Transposon Mutagenesis Identifies Novel Genes Associated with Staphylococcus aureus Persister Formation.
Transposon Mutagenesis Identifies Novel Genes Associated with Staphylococcus aureus Persister Formation.
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转座子诱变鉴定与金黄色葡萄球菌持久形成相关的新基因
DOI：
10.3389/fmicb.2015.01437
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发表时间：
2015
影响因子：
5.2
通讯作者：
Zhang Y
中科院分区：
文献类型：
--
作者：
Wang W;Chen J;Chen G;Du X;Cui P;Wu J;Zhao J;Wu N;Zhang W;Li M;Zhang Y
致病菌的持续存在是造成慢性和持续性感染对抗菌药物治疗难以抵抗的原因。尽管大肠杆菌的持久性形成和存活机制已被广泛研究，但金黄色葡萄球菌的持久性机制在很大程度上仍然未知。在这里，我们筛选了临床耐甲氧西林金黄色葡萄球菌（MRSA）菌株USA500 （ST8）在抗生素压力下的转座子突变文库，并鉴定了13个插入突变导致持久性缺陷的基因。通过评估突变体在暴露于左氧氟沙星和其他几种应激条件下的存活率，进一步证实了这些候选基因。我们发现了13个插入突变体，包括sdhA、sdhB、ureG、mnhG1、fbaA、ctaB、clpX、parE、HOU_0223、HOU_0587、HOU_2091、HOU_2315和HOU_2346，在不同胁迫条件下，这些突变体的持久性数量显著降低，它们映射到氧化磷酸化、TCA循环、糖酵解、细胞周期和ABC转运蛋白的途径中，表明这些基因和途径可能在持久性形成和存活中发挥重要作用。对新构建的ureG、sdhA和sdhB基因敲除菌株及其补充菌株在左氧氟沙星和其他应激条件下的缺陷进行了检测。这些实验结果与筛选结果一致，表明MRSA USA500中这些基因的缺失导致了持久性缺陷。这些发现为金黄色葡萄球菌持续性形成和存活的机制提供了新的见解，并为开发针对持续性的抗生素提供了新的靶点，以改善慢性和持续性感染的治疗。
Pathogenic bacterial persisters are responsible for the recalcitrance of chronic and persistent infections to antimicrobial therapy. Although the mechanisms of persister formation and survival have been widely studied in Escherichia coli, persistence mechanisms in Staphylococcus aureus remain largely unknown. Here, we screened a transposon mutant library of a clinical methicillin-resistant Staphylococcus aureus(MRSA)strain, USA500 (ST8), under antibiotic pressure and identified 13 genes whose insertion mutations resulted in a defect in persistence. These candidate genes were further confirmed by evaluating the survival of the mutants upon exposure to levofloxacin and several other stress conditions. We found 13 insertion mutants with significantly lower persister numbers under several stress conditions, including sdhA, sdhB, ureG, mnhG1, fbaA, ctaB, clpX, parE, HOU_0223, HOU_0587, HOU_2091, HOU_2315, and HOU_2346, which mapped into pathways of oxidative phosphorylation, TCA cycle, glycolysis, cell cycle, and ABC transporters, suggesting that these genes and pathways may play an important role in persister formation and survival. The newly constructed knockout strains of ureG, sdhA and sdhB and their complemented strains were also tested for defect in persisters following exposure to levofloxacin and several other stress conditions. The results from these experiments were consistent with the screening results, which indicated that deletion of these genes in MRSA USA500 leads to persister defect. These findings provide novel insights into the mechanisms of persister formation and survival in S. aureus and offer new targets for the development of persister-directed antibiotics for the improved treatment of chronic and persistent infections.