楚会萌　任亚初　程凯慧　蒋仁新　张云飞　解晓莉　张亮　杨宏军

摘要：牛流行热病毒（ BFFV）是引发世界范围内牛流行热最主要病原之一，给奶牛业的发展造成严重危害。本研究通过RT-PCR对疑似牛流行热奶牛抗凝血样品进行检测，选取阳性样品颅内接种3日龄乳鼠，7日后乳鼠死亡，鼠脑组织通过PCR扩增出BFFV目的条带，乳鼠颅内重复接种3次，乳鼠发病死亡时间明显提前。选取阳性脑组织接种BHK-21细胞，盲传3代后出现明显细胞病变，特异性引物扩增获得目的条带，测序结果与牛流行热病毒序列一致，表明成功分离到一株牛流行热病毒。

关键词：牛流行热病毒;分离与鉴定;BHK - 21细胞;细胞病变

Isolation and Identification of a Bovine Ephemeral Fever VirusChu Huimeng， Ren Yachu， Cheng Kaihui， Jiang Renxin，Zhang Yunfei， Xie Xiaoli， Zhang Liang， Yang Hongjun

Abstract Bovine ephemeral fever virus is one of the most pathogens leading to the bovine ephemeral fe-ver worldwide， which has caused serious harm to the development of dairy industry. In this study， the antico-agulant sample of suspected bovine ephemeral fever from Henan were tested by RT - PCR， then positive sam-ples was inoculated intracerebrally into sucking mice with 3 days of age. After 7 days， the suckling mice died.The target band of BFFV was identified by PCR from rat brain. After repeated intracranial inoculation for 3times， the death time of sucking mice were significantly advanced. Followed by inoculation of positive samplesinto BHK -21 monolayer cells， the CPE was obvious after three blind inoculation. The target band were ob-tained with the specific primers， and the sequencing was consistent with the bovine ephemeral fever sequence.The results showed that a bovine ephemeral fever virus was successfully isolated.

Keywords Bovine ephemeral fever virus; Isolation and identification; BHK -21 cell; Cytopathy

牛流行热（ bovine ephemeral fever，BEF）又名牛暂时热、牛三日热，是由牛流行热病毒（Bovineephemeral fever virus，BEFV）引起的一種牛急性热性传染病[1]。BEFV属于弹状病毒科（Rhabdoviri-dae）暂时热病毒属（Ephemerovims），为单股负链RNA病毒，有囊膜，目前只有一种血清型[2]。1867年，该病在东非地区首次发现，随后在非洲、亚洲和澳大利亚等多个国家和地区流行[3]。1934年，我国江苏省首次报道牛流行热的爆发，1955年以后，大部分省份都有牛流行热发病的报道[4]。牛流行热的临床典型症状为发热，感染牛体温可以升至40℃以上，其他临床表现包括气短、心率快、眼溢、结膜充血、跛行甚至卧地不起等[5]。期间乳产量明显降低，部分怀孕母牛流产，给养牛业造成重大经济损失。2014年农业部印发的《牛羊常见疫病防控技术指导意见》将牛流行热列为重点防控疫病[6]。本实验室从牛场采集疑似患牛流行热的奶牛抗凝血[7]，进行病原分离鉴定，最终证明实验室本次分离到的毒株为牛流行热病毒。……