奶牛源绿脓杆菌内毒素蛋白的提取及免疫原性和保护性研究  

Study on extraction and immunogenicity and protection of endotoxin protein of Pseudomonas aeruginosa in cow

作　　者：曲晓军[1,2] 王金英[1,2] 孙建华[1,2] 于冲[1,2] 夏海华[1,2] 潘钰[1,2] QU Xiaojun;WANG Jinying;SUN Jianhua;YU Chong;XIA Haihua;PAN Yu(Institute of Microbiology,Heilongjiang Academy of Sciences,Harbin 150010,China;Institute of Advanced Technology, Heilongjiang Academy of Sciences,Harbin 150020,China)

机构地区：[1]黑龙江省科学院微生物研究所,哈尔滨150010 [2]黑龙江省科学院高技术研究院,哈尔滨150020

出　　处：《黑龙江畜牧兽医》2019年第7期25-28,共4页Heilongjiang Animal Science And veterinary Medicine

基　　金：黑龙江省院所基本应用技术研究专项(ZNJZ2016SW01);黑龙江省科学院科学研究基金项目(ZS201604)

摘　　要：为了评估绿脓杆菌内毒素蛋白的免疫原性和保护性,试验采用最小致死量(MLD)测定法从奶牛源绿脓杆菌菌株中筛选高毒力菌株,用可使蛋白质与脂多糖分离的合成培养基进行发酵、提取、纯化制备内毒素蛋白,采用考马斯亮蓝法测定内毒素蛋白含量,间接ELISA法和免疫攻毒保护试验对制备的内毒素蛋白进行免疫原性和保护性研究。结果表明:筛选得到1株高毒力绿脓杆菌CPA-009菌株,其MLD为2.0×10~7 cfu/mL;用CPA-009菌株发酵制备出5批次内毒素蛋白,平均含量为44.62%,各批次间差异不显著(P>0.05);免疫组小鼠体内抗体效价随着免疫进程持续升高,最高达到1∶12 500;内毒素蛋白对CPA-009菌株的保护率为100%,对其他3种血清型绿脓杆菌菌株的保护率均在80.0%以上。说明由高毒力绿脓杆菌CPA-009菌株制备的内毒素蛋白具有良好的免疫原性和高保护性。In this test,we used the minimum lethal dose method ( MLD) to screen the highly toxic strain from the Pseudomonas Aeruginosa strain,and used a synthetic medium that can separate the protein from lipopolysaccharide to ferment,extract,and purify the endotoxin protein. Kaomasiliang method was used to determine the content of endotoxin protein. The immunogenicity and protection of endotoxin protein were stud-ied by indirect ELISA and immunoprotective test.The results showed that CPA-009 strain of Pseudomonas aeruginosa with high virulence were selected,and it’s MLD was 2. 0×10^7 cfu /mL.Five batches of endotoxin protein were prepared by liquid fermentation of CPA-009 strain,the average protein content was 44. 62%,there was no significant difference between batches ( P>0. 05).The titer of antibody in immune group in-creased with the progress of immunization,and reached the highest level of 1 ∶12 500.The protection rate of endotoxin protein to CPA-009 strain was 100%,and the protection rate to other 3 kinds of serotype of Pseudomonas aeruginosa were more than 80. 0%. It indicates that endotoxin protein produced by the high virulent strain CPA-009 has good immunogenicity and high protection.

关 键 词：绿脓杆菌 内毒素蛋白 提取 免疫 间接ELISA 攻毒 

分 类 号：S823[农业科学—畜牧学] Q51[农业科学—畜牧兽医]