Species-level identification of clinical staphylococcal isolates based on polymerase chain reaction–restriction fragment length polymorphism analysis of a partial groEL gene sequence

A pair of degenerate primers that amplified, by polymerase chain reaction (PCR), a partial groEL gene sequence (550 bp) was used for the identification of the 12 most common human staphylococcal pathogens. The amplified products were digested by AluI endonuclease, and distinctive PCR restriction fragment length polymorphism (RFLP) patterns for reference strains were obtained. This protocol was validated by the identification of 89 clinical staphylococcal isolates, and the results were compared with those obtained by the reference biochemical identification, showing 100% concordant results. Two species, Staphylococcus aureus and Staphylococcus lugdunensis, showed intraspecies polymorphisms on their PCR-RFLP patterns. All strains were also identified using the API Staph ID test (bioMérieux, Durham, NC) and the MicroScan WalkAway automated system (Dade Behring, West Sacramento, CA). When 17 Staphylococcus isolates were tested in a blind experiment by the PCR-RFLP of the groEL gene method, all strains were also correctly identified. We propose the PCR-RFLP of the groEL gene with AluI as a reliable and reproducible method for identification of Staphylococcus spp.