鸭源鸡杆菌ompW、gtxA双基因缺失株的构建及其相关特性分析  

Construction and characterization of Gallibacterium anatis △ompW△gtxA double deletion strain

作　　者：夏银河 王坤芃 刘盼盼 王新卫[1] 常洪涛[1] 彭志锋[1] 刘红英[1] 陈陆[1] 杨霞[1] 王川庆[1] Yinhe Xia;Kunpeng Wang;Panpan Liu;Xinwei Wang;Hongtao Chang;Zhifeng Peng;Hongying Liu;Lu Chen;Xia Yang;Chuanqing Wang(College of Animal Science and Veterinary Medicine,Henan Agricultural University,Zhengzhou 450002,China)

机构地区：[1]河南农业大学牧医工程学院,河南郑州450002

出　　处：《微生物学报》2020年第10期2311-2322,共12页Acta Microbiologica Sinica

基　　金：河南省科技厅科技攻关项目(172102110046);河南省自然科学基金(162300410153)。

摘　　要：鸭源鸡杆菌(Gallibacterium anatis,G.anatis)是家禽中常见的条件性致病菌,主要引起蛋鸡生殖道疾病,造成产蛋量下降。【目的】为探究RTX样毒素GtxA及外膜蛋白(OmpW)对鸭源鸡杆菌生物学特性和致病力的影响。【方法】本研究采用自然转化法对突变株RZ△ompW进一步缺失gtxA构建突变株RZ△ompW△gtxA,通过分析其生长特性、黏附能力、引起细胞凋亡程度及对小鼠致病性等,探究其与生物学特性及致病性可能的关系。【结果】结果显示,RZ△ompW△gtxA能稳定遗传gtxA的缺失;单双基因突变株溶血活性均消失、与RZ株相比菌落形态及生长速率并未出现显著改变;相比RZ、RZ△ompW和RZ△gtxA,双基因缺失株RZ△ompW△gtxA在不同时段对鸡原代输卵管上皮细胞的黏附能力显著降低(P<0.05),诱导鸡输卵管上皮原代细胞发生凋亡的能力明显减弱,对小鼠的致病力显著降低。【结论】GtxA毒素和外膜蛋白OmpW在鸭源鸡杆菌毒力、黏附宿主细胞及诱导其细胞凋亡中起重要作用,且可能存在明显的协同关系。本研究为鸭源鸡杆菌感染机制的阐明奠定基础。Gallibacterium anatis is a conditional pathogen common in poultry production, and causes the laying hen genital tract diseases, resulting in decreased egg production. [Objective] To understand the biological characteristics and pathogenicity of the virulence factor RTX-like toxin GtxA and outer membrane protein W of G. anatis. [Methods] Based on the G. anatis OmpW gene deletion strain, we successfully constructed the OmpWgtxA double gene deletion strain by natural transformation. The differences in biological characteristics and virulence between single and double gene deletion strains and parental strain were investigated. [Results] The hemolytic activity of the OmpWgtxA double gene deletion strain was inactivated, while the deletion was stable in vitro and in vivo. Compared with gtxA, ompW single gene deletion strain and parental strain Yu-PDS-RZ-1-SLG, the growth of the double gene deletion strain was not significantly altered, while its adhesion ability and lesions of primary chicken oviduct epithelial cells were decreased to some extent;and its ability to induce apoptosis in cells of chicken oviduct epithelium was lessened;its pathogenicity was also reduced. [Conclusion] GtxA toxin and outer membrane protein W were suggested to play important roles in G. anatis virulence, adhesion to host cells and induction of apoptosis and there may be synergies. This study laid the foundation for further revealing the pathogenesis of G. anatis.

关 键 词：鸭源鸡杆菌 RZ△ompW△gtxA缺失株 生物学特性 致病性 

分 类 号：S852.61[农业科学—基础兽医学]