Multilocus analysis of extracellular putative virulence proteins made by group A Streptococcus: Population genetics, human serologic response, and gene transcription

Species of pathogenic microbes are composed of an array of evolutionarily d istinct chromosomal genotypes characterized by diversity in gene content an d sequence (allelic variation). The occurrence of substantial genetic diver sity has hindered progress in developing a comprehensive understanding of t he molecular basis of virulence and new therapeutics such as vaccines. To p rovide new information that bears on these issues, 11 genes encoding extrac ellular proteins in the human bacterial pathogen group A Streptococcus iden tified by analysis of four genomes were studied. Eight of the 11 genes enco de proteins with a LPXTC(L) motif that covalently links Cram-positive virul ence factors to the bacterial cell surface. Sequence analysis of the 11 gen es in 37 geographically and phylogenetically diverse group A Streptococcos strains cultured from patients with different infection types found that re cent horizontal gene transfer has contributed substantially to chromosomal diversity. Regions of the inferred proteins likely to interact with the hos t were identified by molecular population genetic analysis, and Western imm unoblot analysis with sera from infected patients confirmed that they were antigenic. Real-time reverse transcriptase-PCR (TaqMan) assays found that t ranscription of six of the 11 genes was substantially up-regulated in the s tationary phase. In addition, transcription of many genes was influenced by the covR and mga trans-acting gene regulatory loci. Multilocus investigati on of putative virulence genes by the integrated approach described herein provides an important strategy to aid microbial pathogenesis research and r apidly identify new targets for therapeutics research.