent decades, bacterial antibiotic resistance (especially in enterococci) has become a significant problem for human and veterinary medicine. One of the most important antibiotic resistances in enterococci, vancomycin resistance, is encoded by van gene family. Objectives: The aim of this study was to investigate antibiotic resistance to vancomycin in enterococci and thegenes responsible for this resistance. Materials and Methods: Two-hundred and thirty enterococcal isolates from pigs (207 isolates), chickens (15 isolates) and humans (eight isolates) were phenotypically and genotypically tested for resistance to vancomycin by minimum inhibitory concentration (MIC) and polymerase chain reaction (PCR). The van genes were confirmed by gene sequencing. Results: Of the total isolates, 19% were phenotypically resistant to vancomycin, while nearly 15% contained either vanC1 or vanC2 gene. One resistant E. casseliflavus isolate with pig origin (MIC > 8 μg/mL) contained both vanC1 and vanC2 genes. Furthermore, one vanC1 was found in a sensitive E. faecalis isolate of pig origin (MIC ≤ 4 μg/mL) and one vanC2 in a resistant E. faecium isolate of chicken origin (MIC > 32 μg/mL). These genes were not accompanied by other van genes. Other detected genes were vanA in 11 E. faecium isolates of chicken origin (MIC > 32 μg/mL). No vanB genes were found. Gene sequencing results showed 100% identity with GenBank reference genes. Conclusions: The current report is the first report on the detection of vanC1 and vanC2 genes in one enterococcal species with pig origin. This report is important as it proves the horizontal transfer of various vanC genes to one species possibly due to the compatibility class of plasmids. Furthermore, detection of vanC genes in E. faecalis and E. faecium isolates is important as it suggests that resistance to vancomycin in non-motile enterococci can be encoded by several mechanisms.

The first report of the vanC1 gene in Enterococcus faecium isolated from a human clinical specimen.

The vanC1 gene, which is chromosomally located, confers resistance to vancomycin and serves as a species marker for Enterococcus gallinarum. Enterococcus faecium TJ4031 was isolated from a blood culture and harbours the vanC1 gene. Polymerase chain reaction (PCR) assays were performed to detect vanXYc and vanTc genes. Only the vanXYc gene was found in the E. faecium TJ4031 isolate. The minimum inhibitory concentrations of vancomycin and teicoplanin were 2 µg/mL and 1 µg/mL, respectively. Real-time reverse transcription-PCR results revealed that the vanC1 and vanXYc genes were not expressed. Pulsed-field gel electrophoresis and southern hybridisation results showed that the vanC1 gene was encoded in the chromosome. E. faecalis isolated from animals has been reported to harbour vanC1 gene. However, this study is the first to report the presence of the vanC1 gene in E. faecium of human origin. Additionally, our research showed the vanC1 gene cannot serve as a species-specific gene of E. gallinarum and that it is able to be transferred between bacteria. Although the resistance marker is not expressed in the strain, our results showed that E. faecium could acquire the vanC1 gene from different species.

Detection of vanC1 gene transcription in vancomycin-susceptible Enterococcus faecalis.

Here we report the presence and expression levels of the vanC1 and vanC(2/3) genes in vancomycin-susceptible strains of Enterococcus faecalis. The vanC1 and vanC(2/3) genes were located in the plasmid DNA and on the chromosome, respectively. Specific mRNA of the vanC1 gene was detected in one of these strains. Additionally, analysis of the vanC gene sequences showed that these genes are related to the vanC genes of Enterococcus gallinarum and Enterococcus casseliflavus. The presence of vanC genes is useful for the identification of E. gallinarum and E. casseliflavus. Moreover, this is the first report of vanC mRNA in E. faecalis.

Detection of vanC1 gene transcription in vancomycin-susceptible Enterococcus faecalis

Here we report the presence and expression levels of the vanC 1 and vanC 2/3 genes in vancomycin-susceptible strains of Enterococcus faecalis. The vanC 1 and vanC 2/3 genes were located in the plasmid DNA and on the chromosome, respectively. Specific mRNA of the vanC 1 gene was detected in one of these strains. Additionally, analysis of the vanC gene sequences showed that these genes are related to the vanC genes of Enterococcus gallinarum and Enterococcus casseliflavus. The presence of vanC genes is useful for the identification of E. gallinarum and E. casseliflavus. Moreover, this is the first report of vanC mRNA in E. faecalis.

Occurrence of vancomycin-resistant enterococci in turkey flocks

In the present study, the prevalence of vancomycin-resistant enterococci (VRE) in turkeys in the southwest of Germany was investigated. For this purpose, 200 cloacal swab samples and 5 environmental dust samples (tested as a pooled sample) of each of the 20 flocks (10 female and 10 male flocks) included in this study were examined. The VRE could be isolated by means of a procedure combining bacterial cultivation in an enrichment broth and on a selective solid media. Enterococci were identified biochemically and subsequently tested on the presence of the vancomycin resistance genes vanA, vanB (B1/B2/B3), and vanC (C1/C2/C3) using real-time PCR assays. In 54 (27%) turkeys originating from 11 (55%) flocks and in 14 (70%) of the dust samples, exclusively vanA and vanC1 genes could be detected. Of the turkeys examined, 46 were colonized with VRE bearing the resistance gene vanC1 and 8 vanA, originating from 9 and 2 flocks, respectively. None of the birds carried vanB, vanC2, or vanC3 positive VRE. The results obtained from the birds are largely confirmed by the dust samples originating from 4 vanA and 10 vanC1 positive flocks. However, one flock housing animals colonized with vanC1 positive VRE could not be confirmed by the dust samples that revealed vanA bearing VRE. However, in one case vanA and in 3 cases vanC1 carrying VRE could be detected in dust samples of the turkey houses, but not in the turkeys of the associated flock. In 5 flocks the turkeys as well as the dust samples were free of VRE.

Presence of the vanC1 gene in a vancomycin-resistant Enterococcus faecalis strain isolated from ewe bulk tank milk

Presence of the vanC1 gene in a vancomycin-resistant Enterococcus faecalis strain isolated from ewe bulk tank milk

High prevalence of Enterococcus spp. from dogs with otitis externa

Otitis externa (OE) is a frequent disease in the ear canals of dogs. To identify the pathogens causing OE in dogs and to determine their antimicrobial resistances, specimens were collected from animal hospitals in Daejeon. The isolates were examined by morphological and biochemical tests, 16S rRNA analysis and antimicrobial susceptibility tests. We analyzed correlation between the isolated pathogens and external factors of dogs such as breed, age, gender, ear mite, hair in ears and experience with antibiotic therapy. Thirty three strains of bacteria were isolated from 26 of the 68 heads of dogs with OE. The most isolated bacteria were Enterococcus faecalis (E. faecalis) followed by Staphylococcus aureus (Sta. aureus), Sta. pseudointermedius, E. faecium, E. avium and Streptococcus canis (Strep. canis) in order of frequency of occurrence. Isolation frequency of Enterococcus spp. and Staphylococcus spp. were 51.5% and 45.5%, respectively. E. faecalis and E. faecium isolates showed VanB phenotype, which is resistant to vancomycin but sensitive to teicoplanin were 58% and 25%, respectively. Nine isolates among total twelve isolates of E. faecalis were isolated from the dogs treated with antibiotics. There was no methicillin-resistant Sta. aureus (MRSA), but were MR-Sta. pseudointermedius (MRSP) (57.1%) and vancomycin-resistant (VR)-Sta. pseudointermedius (14.3%) (VRSP) showing VanB phenotype. However, vanA, vanB and vanC genes were not detected in VR isolates from the dogs. Taken together, VR-Enterococcus spp. (VRE) is one of the major pathogens in domestic animals, as well as community-and hospital-acquired infection.

Presence of the Resistance GenesvanC1andpbp5in Phenotypically Vancomycin and Ampicillin SusceptibleEnterococcus faecalis

Ampicillin and vancomycin are important antibiotics for the therapy of Enterococcus faecalis infections. The ampicillin resistance gene pbp5 is intrinsic in Enterococcus faecium. The vanC1 gene confers resistance to vancomycin and serves as a species marker for Enterococcus gallinarum. Both genes are chromosomally located. Resistance to ampicillin and vancomycin was determined in 484 E. faecalis of human and porcine origin by microdilution. Since E. faecalis are highly skilled to acquire resistance genes, all strains were investigated for the presence of pbp5 (and, in positive strains, for the penicillin-binding protein synthesis repressor gene psr) and vanC1 (and, in positive strains, for vanXYc and vanT) by using polymerase chain reaction (PCR). One porcine and one human isolate were phenotypically resistant to ampicillin; no strain was vancomycin resistant. Four E. faecalis (3/1 of porcine/human origin) carried pbp5 (MIC=1 mg/L), and four porcine strains were vanC1 positive (minimum inhibitory concentration [MIC]=1 mg/L). Real-time reverse transcriptase (RT)-PCR revealed that the genes were not expressed. The psr gene was absent in the four pbp5-positive strains; the vanXYc gene was absent in the four vanC1-positive strains. However, vanT of the vanC gene cluster was detected in two vanC1-positive strains. To our knowledge, this is the first report on the presence of pbp5, identical with the "E. faecium pbp5 gene," and of vanC1/vanT in E. faecalis. Even if resistance is not expressed in these strains, this study shows that E. faecalis have a strong ability to acquire resistance genes-and potentially to spread them to other bacteria. Therefore, close monitoring of this species should be continued.

Prevalence and antimicrobial resistance pattern of multidrug-resistant enterococci isolated from clinical specimens

Purpose: Vancomycin-resistant enterococci (VRE) pose an emerging problem in hospitals worldwide. The present study was undertaken to determine the occurrence, species prevalence, antibacterial resistance, and phenotypic and genetic characteristics of VRE isolated in Riyadh hospitals, KSA. Materials and Methods: Two hundred and six isolates of enterococcal species were obtained from clinical samples. The antibiotic susceptibility of isolates and minimum inhibitory concentration (MIC) tests for vancomycin and teicoplanin were determined. Molecular typing of VRE isolates was carried out by using pulsed field gel electrophoresis (PFGE) and the resistance genotype was determined by polymerase chain reaction (PCR). Results: VRE accounted for 3.9% of the isolates and were detected mostly in urine, wound and blood specimens isolated from ICU, internal medicine and surgical wards. All strains were identified to species level and were found to consist of E. faecalis (69.2%), E. faecium (11.3%), E. avium (2.1%), E. hirae (0.8%), E. casseliflavus (1.3%) and E. gallinarum (1.3%) species. According to the susceptibility data obtained, 8 (3.9%) out of 206 isolates were found to be VRE (MICs > 32 µg/ml). The vanA, vanB and vanC gene fragments of E. faecalis, E. faecium and E. gallinarum were amplified from isolates and were detected. PFGE patterns of the VRE isolates revealed homogenous patterns with dominant clone suggesting that the strains intrinsic resistance is independent. Conclusions: This study shows an emergence of VRE along with increased rate of multidrug-resistant enterococci in the area of the study. Regular surveillance of antimicrobial susceptibilities should be done regularly and the risk factors should be determined.

Prevalence ofvanCVancomycin-Resistant Enterococci in the Teaching Hospitals of the University of Debrecen, Hungary

Vancomycin-resistant enterococci (VRE) are common nosocomial pathogens; however, until now they have been rarely encountered in Hungary. In the present study, we investigated the prevalence of VRE in the teaching hospitals of the University of Debrecen. Of 7,271 Enterococcus-containing clinical samples collected between 2004 and 2009, we identified 16 VRE. Species-specific polymerase chain reaction was used to detect Enterococcus faecalis, Enterococcus faecium, Enterococcus casseliflavus, and Enterococcus gallinarum. Multiplex polymerase chain reaction was performed to identify the vancomycin resistance genes: vanA, vanB, vanC1/C2, vanD, vanE, and vanG. Restriction digestion with SalI and HindIII was introduced to differentiate the vanC1 and vanC2 genes from each other. Genetic relationships between the strains were investigated by pulsed-field gel electrophoresis. Overall, we identified the vanC1 resistance gene in 14 E. gallinarum and the vanC2 resistance gene in two E. casseliflavus strains. Except for two samples, the isolates had different pulsed-field gel electrophoresis types, suggesting sporadic emergence of the resistant bacteria. In addition, antibiotic resistance profile was determined by E-test. Three E. gallinarum strains proved to be resistant to gentamicin because of the presence of the aacA-aphD gene. Although the prevalence of VRE in Debrecen is rather low, the appearance of multiple resistances is of concern.

Antibiotic resistance in faecal bacteria (Escherichia coli, Enterococcus spp.) in feral pigeons

To determine the presence of antibiotic-resistant faecal Escherichia coli and Enterococcus spp. in feral pigeons (Columba livia forma domestica) in the Czech Republic. Cloacal swabs of feral pigeons collected in the city of Brno in 2006 were cultivated for antibiotic-resistant E. coli. Resistance genes, class 1 and 2 integrons, and gene cassettes were detected in resistant isolates by polymerase chain reaction (PCR). The samples were also cultivated for enterococci. Species status of enterococci isolates was determined using repetitive extragenic palindromic-PCR. Resistance genes were detected in resistant enterococci by PCR. E. coli isolates were found in 203 of 247 pigeon samples. Antibiotic resistance was recorded in three (1·5%, n(E. coli) =203) isolates. Using agar containing ciprofloxacin, 12 (5%, n(samples) =247) E. coli strains resistant to ciprofloxacin were isolated. No ESBL-producing E. coli isolates were detected. A total of 143 enterococci were isolated: Ent. faecalis (36 isolates), Ent. faecium (27), Ent. durans (19), Ent. hirae (17), Ent. mundtii (17), Ent. gallinarum (12), Ent. casseliflavus (12) and Ent. columbae (3). Resistance to one to four antibiotics was detected in 45 (31%) isolates. Resistances were determined by tetK, tetL, tetM, tetO, aac(6')aph(2''), ant(4')-Ia, aph(3')-IIIa, ermB, pbp5, vanA and vanC1 genes. Antibiotic-resistant E. coli and Enterococcus spp. occurred in feral pigeons in various prevalences. Feral pigeon should be considered a risk species for spreading in the environment antimicrobial resistant E. coli and enterococci.

Investigation of antimicrobial susceptibility for enterococci isolated from cats and dogs and the determination of resistance genes by polymerase chain reaction

The aim of this study was to investigate the antimicrobial susceptibility and to determinate the resistance genes by Polymerase Chain Reaction (PCR) among enterococci isolated from healthy cats and dogs in İzmir, Turkey. From a total of 114 rectal swabs, 91 (31 cats and 60 dogs) stains were isolated as enterococci. Twenty eight isolates were identified as Enterococcus faecalis and 63 as Enterococcus faecium with the API 20 IDStrep system. Antimicrobial susceptibilities were determined by disc diffusion. Susceptibility rates of all enterococci strains were found as 100.0%, 95.6%, 94.5%, 87.9%, 84.6%, 30.8%, 29.7%, and 0.0% for teicoplanin, penicillin, vancomycine, chloramphenicol, gentamycin, erythromycin, tetracycline, and clindamicin, respectively. Of 91 enteroccocci isolates 14 (15.4%) were found susceptible to all antimicrobials tested while 77 (84.6%) strains were resistant. All isolates were found resistant to clindamicin while 5 E. faecium strains had an intermediate resistance to vancomycin. None of the intermediate resistant vancomycin strains has any vanA, vanB, or vanC genes. From 91 isolates 4, 11, 14, 63, and 64 were found resistant to penicillin, chloramphenicol, gentamycin, erythromycin, and tetracycline, respectively. Sixty two tetracycline resistant strains had tet(M) gene. From erythromycin resistant strains 2 and 54 had erm(A) and erm(B). Gentamycin resistance was detected for 14 isolates from which 10 had aac(6')-aph(2') genes. Seven strains have the cat gene of 11 chloramphenicol resistant strains. The results revealed that there was a high level of antimicrobial resistance among enterococci isolated from healthy cats and dogs and they pose a potential risk for transmission of enterococci to humans.

Genetic Diversity of the Low-Level Vancomycin Resistance GenevanC-2/vanC-3and Identification of a NovelvanCSubtype (vanC-4) inEnterococcus casseliflavus

An intrinsic low-level vancomycin resistance (VanC phenotype) in Enterococcus casseliflavus is conferred by either of two subtypes of vanC genes, that is, vanC-2 or vanC-3, which are genetically closely related. To know genetic diversity of vanC-2/C-3 genes among E. casseliflavus, nucleotide sequences of vanC-2/C-3 and other genetic components in vanC gene cluster (vanXYc, vanTc, vanRc, and vanSc) were analyzed for nine clinical isolates and four standard strains that showed low-level vancomycin resistance. While the vanC-2/C-3 gene sequences showed 93-100% identities among the strains examined, two genetic groups were discriminated by phylogenetic analysis: one closely related to the previously reported vanC-2 or vanC-3 genes (vanC-2/C-3 genotype) with 98-100% identity, and the other distinct from the vanC-2/C-3 genotype (93-95% identity). The latter group found in three clinical isolates was considered as a new subtype of vanC and tentatively designated as vanC-4. Between strains with the vanC-2/C-3 genotype and those with vanC-4, vanXYc genes were also genetically discriminated with 92-93% identity. Similar sequence diversity was observed for vanTc, vanRc, and vanSc (88-93% identity). Clonal relatedness among the E. casseliflavus strains was investigated by phylogenetic analysis of atpA gene. While among E. casseliflavus strains with vanC-2/C-3 genotype, extremely high sequence identities of atpA were found (98.7% or higher), these strains showed slightly lower identity to those with vanC-4 (94-96%). These two groups of E. casseliflavus strains were also discriminated by genotyping with arbitrarily primed PCR. These findings indicated that among E. casseliflavus there are at least two genetic lineages with the distinct vanC genes, that is, a single subtype including previously known vanC-2/C-3, and a novel subtype vanC-4.

Emergence of the vanA genotype among Enterococcus gallinarum isolates colonising the intestinal tract of patients in a university hospital in Rio de Janeiro, Brazil

We describe the characteristics of seven unusual isolates of vancomycin-resistant enterococci (VRE) carrying both the vanC1 and vanA genes that were detected during a 3-month survey carried out to investigate the occurrence of faecal carriage of VRE. The isolates were identified as Enterococcus gallinarum and showed high-level resistance to both vancomycin and teicoplanin (minimum inhibitory concentrations >256 μg/mL and 64–96 μg/mL, respectively). All seven isolates were also resistant to chloramphenicol, erythromycin and high levels of gentamicin, and showed intermediate susceptibility to both quinolones tested (ciprofloxacin and norfloxacin). Susceptibility to fosfomycin, rifampicin and tetracycline varied among isolates. High-level resistance to gentamicin was associated with the aac(6′ )- aph(2″ ) gene, and resistance to erythromycin was associated with the erm(B) gene. The seven vanA-carrying E. gallinarum isolates had similar pulsed-field gel electrophoresis (PFGE) profiles. The emergence of multiple antimicrobial resistance, including high-level resistance to glycopeptides, among E. gallinarum points out the need to increase awareness for detection and proper characterisation of these microorganisms, as they may represent potential reservoirs of transmissible, clinically significant resistance genes in nosocomial settings.

Nosocomial outbreak of Enteroccocus gallinarum: untaming of rare species of enterococci

An unusual increase in infections caused by vancomycin-resistant Enterococcus gallinarum (VREG) was identified in May 2004, in a Colombian tertiary care teaching hospital. A case-control study was subsequently designed to identify risk factors associated with the development of infections due to these organisms. All VREG isolates were subjected to antimicrobial susceptibility testing, vancomycin resistance gene detection and pulsed-field gel electrophoresis (PFGE) typing. Additionally, the presence of genes associated with an acquired pathogenicity island of E. faecalis and a hyl-like gene of E. faecium was assessed by hybridisation assays. Eleven cases of VREG were identified between May through June 2004. VREG was isolated from blood (N=4), surgical secretions (N=4), paranasal sinus secretion (N=1), lung abscess (N=1) and urine (N=1). Infections with VREG were associated with mucositis, hospitalisation in the haematology ward and surgical unit, length of hospital stay prior to culture and invasive procedures within 30 days prior to the culture. Logistic regression found that female sex and hospitalisation in the surgical unit were independent factors for VREG infection. All isolates were identified as E. gallinarum, harboured the vanC1 gene and exhibited indistinguishable restriction patterns by PFGE. Virulence-associated genes were not detected. This is the first documented hospital-wide outbreak of VREG and highlights the fact that uncommon species of enterococci are capable of nosocomial dissemination.

VanC Gene-Related Intrinsic Teicoplanin Resistance Detected in Enterococcus casseliflavus and E. gallinarum Strains by the BD Phoenix Automated Microbiology System

Enterococcus members are becoming increasingly important agents of human disease, largely because of their resistance to antimicrobial agents. Because of their resistance to penicillins and cephalosporins of several activity ranges, the acquisition of high-level resistance to aminoglycosides, and now the emergence of vancomycin resistance, these bacteria are often involved in serious superinfections among patients receiving broad-spectrum antimicrobial chemotherapy (2). Acquired glycopeptide resistance in Enterococcus species corresponds to five different phenotypes, with VanA and VanB phenotypes being the most prevalent and important clinically. Strains with the vanA genotype characteristically display inducible, transposon-mediated, high-level resistance to both vancomycin (MIC, 64 to 1,000 μg/ml) and teicoplanin (MIC, 16 to 512 μg/ml). Strains with the vanB genotype have acquired inducible resistance to various concentrations of vancomycin (MIC, 4 to 1,000 μg/ml) but remain susceptible to teicoplanin (MIC, 0.5 to 1 μg/ml), although rare vanB strains may also be resistant to the latter antibiotic. Isolates that have the vanC genotype display intrinsic, constitutive, low-level resistance to vancomycin (MIC, 2 to 32 μg/ml) and are susceptible to teicoplanin (MIC, 0.5 to 1 μg/ml). The vanC genotype corresponds to the intrinsic glycopeptide resistance seen in Enterococcus gallinarum, E. casseliflavus, and E. flavescens (2). The antibacterial activities of teicoplanin against gram-positive bacteria, including those expressing resistance to unrelated compounds, are similar to that of vancomycin, but with increased potency, particularly against Streptococcus spp. and Enterococcus spp. Teicoplanin is active against vancomycin resistance caused by VanB and VanC but is not active against VanA resistant strains. Despite the increasing importance of glycopeptide resistance, teicoplanin has proved its clinical worth and continues to have important potential in the treatment of life-threatening gram-positive sepsis (1a). Automated microbiology systems are routinely used for identification and antimicrobial susceptibility testing of a wide spectrum bacteria, including those of the Enterococcus genus. The Phoenix Automated Microbiology System (Becton Dickinson) is one of these systems used in our microbiology laboratory at Yuzuncu Yil University Medical Faculty Hospital. Recently, E. casseliflavus and E. gallinarum strains were isolated from two different urine samples from a patient, and teicoplanin was reported as resistant, although the MIC was ≤1. Upon examination of the preliminary reports, a change from sensitive to resistant for teicoplanin by the automated BDXpert system in accordance with rule 1099 of that system was observed (1). Rule 1099 under the expert trigger rules states that “E. casseliflavus or E. gallinarum is intrinsically low-level resistant to vancomycin and teicoplanin (VanC),” and because of the identified bacteria, the exchange for teicoplanin from sensitive to resistant was made following this rule. Resistance to teicoplanin in Enterococcus species is often seen in E. faecalis and E. faecium strains due to the vanA gene. Identification and antibiotic susceptibility tests for both of the two isolated strains were repeated in order to prevent any misidentifications, but there was not any change in the results. Antimicrobial testing of teicoplanin by use of the Kirby-Bauer disc diffusion method was performed on both of the isolated strains, and they were both detected as sensitive to teicoplanin. E. casseliflavus and E. gallinarum strains have the chromosomal nontransferable vanC gene, and they are intrinsically low-level resistant to vancomycin; however, they are sensitive to teicoplanin. Researchers in clinical microbiology laboratories using automated systems for antimicrobial susceptibility tests should be aware of this situation, and the result needs to be verified using a reference method before reporting the isolate as resistant to teicoplanin.