Emergence of vanA gene among vancomycin-resistant enterococci in a tertiary care hospital of North - East India.

Background & objectives: Vancomycin-resistant enterococci (VRE) have become one of the most challenging nosocomial pathogens with the rapid spread of the multi-drug resistant strain with limited therapeutic options. It is a matter of concern due to its ability to transfer vancomycin resistant gene to other organisms. The present study was undertaken to determine the emergence of vancomycin-resistant enterococci and the vanA gene among the isolates in a tertiary care hospital of North-East India. Methods: A total of 67 consecutive enterococcal isolates from different clinical samples were collected and identified by using the standard methods. Antibiogram was done by disk diffusion method and VRE was screened by the disk diffusion and vancomycin supplement agar dilution method. The minimum inhibitory concentration (MIC) value for vancomycin was determined by E-test. The VRE isolates were analyzed by PCR for vanA gene. Results: A total of 54 (81%) Enterococcus faecalis and 13 (19%) E. faecium were detected among the clinical isolates and 16 (24%) were VRE. The VRE isolates were multidrug resistant and linezolid resistance was also found to be in three. MIC range to vancomycin was 16-32 μg/ml among the VRE. The vanA gene was found in nine of 16 VRE isolates. Interpretation & conclusions: Emergence of VRE and presence of vanA in a tertiary care hospital setting in North-East India indicate toward a need for implementing infection control policies and active surveillance.

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.

Molecular Diversity of vanA Gene Cluster in Vancomycin-resistant Enterococci Isolated in One University Hospital

BACKGROUND: The vanA gene is the most frequently encountered gene among isolates, causing vancomycin-resistant Enterococci (VRE) infections in humans, and it is part of the transposable element Tn1546. Knowledge of the diversity of Tn1546 is important to distinguish between the dissemination of a single VRE clone and the transmission of a particular Tn1546 type through a genetically divergent population of enterococci. Recently, we studied molecular diversity of Tn1546-related elements in enterococci isolated in one hospital to facilitate understanding of the molecular epidemiology of vancomycin resistance. METHODS: Nineteen VanA-type VRE clinical isolates, collected in one university hospital during 1997 and 1999, were investigated for characteristics such as antibiotic resistance, structure of vanA gene cluster and genomic DNA type by means of antibiotic susceptibility test, PCR amplified length polymorphism of vanA gene cluster and pulse-field gel electrophoresis (PFGE), respectively. RESULTS: Nine (A, B1 to B5, C, D and E) different vanA gene cluster types were identified. Three isolates were grouped into vanA gene cluster type A, similar to that of Tn1546 prototype, and twelve isolates were grouped into type B that has an insertion of IS1216V at vanX-Y intergenic region. Type B was further subdivided into B1 to B5 according to the size variation of vanX-Y intergenic region, which was resulted from the insertion of IS1216V and deletions associated with the insertion. Both vanY and vanZ were deleted in three isolates, suggesting that these genes are not essential for vancomycin resistance. Notably, three E. faecalis and three E. faecium strains isolated during the same period were shown to carry the same vanA gene cluster, showing intergenic transmission of vancomycin resistance. CONCLUSION: The presence of variable types of vanA gene cluster among VRE strains isolated in one hospital suggests that several evolutionary changes of vanA gene clusters have occurred during the horizontal spread of resistance gene in the hospital environment. This approach may be useful for monitoring the evolution of VanA resistance.

Molecular Diversity of vanA Gene Cluster in Vancomycin-resistant Enterococci Isolated in One University Hospital

BACKGROUND: The vanA gene is the most frequently encountered gene among isolates, causing vancomycin-resistant Enterococci (VRE) infections in humans, and it is part of the transposable element Tn1546. Knowledge of the diversity of Tn1546 is important to distinguish between the dissemination of a single VRE clone and the transmission of a particular Tn1546 type through a genetically divergent population of enterococci. Recently, we studied molecular diversity of Tn1546-related elements in enterococci isolated in one hospital to facilitate understanding of the molecular epidemiology of vancomycin resistance. METHODS: Nineteen VanA-type VRE clinical isolates, collected in one university hospital during 1997 and 1999, were investigated for characteristics such as antibiotic resistance, structure of vanA gene cluster and genomic DNA type by means of antibiotic susceptibility test, PCR amplified length polymorphism of vanA gene cluster and pulse-field gel electrophoresis (PFGE), respectively. RESULTS: Nine (A, B1 to B5, C, D and E) different vanA gene cluster types were identified. Three isolates were grouped into vanA gene cluster type A, similar to that of Tn1546 prototype, and twelve isolates were grouped into type B that has an insertion of IS1216V at vanX-Y intergenic region. Type B was further subdivided into B1 to B5 according to the size variation of vanX-Y intergenic region, which was resulted from the insertion of IS1216V and deletions associated with the insertion. Both vanY and vanZ were deleted in three isolates, suggesting that these genes are not essential for vancomycin resistance. Notably, three E. faecalis and three E. faecium strains isolated during the same period were shown to carry the same vanA gene cluster, showing intergenic transmission of vancomycin resistance. CONCLUSION: The presence of variable types of vanA gene cluster among VRE strains isolated in one hospital suggests that several evolutionary changes of vanA gene clusters have occurred during the horizontal spread of resistance gene in the hospital environment. This approach may be useful for monitoring the evolution of VanA resistance.

Molecular Relationship of vanA Glycopeptide Resistance Gene in Enterococci from Hospitalized Patients and Poultry in Korea / 감염

BACKGROUND: Vancomycin-resistant enterococci (VRE) with vanA gene have been reported as a significant nosocomial pathogen. The vanA gene cluster (Tn1546) located on mobile DNA elements is known to be transferable from VRE to other enterococci. The purpose of this study was to investigate the genetic relationship between the vanA VRE strains isolated from hospitalizd patients and poultry. METHODS: Total 145 isolates, including 58 E. faecium, 12 E. faecalis, 3 E. casseliflavus, and 4 E. gallinarum from humans and 68 E. faecium from poultry, were studied. Antimicrobial susceptibility tests were done by disk diffusion or agar dilution methods and molecular epidemiological analysis was performed by pulsed-field gel electrophoresis (PFGE). The internal and structural regions of vanA gene cluster were analyzed by PCR fragment length polymorphism, restriction enzyme, and sequencing of Orf2D region and vanXY intergenic region. The point mutation at Tn1546 nucleotide position 8234 (G->T) within the vanX gene was screened with DdeI restriction enzyme. RESULTS: The antibiotic resistance patterns of human isolates were different from those of poultry. PFGE patterns revealed high heterogeneity. Three PCR fragment length patterns in the vanA gene cluster were found : (I) PCR amplicon of the same size as prototype (E. faecium BM4147) in 17% of human isolates and 100% of poultry ones; (II) PCR amplicon for vanXY intergenic region due to an insertion between vanX and vanY genes in 2.5% of human isolates; (III) the insertions in vanX-vanY intergenic and Orf2 regions in 81% of human isolates. The T type in vanX gene of human and poultry isolates was not found. CONCLUSION: Despite the diverse PFGE patterns, 81% of human and all of poultry isolates belonged to vanA gene cluster type III and I, respectively. These results indicate that the horizontal spread of vanA gene is occurring among genetically diverse strains of VRE in Korea.