一株牦牛源G6P[11]型牛轮状病毒的分离鉴定及部分基因的序列分析

doi:10.11843/j.issn.0366-6964.2019.03.014

摘要/Abstract

摘要：

本研究的目的是分离、鉴定牦牛源牛轮状病毒（bovine rotavirus，BRV）。将经RT-PCR检测BRV阳性的牦牛腹泻粪便样本接种MA-104细胞，进行病毒分离和鉴定，并对其VP4、VP6和VP7完整基因进行测序，分析其分子特征。结果显示：病毒盲传3代后出现细胞病变，至第7代后出现细胞病变的时间稳定，经蚀斑纯化后测得病毒滴度为10^8.39TCID₅₀·mL^-1。分离株经RT-PCR、间接免疫荧光和电镜观察证实为BRV，命名为HY-1株；扩增HY-1株VP4、VP6和VP7完整基因序列，分析表明HY-1株为G6P[11]I2型；系统发育树显示，HY-1株VP4、VP6和VP7节段分别与中国牛源DQ-75株和印度牛源M-1和RUBV319株遗传演化关系最近，可能为重配毒株。与国内的G6型和P[11]型BRV毒株相比，HY-1株的VP4和VP7蛋白重要氨基酸位点变化较大。成功分离得到一株牦牛源BRV，基因型为G6P[11]型，为我国首次报道。

Abstract:

The aim of this study was to isolate and identify bovine rotavirus (BRV) from the diarrhea stool samples of Yak. BRV-positive samples detected by RT-PCR was used to inoculate into MA-104 cells for virus isolation and identification, and the complete genes of VP4, VP6 and VP7 were sequenced to determine their molecular characteristics. The cytopathic effect occurred after three continuous blind passages, and the stable time that cytopathic effect occurred was found after seven passages. The virus titer was 10^8.39TCID₅₀·mL^-1 after plaque purification. The isolate, named HY-1 strain, was confirmed to be BRV by RT-PCR, indirect immunofluorescence and electron microscopy. The complete VP4, VP6 and VP7 genes of HY-1 strain were sequenced successfully, showing that HY-1 strain belong to G6P[11] I2 genotype. The phylogenetic analysis showed that the complete VP4, VP6 and VP7 segments of HY-1 strain were closely genetic related to Chinese bovine DQ-75 strain and Indian bovine M-1 and RUBV319 strains, respectively, which indicated that HY-1 strain may be a reassortant strain. Compared with the domestic G6 and P[11] BRV strains, VP4 and VP7 of HY-1 isolate had significant variation in the important amino acid regions. In this study, a BRV strain was successfully isolated from yak with G6P[11] genotype. To the best of our knowledge, this is the first report on the identification of G6P[11] BRV in China.

中图分类号:

S852.659.4

李然, 汤承, 罗雪, 岳华. 一株牦牛源G6P[11]型牛轮状病毒的分离鉴定及部分基因的序列分析[J]. 畜牧兽医学报, 2019, 50(3): 592-601.

LI Ran, TANG Cheng, LUO Xue, YUE Hua. Isolation and Gene Sequence Analysis of G6P[11] Bovine Rotavirus HY-1 Strain from Yak[J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2019, 50(3): 592-601.

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