鸡白痢沙门菌诱导Th2应答的抗原发掘及免疫机制研究

Pullorum disease is the key bacterial infectious disease of poultry, and costs persistent damage in poultry industry in China. As few studies were performed to reveal the pathogenesis and immune mechanism of Salmonella enterica serovar Pullorum (Salmonella Pullorum), it is not conductive to prevent and control the spread of disease. Salmonella Pullorum is the host-restricted serovar, which can cause systemic infection to chicken with high mortality rate to young chicks and display persistent infection in adult chickens accompanied by vertical transmission. Recent studies showed the Salmonella Pullorum is inclined to induce the specific Th2-bias immune response with difference to the combined Th1 and Th2 immune response caused by host-ranged Salmonella enterica serovar Enteritidis (Salmonella Enteritidis). So the project choose the immune responses caused by different genotypes of Salmonella Pullorum as the breakthrough point, and analyze the characteristic of immune response caused by Salmonella Pullorum in host (avian cells and chicken). On this basis, the antigens inducing Th2 immune response will be screened from transposon mutagenesis library of Samonella Pullorum. Following with structure and function analysis of these effector proteins, we will analyze the signal transduction pathway and immune response caused by them. Finally to systemically reveal the roles played by these antigens in persistent infection or host restriction for Salmonella Pullorum. This study will provide new knowledge, new target and new technology for the control and eradication of Pullorum disease.

鸡白痢对我国养禽业危害巨大。目前对鸡白痢沙门菌的分子致病与免疫机制研究甚少，不利于该病的防控。鸡白痢沙门菌为宿主限制性致病菌，感染鸡后可引起全身性感染，对雏鸡致死率高，对成年鸡可造成持续感染，引发垂直传播。前期的研究发现鸡白痢沙门菌呈现独特的诱导Th2倾向的免疫应答，明显不同于宿主谱广的肠炎沙门菌等诱导的Th1/Th2混合型免疫应答，但对其分子免疫机制及致病中的作用亟待揭示。本项目以不同基因组型鸡白痢沙门菌感染后引起的免疫应答分析为切入点，揭示该菌在感染宿主（细胞和鸡）的过程中引起的免疫应答特征。在此基础上，从鸡白痢沙门菌的转座突变体库中发掘诱导Th2免疫应答的新抗原；通过对这些抗原结构与功能解析，探明其作用的信号通路和Th2免疫效应机制，探究它们在持续感染或宿主特异性致病中的作用。本研究将为鸡白痢防控和净化提供新知识、新靶点和新技术基础。

鸡白痢是危害我国养禽业的重要细菌性疾病之一，对鸡白痢沙门菌的分子免疫机制研究的匮乏不利于该病的防控。本项目基于前期发现的鸡白痢沙门菌诱导宿主Th2倾向性免疫应答的特征，利用细胞感染模型发掘诱导Th2应答的新抗原；通过对抗原编码基因的缺失株引起的免疫应答分析和对抗原结构与功能的解析探明这些抗原作用的信号通路和分子免疫机制，探究它们在鸡白痢沙门菌宿主特异性致病中的作用，为鸡白痢的防控净化提供新认识、新靶点和新技术基础。.本项目首先从比较基因组学水平揭示了全球鸡白痢沙门菌的遗传演化规律，将全球鸡白痢沙门菌分为四个克隆群，揭示了鸡白痢沙门菌的全球传播与历史事件“母鸡热”相关联，并阐明了可自发接合转移的IncX1和IncN型MDR质粒的出现和扩散是我国近20年鸡白痢沙门菌耐药性显著上升的主要分子机制。随后，我们建成鸡白痢沙门菌与宿主细胞和禽互作研究的技术平台，建立了鸡白痢沙门菌与禽原代巨噬细胞及原代T细胞的感染互作模型，揭示了鸡白痢沙门菌诱导宿主Th2倾向性免疫应答的分子基础，挖掘出10个Th2应答相关基因。同时，我们创建了鸡白痢沙门菌的比较基因组学分析新平台，联合基于cuSNPs的分析与筛选鸡白痢沙门菌诱导Th2应答相关抗原的“Duo”软件，揭示启动子区域的点突变会影响效应蛋白的表达及其参与引起的免疫应答。重点发现鸡白痢沙门菌IpaJ蛋白为T3SS效应蛋白，并解析了其受转录调控因子ItrA的直接调控；揭示了IpaJ蛋白为半胱氨酸蛋白酶家族成员之一，通过阻止NF-κB与MAPK信号通路的活化抑制宿主Th1应答的分子免疫机制，丰富了对鸡白痢沙门菌宿主限制性的认识。此外，我们确定了鸡白痢沙门菌减毒株SKLZΔspiC可以研发成为防控鸡白痢的减毒活疫苗，正在积极申报兽药证书；并且建立了鸡白痢沙门菌基因组等重要数据库，保存有鸡白痢沙门菌菌株1000多株。本项目深化了对鸡白痢沙门菌诱导宿主Th2倾向性免疫应答机制的认识，对该病的防控新技术研究具有重要价值。