Evaluation of the iNtRON VRE vanA/vanB Real-Time PCR Assay for Detection of Vancomycin-Resistant Enterococci

BACKGROUND: Recently, the iNtRON VRE vanA/vanB real-time PCR (iNtRON; iNtRON Biotechnology, Korea) assay, a multiplex real-time PCR method, was introduced. In this prospective study, we compared the iNtRON assay with the Seeplex VRE ACE detection kit (Seeplex; Seegene, Korea), a conventional multiplex PCR assay. METHODS: A chromogenic agar-based culture, in which pre-selected vancomycin-resistant enterococci (VRE) was grown and subsequently plated on blood agar with vancomycin disks, was regarded as the reference method. A total of 304 consecutive rectal swab specimens were tested for VRE by culture and by iNtRON and Seeplex PCR assays. For the PCR assays, specimens were enriched for 16-24 hr before PCR. RESULTS: VRE were isolated from 44 (14.5%) specimens by chromogenic agar-based culture. The clinical sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) of the iNtRON assay were 100% (95% confidence interval: 89.8%-100%), 99.2% (96.9%-99.9%), 95.6% (83.6%-99.2%), and 100% (98.2%-100%), respectively, while those of the Seeplex assay were 97.7% (86.2%-99.9%), 99.6% (97.5%-99.9%), 97.7% (86.2%-99.9%), and 99.6% (97.5%-99.9%), respectively. The iNtRON assay had a detection limit of 3,159 copies/microL and 13,702 copies/microL for the vanA and vanB genes, respectively. No cross-reactivity was observed in 11 non-VRE bacterial culture isolates. CONCLUSIONS: The overall performance of the iNtRON assay was comparable to that of a chromogenic agar-based culture method for prompt identification of VRE-colonized patients in hospitals. This assay could be an alternative or supportive method for the effective control of nosocomial VRE infection.

Reduced susceptibility to vancomycin and biofilm formation in methicillin-resistant Staphylococcus epidermidis isolated from blood cultures

This study aimed to correlate the presence of ica genes, biofilm formation and antimicrobial resistance in 107 strains of Staphylococcus epidermidis isolated from blood cultures. The isolates were analysed to determine their methicillin resistance, staphylococcal cassette chromosome mec (SCCmec) type, ica genes and biofilm formation and the vancomycin minimum inhibitory concentration (MIC) was measured for isolates and subpopulations growing on vancomycin screen agar. The mecA gene was detected in 81.3% of the S. epidermidis isolated and 48.2% carried SCCmec type III. The complete icaADBC operon was observed in 38.3% of the isolates; of these, 58.5% produced a biofilm. Furthermore, 47.7% of the isolates grew on vancomycin screen agar, with an increase in the MIC in 75.9% of the isolates. Determination of the MIC of subpopulations revealed that 64.7% had an MIC ≥ 4 μg mL-1, including 15.7% with an MIC of 8 μg mL-1 and 2% with an MIC of 16 μg mL-1. The presence of the icaADBC operon, biofilm production and reduced susceptibility to vancomycin were associated with methicillin resistance. This study reveals a high level of methicillin resistance, biofilm formation and reduced susceptibility to vancomycin in subpopulations of S. epidermidis. These findings may explain the selection of multidrug-resistant isolates in hospital settings and the consequent failure of antimicrobial treatment.

Antimicrobial susceptibility patterns, emm type distribution and genetic diversity of Streptococcus pyogenes recovered in Brazil

Streptococcus pyogenes is responsible for a variety of infectious diseases and immunological complications. In this study, 91 isolates of S. pyogenes recovered from oropharynx secretions were submitted to antimicrobial susceptibility testing, emm typing and pulsed-field gel electrophoresis (PFGE) analysis. All isolates were susceptible to ceftriaxone, levofloxacin, penicillin G and vancomycin. Resistance to erythromycin and clindamycin was 15.4%, which is higher than previous reports from this area, while 20.9% of the isolates were not susceptible to tetracycline. The macrolide resistance phenotypes were cMLSB (10) and iMLSB (4). The ermB gene was predominant, followed by the ermA gene. Thirty-two emm types and subtypes were found, but five (emm1, emm4, emm12, emm22, emm81) were detected in 48% of the isolates. Three new emm subtypes were identified (emm1.74, emm58.14, emm76.7). There was a strong association between emm type and PFGE clustering. A variety of PFGE profiles as well as emm types were found among tetracycline and erythromycin-resistant isolates, demonstrating that antimicrobial resistant strains do not result from the expansion of one or a few clones. This study provides epidemiological data that contribute to the development of suitable strategies for the prevention and treatment of such infections in a poorly studied area.

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 vanC1gene. 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 vanC1and 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 vanC1gene. However, this study is the first to report the presence of the vanC1gene in E. faecium of human origin. Additionally, our research showed the vanC1gene 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 vanC1gene from different species.

Presence of virulence factors in Enterococcus faecalis and Enterococcus faecium susceptible and resistant to vancomycin

Despite the increasing importance of Enterococcus as opportunistic pathogens, their virulence factors are still poorly understood. This study determines the frequency of virulence factors in clinical and commensal Enterococcus isolates from inpatients in Porto Alegre, Brazil. Fifty Enterococcus isolates were analysed and the presence of the gelE, asa1 and esp genes was determined. Gelatinase activity and biofilm formation were also tested. The clonal relationships among the isolates were evaluated using pulsed-field gel electrophoresis. The asa1, gelE and esp genes were identified in 38%, 60% and 76% of all isolates, respectively. The first two genes were more prevalent in Enterococcus faecalis than in Enterococcus faecium, as was biofilm formation, which was associated with gelE and asa1 genes, but not with the esp gene. The presence of gelE and the activity of gelatinase were not fully concordant. No relationship was observed among any virulence factors and specific subclones of E. faecalis or E. faecium resistant to vancomycin. In conclusion, E. faecalis and E. faecium isolates showed significantly different patterns of virulence determinants. Neither the source of isolation nor the clonal relationship or vancomycin resistance influenced their distribution.

Detection of vanC 1 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.

Evaluation of Vancomycin Resistance 3 Multiplexed PCR Assay for Detection of Vancomycin-Resistant Enterococci from Rectal Swabs

BACKGROUND: Active screening for vancomycin-resistant enterococci (VRE) using rectal specimens is recommended to limit the spread of antimicrobial resistance within certain high-risk populations. We evaluated the diagnostic performance of Vancomycin Resistance 3 Multiplexed Tandem PCR assay (AusDiagnostics, Australia), a rapid multiplex real-time PCR assay that detects vanA and/or vanB. METHODS: Two-hundred-and-eleven rectal swabs from Hematology and Oncology unit were submitted for VRE surveillance via direct detection of vanA and/or vanB by culture and by using Vancomycin Resistance 3 Multiplexed Tandem PCR assay. Enterococci were identified to the species level by using standard biochemical tests and BD Phoenix Automated Microbiology System (BD Diagnostic Systems, USA). Vancomycin susceptibility of enterococci was determined using Etest (BioMerieux, France). RESULTS: Compared to the culture method, Vancomycin Resistance 3 Multiplexed Tandem PCR assay had a sensitivity of 84.0%, specificity of 98.8%, positive predictive value (PPV) of 91.3%, and negative predictive value (NPV) of 97.6%. The assay failed to detect 18 (8.5%) specimens because of the presence of PCR inhibitors; of the remaining 193 specimens, 25 (12.9%) were positive, 23 for vanA, and 2 for vanB. Although both sensitivity and specificity for vanA VRE was 100% compared to the culture method, all vanB-positive specimens tested negative by VRE culture. CONCLUSIONS: Vancomycin Resistance 3 Multiplexed Tandem PCR assay is a rapid and laborsaving option for VRE surveillance for direct use on rectal swabs. However, the high rate of PCR failure owing to the inhibitors in the specimens and the low specificity for vanB should be considered when interpreting the results.

Emergence and characterisation of vanB vancomycin-resistant Enterococcus faecalis in Rio de Janeiro, Brazil

Here we describe the detection and characterisation of three isolates of vancomycin-resistant VanB-type Enterococcus faecalis. Sequence analysis suggested that these isolates harboured the vanB1 gene. The isolates were susceptible to the majority of antimicrobial agents tested, with the exception of chloramphenicol, erythromycin and vancomycin, and showed distinct profiles of high-level resistance to aminoglycosides. Analysis of the clonal relatedness of the vanB E. faecalis isolates showed similar pulsed-field gel electrophoresis profiles. To our knowledge, this is the first report of the occurrence of enterococcal strains carrying vanB genes in Brazil.

Susceptibilidad antimicrobiana y bases genéticas de la resistencia de cepas de Enterococcus causantes de infecciones en Cuba / Antimicrobial susceptibility and genetic bases for resistance of infectioncausingEnterococcus strains in Cuba

Objective. To identify infection-causing Enterococcus species in Cuban hospitalsand determine their susceptibility to antimicrobial drugs, as well as their resistance mechanisms. Methods. A total of 687 Enterococcus isolates from 30 Cuban hospitals in nine provinces of the country were studied over the period 2000–2009. The species were identified using both the conventional method and the automatic API® system.The minimum inhibitory concentration was determined for 13 antimicrobial drugs following the standards recommended by the Clinical Laboratory and Standards Institute. The polymerase chain reaction technique was used to characterize the genes that were resistant to aminoglycosides, erythromycin, tetracycline, andglucopeptides. The presence of beta-lactamase was determined by the chromogenic cephalosporin test. Results. The most prevalent species were Enterococcus faecalis (82.9%) and E. faecium (12.2%). Resistance to glucopeptides (1.0%) was mediated by the vanA and vanB genes. The strains resistant to ampicillin (6%) did not produce beta-lactamases. A high percentage of resistance to aminoglycosides was observed. Gentamicin (31.0%) and streptomycin and amikacin (29.1%) were mediated by the aac(6’)Ie-aph(2”)Ia, aph(3’)-IIIa, ant(6)Ia, and ant(3”)(9) genes. A correlation was found between resistance to tetracycline (56.0%) and presence of the tet(M) (75.1%) and tet(L) genes (7.0%), while resistance to erythromycin (34.1%) was due to the erm(B) gene (70.9%). Conclusions. Resistance to vancomycin is infrequent in Cuba, as opposed to a high level of resistance to aminoglycosides, which may be indicative of treatment failures. The microbiology laboratory is a cornerstone of Enterococcus infectionsurveillance, along with ongoing monitoring of the susceptibility of these infections to antimicrobial drugs at a time when resistance of this microorganism is on the rise.

Objetivo. Identificar las especies de Enterococcus causantes de infecciones en hospitales cubanos, su susceptibilidad a los antimicrobianos y sus mecanismos de resistencia.Métodos. Se estudiaron 687 aislamientos de Enterococcus procedentes de 30 hospitalescubanos de nueve provincias del país durante el período de 2000 a 2009. La identificación de las especies se realizó mediante el método convencional y sistema automatizado API®. Laconcentración inhibitoria mínima se determinó para 13 antimicrobianos según las recomendaciones del Instituto de Estándares Clínicos y de Laboratorio. Se determinaron los genes de resistencia a aminoglucósidos, eritromicina, tetraciclina y glucopéptidos mediante reacciónen cadena de la polimerasa. La presencia de betalactamasa se determinó por el método de lacefalosporina cromógena. Resultados. Las especies más prevalentes fueron Enterococcus faecalis (82,9%) y Enterococcus faecium (12,2%). La resistencia a los glucopéptidos (1,0%) estuvo mediada por los genes vanA y vanB y las cepas resistentes a ampicilina (6%) no produjeron betalactamasas. Se observó un alto porcentaje de resistencia a los aminoglucósidos: gentamicina (31,0%) y estreptomicina y amikacina (29,1%) mediada por los genes aac(6’)Ie-aph(2”)Ia, aph(3’)-IIIa, ant(6)Ia, ant(3”)(9). Hubo correlación entre la resistencia a tetraciclina (56,0%) y la presencia de los genes tet(M) (75,1%) y tet(L) (7,0%), mientras que la resistencia a eritromicina (34,1%) obedeció al gen erm(B) (70,9%).Conclusiones. La resistencia a vancomicina es infrecuente en Cuba, a diferencia del alto nivel de resistencia a los aminoglucósidos, que sugiere posibles fracasos terapéuticos. El laboratorio de microbiología constituye un pilar fundamental de la vigilancia de las infecciones por cepas de Enterococcus y el monitoreo continuo de su susceptibilidad a los antimicrobianos,dado el incremento de la resistencia de ese microorganismo en el tiempo.

Vancomycin resistance among methicillin resistant Staphylococcus aureus isolates from intensive care units of tertiary care hospitals in Hyderabad.

Background & objectives: Multidrug resistant methicillin-resistant Staphylococcus aureus (MRSA) is a major cause of nosocomial and community acquired infections and is on the rise. The glycopeptide vancomycin has been proposed as the drug of choice for treating such infections. The present study aimed at identifying the vancomycin resistance both phenotypically and genotypically among the MRSA isolates from two tertiary care hospitals in Hyderabad, south India. Methods: MRSA were isolated and identified from different clinical samples collected from ICUs of tertiary care hospitals in Hyderabad using conventional methods. Antibiogram of the isolates and vancomycin MIC were determined following CLSI guidelines. vanA was amplified by PCR using standard primers. Results: All vancomycin resistant S. aureus (VRSA) isolates were MRSA. The VRSA isolates were positive for vanA gene, except one which was negative. All VRSA had a vancomycin MIC in the range of 16-64 mg/l. Interpretation & conclusions: The increase in vancomycin resistance among MRSA and excessive use of antimicrobial agents have worsened the sensitivity. Larger studies need to be done in various geographical regions of the country to better define the epidemiology, mechanism of vancomycin resistance in S. aureus and its clinical implications.

Rapid detection of Van genes in rectal swabs by real time PCR in Southern Brazil / Rápida detecção de genes Van em swabs retais por PCR real-time na região sul do Brasil

INTRODUCTION: Laboratory-based surveillance is an important component in the control of vancomycin resistant enterococci (VRE). METHODS: The study aimed to evaluate real-time polymerase chain reaction (RT-PCR) (genes vanA-vanB) for VRE detection on 115 swabs from patients included in a surveillance program. RESULTS: Sensitivity of RT-PCR was similar to primary culture (75 percent and 79.5 percent, respectively) when compared to broth enriched culture, whereas specificity was 83.1 percent. CONCLUSIONS: RT-PCR provides same day results, however it showed low sensitivity for VRE detection.

INTRODUÇÃO: Vigilância com base em detecção laboratorial é um componente importante no controle de enterococos resistentes a vancomicina (ERV). MÉTODOS: Avaliamos procedimento da reação em cadeia da polimerase real time (PCR-RT) (genes vanA-vanB) para detecção de ERV em 115 swabs de pacientes incluídos em um programa de vigilância. RESULTADOS: A sensibilidade do RT-PCR foi semelhante a da cultura primária (75 por cento e 79,5 por cento, respectivamente) quando comparada com a cultura em caldo enriquecido, enquanto a especificidade foi de 83,1 por cento. CONCLUSÕES: O RT-PCR fornece resultados no mesmo dia, contudo mostra baixa sensibilidade para a detecção de VRE.

Vancomycin-dependent Enterococcus faecium vanA: characterization of the first case isolated in a university hospital in Brazil

In this study, we report the characterization of a strain of Enterococcus faecium vanA, which grows only in the presence of vancomycin (VDEfm-UEL). The bacterium was isolated from the feces of a female patient who had undergone surgical treatment of Reinke’s edema and was receiving intravenous vancomycin therapy for infection with methicillin/oxacillin-resistant Staphylococcus aureus, a postoperative complication. Antimicrobial dependence was further confirmed by the vancomycin E-test. VDEfm-UEL was also shown to be resistant to ampicillin, ciprofloxacin, chloramphenicol, erythromycin, levofloxacin, penicillin, rifampicin, and teicoplanin. The putative virulence genes efaA, gelE and esp were detected by PCR. The ddl gene from VDEfm-UEL was cloned and sequenced. Vancomycin dependence seems to be associated with the insertion of a nucleotide in that sequence, which results in a frame-shift mutation, introducing a premature stop codon. This is the first report of vancomycin-dependent E. faecium isolation in a university hospital in Brazil.

First report of vancomycin-resistant Enterococcus faecalis in Uberaba, Minas Gerais state

In this study we report the first isolation of VanA-type vancomycin-resistant Enterococcus faecalis strains from two different patients hospitalized in the same intensive care unit at the hospital of Universidade Federal do Triângulo Mineiro (UFTM), Uberaba, Minas Gerais, Brazil.

Detection of Vancomycin-resistant Enterococci using Multiplex Real-time PCR Assay and Melting Curve Analysis / 대한진단검사의학회지

BACKGROUND: We developed and evaluated the utility of a multiplex real-time PCR assay that uses melting curve analysis and allows simultaneous identification of vancomycin-resistant genotypes and clinically relevant enterococci. METHODS: The specificity of the assay was tested using 4 reference strains of vancomycin-resistant enterococci (VRE) and 2 reference strains of vancomycin-susceptible enterococci. Ninety-three clinical isolates of enterococci with different glycopeptide-resistant phenotypes were genotyped and identified using a multiplex real-time PCR assay and melting curve analysis. RESULTS: Representative melting curves were obtained for Enterococcus faecium, Enterococcus faecalis, vanA-containing E. faecium, vanB-containing E. faecalis, Enterococcus gallinarum, and Enterococcus casseliflavus. Phenotypic and genotypic analysis of the isolates revealed same results for 82 enterococcal isolates, while in 4 isolates, the glycopeptide-resistant phenotypes were inconsistent with the glycopeptide-resistant genotypes and in the 4 other isolates, species could not be accurately identified. Three isolates with mixed strains, which were detected by the PCR assay, could not be correctly identified using phenotypic methods. CONCLUSIONS: VRE genotyping and identification of clinically relevant enterococci were rapidly and correctly performed using multiplex real-time PCR assay and melting curve analysis.

Rapid detection of Vancomycin-Resistant Enterococci (VRE) in rectal samples from patients admitted to intensive care units

The reduction in time required to identify vancomycin-resistant enterococci (VRE) has gained increased importance during hospital outbreaks. In the present study, we implemented a laboratory protocol to speed up the VRE screening from rectal samples. The protocol combines a medium for selective VRE isolation (VREBAC®, Probac, São Paulo) and a multiplex PCR for detection and identification of vanA and vanB resistance genes. The screening performance was analyzed in 114 specimens collected from four intensive care units. The swabs were collected at two periods: (1) during a VRE outbreak (February 2006, n=83 patients) and (2) at the post-outbreak period, after adoption of infection control measures (June 2006, n=31 patients). Forty-one/83 VRE (49.4 percent) and 3/31(9.7 percent) VRE were found at the first and second period, respectively. All isolates harbored the vanA gene. In both periods, detection of the gene vanA parallels to the minimum inhibitory concentration values of >256 µg/mL and >48 µg/mL for vancomycin and teicoplanin, respectively. Multiplex PCR and conventional methods agreed in 90.2 percent for enterococci identification. Besides this accuracy, we also found a remarkable reduction in time to obtain results. Detection of enterococcal species and identification of vancomycin resistance genes were ready in 29.5 hours, in comparison to 72 hours needed by the conventional methods. In conclusion, our protocol identified properly and rapidly enterococci species and vancomycin-resistance genes. The results strongly encourage its adoption by microbiology laboratories for VRE screenning in rectal samples.

Linezolid vancomycin resistant Enterococcus isolated from clinical samples in Tehran hospitals.

Background: Vancomycin-resistant enterococci pose an emerging health risk. The limitation in therapeutic options has resulted in the development of new drugs such as quinupristin/ dalfopristin and linezolid. Aim, Setting and Design: This study investigated the species prevalence and antibacterial resistance among enterococci isolated in selected Tehran hospitals. Materials and Methods: Between March 2006 and August 2007, 200 enterococcal isolates from urine, blood, stool and wound were recovered in 2 teaching hospitals of Tehran province. Susceptibility of all isolates was tested against vancomycin, teicoplanin and linezolid antibiotics by disk diffusion and agar dilution method. Results and Conclusion: Seventeen (8.5%), 6 (3%) and 4 (2%) of the isolates were resistant to vancomycin, teicoplanin and linezolid, respectively. Within the vancomycin-resistant isolates, 6 (35.2%), 4 (25%) and 1 (5.88%) showed vanA, vanB and vanC genotype patterns, respectively. Four (23.5%) of VRE isolates were resistant to linezolid with minimum inhibitory concentrations between 16 and 32 µg/mL. Two linezolid vancomycin resistant enterococci were E. faecium.

Molecular characterization of vancomycin-resistant Enterococci strains eight years apart from its first isolation in São Paulo, Brazil / Caracterização molecular de espécies de Enterococci resistentes à vancomicina oito anos após seu primeiro isolamento em São Paulo, Brasil

E. faecium was the first reported VRE species, carrying the vanA gene in Brazil. In spite of this, vancomycin-resistant E. faecalis has become the predominant species in Brazilian hospitals. The aim of this study was to evaluate the genetic relatedness of VREs isolated in a Brazilian teaching hospital eight years apart from its first isolation. We analyzed 38 VRE strains obtained from 81 surveillance cultures of patients admitted to the four largest intensive care units in Hospital São Paulo in February, 2006. Presence of the vanA gene was assayed by PCR and PFGE analysis was used for molecular characterization. All VRE strains carried the vanA gene. Two distinct clonal groups were observed among vancomycin-resistant E. faecalis. Vancomycin-resistant E. faecium belonged to five distinct clones were demonstrated by molecular typing. All of these clones were different from the first vancomycin-resistant enterococci clone isolated eight years ago in our hospital.

E. faecium contendo o gene vanA foi a primeira espécie de VRE descrita, no Brasil. Apesar disto, E. faecalis resistente a vancomicina tem se tornado a espécie predominante nos hospitais brasileiros.O objetivo desse estudo foi avaliar a relação genética de VREs isolados em um hospital de ensino brasileiro após oito anos de seu primeiro isolamento. Analisamos 37 isolados de VRE obtidos de 81 culturas de vigilância de pacientes admitidos nas quatro maiores Unidades de Tratamento Intensivo em Fevereiro de 2006. A presença do gene vanA foi analisada por PCR e a caracterização molecular por PFGE. Todas as amostras VRE carreavam o gene vanA. Entre os E. faecalis vancomicina-resistentes, dois distintos grupos clonais foram observados. E. faecium resistente a vancomicina pertencentes a cinco clones distintos foram demonstrados por tipagem molecular. Todos esses clones foram diferentes do primeiro clone de enterococo resistente a vancomicina isolado oito anos atrás em nosso hospital.

Molecular characterization of enterococci harboring genotype and phenotype incongruence related to glycopeptide resistance isolated in Brazilian hospitals

Three Enterococcus faecalis and one Enterococcus faecium strains were characterized by plasmid profile, pulsed-field gel electrophoresis (PFGE) and determination of antimicrobial minimal inhibitory concentrations. VanA elements were characterized by Long PCR, overlapping PCR and DNA sequencing. Enterococcal strains showed resistance to vancomycin and harbored the vanA gene, and three these were teicoplanin susceptible while one showed intermediate resistance to teicoplanin. Two E. faecalis strains showed indistinguishable PFGE profile while the third was unrelated. E. faecalis strains showed a deletion in the right terminal region of the Tn1546-like element. The E. faecium strain showed an insertion element in the vanXY intergenic region. Mutations in VanA elements were not found. Rearrangements in the VanA element could be responsible for incongruities in genotype and phenotype in these strains.

[Phenotypic and genotypic characterization of vancomycin-resistant Enterococci from clinical isolates]

Although Enterococci are part of the normal flora of the gastrointestinal tract in humans, they cause infections under certain circumstances. Vancomycin Resistant Enterococci [VRE] cause serious problems resulting in limited therapeutic options in hospitalized patients. In this study we examined the VRE isolated from clinical specimens to determine the prevalence of vanA and vanB phenotypes. Thirty-two vancomycin resistance Enterococci isolates cultured from clinical samples were investigated. Resistance of isolates to vancomycin, teicoplanin, tetracycline, gentamicin, erythromycin and ciprofloxacin were determined by disk diffusion method. MIC of vancomycin for all strains was determined using by micro-dilution method. Existence of vanA and vanB genes was checked by PCR. Using microbroth dilution assay, 25 and 5 isolates appeared as vanA and vanB phenotypes respectively. All isolates were resistance to ciprofloxacin. Resistance to erythromycin, tetracycline and gentamicin were detected in 96.87%, 81.25% and 78.12% of isolates respectively. vanA and vanB genes were found in 25 and 6 isolates respectively. Co-existence of vanA and vanB were found in 13 isolates using PCR. Our results showed that 12 and 6 of the strains are phenotypically and genotypically vanA and vanB respectively. Although 13 of 25 isolates [52%] showed vanA phenotype, they have both vanA and vanB genes. With the possibility of genotypically alteration in enterococci, it seems that these isolates acquired vanB gene through conjugation