新疆牛蓝舌病毒的分离和鉴定

Isolation and identification of bluetongue virus from cattle in Xinjiang

为监测新疆地区自然环境中蓝舌病流行情况，并分离蓝舌病毒（BTV），本研究在巴州尉犁县设置10头牛为哨兵动物。自2016年5月～11月期间每周采集哨兵动物的血清和抗凝血，各采集了280份。用竞争ELISA（C-ELISA）方法对所有血清样本进行BTV抗体检测，并将所有抗凝血样本接种于C6／36、BHK21和Vero细胞，盲传5代，有4份细胞出现病变，对这些病变细胞液进行BTV抗原捕获ELISA（Ac—ELISA）检测、BTV血清型群特异片段VP7基因PCR鉴定和免疫电镜观察。结果显示，所有细胞病变样本的BTV抗原检测均为阳性；经RT—PCR扩增，在1156bp处均获得目的基因片段；免疫电镜观察到蓝舌病毒粒子。上述结果表明，本研究在新疆首次成功分离到了牛源BTV。

To monitor the prevalence of bluetongue in the natural environment in Xinjiang and to isolate the blue-tongue virus（BTV）,in this study,we set ten cattle as sentinel animals in Yuli County, Bazhou. Sentinel animal serum and anticoagulant were collected weekly from May to November,2016,and 280 samples were collected respectively. All serum samples were subjected to BTV antibody detection by the competitive ELISA （c-ELISA） method. All anticoagulated blood samples were inoculated into C6/36, BHK21,and Vero cells. The cells were blindly transmitted for 5 generations and 4 cells developed lesions.The BTV antigen capture ELISA（Ac-ELISA）was performed on these cell fluids,and the BTV serogroup-specific fragment VP7 gene was identified by PCR,and the immunoelectron microscopy was performed. In result,all of the cytopathogenic samples were positive for BTV antigen detection. The target gene fragment of 1 156 bp was obtained by RT-PCR. Bluetongue virus particles were observed by the immunoelectron microscopy. In conclusion,this study successfully isolated cattle source BTV in Xinjiang for the first time.