Department of Microbiology, Harvard Medical School, Boston, Massachusetts, United States of America.
School of Pharmacy, Sungkyunkwan University, Suwon, Republic of Korea.
PLoS Pathog. 2019 Nov 18;15(11):e1007862. doi: 10.1371/journal.ppat.1007862. eCollection 2019 Nov.
Antibiotic-resistant Staphylococcus aureus remains a leading cause of antibiotic resistance-associated mortality in the United States. Given the reality of multi-drug resistant infections, it is imperative that we establish and maintain a pipeline of new compounds to replace or supplement our current antibiotics. A first step towards this goal is to prioritize targets by identifying the genes most consistently required for survival across the S. aureus phylogeny. Here we report the first direct comparison of multiple strains of S. aureus via transposon sequencing. We show that mutant fitness varies by strain in key pathways, underscoring the importance of using more than one strain to differentiate between core and strain-dependent essential genes. We treated the libraries with daptomycin to assess whether the strain-dependent differences impact pathways important for survival. Despite baseline differences in gene importance, several pathways, including the lipoteichoic acid pathway, consistently promote survival under daptomycin exposure, suggesting core vulnerabilities that can be exploited to resensitize daptomycin-nonsusceptible isolates. We also demonstrate the merit of using transposons with outward-facing promoters capable of overexpressing nearby genes for identifying clinically-relevant gain-of-function resistance mechanisms. Together, the daptomycin vulnerabilities and resistance mechanisms support a mode of action with wide-ranging effects on the cell envelope and cell division. This work adds to a growing body of literature demonstrating the nuanced insights gained by comparing Tn-Seq results across multiple bacterial strains.
耐抗生素金黄色葡萄球菌仍然是美国抗生素耐药相关死亡的主要原因。鉴于多药耐药感染的现实情况，我们必须建立和维持新化合物的管道，以替代或补充我们现有的抗生素。实现这一目标的第一步是通过确定在金黄色葡萄球菌系统发育中最一致地生存所需的基因来确定优先目标。在这里，我们通过转座子测序首次直接比较了多种金黄色葡萄球菌菌株。我们表明，在关键途径中，突变体的适应性因菌株而异，这强调了使用不止一种菌株来区分核心基因和菌株依赖性必需基因的重要性。我们用达托霉素处理文库，以评估菌株依赖性差异是否会影响生存的重要途径。尽管在基因重要性方面存在基线差异，但包括脂磷壁酸途径在内的几种途径在达托霉素暴露下均能持续促进生存，这表明存在核心弱点，可以利用这些弱点使达托霉素不敏感的分离物重新敏感。我们还证明了使用具有向外启动子的转座子来过表达附近基因以识别临床相关获得性功能耐药机制的优点。总之，达托霉素的脆弱性和耐药机制支持一种作用模式，对细胞包膜和细胞分裂具有广泛的影响。这项工作增加了越来越多的文献，证明了通过比较多个细菌菌株的 Tn-Seq 结果获得的细致入微的见解。
