Fisheries College, Key Laboratory of Health Mariculture for the East China Sea, Ministry of Agriculture and Rural Affairs, Jimei University, Xiamen, China.
J Fish Dis. 2022 Nov;45(11):1609-1621. doi: 10.1111/jfd.13684. Epub 2022 Jul 12.
Aeromonas hydrophila infections are common in aquaculture. Our previous studies found that the A. hydrophila B11 strain can survive in fish macrophages for at least 24 h and the two-component system EnvZ/OmpR may be involved in intracellular survival. To reveal the role and mechanism of the two-component system EnvZ/OmpR in intracellular survival of A. hydrophila, the genes of envZ/ompR were silenced by shRNAi. The results showed that the survival rates of the envZ-RNAi and ompR-RNAi strains were only 2.05% and 3.75%, respectively, which were decreased by 91% and 83.6% compared with that of the wild-type strain. The escape ability of envZ-RNAi and ompR-RNAi was also decreased by 51.4% and 19.7%, respectively. The comparative transcriptome analysis revealed that the functional genes directly related to bacterial intracellular survival mainly included the genes related to anti-stress capacity, and the genes related to Zn and Mg transport. Further research confirmed that two-component system EnvZ/OmpR can regulate the expression of the important molecular chaperones, such as groEL, htpG, dnaK, clpB and grpE. The expression of these molecular chaperones in wild-type strain was up-regulated with the increase in H O concentrations, while the expression of these molecular chaperones in silent strains did not change significantly. Cells that phagocytosed wild-type strain had higher ROS content than cells that phagocytosed silent strains. Two-component system EnvZ/OmpR could also regulate zinc transporter (znuA, znuB, znuC) and zinc efflux protein (zntA) to maintain zinc homeostasis in cells, thus affecting the ability of bacteria to survive in phagocytes. Moreover, two-component system EnvZ/OmpR could affect the growth and intracellular survival of A. hydrophila by regulating the expression of MgtA, MgtC and MgtE and participating in bacterial Mg homeostasis in fish macrophages.
嗜水气单胞菌感染在水产养殖中很常见。我们之前的研究发现，A. hydrophila B11 菌株至少可以在鱼巨噬细胞中存活 24 小时，而双组分系统 EnvZ/OmpR 可能参与细胞内生存。为了揭示双组分系统 EnvZ/OmpR 在 A. hydrophila 细胞内生存中的作用和机制，我们通过 shRNAi 沉默了 envZ/ompR 基因。结果表明，envZ-RNAi 和 ompR-RNAi 菌株的存活率分别仅为 2.05%和 3.75%，与野生型菌株相比分别下降了 91%和 83.6%。envZ-RNAi 和 ompR-RNAi 的逃逸能力也分别下降了 51.4%和 19.7%。比较转录组分析表明，与细菌细胞内生存直接相关的功能基因主要包括与抗应激能力相关的基因，以及与 Zn 和 Mg 转运相关的基因。进一步的研究证实，双组分系统 EnvZ/OmpR 可以调节重要分子伴侣的表达，如 groEL、htpG、dnaK、clpB 和 grpE。随着 H2O2 浓度的增加，野生型菌株中这些分子伴侣的表达上调，而沉默菌株中这些分子伴侣的表达没有明显变化。吞噬野生型菌株的细胞的 ROS 含量高于吞噬沉默菌株的细胞。双组分系统 EnvZ/OmpR 还可以通过调节锌转运蛋白（znuA、znuB、znuC）和锌外排蛋白（zntA）来维持细胞内的锌稳态，从而影响细菌在吞噬细胞中的生存能力。此外，双组分系统 EnvZ/OmpR 通过调节 MgtA、MgtC 和 MgtE 的表达和参与鱼类巨噬细胞中细菌 Mg 稳态来影响 A. hydrophila 的生长和细胞内生存。
