QX型传染性支气管炎病毒病毒样颗粒的制备与初步评价  

Preparation and Preliminary Evaluation of Virus-like Particles of QX Genotype Infectious Bronchitis Virus

作　　者：张梦寒 程敏 陈杨 薄宗义 张成成 郭梦娇 吴艳涛[1,2] 张小荣 ZHANG Menghan;CHENG Min;CHEN Yang;BO Zongyi;ZHANG Chengcheng;GUO Mengjiao;WU Yantao;ZHANG Xiaorong(College of Veterinary Medicine,Yangzhou University,Yangzhou,Jiangsu 225009;Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses,Yangzhou,Jiangsu 225009)

机构地区：[1]扬州大学兽医学院,江苏扬州225009 [2]江苏高校重要动物疫病与人兽共患病协同创新中心,江苏扬州225009

出　　处：《中国家禽》2024年第5期46-55,共10页China Poultry

基　　金：国家重点研发计划项目子课题(2022YFD1801004-6);财政部和农业农村部:国家现代农业产业技术体系资助项目(CARS-40-K16);江苏高校优势学科建设工程资助项目(2022年)。

摘　　要：研究旨在制备一种针对QX型传染性支气管炎病毒(Infectious bronchitis virus,IBV)的病毒样颗粒(Virus-like particle,VLP)疫苗并对其免疫保护效果进行初步评价。试验首先构建了分别表达QX型IBV S蛋白(QXLS)、M蛋白(QXLM)和E蛋白(QXLE)的三种重组杆状病毒rBacmid-QXLS、rBacmid-QXLM和rBacmid-QXLE,并通过间接免疫荧光试验(IFA)和West-ern blot鉴定目的蛋白的表达;进一步将3种重组杆状病毒通过适当比例共感染Sf9细胞获得VLP,经蔗糖密度梯度离心纯化后使用透射电子显微镜观察鉴定VLP的组装效果;最后将纯化的VLP与氢氧化铝佐剂混合制成疫苗免疫SPF鸡进行免疫保护效果评价。结果显示:S、M和E三种目的蛋白均能成功表达,透射电子显微镜观察可见组装形成的VLP呈现典型的椭圆形病毒样结构,直径为80~120 nm;单独VLP疫苗免疫具有一定的免疫保护效果,QXL87+VLP疫苗联合免疫保护效果较好,可有效减轻临床病变、降低气管和泄殖腔排毒量及组织器官载毒量。研究表明,试验已经成功制备出QX型IBV VLP,且具有良好的免疫原性,为后续进一步研制基于VLP技术的QX型IBV基因工程疫苗奠定了基础。This study aimed to prepare a virus-like particle(VLP)vaccine against QX genotype infectious bronchitis virus(IBV)and evaluate its immunoprotection potential.Three recombinant baculoviruses(rBacmid QXLS,rBacmid QXLM,and rBacmid QXLE)were constructed,each expressing a different IBV QX structural protein(S,M,and E,respectively).The expression of the target proteins was confirmed by indirect immunofluorescence assay(IFA)and Western blot.The Sf9 cells were co-infected with the three recombinant baculoviruses in the appropriate ratio to obtain VLPs,which were purified by sucrose density gradient centrifugation and observed using transmission electron microscopy(TEM)to assess the assembly of the VLPs.The purified VLPs were combined with aluminium hydroxide adjuvant to form a vaccine,which was then used to immunize SPF chickens to evaluate its immunoprotection.The results showed the three target proteins were successfully expressed,and the TEM observation revealed that the assembled VLPs had a typical coronavirus-like structure,with a diameter of 80 to 120 nm.The immunization of individual VLP vaccine had a certain degree of immune protection effect,and the combination immunization of QXL87 and VLP vaccine had a better immune protection effect,which could effectively reduce clinical lesions,tracheal and cloacal discharge,virus load in tissue and organ.It was demonstrated that IBV QX VLP had been successfully prepared and had good immunogenicity,thus laying the groundwork for the further development of IBV QX genetic engineering vaccines based on VLP technology.

关 键 词：IBV QX型 病毒样颗粒 重组杆状病毒 

分 类 号：S855.3[农业科学—临床兽医学]