Genome-wide association analysis provides insights into the genomics and extracellular expression of <i>Staphylococcus aureus</i> proteases

Extracellular proteases are a class of <i>Staphylococcus aureus</i> virulence factors that thwart the immune system, promote nutrient acquisition, and shape the activity of virulence determinants. <i>S. aureus</i> displays considerable phenotypic and genotypic variation within clinical important lineages, giving rise to diverse infection types. Therefore, understanding how protease expression influences pathogenicity requires consideration of the underlying genes and their regulation in natural populations. In this study we determined the protease activity of 134 USA300 S. aureus isolates from clinical infections and asymptomatic carriage. In high-throughput casein hydrolysis assays, bloodstream infection isolates had significantly lower protease activity than carriage isolates. To identify the genetic variation underlying this variation in protease expression, we employed a <i>k</i>-mer-based genome wide association study, identifying 68 genes with polymorphisms significantly associated with proteolytic activity. Population-scale genomic variation was compared with strains from a sequenced- defined transposon library, validating the function of 27 loci that were significantly associated with decreased protease expression. Associated genes included known protease-regulating genes, including<i> agrA</i>, but most were novel. These included genes linked to central metabolism, permeases, transporters and membrane proteins. Characterizing the complexity of protease regulation and expression will enhance our fundamental understanding of <i>S. aureus</i> virulence which may result in improved treatment options for problematic clinical <i>S. aureus</i> infections.胞外蛋白酶是金黄色葡萄球菌（<i>Staphylococcus aureus</i>）的一类毒力因子，它们能抑制免疫系统、促进营养获取并调控毒力决定因子的活性。<i>S. aureus</i>在临床重要谱系中表现出显著的表型和基因型变异，从而导致多种感染类型。因此，要理解蛋白酶表达如何影响致病性，需考虑自然群体中相关基因及其调控机制。本研究测定了134株来自临床感染和无症状携带的USA300型<i>S. aureus</i>分离株的蛋白酶活性。在高通量酪蛋白水解实验中，血流感染分离株的蛋白酶活性显著低于携带株。为了确定蛋白酶表达变异背后的遗传变异，我们采用了基于<i>k</i>-mer的全基因组关联研究（genome wide association study），鉴定出68个具有与蛋白水解活性显著相关的多态性基因。我们将群体水平的基因组变异与序列明确的转座子文库（sequenced-defined transposon library）菌株进行比较，验证了27个与蛋白酶表达降低显著相关的基因座的功能。相关基因包括已知的蛋白酶调控基因（如<i>agrA</i>），但大多数为新发现的基因。这些基因涉及中心代谢、通透酶、转运蛋白和膜蛋白相关基因。阐明蛋白酶调控和表达的复杂性将加深我们对<i>S. aureus</i>毒力的基础理解，这可能为棘手的临床<i>S. aureus</i>感染提供更好的治疗方案。