副猪嗜血杆菌荚膜多糖合成调节机理及其对致病力的影响

项目名称： 副猪嗜血杆菌荚膜多糖合成调节机理及其对致病力的影响

项目编号： No.31201934

项目类型： 青年科学基金项目

立项/批准年度： 2013

项目学科： 畜牧学与草地科学、兽医学、水产学

项目作者： 李军星

作者单位： 浙江省农业科学院

项目金额： 23万元

中文摘要： 本课题以严重危害养猪业的副猪嗜血杆菌（HPs）的潜在毒力因子荚膜多糖（CPS）为研究对象，探索致病菌株的CPS转录水平的调控机理及其对HPs致病力的影响。拟构建CPS输出基因Wza缺失株和BadM/Rrf2型转录调节因子缺失株，通过比较缺失株及其亲本菌株CPS表达量及其与脂多糖合成基因簇中14个编码序列转录的相关性，鉴定参与CPS合成的相关基因，并分析Wza及BadM/Rrf2在CPS基因转录调控中的作用；构建启动子检测质粒，鉴定CPS合成相关基因的启动子；通过分析BadM/Rrf2型转录调节因子对CPS合成相关基因调控序列的结合能力，鉴定其靶基因及靶位点，阐明其对CPS合成的调控机理；通过比较荚膜缺失株及其亲本菌株对肺泡巨噬细胞的敏感性、气管上皮细胞的黏附能力及血清抗性等差异，结合其对易感仔猪的致病性强弱，分析CPS在HPs致病过程各重要环节中的作用,为揭示HPs致病机理提供新的依据。

中文关键词： 副猪嗜血杆菌；遗传操作系统；转录调节因子IscR；荚膜多糖输出蛋白Wza；荚膜多糖

英文摘要： This research will focus on capsular polysaccharide (CPS), a potential virulence factor, of haemophilus parasuis (HPs) which is an important bacterial agent involved in porcine industry. Mechanism of biosynthesis regulation at transcriptional level and effect on pathogenicity of haemophilus parasuis capsular polysaccharide will be explored. CPS export gene Wza deficient mutant and BadM/Rrf2 transcriptional regulator gene deficient mutant of pathogenic HPs will be constructed. The transcription of 14 coding sequences (CDS) in LPS biosynthetic regeion will be detected under capsule-inducing culture conditions, and genes invovled in CPS biosynthesis will be determined based on the relationship between gene transcription and CPS production. Regulation effect of Wza and BadM/Rrf2 transcriptional regulator on CPS biosynthesis will be analyzed through comparison of CPS production in both mutant and wild-type HPs. A promoter detection plasmid with green fluorescence protein coding sequence as reporter gene will be constructed, and the promoter of genes involved in CPS biosynthesis will be determined. Combination capacity of BadM/Rrf2 transcriptional regulator to the regulatory sequence of genes involved in CPS biosynthesis will be detected, which will allow the determination of target gene and the target site of combina

英文关键词： Haemophilus parasuis；genetic manipulation system；transcriptional regulator IscR；capsular polysaccharide export protein Wza；Capsular polysaccharide