Species Identification of Coagulase Negative Staphylococci by Polymerase Chain Reaction-Restriction Fragment Length Polymorphism of Heat Shock Protein 60 Gene

oh, Eun Jee; Jang, Jung Do; Park, Yeon Joon; Kim, Sun Moo; Kim, Byung Kee

Abstract

BACKGROUND: An accurate and rapid method for specise identification of coagulase negative staphylococci(CNS) has been increasingly necessary for the clinical significance and planning the management of patients with staphylococcal infections. Recently, it has been reported that there is a highly conserved area on their 60KDa heat shock protein(HSP60) gene sequences between the interspecies of CNS and it can be amplified by a set of universal degenerate primer. This led us our attention to focus on whether the PCR-based RFLP method using Mse / restriction enzyme could be a useful tool for the species identification of CNS.
METHODS
In the present study, we performed PCR-based RFLP analysis using a set of degenerate primers covering HSP60 and Mse / restriction enzyme on the reference strains and 25 clinical isolates(10 of S. epidermidis, 10 of S. haemolyticus, 4 of S. lugdunensis and 1 of S. warneri) which were previously identified by the API-STAPH, Vitek GPI card and/or with conventional biochemical test. RESULT: All the seven reference strains revealed that each strain has a distinct electrophoresed band patterns with combination of different number (up to 8) and size of fragments. And these distinct band patterns showed remarkable concordance with the seven reference strains and 25 clinical isolates.
CONCLUSION
These result strongly suggest that the PCR-RFLP method using degenerate primers covering the HSP60 gene and Mse / digestion enzyme offer a convenient and accurate tool for species-specific identification of CNS.