Fisheries College, Key Laboratory of Healthy Mariculture for the East China Sea, Ministry of Agriculture, Jimei University, Xiamen, Fujian, China.
Third Institute of Oceanography, State Oceanic Administration, Xiamen, Fujian, China.
Microbiologyopen. 2017 Dec;6(6). doi: 10.1002/mbo3.517. Epub 2017 Jul 25.
Adhesion is a key virulence factor of pathogens and can be affected by the environment. Our previously research with RNA-seq indicated that mcp, aer, cheB, and cheV might play roles in the regulation of adhesion in Vibrio alginolyticus (ND-01). In order to determine whether and how environmental factors affect adhesion through these genes, gene silencing was performed followed by quantitative real-time PCR (qRT-PCR), RNAi, transmission electron microscopy, and adhesion, capillary, and motility assays to verify how these genes influence adhesion. Silencing these genes led to deficiencies in adhesion, chemotaxis, flagellar assembly, and motility. The expression levels of cheA, cheW, and cheY, which are important genes closely related to the functions of mcp, aer, cheV, and cheB, were significantly downregulated in all of the RNAi groups. The expression of mcp, aer, cheV, and cheB under different gradients of temperature, pH, and salinity and after starvation for various durations was also detected, which showed that these genes were sensitive to certain environmental stresses, particularly pH and starvation. Our results indicated that mcp, aer, cheB, and cheV: (1) are necessary for ND-01 adhesion; (2) play key roles in the bacterial chemotaxis pathway by controlling the expression of downstream genes; (3) might affect adhesion by impacting motility, though motility is not the only route through which adhesion is affected; and (4) contribute to the regulation of ND-01 adhesion in natural environments with different temperatures, pH levels, and salinities as well as after various starvation periods.
黏附是病原体的一个关键毒力因子,它可以受到环境的影响。我们之前的 RNA-seq 研究表明,mcp、aer、cheB 和 cheV 可能在弧菌(ND-01)的黏附调控中发挥作用。为了确定环境因素是否以及如何通过这些基因影响黏附,我们进行了基因沉默实验,然后进行了定量实时 PCR(qRT-PCR)、RNAi、透射电子显微镜以及黏附、毛细和运动测定,以验证这些基因如何影响黏附。这些基因的沉默导致黏附、趋化性、鞭毛组装和运动能力缺陷。与 mcp、aer、cheV 和 cheB 功能密切相关的重要基因 cheA、cheW 和 cheY 的表达水平在所有 RNAi 组中均显著下调。还检测了 mcp、aer、cheV 和 cheB 在不同温度、pH 值和盐度梯度下以及在不同时间饥饿后的表达情况,结果表明这些基因对某些环境压力敏感,尤其是 pH 值和饥饿。我们的研究结果表明,mcp、aer、cheB 和 cheV:(1) 是 ND-01 黏附所必需的;(2) 通过控制下游基因的表达在细菌趋化途径中发挥关键作用;(3) 可能通过影响运动性来影响黏附,尽管运动性不是影响黏附的唯一途径;(4) 有助于在不同温度、pH 值和盐度的自然环境以及在不同的饥饿期后调节 ND-01 的黏附。