Section of Critical Care, Department of Pediatrics, University of Chicago, Chicago, Illinois, United States of America.
PLoS One. 2010 Dec 2;5(12):e15177. doi: 10.1371/journal.pone.0015177.
CA-MRSA infection, driven by the emergence of the USA300 genetic background, has become epidemic in the United States. USA300 isolates are hypervirulent, compared with other CA- and HA-MRSA strains, in experimental models of necrotizing pneumonia and skin infection. Interestingly, USA300 isolates also have increased expression of core genomic global regulatory and virulence factor genes, including agr and saeRS. To test the hypothesis that agr and saeRS promote the observed hypervirulent phenotype of USA300, isogenic deletion mutants of each were constructed in USA300. The effects of gene deletion on expression and protein abundance of selected downstream virulence genes were assessed by semiquantitative real-time reverse-transcriptase PCR (qRT-PCR) and western blot, respectively. The effects of gene deletion were also assessed in mouse models of necrotizing pneumonia and skin infection. Deletion of saeRS, and, to a lesser extent, agr, resulted in attenuated expression of the genes encoding α-hemolysin (hla) and the Panton-Valentine leukocidin (lukSF-PV). Despite the differences in hla transcription, the toxin was undetectable in culture supernatants of either of the deletion mutants. Deletion of agr, but not saeRS, markedly increased the expression of the gene encoding protein A (spa), which correlated with increased protein abundance. Each deletion mutant demonstrated significant attenuation of virulence, compared with wild-type USA300, in mouse models of necrotizing pneumonia and skin infection. We conclude that agr and saeRS each independently contribute to the remarkable virulence of USA300, likely by means of their effects on expression of secreted toxins.
CA-MRSA 感染，由 USA300 遗传背景的出现所驱动，已在美国流行。与其他 CA 和 HA-MRSA 菌株相比，USA300 分离株在实验性坏死性肺炎和皮肤感染模型中具有更高的毒力。有趣的是，USA300 分离株还增加了核心基因组全局调节和毒力因子基因的表达，包括 agr 和 saeRS。为了检验 agr 和 saeRS 促进 USA300 观察到的高毒力表型的假设，我们在 USA300 中构建了每个基因的同源缺失突变体。通过半定量实时逆转录 PCR（qRT-PCR）和 Western blot 分别评估基因缺失对选定下游毒力基因表达和蛋白丰度的影响。还在坏死性肺炎和皮肤感染的小鼠模型中评估了基因缺失的影响。saeRS 和 agr 的缺失（程度较小）导致编码α-溶血素（hla）和 Panton-Valentine 白细胞毒素（lukSF-PV）的基因表达减弱。尽管 hla 转录存在差异，但两种缺失突变体的培养上清液中均未检测到毒素。agr 的缺失，但不是 saeRS 的缺失，显著增加了编码蛋白 A（spa）的基因的表达，这与蛋白丰度的增加相关。与野生型 USA300 相比，每种缺失突变体在坏死性肺炎和皮肤感染的小鼠模型中均表现出明显的毒力减弱。我们得出结论，agr 和 saeRS 各自通过对分泌毒素表达的影响，独立地促成了 USA300 的显著毒力。
