基于十八年跨度的乙型肝炎病毒基因型I分子进化研究  

A study based on the molecular evolution of genotype I hepatitis B virus over an 18-year span

作　　者：黄小倩 陈钦艳[2] 蒋智华[2] 贾蕙华 胡雪 胡莉萍[2] 张陆娟[2] 方钟燎[1,2] HUANG Xiaoqian;CHEN Qinyan;JIANG Zhihua;JIA Huihua;HU Xue;HU Liping;ZHANG Lujuan;FANG Zhongliao(School of Preclinical Medicine,Guangxi Medical University,Nanning,Guangxi 530021,China;Guangxi Zhuang Autonomous Region Center for Disease Prevention and Control,Guangxi Key Laboratory for the Prevention and Control of Viral Hepatitis,Nanning,Guangxi 530028,China;The Animal Husbandry Research Institute of Guangxi Zhuang Autonomous Region,Nanning,Guangxi 530001,China;Guangxi Vocational University Of Agriculture,Nanning,Guangxi 530007,China)

机构地区：[1]广西医科大学基础医学院,广西南宁530021 [2]广西壮族自治区疾病预防控制中心广西病毒性肝炎防制研究重点实验室,广西南宁530028 [3]广西壮族自治区畜牧研究所,广西南宁530001 [4]广西农业职业技术大学,广西南宁530007

出　　处：《应用预防医学》2024年第2期71-78,84,共9页Applied Preventive Medicine

基　　金：国家自然科学基金(81860595)。

摘　　要：目的 探索乙型肝炎病毒(hepatitis B virus,HBV)基因型I长期进化特征。方法 从广西隆安研究队列中选取已明确感染基因型I的HBsAg无症状携带者作为研究对象,检测其血清HBV标志物和病毒载量。使用酚氯仿法提取HBV DNA,采用巢式PCR扩增HBV全基因组,进行克隆测序,运用BioEdit、Mega等软件对测序结果进行生物信息学分析。结果 共纳入9名研究对象,获得279条全长基因序列。系统进化树分析结果显示,22.2%(2/9)的研究对象HBV基因型发生了转换,从基因型I1转换成基因型C1。2004年所有研究对象HBV毒株均发生X基因核心基因启动子(BCP)双突变(nt1762A→T/1764 G→A);88.9%(8/9)的研究对象HBV毒株发生PreC基因nt1896(G→A)终止密码突变。多数研究对象发生免疫逃避突变和耐药突变,共检出18种免疫逃逸突变和6种多重耐药突变。研究对象GD056、XW511、GA135、XX55、CZ647、TX119、WX058和YF201携带的毒株免疫逃逸突变率分别为:69.6%、7.5%、96.7%、94.4%、11.8%、4.7%、100.0%和100.0%;研究对象GA135、XW511、WX058、XX55、YF201、CZ647和TX119携带的毒株耐药突变率分别为:6.7%、2.5%、3.8%、50.0%、4.5%、2.9%和4.7%。66.7%、55.6%和44.4%的研究对象2004年和/或2022年携带的HBV毒株分别在PreS/S基因、X基因和PreC/C基因受到正向选择压力,所有研究对象携带的HBV毒株在P基因以负向选择压力占主导。结论 在自然感染进程中,HBV基因型I可以发生基因型转换,基因型I在一些常见的临床相关热点突变上突变率较高,部分毒株可出现免疫逃避突变和耐药突变,HBV基因组各个开放读码框所受的选择压力不相同。Objective To explore the long-term evolutionary characteristics of hepatitis B virus(HBV)genotype I.Methods Asymptomatic HBsAg carriers with confirmed genotype I infection were selected as study subjects from the Guangxi Long’an cohort.Their HBV serum markers and viral load were detected.HBV DNA was extracted by phenol/chloroform. The whole HBV genome was amplified using nested PCR for cloningsequencing. The data were analyzed by BioEdit, Mega, and other software to conduct bioinformatics analysis ofthe sequencing results. Results A total of 9 research subjects were included, and 279 full-length gene sequenceswere obtained. Phylogenetic tree analysis showed that 22.2% (2/9) study subjects had HBV genotype conversion,from genotype I1 to genotype C1. In 2004, all HBV strains studied had double mutations (nt1762 A→T/1764G→A) in the X gene core gene promoter (BCP). 88.9% (8/9) of the HBV strains studied had the nt1896 (G→A)stop codon mutation in the PreC gene. Most of the study subjects had immune escape mutations and drugresistance mutations, and a total of eighteen immune escape mutations and six multidrug resistance mutationswere detected. The immune escape mutation rates of strains in subject GD056, XW511, GA135, XX55, CZ647,TX119, WX058, and YF201 were 69.6%, 7.5%, 96.7%, 94.4%, 11.8%, 4.7%, 100.0%, and 100.0%, respectively.The drug-resistant mutation rates of strains in subjects GA135, XW511, WX058, XX55, YF201, CZ647, andTX119 strains were 6.7%, 2.5%, 3.8%, 50.0%, 4.5%, 2.9%, and 4.7%, respectively. 66.7%, 55.6%, and 44.4%of the 2004 and/or 2022 HBV strains were subjected to positive selection pressure on preS/S gene, X gene, andpreC/C genes, respectively;the HBV strains carried by all study subjects were dominated by negative selectionpressure on the P gene. Conclusions During the natural infection process, HBV genotype may undergo genotypeconversion. Genotype I has a higher mutation rate in some common clinically relevant hotspot mutations. Immuneescape mutations and drug resistance mutations may occ

关 键 词：乙型肝炎病毒(hepatitis B virus HBV) 基因型I 克隆测序 进化 

分 类 号：R373.21[医药卫生—病原生物学]