摘要
为了分析鸡源致病性大肠杆菌地方流行株外膜蛋白(OMPs)免疫保护性,本研究采用Sarkosyl法提取10株鸡源致病性大肠杆菌地方流行株的OMPs,进行浓度测定和超微结构观察,经SDS-PAGE和western blot分析其免疫原性。结果显示,制备的OMPs浓度为1026.632μg/mL^1715.589μg/m L;电镜结果显示其超微结构呈双层泡状的C形或O形;SDS-PAGE分析显示其大小集中在25 ku^55 ku;动物免疫结果显示其具有一定的免疫原性。用提取的OMPs免疫小鼠和雏鸡后,以鸡源致病性大肠杆菌强毒株EC-9攻毒,结果显示:OMPs对小鼠和雏鸡均具有较好保护力,对小鼠保护率为70%,对鸡的保护率为80%;间接EILSA法检测免疫小鼠后的抗体效价最高为1:400,免疫雏鸡后抗体效价最高为1∶800;MTT法检测免疫小鼠后的T淋巴细胞转化率最高为1.582,免疫雏鸡后的T淋巴细胞转化率最高为2.215;常规法分析免疫小鼠脾脏指数为5.334,雏鸡脾脏指数为3.981,表明OMPs可以诱导小鼠和雏鸡产生良好的体液免疫应答和细胞免疫应答。本研究提取的鸡源致病性大肠杆菌地方流行株Omps对小鼠和雏鸡均具有一定的免疫保护性,为致病性大肠杆菌的亚单位疫苗的研制奠定基础。
In order to investigate the immunoprotection of outer membrane proteins(OMPs)of endemic strains of chicken-derived E.coli,this study used Sarkosyl method to extract OMPs of 10 local strains of chicken-borne pathogenic coliforms for concentration determination and ultrastructural observation,followed by analyzing their immunogenicity with SDS-PAGE and western blot.The results showed that the concentration of OMPs was between 1,026.632μg/m L and 1,715.589μg/m L,and the ultrastructure was C-shaped or O-shaped bilayer bubbles.The size of immunoreactiveprotein was between 25 ku and 55 ku,and they are immunogenic.After immunizing mice and chicks with OMPs extracted,they were challenged with EC-9.The results showed that OMPs had good protection against mice and chicks.The protection rateswere 70%on mice,and 80%on chickens,respectively.The OMPs can induce good humoral and cellular immune responses in mice and chicks.Indirect EILSA method was used to detect specific antibody titers.The highest antibody titers was 1∶400 in immunized mice and 1:800 in immunized chickens.MTT method was used to detect T lymphocyte transformation rate in peripheral blood.The highest T lymphocyte transformation rate was 1.582 in immunized mice and 2.215 in immunized chickens.The spleen index of immune organs was 5.334 in mice and3.981 in chickens.In conclusion,the OMPs prepared in this study hadimmunoprotective effects on mice and chicken,and this study laid a foundation for the development of subunit vaccine of pathogenic E.coli.
作者
张艳英
张召兴
郭家媚
李蕴玉
李佩国
索勋
ZHANG Yan-ying;ZHANG Zhao-xing;GUO Jia-mei;LI Yun-yu;LI Pei-guo;SUO Xun(Agricultural University of China,Beijing 10094,China;Hebei Key Laboratory of Preventive Veterinary Medicine,Hebei Normal University of Science&Technology,Qinhuangdao 066004,China;Department of Animal Husbandry and Veterinary Medicine,Hebei Vocational College of Tourism,Chengde 067000,China)
出处
《中国预防兽医学报》
CAS
CSCD
北大核心
2019年第11期1141-1146,共6页
Chinese Journal of Preventive Veterinary Medicine
基金
河北省现代农业产业技术体系蛋鸡产业创新团队岗位资助项目(HBCT2018150206)
河北省高等院校科学技术研究项目(QN20131123,ZD2019092)
关键词
鸡
致病性大肠杆菌
外膜蛋白制备
免疫原性
chicken
pathogenic Escherichia coli
Outer membrane protein preparation
Immunogenicity
