Defining novel mechanisms of clonal emergence in Group A Streptococcus

点击翻译按钮获取中文摘要

PROJECT SUMMARY The emergence and spread of closely related strains or clones are characteristic of many bacteria causing serious disease in humans. The major human pathogen group A Streptococcus (GAS) displays such behavior and has long been a model organism for studying clonal emergence in bacteria. GAS is divided into emm types based on variation in the emm gene which encodes for the cell-surface, anti-phagocytic M protein. The present paradigm is that GAS clonal emergence occurs due to genetic recombination events which either allow for acquisition of a novel virulence factor or for increased production of existing virulence factors, particularly those encoded by the nga-slo operon. By sequencing >1,000 emm4 strains from diverse temporal and geographic sources, we have identified that a new emm4 clone has replaced previously circulating emm4 strains over the past decade. The “emergent” strains have not undergone significant genetic recombination, do not contain new virulence factor encoding genes, and have significantly lower transcript levels of the nga-slo operon relative to the “replaced” strains. However, emergent emm4 GAS are more virulent than replaced emm4 strains in both animal models and during growth in human blood. Thus, this newly identified clonal emergence does not fit the current understanding of GAS clonal emergence. It is the goal of this R21 proposal to begin to establish novel mechanisms underlying the proliferation of emergent emm4 GAS. In specific aim 1, we will determine whether emergent emm4 strains have increased colonization/transmission capacities relative to replaced strains. This aim will employ both primary human cells as well as a newly established animal model of GAS transmission. In specific aim 2, we will leverage our existing transcriptomic data which show that emergent strains have significantly higher transcript levels of genes encoding proteins putatively involved in cell surface oxidative stress response and peptidoglycan turnover. We will determine whether the emergent GAS strains have augmented resistance to oxidative stress and to challenge by human neutrophils, which utilize reactive oxygen species as a major killing mechanism. Moreover, cell wall differences between emergent and replaced strains will be explored using complementary imaging techniques and by testing susceptibility to cell-envelope active innate antimicrobials. The specific role of particular genes in observed phenotypic differences will be assessed using either an insertional mutagenesis approach or by modifying gene expression when the candidate genes are essential. These studies have been devised to facilitate the subsequent design and execution of downstream investigations of the molecular underpinning of bacterial epidemics, a key aspect of pathogenesis for a wide variety of medically important pathogens.