基于F和HN蛋白的基因Ⅶ型新城疫病毒嵌合疫苗的构建及免疫效力评估  

Construction and Immune Efficacy Evaluation of a GenotypeⅦNewcastle Disease Virus Chimeric Vaccine Based on F and HN Proteins

作　　者：李丽 王国康 商雨[1,2,3] 徐英英 冯贺龙 汪宏才 张蓉蓉[1,2,3] 曾哲 罗青平[1,2,3] 邵华斌 曾驰[4] 温国元 LI Li;WANG Guokang;SHANG Yu;XU Yingying;FENG Helong;WANG Hongcai;ZHANG Rongrong;ZENG Zhe;LUO Qingping;SHAO Huabin;ZENG Chi;WEN Guoyuan(Institute of Animal Husbandry and Veterinary,Hubei Academy of Agricultural Sciences,Wuhan 430064,China;Key Laboratory of Prevention and Control Agents for Animal Bacteriosis,Ministry of Agriculture and Rural Affairs,Wuhan 430064,China;Hubei Provincial Key Laboratory of Animal Pathogenic Microbiology,Wuhan 430064,China;School of Biology and Pharmaceutical Engineering,Wuhan Polytechnic University,Wuhan 430023,China)

机构地区：[1]湖北省农业科学院畜牧兽医研究所,武汉430064 [2]农业农村部畜禽细菌病防治制剂创制重点实验室,武汉430064 [3]畜禽病原微生物学湖北重点实验室,武汉430064 [4]武汉轻工大学生物与制药工程学院,武汉430023

出　　处：《中国畜牧兽医》2022年第11期4346-4355,共10页China Animal Husbandry & Veterinary Medicine

基　　金：湖北省自然科学基金杰出青年人才项目(2019CFA071);国家自然科学基金面上项目(31873018);国家现代农业产业技术体系(CARS-41-G13);湖北省农业科技创新中心项目(2019-620-000-001-017);湖北省科技创新项目重大专项(2020ABA016)。

摘　　要：【目的】试验旨在构建一种基因Ⅶ型新城疫病毒(Newcastle disease virus,NDV)嵌合疫苗,并对其免疫效力进行评估。【方法】利用反向遗传学技术,以含有禽偏禽腮腺炎病毒2型(Avian metaavulavirus-2,AMAV-2)Y2株基因组的重组质粒pT7-Y2为模板,将Y2株的F和HN蛋白的胞外区替换为基因Ⅶ型NDV HB0901株的F和HN蛋白的胞外区,将HB0901株的F蛋白裂解位点突变为LaSota弱毒株的F蛋白裂解位点,构建嵌合重组病毒rY2-FHNR株。对rY2-FHNR株的增殖特性、致病力及遗传稳定性等生物学特性进行检测,并通过接种2周龄SPF鸡评估rY2-FHNR株的免疫原性及其免疫血清与Y2株的交叉反应性,利用NDV NP蛋白的间接ELISA方法对免疫血清进行检测,验证其鉴别诊断效果。【结果】试验成功获得了嵌合重组病毒rY2-FHNR株,生物学特性检测结果显示,rY2-FHNR株在鸡胚中的增殖滴度和致病性符合弱毒特征,其鸡胚传代的遗传稳定性良好。rY2-FHNR株可诱导机体产生针对NDV的抗体,且免疫血清不与rY2株抗原发生交叉反应。通过NDV NP ELISA抗体检测方法可以实现区分疫苗免疫与野毒感染。【结论】本试验研发了一种基因Ⅶ型NDV嵌合候选疫苗,为基因Ⅶ型NDV的监测、防控和净化提供了技术支撑。【Objective】This study was aimed to construct a genotypeⅦNewcastle disease virus(NDV)chimeric vaccine,and access its immune efficacy.【Method】The F and HN protein ectodomains of recombinant plasmid pT7-Y2 of the Avian metaavulavirus-2(AMAV-2)Y2 strain were replaced with the corresponding F and HN protein ectodomains of genotypeⅦNDV HB0901 strain by utilizing reverse genetic technology.Then F protein cleavage site of HB0901 strain was transformed into that of lentogenic LaSota strain,and the chimeric virus was recovered and named rY2-FHNR.The growth characteristics,pathogenicity and genetic stability of rY2-FHNR strain were evaluated,then 2-week-old SPF chickens were immunized with rY2-FHNR to determine the immunogenicity and cross-reactivity between immune serum and Y2 strain.The immune serum was identified by indirect NDV NP ELISA to verify its diagnosis effect.【Result】Chimeric recombinant virus strain rY2-FHNR was constructed successfully.The biological characteristics results showed that the proliferation titer and pathogenicity in chicken embryos of rY2-FHNR strain were avirulent,while the chimeric virus was genetic stable when serially passaged in chicken embryos.Immunization with rY2-FHNR stimulated antibodies against NDV,and the immune serum had no cross-reactivity with rY2 strain.Through indirect NDV NP ELISA could distinguish infected animals from vaccinated animals.【Conclusion】This study developed a genotypeⅦNDV chimeric vaccine candidate,which could be used as chimeric vaccine,provided technological support for the monitoring,prevention and eradication of genotypeⅦNDV.

关 键 词：新城疫病毒(NDV) 基因Ⅶ型 禽偏禽腮腺炎病毒血清2型(AMAV-2) 反向遗传技术 嵌合疫苗 

分 类 号：S852.657[农业科学—基础兽医学]