鸢尾黄素抗产气荚膜梭菌鸡坏死性肠炎的作用及机制

Clostridium perfringens, an important zoonosis, is the main cause agent of avian necrotic enteritis and has become one of the major threaten pathogens in the poultry industry. In China, antibiotics have been used extensively in controling C. perfringens infections. However, the withdrawal times resultes in a rised rate of necrotic enteritis in flocks and affects the performance of broilers, which calls for the development of safe and effective alternatives to antibiotics. As adherence organelles, Type IV pilins（TFP） are important for C. perfringens’ adherence. Recent evidences suggested that the mutant lacking functional TFP displayed attenuated adherence to host cells. Based on the phenotype of TFP systems in C. perfringens, we have identified tectorigenin and other natural compounds exhibited potent inhibition against TFP-linked functions. The aims of this project are to analyze the mechanism of tectorigenin-mediated inhibition of C. perfringens TFP systems by identifying its targets and to elucidate the biochemical and structural basis of such inhibition. The chicken disease model was further used to evaluate the protection of tectorigenin against necrotic enteritis. Our results will provide several effective anti-virulence compounds targeting TFP systems of C. perfringens, which should facilitate the development of novel drugs and new strategies for C. perfringens infections.

产气荚膜梭菌是兽医临床上重要的条件致病菌，可引起鸡及其它家禽的坏死性肠炎，严重危害养禽业的发展。目前我国主要是通过在饲料中添加抗生素来控制细菌感染，但由于大多数抗生素都有较长的休药期，停药后鸡坏死性肠炎的发病率急剧增加，因此急需开发安全有效的抗生素替代药物来控制本病。已有研究报道产气荚膜梭菌四型菌毛（TFP）对其粘附功能起决定性作用，如将TFP缺失，该菌对机体细胞的粘附能力显著下降从而失去致病力。本课题组前期研究通过滑动运动试验和生物被膜形成等试验建立了TFP抑制剂筛选平台，并发现鸢尾黄素和异土木香内酯等多种天然化合物具有较好的抑制活性。本研究拟在此基础上应用分子遗传、生化和生物物理等技术手段分析活性化合物抑制TFP的机制并进行靶点确证；通过肉鸡感染模型评价活性化合物的治疗效果。以期获得多个对鸡坏死性肠炎有较好治疗作用的中药成分，为控制产气荚膜梭菌感染提供候选中药药材和新型抗感染策略。

产气荚膜梭菌是兽医临床上重要的条件致病菌，可引起鸡及其它家禽的坏死性肠炎，严重危害养禽业的发展。目前我国主要是通过在饲料中添加抗生素来控制细菌感染，但由于大多数抗生素都有较长的休药期，停药后鸡坏死性肠炎的发病率急剧增加，因此急需开发安全有效的抗生素替代药物来控制本病。已有研究报道产气荚膜梭菌四型菌毛（TFP）对其粘附功能起决定性作用，如将TFP缺失，该菌对机体细胞的粘附能力显著下降从而失去致病力。本项目通过滑动运动试验和生物被膜形成等试验建立了TFP抑制剂筛选平台，成功筛选获得鸢尾黄素、穗花杉双黄酮和毛蕊花糖苷等多种具有抑制产气荚膜梭菌TFP系统功能的抑制剂，体外抗菌活性分析、TFP功能分析、细胞感染试验和体内动物感染试验等验证了活性活性化合物在无抗菌活性浓度范围内即可有效抑制TFP功能，体外和体内降低细菌的致病力，RT-PCR和肠道菌群分析发现活性化合物抑制TFP形成，降低细菌的粘附和定植，在分子水平上阐明了天然化合物以TFP为靶标发挥抗产气荚膜梭菌感染的作用机制。另外，在此基础上，本项目进一步进行了防治肉鸡坏死性肠炎新兽药川射干口服液的创制。为家禽临床养殖坏死性肠炎的防控提供了新型抗感染策略和潜在的新兽药。