Zhou Xiujuan, Liu Bin, Shi Chunlei, Shi Xianming
MOST-USDA Joint Research Center for Food Safety, School of Agriculture & Biology, and State Key Lab of Microbial Metabolism, Shanghai Jiao Tong University, Shanghai, China.
College of Food Science and Engineering, Northwest Agriculture & Forestry University, Shaanxi, Yangling, China.
PLoS One. 2014 Sep 11;9(9):e106708. doi: 10.1371/journal.pone.0106708. eCollection 2014.
Several molecular detection marker genes specific for a number of individual Salmonella serogroups have been recently identified in our lab by comparative genomics for the genotyping of diverse serogroups. To further understand the correlation between serotype and genotype, the function of a Salmonella serogroup-C1-specific gene (SC_2092) was analyzed in this study. It was indicated from the topological prediction using the deduced amino acid sequence of SC_2092 that this putative protein was highly similar to the confirmed Wzx flippases. Furthermore, SDS-PAGE revealed that lipopolysaccharide (LPS) biosynthesis, specifically O-antigen synthesis, was incomplete in an SC_2092 in-frame deletion mutant, and no agglutination reaction with the O7 antibody was exhibited in this mutant. Therefore, it was revealed that this Salmonella serogroup-C1-specific gene SC_2092 encoded a putative flippase, which was required for O7-polysaccharide biosynthesis, and was designated here as wzxC1. Subsequently, the effects of the deletion of wzxC1 on bacterial motility and sodium chloride (NaCl) tolerance were evaluated. The wzxC1 mutant lacked swarming motility on solid surfaces and was impaired in swimming motility in soft agar. Moreover, microscopic examination and RT-qPCR exhibited that an increased auto-aggregation and a strong defect in flagella expression, respectively, were responsible for the reduced motility in this mutant. In addition, the wzxC1 mutant was more sensitive than the wild-type strain to NaCl, and auto-aggregation of mutant cells was observed immediately up on the addition of 1% NaCl to the medium. Interestingly, the motility deficiency of the mutant strain, as well as the cell agglomeration and the decrease in flagellar expression, were relieved in a NaCl-free medium. This is the first study to experimentally demonstrate a connection between a Salmonella serogroup specific gene identified by comparative genomics with the synthesis of a specific O-antigen biosynthesis. Also, our results show that the mutation of wzxC1 triggers a NaCl-dependent motility deficiency.
最近，我们实验室通过比较基因组学鉴定出了一些针对多个沙门氏菌血清群的分子检测标记基因，用于不同血清群的基因分型。为了进一步了解血清型与基因型之间的相关性，本研究分析了沙门氏菌血清群C1特异性基因（SC_2092）的功能。使用SC_2092推导的氨基酸序列进行的拓扑预测表明，该推定蛋白与已确认的Wzx翻转酶高度相似。此外，SDS-PAGE显示，在SC_2092框内缺失突变体中，脂多糖（LPS）生物合成，特别是O抗原合成不完整，并且该突变体未表现出与O7抗体的凝集反应。因此，揭示了这个沙门氏菌血清群C1特异性基因SC_2092编码一种推定的翻转酶，它是O7多糖生物合成所必需的，在此被命名为wzxC1。随后，评估了wzxC1缺失对细菌运动性和氯化钠（NaCl）耐受性的影响。wzxC1突变体在固体表面缺乏群体运动性，在软琼脂中的游动运动性受损。此外，显微镜检查和RT-qPCR显示，分别是自聚集增加和鞭毛表达严重缺陷导致了该突变体运动性降低。此外，wzxC1突变体比野生型菌株对NaCl更敏感，在培养基中加入1%NaCl后立即观察到突变体细胞的自聚集。有趣的是，在无NaCl培养基中，突变菌株的运动性缺陷以及细胞聚集和鞭毛表达减少的情况得到缓解。这是第一项通过实验证明比较基因组学鉴定出的沙门氏菌血清群特异性基因与特定O抗原生物合成之间联系的研究。而且，我们的结果表明wzxC1的突变引发了NaCl依赖性运动性缺陷。
