蠓虫源盖塔病毒(SZC30)结构基因分子生物学特征分析

doi:10.16431/j.cnki.1671-7236.2021.04.001

摘要/Abstract

摘要： 为了解云南蠓虫源盖塔病毒（GETV） SZC30株分子特征及其与国内外其他媒介和宿主动物中分离病毒的遗传进化关系，本研究采用5对盖塔病毒特异引物对2013年首次在云南省蠓虫中分离的盖塔病毒SZC30株结构基因进行RT-PCR扩增，并对扩增产物进行测序；采用DNAStar软件中SeqMan进行序列拼接，获得SZC30株病毒结构基因序列长3 762 nt，编码衣壳蛋白（C）、E1、E2、E3和6K蛋白，序列长度分别为804、1 317、1 266、192和183 nt，编码蛋白长度分别为268、438、422、64和61个氨基酸。C、E1和E2基因系统进化分析显示，SZC30株与1955-2018年不同地域、宿主分离的27株盖塔病毒分离株形成Ⅰ、Ⅱ、Ⅲ和Ⅳ 4个进化分支；SZC30株与中国、韩国和日本蚊虫和动物分离株位于Ⅲ进化分支内，核苷酸同源性最高，在98.0%以上，氨基酸同源性在98.9%以上，亲缘关系较近；而与马来西亚、俄罗斯等蚊虫分离株位于不同进化分支，核苷酸同源性低于97.6%，亲缘关系较远；与马来西亚蚊虫分离株（GenBank登录号：AF339484）、中国海南和云南蚊虫分离株（GenBank登录号：EU015061和KY434327）在C、E1和E2蛋白存在31个氨基酸差异位点，而与日本蚊虫分离株（GenBank登录号：LC152056）、中国猪分离株（GenBank登录号：MG865966和MG865969）氨基酸位点无差异，且同源性为100%。SZC30株与27株盖塔病毒在E1、E2蛋白上存在2个潜在糖基化位点和3个跨膜区；T细胞抗原表位分析结果显示，SZC30株与分离自蚊、猪、狐、牛和马等的盖塔病毒分离株均存在表位差异，其中，E1、E2蛋白发现较多表位差异。以上结果提示，蠓虫源盖塔病毒与大多数蚊虫和动物分离毒株同源性高、遗传进化关系近，且氨基酸位点、糖基化位点、跨膜区结构等分子特征相似，提示蠓虫可能作为一种潜在的传播媒介参与了当地盖塔病毒的传播扩散。

关键词: 蠓虫; 盖塔病毒; 结构基因; 序列分析

Abstract: To investigate the molecular characteristics of a Getah virus (GETV) strain SZC30 isolated from midge in Yunnan province and its phylogenetic relationship with GETV isolated from other vectors and host animals at home and abroad,the structural genes sequencing of a GETV strain SZC30 isolated from midge in Yunnan province for the first time in 2013 were amplified by 5 pairs of specific primers of GETV RT-PCR reactions.The sequenced cDNA fragments were analyzed by DNAStar software (SeqMan) and spliced to a complete structural genes.The structural genes of SZC30 strain was 3 762 nt in length and encoded capsid protein (C),E1,E2,E3 and 6K proteins with the length of 804,1 317,1 266,192 and 183 nt,and encoding 268,438,422,64 and 61 amino acids,respectively.Phylogenetic analysis based on C,E1 and E2 genes showed that SZC30 strain and other 27 strains of GETV isolated from different regions and hosts from 1955 to 2018 gradually evolved into four distinct evolutionary populations,including groupsⅠ,Ⅱ,Ⅲ and Ⅳ.The SZC30 strain was clustered within group Ⅲ with other GETV isolates from mosquitoes or animals in China,Korea and Japan,with nucleotide homology of more than 98.0%,amino acid homology of more than 98.9%,which indicated more related relationship between those GETV.However,SZC30 strain and other GETV isolates from mosquitoes in Malaysia,Russia were from a different evolutionary branch,and the homology of nucleotides were lower than 97.6%,which indicated that SZC30 strain had a far relationship with those GETV isolates from mosquitoes.Compared with other GETV isolates from mosquitoes in Malaysia (GenBank accession No.:AF339484),Hainan and Yunnan (GenBank accession No.:EU015061 and KY434327),there were 31 amino acid difference sites on C,E1 and E2 proteins of GETV strain SZC30 isolated from midge,but there were no difference with other GETV isolates from mosquitoes in Japan (GenBank accession No.:LC152056) and pigs in China (GenBank accession No.:MG865966 and MG865969),and the homology of amino acids were 100%.There were 2 potential glycosylation sites and 3 transmembrane domains on both of E1 and E2 proteins between SZC30 strain and other 27 strains of GETV.The analysis of antigen epitopes of T cells predicted that there were epitope differences between SZC30 strain and GETV isolated from mosquitoes,pigs,foxes,cattle and horses,and more epitope differences were found in E1 and E2 proteins.These results suggested that GETV isolated from midge had high homology and close phylogenetic relationship with most GETV isolated from other vectors and host animals and had similar similar molecular characteristics such as amino acid difference sites,potential glycosylation sites and transmembrane domains,suggesting that midge might be involved in the spread of local GETV as a potential vector.

Key words: midge; Getah virus (GETV); structural genes; sequence analysis

中图分类号:

S852.65⁺9.6

李楠, 何于雯, 左媛媛, 孟锦昕, 徐天刚, 王静林. 蠓虫源盖塔病毒(SZC30)结构基因分子生物学特征分析[J]. 中国畜牧兽医, 2021, 48(4): 1149-1160.

LI Nan, HE Yuwen, ZUO Yuanyuan, MENG Jinxin, XU Tiangang, WANG Jinglin. Analysis of Molecular Biological Characteristics of Structural Genes of Getah Virus (SZC30) Isolated from Midge[J]. China Animal Husbandry and Veterinary Medicine, 2021, 48(4): 1149-1160.

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