肠炎沙门菌sipA基因缺失株C50336ΔsipA的构建及鉴定

Construction and characterization of a sipA mutant of Salmonella enteritidis C50336 strain

为研究sipA基因对肠炎沙门菌(Salmonella enteritidis)的生物学特性的影响,利用λ-Red同源重组系统构建肠炎沙门菌C50336的sipA基因缺失株,同时将sipA基因克隆至pBR322质粒,将回补质粒电转化至缺失株,成功构建了回补株。通过比较肠炎沙门菌野生株、缺失株以及回补株在生长特性、生化特性、遗传稳定性、对小鼠的半数致死量以及对Caco-2细胞的黏附侵袭能力的差异,发现肠炎沙门菌sipA基因缺失株具有良好的遗传稳定性;与野生株及回补株相比,其生长特性和生化特性未发生明显改变,对8周龄小鼠的毒力没有显著变化,不影响细菌对Caco-2细胞的黏附能力,但对Caco-2细胞的侵袭能力明显下降。上述研究结果为进一步研究sipA基因在肠炎沙门菌感染过程中的功能奠定了基础。

To investigate the role of sipA gene from Salmonella enteritidis in biological characteristics,the sipA mutant of Salmonella enteritidis was constructed by λ-Red recombinant system and the corresponding complementary strain C50336ΔsipApsipA was also developed using pBR322 plasmid to carry the gene.The characterization including growh property,biochemical property,genetic stability,and median lethal dose（LD50） was then compared among the three strains. In addition,the adhesion and invasion ability to Caco-2 cells by C50336ΔsipA was also determined to evaluate the role of sipA in Salmonella enteritidis infection. Among the three strains,there were no obviously differences in growth property,biochemical properties and virulence. The pass-generation test showed that the hereditary property of the mutant strain was stable. The infection analysis showed that the adhesion ability to Caco-2 cells by C50336ΔsipA was similar to wild-type strain,but the invasion ability was significantly decreased. The research provided the foundation for studying the role of sipA in the process of Salmonella enteritidis infection.