高致病性2型猪链球菌IV型样分泌系统virB1-89K基因的敲除及其对毒力的影响  

Construction and virulence evaluation of the virB1-89K gene knockout mutant of type Ⅳ-like secretion system of Streptococcus suis serotype 2

作　　者：钟秋[1] 李明[1] 赵岩[1] 陈恬[1] 殷素鹏[1] 王敏[1] 饶贤才[1] 谭银玲[1] 胡福泉[1] 

机构地区：[1]第三军医大学基础部微生物学教研室,重庆400038

出　　处：《微生物学报》2013年第3期276-283,共8页Acta Microbiologica Sinica

基　　金：国家自然科学基金(30901282)~~

摘　　要：【目的】构建高致病性2型猪链球菌IV型样分泌系统virB1-89K基因的敲除株和互补株,研究virB1-89 K基因缺失对细菌毒力的影响。【方法】通过同源重组技术敲除virB1-89 K基因,多重PCR筛选敲除株并测序鉴定。再将virB1-89K基因克隆到穿梭质粒pSET1后转入virB1-89K敲除株中,构建互补株。比较野生株05ZYH33、突变株△virB1-89K和互补株CvirB1-89K三者基本生物学特性的差异,小鼠实验分析virB1-89K基因敲除后对细菌毒力的影响。【结果】成功构建突变株△virB1-89K和互补株CvirB1-89K,在基本生物学性状无明显改变的情况下,敲除株的毒性降低到野生株的30%,互补株可恢复其毒性。【结论】virB1-89K基因作为2型猪链球菌高致病性菌株05ZYH33的IV样分泌系统的重要组分,与其高致病性密切相关。[ Objective] To construct the virB1-89K gene knockout mutant and its complementary strain of Streptococcus suis serotype 2 （SS2） highly virulent strain 05ZYH33 and evaluate the role of virB1-89K in the pathogenesis of SS2. [Methods] The virB1-89K gene was knocked out by homologous recombination, then muhiple-PCR and sequence analysis were used to identify the knockout strain AvirB1-89K. The virB1-89K gene and its upstream promoter were cloned into the E. coli-S, suis shuttle vector pSET1, and the recombinant plasmid was electrotransformed into the AvirBl-89K mutant to generate the complementary strain CvirB1-89K. The effects of virB1-89K deletion on the basic biological characteristics and virulence of SS2 were then determined in this study. [ Results] The isogenic mutant AvirB1-89K and its complementary strain CvirB1-89K were successfully constructed. No significant differences in biological characteristics were found among the three strains. However, the virulence of the AvirB1-89K mutant was reduced to 30% of the wild- type level and functional complementation of virB1-89K restored its pathogenicity. [ Conclusion] The virB1-89K gene plays an important role in the pathogenesis of S. suis 2 infection.

关 键 词：2型猪链球菌 Ⅳ型样分泌系统 virB1基因 基因敲除 

分 类 号：R378[医药卫生—病原生物学]