表达PEDV S蛋白重组仙台病毒的构建及其免疫原性研究  

Construction and immunogenicity of a recombinant Sendai virus expressing S protein of PEDV

作　　者：邹静娴 孟慧 潘朔楠 陈振海 牟春晓 ZOU Jingxian;MENG Hui;PAN Shuonan;CHEN Zhenhai;MOU Chunxiao(College of Veterinary Medicine,Yangzhou University,Yangzhou 225009,China;Jiangsu Co-Innovation Center for Prevention and Control of Important Animal Diseases and Zoonosis,Yangzhou University,Yangzhou 225009,China)

机构地区：[1]扬州大学兽医学院,江苏扬州225009 [2]扬州大学江苏高校动物重要疫病与人兽共患病防控协同创新中心,江苏扬州225009

出　　处：《扬州大学学报（农业与生命科学版）》2023年第4期43-50,67,共9页Journal of Yangzhou University：Agricultural and Life Science Edition

基　　金：江苏省农业科技自主创新资金项目[CX(21)2014];广西重点研发计划项目(桂科AB21238003);江苏省高校优势学科建设工程项目(PAPD)。

摘　　要：为构建表达猪流行性腹泻病毒(PEDV)S蛋白的重组仙台病毒(SeV),并评价其诱导的特异性免疫应答反应,利用PCR扩增得到PEDV中国流行株的全长S基因,构建含S基因的重组仙台病毒感染性克隆质粒(pBeloBAC-SeV-PEDV-S),并拯救重组病毒SeV-PEDV-S。对重组病毒进行鉴定并测定其生长曲线以确定成功获得表达PEDV S蛋白的重组仙台病毒。将表达S蛋白的仙台病毒免疫小鼠;间接免疫荧光和Western blot试验检测血清中特异性抗体的产生;利用脾脏淋巴细胞分离和流式细胞技术检测特异性T淋巴细胞增殖反应;对血清中的中和抗体效价进行评价。结果表明:重组仙台病毒能特异性表达PEDV S蛋白,且与野生型仙台病毒有一致的生长曲线;免疫小鼠后,重组仙台病毒能有效刺激小鼠产生特异性抗体,诱导较高的中和抗体水平,且能显著促进PEDV特异性T淋巴细胞增殖。综上,该试验为进一步研究重组仙台病毒在猪体上的免疫反应奠定了基础,也为新一代安全有效的PEDV活载体疫苗的研发提供了策略。To construct the recombinant Sendai virus(SeV)expressing S protein of porcine epidemic diarrhea virus(PEDV)and evaluate the immunogenicity of this recombinant virus.The full length S gene of PEDV was amplified and cloned into the plasmid pBeloBAC-SeV to get pBeloBAC-SeV-PEDV-S,and then the recombinant virus SeV-PEDV-S was rescued.The recombinant virus was identified and the growth curve was determined.After the mice immunized with SeV-PEDV-S,the specific antibody of PEDV in serum was detected by indirect immunofluorescence and Western blot.The proliferation of PEDV specific T lymphocyte was analyzed through spleen lymphocyte isolation and flow cytometry.In addition,the neutralizing antibody titer in serum was also evaluated.The results showed that the recombinant virus SeV-PEDV-S had the similar growth curve with the wild SeV.SeV-PEDV-S effectively stimulated the PEDV specific antibodies,induced a high level of neutralizing antibody titers,and significantly promoted the proliferation of PEDV specific T lymphocytes in the mice.This study provided the theoretical basis for further research on the immune response of SeV-PEDV-S in pigs,and also provided strategies for the development of safe and effective PEDV live vector vaccine.

关 键 词：猪流行性腹泻 S蛋白 重组仙台病毒 免疫原性 

分 类 号：S858.28[农业科学—临床兽医学]