Application of multilocus sequence analysis (MLSA) for rapid identification of Enterococcus species based on rpoA and pheS genes.

The aim of this study was to evaluate the use of RNA polymerase alpha subunit (rpoA) and phenylalanyl-tRNA synthase (pheS) gene sequences as species identification tools for enterococci. Ninety-six representative strains comprising all currently recognized Enterococcus species were examined. rpoA gene sequences generated a robust classification into species groups similar to the one based on 16S rRNA gene sequence analysis. On the other hand, the pheS gene is a fast-evolving clock even better suited for species delineation than the rpoA gene, but not for recognition of species groups within Enterococcus as determined by both rpoA and 16S rRNA genes. All enterococcal species were clearly differentiated on the basis of their rpoA and pheS sequences. Evaluation of intraspecies variation showed that both rpoA and pheS genes have a high degree of homogeneity among strains of the same species. Strains of the same enterococcal species have at least 99% rpoA and 97% pheS gene sequence similarity, whereas, different enterococcal species have at maximum 97% rpoA and 86% pheS gene sequence similarity. It was concluded that both genes can be used as reliable tools for identification of clinical and environmental species of Enterococcus and are efficient screening methods for the detection of novel species. The sequence data obtained in this study were compared to the available atpA and 16S rRNA gene sequences. The MLSA approach to Enterococcus taxonomy provides portable, highly reproducible data with lower costs for rapid identification of all enterococcal species.