牛肠道病毒2型的分离与鉴定  

Isolation and Identification of Bovine Enterovirus Type 2

作　　者：彭小薇[1] 董浩[1] 吴丹[2] 张鹤晓[2] 吴清民[1] 吕艳丽[1] 

机构地区：[1]中国农业大学动物医学院,农业部动物流行病和人畜共患病重点实验室,北京100193 [2]北京出入境检验检疫局,北京100026

出　　处：《畜牧兽医学报》2013年第5期823-828,共6页ACTA VETERINARIA ET ZOOTECHNICA SINICA

摘　　要：本研究从北京某牛场发生严重腹泻的泌乳奶牛采集直肠拭子,接种MDBK细胞,分离获得1株病毒,命名为BJ001。理化特性鉴定结果表明,该病毒大小约30nm,为无囊膜的RNA病毒;用随机PCR方法对氯化铯梯度离心后的各区带扩增,扩增产物凝胶电泳后,回收"呈彗尾样拖带现象"最明显的区带2的500～1 500bp的片段进行序列测定与分析,结果显示,扩增片段与GenBank中已知的牛肠道病毒2型相似性较高;用BEV2型特异性引物对病毒进行RT-PCR扩增,并将扩增产物进行序列测定与分析,结果表明,其与BEV2相似性最高可达93%。故该病毒被确定为牛肠道病毒2型。将该病毒接种2头成年健康奶牛,均未出现肉眼可见的临床症状,但病毒可在接种奶牛体内持续存在,并产生较高滴度抗体。In this study, an unknown virus was isolated from rectal swabs of cattle with severe diarrhea and hemorrhagic dysentery in Beijing. The unknown virus was identified as non-enveloped particles about 30 nm in diameter and containing RNA genome by physicochemical characteristics analysis. All of four layers which were purified by cesium chloride density gradient centrifugation were amplified with random polymerase chain reaction （PCR）, and the products from layer 2 ranged mainly from 500 to 1 500 bp, showing comet tail-shaped entrainment phenomena, were purified and sequenced. The sequence blast results showed that the virus shared high gene homology with bovine enterovirus type 2. Therefore, specific primers of bovine enterovirus type 2 was used further to identify the virus, and the unknown virus was finally verified exactly belonging to BEV2 （93% nucleotide identity）. Two healthy adult cattle were inoculated with the virus as experiment animals, but no clinical symptoms appeared, although antigen and serum antibody were positive after challenge.

关 键 词：牛肠道病毒2型 分离与鉴定 理化特性 随机PCR 

分 类 号：S852.659.6[农业科学—基础兽医学]