牛环曲病毒云南株纤突蛋白基因分子特征及进化分析

doi:10.16431/j.cnki.1671-7236.2024.11.029

摘要/Abstract

摘要： 【目的】了解牛环曲病毒(Bovine torovirus，BToV)在云南部分地区牛群中的流行及其S基因遗传进化情况。【方法】采集云南省内13个地区15个牛场不同年龄牛的粪便样品655份，利用RT-PCR方法检测BToV，并对其中6株BToV阳性样品进行S基因扩增、克隆、测序及遗传进化分析。【结果】 RT-PCR检测结果显示，655份牛粪便样品检出71份BToV阳性，总阳性率为10.8%；其中腹泻样品阳性率为21.2%(59/278)，非腹泻样品阳性率为3.2%(12/377)；1周龄、1周龄～1月龄、1～6月龄和1岁及上样品的阳性率分别为15.0%(3/20)、17.7%(20/113)、14.0%(45/321)和1.5%(3/201)。相似性分析结果显示，6条BToV S基因序列相似性为96.0%～99.8%，且与原始毒株Breda1的相似性为94.7%～96.0%，与国内流行毒株的相似性为95.2%～99.7%。遗传进化分析结果显示，6条S基因均与原始毒株Breda1不在同一大分支，与国内四川、河南毒株及土耳其毒株形成一个大分支，表明云南省BToV毒株与目前国内流行毒株一致，亲缘关系近。氨基酸突变位点分析结果显示，S基因存在少量独特的氨基酸突变位点。重组分析结果显示，BTOV-China/YN03-2021、BTOV-China/YN01-2021和BTOV-China/YN06-2022为重组序列,其中重组序列BTOV-China/YN06-2022得分最高，重组区域为1 438～1 609 bp区间，主要亲本为BTOV-China/YN05-2021；重组序列BTOV-China/YN03-2021，重组区域位于1 668～3 569 bp区域，主要亲本为土耳其毒株BToV-HT2-TUR(MG957146.1)；重组序列BTOV-China/YN01-2021，重组区域位于261～2 552 bp，主要亲本为土耳其毒株BToV-HT2-TUR(MG957146.1)。基因选择压力分析结果显示，BToV S基因的进化方式主要以中性选择为主，但同时也存在净化选择和正向选择的压力影响。【结论】本研究揭示了BToV已存在于云南省部分地区牛群体中且防控形势较严峻，可为中国研究BToV流行病学和BToV S基因遗传进化关系提供参考，同时也为云南地区制定BToV感染的防控措施提供理论依据。

关键词: 牛环曲病毒(BToV); S基因; 分子特征; 进化分析

Abstract: 【Objective】 This study was aimed to investigate the prevalence of Bovine torovirus (BToV) and the genetic evolution of the S gene in cattle herds in some areas of Yunnan province. 【Method】 655 faecal samples were collected from cattle of different ages from 15 cattle farms in 13 regions of Yunnan province,and BToV was detected by RT-PCR.6 representative positive samples were subjected to the S gene amplification,cloning,sequencing and genetic evolution analysis. 【Result】 RT-PCR results showed that there were 71 BToV positive samples were detected for the 655 samples with a total detection rate of 10.8%.The detection rate of diarrhea and non-diarrhea samples were 21.2% (59/278) and 3.2% (12/377),respectively.The detection rate of the samples from cattle at 1 week old,1 week to 1 month old,1 to 6 months old,and over 1 year old were 15.0% (3/20),17.7% (20/113),14.0% (45/321)and 1.5% (3/201) respectively.The results of similarity analysis showed that the six S gene sequences with a nucleotide similarity from 96.0% to 99.8%,and nucleotide similarity with the original strain Breda1 was from 94.7% to 96.0%.The similarity with domestic epidemic strains was 95.2% to 99.7%. Genetic evolutionary analysis showed that the six S gene were not in the same branch with the original strain Breda1,and formed a branch with the domestic strains in Sichuan and Henan as well as the Turkish strains,indicating that the BToV strains prevalent in Yunnan province were consistent with those of the currently prevalent strains in China,and that they were closely related to the domestic strains.Amino acid mutation site analysis showed that there were a small number of unique amino acid mutation sites of S gene in Yunnan province.The results of recombination analysis showed that BTOV-China/YN03-2021,BTOV-China/YN01-2021 and BTOV-China/YN06-2022 were predicted to be recombinant sequences,in which BTOV-China/YN06-2022 had the highest recombination score,with recombination region was located in 1 438 to 1 609 bp,and the main parent was strain BTOV-China/YN05-2021.The recombination region of BTOV-China/YN03-2021 was located in the 1 668 to 3 569 bp region,and the main parent was the Turkish strain BToV-HT2-TUR.The recombination region of BTOV-China/YN01-2021 was located in 261 to 2 552 bp,and the main parent was the Turkish strain BToV-HT2-TUR.The results of gene selection pressure analysis showed that the evolutionary mode of S gene was mainly dominated by neutral selection,there were the influence of purifying selection and positive selection pressure,too. 【Conclusion】 This study revealed that BToV had existed in Yunnan province cattle populations and the situation of prevention and control was relatively serious,which provided a reference for the study of epidemiology for BToV and the genetic evolutionary relationship of the spike protein gene in China,as well as a theoretical basis for the development of preventive and control measures for BToV infection in Yunnan province.

Key words: Bovine torovirus (BToV); S gene; molecular characterization; evolutionary analysis

中图分类号:

S852.65⁺3

耿娟, 曲伟杰, 张晓梅, 魏志杰, 钟德凯, 王鑫文, 李文贵, 张双玲, 李金存, 张振兴, 张以芳, 郑锦玲, 宋建领. 牛环曲病毒云南株纤突蛋白基因分子特征及进化分析[J]. 中国畜牧兽医, 2024, 51(11): 4943-4955.

GENG Juan, QU Weijie, ZHANG Xiaomei, WEI Zhijie, ZHONG Dekai, WANG Xinwen, LI Wengui, ZHANG Shuangling, LI Jincun, ZHANG Zhenxing, ZHANG Yifang, ZHENG Jinling, SONG Jianling. Molecular Characteristics and Evolutionary Analysis of the Spike Protein Gene of Bovine Torovirus Strain from Yunnan[J]. China Animal Husbandry and Veterinary Medicine, 2024, 51(11): 4943-4955.

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