牦牛源牛冠状病毒部分基因的扩增、序列分析及病毒分离鉴定

doi:10.11843/j.issn.0366-6964.2019.02.012

摘要/Abstract

摘要：

本试验的目的是对青藏高原地区牦牛进行牛冠状病毒（bovine coronavirus，BCoV）的分子流行病学调查，并分离牦牛源BCoV。采用RT-PCR方法检测西藏、青海、四川、云南的犊牦牛腹泻粪便中BCoV，并扩增其S、HE及N基因片段；选取阳性样本进行BCoV分离。结果显示：从336份犊牦牛腹泻粪便中检出232份BCoV阳性，检出率为69.05%。序列分析和系统发育分析显示，本试验克隆的32个BCoV阳性样本中的S1亚基序列、HE基因片段和N基因片段均有独特的遗传进化趋势；首次成功分离到1株牦牛源BCoV，蚀斑纯化后病毒TCID₅₀为10^-7.17·0.1 mL^-1，鉴定结果显示该毒株S基因发生了重组事件。本试验结果表明青藏高原牦牛源BCoV感染率很高，且有独特的遗传进化趋势。

Abstract:

The aim of this study was to investigate molecular prevalence of bovine coronavirus (BCoV) in yak in Qinghai-Tibetan Plateau and to isolate BCoV from clinical samples. RT-PCR assay was used to detect BCoV from 336 diarrhea feces samples of yak from Tibet, Qinghai, Sichuan and Yunnan, and the S, HE, and N gene fragments were further amplified, moreover, positive samples were used to isolate BCoV strain. Results were as follows:Approximate 69.05% (232/336) diarrheic samples were detected BCoV positive by RT-PCR. Sequence and phylogenetic analysis showed that BCoV S1 subunit sequence, HE gene fragment and N gene fragment from 32 clinical samples in this study had unique evolutionary features. Notably, a BCoV strain of yak was successfully isolated firstly, the TCID₅₀ of this strain was 10^-7.17·0.1 mL^-1,and a recombinant event was predicted in S gene of this strain. These results indicate that BCoV widely circulates in yak in the Qinghai-Tibetan Plateau of China and exhibit unique evolution.

中图分类号:

S852.659.6

何琪富, 汤承, 郭紫晶, 谭烁, 张斌, 岳华. 牦牛源牛冠状病毒部分基因的扩增、序列分析及病毒分离鉴定[J]. 畜牧兽医学报, 2019, 50(2): 343-353.

HE Qifu, TANG Cheng, GUO Zijing, TAN Shuo, ZHANG Bin, YUE Hua. Amplification, Sequence Analysis of Bovine Coronavirus Genes and Isolation of the Viruses from Yak[J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2019, 50(2): 343-353.

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