云南省墨江县库蠓中一株西藏环状病毒的分离及全基因组序列分析  

Isolation and Genomic Characterization of A Strain of Tibet Orbivirus from Culicoides Species Collected in Mojiang County,Yunnan Province,China

作　　者：杨振兴[1] 孟锦昕 何于雯 李楠[1] 王静林[1] YANG Zhenxing;MENG Jinxin;HE Yuwen;LI Nan;WANG Jinglin(Yunnan Tropical and Subtropical Animal Virus Disease Laboratory,Yunnan Animal Science and Veterinary Institute,Kunming650224,China)

机构地区：[1]云南省畜牧兽医科学院云南省热带亚热带动物病毒病重点实验室,昆明650224

出　　处：《病毒学报》2022年第6期1419-1428,共10页Chinese Journal of Virology

基　　金：云南省科技厅重点基金(项目号:2019FA015),题目:云南省蓝舌病毒传播媒介库蠓及其唾液转录组的研究;云南省科技厅重点基金(项目号:202201AS070062),题目:蚊虫特异性Negwotan病毒对乙型脑炎病毒传播影响的研究;云南省程功专家工作站(项目号:202005AF150034);云南省科技人才和平台计划项目(项目号:2018HB046),题目:云南省中青年学术和技术带头人后备人才;中央引导地方科技发展资金项目(项目号:202207AB110006),题目:云南省虫媒病毒病防控技术科技创新平台建设;昆明医科大学科技创新团队建设项目(项目号:CXTD202111),题目:昆明医科大学艾滋病共感染传染性疾病诊疗科技创新团队。

摘　　要：西藏环状病毒(Tibet orbivirus,TIBOV)于2009年首次从中国西藏自治区采集的圆斑按蚊中分离出,目前在中国的云南省、广东省、湖南省和临国日本均有分离报道。2017年我们从云南省墨江县采集的库蠓样品中分离到一株病毒(MJC1-7),接种BHK-21和C6/36细胞后均可产生聚集、皱缩和脱落等明显细胞病变。1%的琼脂糖凝胶电泳显示,病毒基因组为十节段的双链RNA,呈现“3-3-3-1”的带型。通过病毒全长cDNA扩增和测序获取MJC1-7毒株的全基因组核苷酸序列,病毒基因组全长为19260 bp(包括编码区18495 bp和非编码区735 bp),由Seg-1(3950 bp)至Seg-10(832 bp)的10个基因节段构成,可编码6165个氨基酸残基。对环状病毒保守的VP1(Pol)、VP3(T2)和VP7(T13)蛋白氨基酸序列及系统发育分析显示,MJC1-7毒株与TIBOV亲缘关系最近,氨基酸序列相似度为95.4%~99.7%。对决定环状病毒血清型VP2(OC1)蛋白氨基酸序列与系统发育分析显示,MJC1-7与云南(SX-2017a、DH13C120和YN15-283-01)和广东(D181/2008)分离的TIBOV毒株聚为一簇,氨基酸序列相似度为97.3%~98.6%,而中国西藏毒株(XZ0906)和日本毒株(KSB-8/C/09和KSB-3/C/10)形成了另外3个独立的进化分支,MJC1-7与其相似度仅为27.8%~41.8%,以上结果表明MJC1-7毒株与云南和广东毒株有更近的亲缘关系,而中国西藏和日本毒株可能为Seg-2(OC1)基因变异较大毒株。本研究报道了一株TIBOV在我国云南省墨江县库蠓上的分离以及全基因组序列分析,研究结果进一步丰富了我们对TIBOV的认知,为深入研究我国TIBOV的分布、致病性与病原学检测技术提供了基础数据。Tibet Orbivirus(TIBOV)was first isolated from mosquitoes(Anopheles maculatus)collected in Motuo County(Tibet,China)in 2009.The TIBOV has been isolated in the Yunnan,Guangdong and Hunan Provinces of China and Japan.In 2017,a strain of the TIBOV(MJC1-7)was isolated from Culicoides collected in Mojiang City(Yunnan Province,China).After inoculation with baby hamster kidney(BHK-21)cells and Aedes albopictus(C6/36)cells,significant cytopathic effects such as aggregation,contraction and shedding were observed.Agarose-gel electrophoresis indicated a genome consisting of 10 segments of double-stranded RNA molecules that demonstrated a 3–3–3–1 pattern.A full-length complementary-DNA copy of the genome was amplified and sequenced.The size of the virus genome was 19,260 bp(coding region of 18,495 bp and non-coding region of 735 bp)and consisted of 10 genes from segment-1(3,950 bp)to segment-10(832bp),and encoding 6,165 amino-acid residues.Phylogenetic analyses of the viral RNA-dependent RNA polymerase(Pol),sub-core-shell(T2),and outer core(T13)proteins revealed that MJC1-7 clustered with the TIBOV,and that the amino-acid sequences of the MJC1-7 virus shared 95.4%-99.7%identity with other strains of the TIBOV.Phylogenetic analyses of VP2(OC1)protein(which determines the serotype of circoviruses)showed that MJC1-7 clustered with TIBOV strains isolated from Yunnan(SX-2017A,DH13C120 and YN15-283-01)and Guangdong(D181/2008),and that the amino-acid sequence of MJC1-7virus shared 97.3%-98.6%identity with them.However,the other TIBOV strains,Tibetan(XZ0906)and Japanese(KJB-8/C/09 and KJB-3/C/10),formed another three independent evolutionary branches in the phylogenetic tree of OC1,and OC1 of the MJC1-7 virus shared only 27.8%-41.8%identity with them.These results indicated that the MJC1-7 strain had a close genetic relationship with TIBOV strains from Yunnan and Guangdong,but other TIBOV strains from Tibet and Japan had large variations in Segment-2(OC1).We reported the isolation and genome sequence of a TIBOV strain from Culi

关 键 词：库蠓 病毒分离 西藏环状病毒 序列分析 

分 类 号：S852.65[农业科学—基础兽医学]