副猪嗜血杆菌HbpA蛋白功能及其在致病中的作用研究

项目名称： 副猪嗜血杆菌HbpA蛋白功能及其在致病中的作用研究

项目编号： No.31201907

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： 张艳禾

作者单位： 中国农业科学院哈尔滨兽医研究所

项目金额： 24万元

中文摘要： 副猪嗜血杆菌（Hps）是严重危害全球养猪业的重要病原菌，但其摄取血红素铁的能力与致病机制的关系不清楚。研究发现血红素结合蛋白（HbpA）是铁摄取系统的重要组成部分，具有结合宿主体内血红素功能并且是主要毒力因子。体外试验鉴定Hps的一个具有免疫原性的HbpA蛋白，但其在体内的作用仍然不详。本项目拟以本课题组在Hps中新发现的HbpA蛋白为研究对象，先采用Heme-agarose结合方法验证HbpA蛋白的精确功能，结合免疫电镜确定Hps结合血红素的细胞定位；再构建hbpA基因缺失突变株，进行细胞感染存活试验、病理学观察和组织细菌分布和定量分析，明确缺失hbpA基因突变株与Hps增殖水平的关系以及对Hps致病力的影响，最终结合转录组测序分析差异表达基因阐明HbpA蛋白在致病过程中的作用。研究结果将丰富Hps与宿主特异适应性的分子致病机理，也为Hps阻断性药物治疗设计提供理论依据。

中文关键词： 副猪嗜血杆菌；血红素结合蛋白；蛋白功能；毒力因子；致病性

英文摘要： Haemophilus parasuis(Hps) is the most important pathogenic bacteria which is severely harmful to pig farming industry. However, the pathogenic mechanism and its ability to acquire heme iron have not been explored. In vitro study has proved that HbpA from Hps is a secreted, immunogenic protein. Nevertheless, the in vivo function is still unknown. This project will be carried out with following approaches based on the new identified HbpA protein by our team. First, the accurate function of HbpA protein will be identified using Heme-Agarose binding assay Cellular localization of binding between heme and Hps will be studied by immunoelectron microscopy. Construction of HbpA gene mutation, performing cell infection test, pathological observation and distribution and quantification of the bacteria could clarify the effect of HbpA protein in regarding to the survival and reproduction; and the Pathogenicity of Hps. Finally, the function of HbpA protein in the process of Pathogenicity will be illustrated with above study combined with analysis on the differentially expressed genes by transcriptomic sequencing. The research data will not only enrich molecular basis of the pathogenesis of specific adaption of Hps to the host, but also provide theoretical foundation for designing drugs against Hps.

英文关键词： Haemophilus parasuis；heme-binding peotein；protein function；virulence factor；pathogenisis of infection