Study of Bacterial Resistance in Clinical Isolates of Staphylococcus aureus by Comparison of Conventional in-vitro Methods with PCR & RFLP Based Genotypic Methods

Methicillin Resistant S. aureus is an increasingly common cause of nosocomial infections, causing severe morbidity and mortality worldwide and accounting for more than 50% of all S. aureus diseases. S. aureus used in our study were procured isolates from the microbiology lab of a corporate hospital. Reference culture of S. aureus NCIM 5021 was obtained from NCIM, Pune. Clinical isolates were subjected to antimicrobial susceptibility testing by standard antibiotic disc (Cefoxitin 30 µg). The selected clinical strains were tested for sensitivity to different antibiotics by Kirby Bauer disc diffusion method. The MIC was determined by two fold serial dilution in broth individually against various antibacterial agents. That was followed by a study on a combination of Ciprofloxacin & Linezolid by two dimensional checkerboard method of synergism testing against the clinical isolates and reference S. aureus NCIM culture. FIC index of combination indicated synergism against MRSA. MRSA was used for PCR based study of the resistant gene namely mecA. PCR-RFLP method was used to study 16S rRNA by restriction digestion using Taq 1. This is indicative of the possible serological differences among clinical isolates and was compared to NCIM culture based on restriction fragments.