C基因型牛副流感3型病毒的分离鉴定  

Isolation and Identification of BPIV3 Genotype C

作　　者：冉旭华[1] 张峣[1] 曹思[1] 卢元赫 张玲玲[1] 刘阳阳[1] 倪宏波[1] 朱战波[1] 闻晓波[1] 

机构地区：[1]黑龙江八一农垦大学预防兽医学省重点实验室,大庆163319

出　　处：《中国畜牧兽医》2016年第5期1368-1373,共6页China Animal Husbandry & Veterinary Medicine

基　　金：黑龙江八一农垦大学研究生创新科研项目(YJSCX2015-Y19);黑龙江省大学生创新创业训练计划项目(201510223020);"十二五"农村领域国家科技计划课题(2012BAD12B03-3);黑龙江省农垦总局"十二五"重点科技计划项目(HNK125B-11-08A;HNK125B-11-02)

摘　　要：为了对宁夏地区患有呼吸系统疾病的舍饲牛进行病原鉴定,试验主要利用RT-PCR方法对样品进行牛副流感3型病毒(BPIV3)M基因序列扩增,将扩增产物连接在pMD18-T载体后转化到大肠杆菌DH5α感受态细胞中进行亚克隆。通过氨苄青霉素平板筛选,将鉴定为阳性的克隆菌进行核苷酸序列测定并利用分子生物学软件与GenBank上参考序列进行同源性比对。测序结果表明,从样品中分离到了1株BPIV3,并命名为NX49,其M基因全长为1 056bp;进化分析表明,NX49隶属于BPIV3C基因型,其M基因与中国山东C型分离株SD0835具有较高同源性,核苷酸同源性为99.4%;理化分析表明,该毒株对温度、酸及有机物均敏感,高温孵育下Mg2+对该病毒无保护力;血凝试验表明,NX49对豚鼠红细胞凝集效价仅为1∶4,且仅在4℃孵育时出现凝集反应。本研究成功分离得到一株BPIV3C型毒株,这将有助于中国BPIV3分子进化规律及病毒流行特点的进一步研究。To identify the infection agents from Ningxia Hui Autonomous region,where feedlot cattle indicated bovine respiratory disease complex （BRDC）,the M gene of the bovine parainfluenza virus type 3 was amplified by RT-PCR.The PCR product was ligated to pMD18-T vector and cloned to E.coli DH5α.The positive clones were sequenced and compared with the reference strains in GenBank by the molecular biology software.Sequence alignment results showed that a BPIV3 strain was isolated from the samples and named NX49,the M gene of NX49 included 1 056 nucleotides.Evolutionary analysis showed that the NX49 belonged to BPIV3 C genotype and shared 99.4％ nucleotide identity with that of the SD0835 isolated in Shandong province.The characterization of the NX49 demonstrated that it was sensitive to temperature,acid and organic matter.The presence of Mg2＋ showed no protection against the treatment at high temperature.The HA test suggested that the NX49 enables to agglutinate the guinea pig RBC at 4 ℃ and the titer was 1 ∶ 4.The study isolated a BPIV3 genotype C strain successfully,which facilitate the study of molecular evolution and epidemiology of BPIV3 in China.

关 键 词：牛副流感3型病毒 C基因型 分离 鉴定 同源性比对 

分 类 号：S852.65[农业科学—基础兽医学]