陕西榆林某羊场羊口疮病毒的分离鉴定

摘要

In order to obtain a wild strain of Orf virus in Yulin,samples of the lip scab from a lamb with clinical symptoms of Orf in a goat farm in Yulin city were collected.After grinded and repeated freezing and thawing,the supernatant was inoculated in calf testis primary cells by passage culture,and then the virus was identified as Orf virus through the observation of cytopathic effects,PCR detection and animal regres-sion test.The results suggested that calf testis primary cells infected with scab by grinding fluid displayed cytopathic effects at the fourth passages,and the virus titer was 107.66 TCID50/mL.B2L gene of isolated vi-rus was cloned and sequenced,and the homologous analysis revealed an identity of 99% with Gansu (KC485343.1)strain and Fujian (KC568399.1)strain.The 2 month-old lambs were inoculated with the cell cultures for the infection experiment,and typical Orf clinic symptoms such as papules and pustules ap-peared in lips and inguen after virus inoculation.In conclusion,a wild strain of Orf virus in Yulin was isola-ted in this study.%为获得陕西榆林地区羊口疮病毒野毒株，以榆林地区某羊场疑似羊口疮（Orf）的山羊口唇部结痂为材料，将结痂研磨悬液接种于犊牛睾丸原代细胞进行传代培养，通过特征性细胞病变（CPE）观察、PCR检测、动物回归试验等对分离的毒株进行鉴定。结果表明，经研磨的病料悬液接种于犊牛睾丸原代细胞后，盲传至第4代时产生细胞病变，测得病毒滴度为107．66 TCID50／mL。对分离毒株的 B2L 基因克隆测序、同源性对比分析结果显示，该毒株与与国内报道的多株羊口疮毒株核苷酸序列均具有较高的同源性，其中与甘肃株（KC485343．1）福建株（KC568399．1）的同源性均达到99％。用纯化病毒液划痕接种2月龄羔羊后，在其唇部和腹股沟内侧出现丘疹和脓疱等 ORFV 感染的典型的羊口疮症状。表明成功获得了羊口疮病毒陕西榆林株。