Detection and molecular characterization of vancomycin resistant Staphylococcus aureus from clinical isolates

Abstract

Until now, few strains of vancomycin resistant Staphylococcus aureus (VRSA) have been reported worldwide. The disk diffusion method for determination of vancomycin sensitivity frequently misclassifies intermediately susceptible isolates to fully susceptible. However, non-automated minimum inhibitory concentration (MIC) detection methods are the gold standards. In the present study, 439 Gram positive clinical isolates were collected. Among them, 220 Staphylococcus aureus were identified. Multiplex polymerase chain reaction (PCR) detection method for vancomycin resistant S. aureus (VRSA) was developed for detection of both vanA gene (for vancomycin resistance) and nuc gene (specific for S. aureus) in a single step PCR compared to conventional non-automated disc diffusion and MIC detection methods. Molecular typing of VRSA isolates was performed using randomly amplified polymorphic (RAPD) DNA assay technique. The results show 10 VRSA isolates detected by disc diffusion method and MIC determination. Five out of them harbored vanA gene that were detected by multiplex PCR and most of them showed low clonal diversity.

Key words: Molecular characterization, multiplex polymerase chain reaction (PCR), vancomycin resistant Staphylococcus aureus (VRSA), vanA gene, nuc gene, randomly amplified polymorphic DNA (RAPD) assay technique.