山东牛轮状病毒的分离鉴定及其卵黄抗体应用试验  

Isolation and Identification of Bovine Rotavirus from Calf in Shandong Province and Application of Yolk Antibody

作　　者：秦晓冰[1] 周建梅 张洪亮[1] 单虎[1] QIN Xiao-bing;ZHOU Jian-mei;ZHANG Hong-liang;SHAN Hu(Shandong Key Laboratory of Preventive Veterinary Medicine,Qingdao Agricultural University,Tsingtao 266109,China)

机构地区：[1]青岛农业大学山东省预防兽医学重点实验室,山东青岛266109

出　　处：《中国兽医杂志》2020年第11期6-10,I0002,共6页Chinese Journal of Veterinary Medicine

基　　金：山东省兽药诊断试剂工程技术研究发展计划;山东省高等学校优势学科人才团队培育计划。

摘　　要：为研究卵黄抗体对牛轮状病毒(BRV)的预防与治疗效果,本试验收集山东某牛场的腹泻粪便样本接种MA104细胞进行病毒分离、纯化,采用RT-PCR和电镜进行鉴定。将分离株增殖灭活制成油苗多次免疫产蛋鸡,收集高免鸡蛋,用水稀释-盐析法纯化卵黄抗体(IgY),用琼扩法测定抗体效价,进行安全性检测。选10头1日龄BRV阴性的犊牛,随机均分为2组,人工感染BRV,进行犊牛口服IgY疗效观察。结果显示:病毒盲传至第8代后出现稳定细胞病变,经蚀斑纯化后测得病毒滴度为10-5.62 TCID 50/mL;分离毒株经RT-PCR和电镜观察证实为BRV,命名为SD-LX株。SD-LX株灭活油苗免疫蛋鸡后得到滴度为1∶64的IgY,该抗体对犊牛感染BRV的保护率与对照组相比为100%,治愈率为40%。本试验成功分离到1株BRV,该毒株油苗免疫蛋鸡制成的IgY对犊牛BRV有良好的保护作用。In order to study the preventive and therapeutic effects of yolk antibodies on bovine rotavirus(BRV),diarrhea stool samples from a cattle farm in Shandong province were collected to inoculate MA104 cells for virus isolation and purification in this test.The isolated virus was identified by RT-PCR and electron microscope.The isolated strains were proliferated and inactivated to make oil-emulsion BRV vaccine for multiple immunization of laying hens.The antibody titer of collected eggs of hyper-immunity was tested by double agar diffusion.The hyper-immunized yolk antibodies(IgY)were purified by chloroform extraction and salt precipitation with ammonium sulfate and tested for safety.Ten 1-day-old BRV-negative calves were randomly divided into 2 groups and artificially infected with BRV to observe the effect of oral IgY on calves.The results showed that the stable time that cytopathogenic effect occurred was found after 8 continuous blind passages.The virus titer was 10-5.62 TCID 50/mL after plaque purification.The isolate,named SD-LX strain,was confirmed to be BRV by RT-PCR and electron microscopy.After laying hens were immunized by the inactivated oil-emulsion from SD-LX strain,IgY with a titer of 1∶64 was obtained.The antibody’s protective rate against BRV infection in calves was 100%compared with the control group,while the cure rate was 40%.A bovine rotavirus was successfully isolated in the trial.The antibody had obvious curative effect against BRV.

关 键 词：牛轮状病毒 分离鉴定 卵黄抗体 犊牛保护试验 

分 类 号：S857.3[农业科学—临床兽医学]