Use of Triplex PCR for Rapid Detection of PVL and Differentiation of MRSA from Methicillin Resistant Coagulase Negative Staphylococci

Abstract

Methicillin-Resistant Staphylococcus aureus (MRSA) has become a major public health problem in both hospitals and communities. Panton - Valentine Leucocidin (PVL) has been reported to be an important marker for the highly pathogenic community acquired S. aureus infections. A rapid detection of these MRSA is very important for its treatment. The specific detection of MRSA is always a problem due to the prevalence of methicillin resistance among the coagulase negative Staphylococci. Hence, this study was done to develop a rapid triplex PCR for the detection of PVL positive MRSA and for the simultaneous differentiation of MRSA from Coagulase Negative Staphylococci (CoNS). We developed a triplex PCR for the specific detection of PVL positive Community Acquired (CA) - MRSA and for its simultaneous differentiation from the coagulase negative Staphylococci. We used PCR for targeting the fem A gene which is specific for S. aureus, mecA which is specific for methicillin-resistance and luk - PV which is specific for the PVL toxin. The method was evaluated with a total of 100 clinical isolates of Staphylococcus spp. The triplex PCR was successfully standardized by using the reference strains and it was evaluated by using clinical strains. The method was found to be rapid, highly sensitive (100%), specific (99%) and cost effective. Triplex PCR can be used as a diagnostic tool for the detection of the highly pathogenic strains of CA-MRSA.