Increasing clinical significance of coagulase-negative staphylococci requires effective methods for species identification and genotyping. In this study, six housekeeping genes (femA, ftsZ, gap, pyrH, rpoB, and tuf) with extensive allelic polymorphisms were identified and evaluated to develop a comprehensive multilocus sequence typing (MLST) scheme. Selected primers were capable of amplification of the six loci from all of the 180 Staphylococcus strains belonging to 18 different species. Sequence analysis of each locus (44-63 alleles) revealed higher nucleotide diversity than 16S rRNA (28 alleles). Phylogenetic analysis of the concatenated sequences (3054 bp) of the six loci provided accurate species identification and highly discriminatory typing for all the strains. Multilocus allelic analysis of the 180 Staphylococcus strains generated 103 different sequence profiles, suggesting high genetic diversity of the strains. For example, 30 S. aureus, 37 S. epidermidis, 32 S. haemolyticus, and 14 S. hominis strains were typed into 15, 21, 11, and 10 sequence profiles, respectively. Compared with published MLST schemes that restrict on a few particular species, this new scheme both achieved similar discrimination for typing S. aureus, S. epidermidis, S. haemolyticus, and S. hominis and provided sufficient discriminatory power for typing additional opportunistic species, such as S. cohnii, S. capitis, and S. warneri. Importantly, the comprehensive MLST scheme for Staphylococcus strains provides a better genotyping tool for understanding the phylogeny of coagulase-positive Staphylococcus aureus strains.
Keywords: MLST; genotyping; housekeeping gene; identification.