Abstract:

Aeromonas veronii is an important pathogen in the co-occurrence of humans and fish, causing significant mortality in various aquatic organisms, posing a major threat to aquaculture and public health care. The type Ⅲ secretion system (T3SS) is one of the most critical virulence mechanisms in determining the pathogenicity of Aeromonas species. The transcription factor ExsA of AraC family serves as the main regulatory factor and activates T3SS assembly and effector protein secretion in response to changes in environmental conditions. In order to explore the effect of exsA on the pathogenicity of the A. veronii, this study constructed an exsA gene knockout strain using homologous recombination strategy and tested its biofilm formation, adhesion capabilities, as well as the cytotoxicity of the spent cultural supernatant. The results showed that the deletion of the exsA gene in A. veronii led to a significant decrease of its biofilm formation capability, its adhesion to ex vivo host tissues, and the cytotoxicity of the spent culture supernatants. Compared with the wild type, the ability of the exsA knockout strain to form biofilm was reduced by 1.5 folds. The adhesion to epithelial cells was not significantly changed in exsA knockout strain as compared with the wild type, while the adhesion to ex vivo mouse intestinal tissues decreased by 3.4 folds and the cytotoxicity of bacterial culture supernatants decreases by 1.2 folds. These results proved that exsA plays an important regulatory role in the virulence mechanism of bacterial pathogen, laying a solid foundation for further research on the function of T3SS in the pathogenic mechanism of A. veronii and provided clues for establishing strategies for the control of the aquatic infection.

Key words: Aeromonas veronii, T3SS, exsA, biofilm, adhesion, virulence

摘要：

维氏气单胞菌是一种重要的人鱼共患病原菌，可以引起多种水产生物的大量死亡，对水产养殖业以及公共卫生安全有重大危害。Ⅲ型分泌系统（type Ⅲ secretion system， T3SS）是决定气单胞菌属致病性最关键的毒力机制之一，AraC家族转录因子ExsA作为主调控因子，响应环境条件变化，激活T3SS组装及效应蛋白分泌。为了探究ExsA对维氏气单胞菌致病性的影响，通过同源重组策略构建exsA基因敲除菌株，并检测其生物膜形成能力、黏附性及培养后上清的细胞毒性。结果显示，维氏气单胞菌中exsA基因缺失导致其生物膜形成能力、对宿主离体组织的黏附能力以及培养后上清液的细胞毒性显著下降。与野生型相比，exsA敲除株形成生物膜的能力降低1.5倍，对上皮细胞的黏附能力无显著性变化，但对小鼠离体肠道组织的黏附能力降低3.4倍，细菌培养后上清的细胞毒性下降1.2倍。研究结果表明ExsA对于维氏气单胞菌的毒力机制有重要调控作用，为进一步阐明T3SS对于病原菌的致病机制以及指导水生生物微生物感染防治策略奠定坚实基础。

关键词: 维氏气单胞菌, Ⅲ型分泌系统, exsA, 生物膜, 粘附能力, 毒力