副溶血弧菌VP2918基因缺失株的构建及功能研究

Gene deletion and functional analysis of vp2918 gene in Vibrio parahemolyticus

摘要:

[目的] 以副溶血弧菌VP2918为研究对象，研究其对副溶血弧菌的生物学特性和致病性的影响。[方法] 利用同源重组技术构建了vp2918基因的基因缺失株（Δvp2918）和互补株（CΔvp2918），并对野生株、缺失株和互补株的细菌生长曲线、运动性、生物被膜形成能力、对HeLa细胞的黏附能力、细胞毒性、对小鼠的致死率和组织载菌量进行分析。[结果] 缺失vp2918基因不影响副溶血弧菌的生长特性、运动性、生物被膜形成能力以及对HeLa细胞的黏附能力。但与野生株相比，Δvp2918对HeLa细胞的毒性作用显著降低；感染Δvp2918的小鼠症状明显减轻，存活率更高；Δvp2918在小鼠脾脏和肝脏中的载菌量显著低于野生株，互补株毒力基本恢复至野生株水平。[结论] vp2918不参与副溶血弧菌的运动性和生物被膜形成能力等过程，但与该菌的致病性相关，为潜在的毒力因子。

Abstract:

[Objective] By using comparative proteomics, we identified several potential T6SS-2 effectors, including VP2918. The aim of this study is to elucidate the role of VP2918 in biological characteristics and pathogenicity of Vibrio parahaemolyticus. [Methods] The vp2918 gene deletion mutant strain (Δvp2918) and complementary strain (CΔvp2918) were constructed via homologous recombination technology. Subsequently, the growth characteristics, biofilm formation ability, motility, adhesion and cytotoxicity to HeLa cells, and mice lethality rate, bacterial colonization were analyzed in the wild-type strain (WT), Δvp2918 and CΔvp2918. [Results] The biological characteristics analysis in vitro showed that there was no significant difference in growth curve, motility, biofilm formation ability and adhesion ability to HeLa cells among WT, Δvp2918 and CΔvp2918. However, compared with WT, the toxicity of Δvp2918 to HeLa cells was significantly weakened. Animal experimental results showed that compared with the mice infected with WT, the mice infected with Δvp2918 showed slighter symptoms and higher survival rate. The bacterial loads of Δvp2918 were significantly lower than that of WT in spleen and liver of infected mice, whereas the complementation strain restored the virulence to resemble that of WT. [Conclusion] The VP2918 was not associated with the motility and biofilm formation ability, but it was a potential virulence factor and associated with the pathogenic of V. parahaemolyticus.