Reconstruction of the Vancomycin-Susceptible Staphylococcus aureus Phenotype From a Vancomycin-Intermediate S. aureus XN108
Reconstruction of the Vancomycin-Susceptible Staphylococcus aureus Phenotype From a Vancomycin-Intermediate S. aureus XN108
复制标题
从万古霉素中间金黄色葡萄球菌 XN108 重建万古霉素敏感金黄色葡萄球菌表型
DOI：
10.3389/fmicb.2018.02955
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发表时间：
2018-11-28
影响因子：
5.2
通讯作者：
Rao, Xiancai
中科院分区：
文献类型：
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作者：
Peng, Huagang;Rao, Yifan;Rao, Xiancai
万古霉素中间金黄色葡萄球菌 (VISA) 的出现引起了全世界的医疗保健关注。 VISA 通常与多种基因变化有关。然而，这些变化对 VISA 表型的相对贡献尚未完全确定。我们对 VISA XN108 进行了表征，万古霉素 MIC 为 12 μg/ml。基因组比较显示，WalK(S221P)、GraS(T136I) 和 RpoB(H481N) 突变可能导致 XN108 的 VISA 表型。本研究对上述突变进行逐步治愈，并比较XN108及其衍生物之间的表型。我们构建了 4 个同基因突变株：XN108-WalK(P221S)（称为 K65）、XN108-GraS(I136T)（称为 S65）、XN108-RpoB(N481H)（称为 B65）和 XN108-WalK(P221S)/GraS(I136T)（称为KS65)，使用源自万古霉素敏感金黄色葡萄球菌分离株 DP65 的天然等位基因进行等位基因替换实验。抗菌药物敏感性测试显示，K65 和 S65 表现出万古霉素耐药性降低，而 B65 与野生型 XN108 相比差异可忽略不计。依次引入WalK(P221S)和GraS(I136T)将XN108完全转变为VSSA表型。透射电镜和自溶测定表明细胞壁增厚和自溶减少与万古霉素耐药水平的变化有关。与XN108相比，K65显示出577个差异表达基因（DEG），而KS65则显示出555个差异表达基因（DEG）。其中，390个DEG在K65和KS65中常见，包括负责柠檬酸循环和细菌自溶的上调基因，以及涉及肽聚糖生物合成和磷壁酸修饰的下调基因。总之，VSSA 表型可以从 VISA 菌株 XN108 完全重建。 WalK(S221P) 和 GraS(T136I) 突变可能协同作用，赋予 XN108 万古霉素耐药性。
The emergence of vancomycin-intermediate Staphylococcus aureus (VISA) has raised healthcare concerns worldwide. VISA is often associated with multiple genetic changes. However, the relative contributions of these changes to VISA phenotypes are incompletely defined. We have characterized VISA XN108 with vancomycin MIC of 12 μg/ml. Genome comparison revealed that WalK(S221P), GraS(T136I), and RpoB(H481N) mutations possibly contributed to the VISA phenotype of XN108. In this study, the above mutations were stepwise cured, and the phenotypes between XN108 and its derivates were compared. We constructed four isogenic mutant strains, XN108-WalK(P221S) (termed as K65), XN108-GraS(I136T) (termed as S65), XN108-RpoB(N481H) (termed as B65), and XN108-WalK(P221S)/GraS(I136T) (termed as KS65), using the allelic replacement experiments with the native alleles derived from a vancomycin-susceptible S. aureus isolate DP65. Antimicrobial susceptibility test revealed K65 and S65 exhibited decreased vancomycin resistance, whereas B65 revealed negligibly differed when compared with the wild-type XN108. Sequentially introducing WalK(P221S) and GraS(I136T) completely converted XN108 into a VSSA phenotype. Transmission electronic microscopy and autolysis determination demonstrated that cell wall thickening and decreasing autolysis were associated with the change of vancomycin resistance levels. Compared with XN108, K65 exhibited 577 differentially expressed genes (DEGs), whereas KS65 presented 555 DEGs. Of those DEGs, 390 were common in K65 and KS65, including those upregulated genes responsible for citrate cycle and bacterial autolysis, and the downregulated genes involved in peptidoglycan biosynthesis and teichoic acid modification. In conclusion, a VSSA phenotype could be completely reconstituted from a VISA strain XN108. WalK(S221P) and GraS(T136I) mutations may work synergistically in conferring vancomycin resistance in XN108.