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福建省慢性HBV感染者HBV基因S区、基本核心启动子区和前C区基因变异特征分析

李东1 杨秀惠1,2 陈致飞1 张苏晗1 郑凝旋1 潘伟毅1,2 周勇1,2 郑奎城1,2

(1.福建省疾病预防控制中心福建省人兽共患病重点实验室, 福州 350001)
(2.福建医科大学公共卫生学院教学基地, 福州 350001)
【创新点】文章根据前期研究选取了HBV基因S区,基本核心启动子区(BCP)及前C区的序列变异作为讨论重点,先前的文章多是研究其中的一个基因功能区。本文共同讨论了三个功能区,并发现这些变异可能造成病毒抗原表达,疫苗逃逸,患者病症改变等,应加强发生这些变异患者的监测。

【摘要】为研究福建省慢性HBV感染者HBV基因多样性及变异规律,了解该人群HBV-DNA的病毒学特征。收集慢性HBV感染者血清标本,通过巢式PCR法扩增其HBV基因序列,比对NCBI数据库中标准基因型序列,分析HBV基因S区,基本核心启动子区(BCP)及前C区的序列变异情况,并对这些变异可能造成的病毒抗原表达,疫苗逃逸,患者病症改变等情况进行探讨。最终成功扩增82例HBV全长基因序列,其中B基因型56例,C基因型26例。基因组特定功能区序列分析发现慢性HBV感染者HBV基因在S区(23.2%)、BCP区(61.0%)和前C区(29.3%)均出现了不同程度的变异。其中主蛋白(HBsAg)主要抗原决定簇a决定簇45.8%位点出现了变异,这些变异位点中包括与肝炎重症化及免疫逃逸密切相关的位点(aa126、aa129、aa145等)。位点G1896A(19.5%),G1764A(11.0%)和A1762T(9.8%)依然是BCP/前C区的主要突变位点。而位点A1752G(25.6%)高突变率的出现在BCP区应引起关注。此外位点G1764A(χ2=5.742,P=0.030)、A1896G(χ2=14.392,P=0.000)以及A1762T/G1764A(χ2=7.289,P=0.012)的突变更容易发生在HBeAg阴性的样品中;而位点A1846T(χ2=11.882,P=0.003)、A1762T(χ2=6.561,P=0.038)和A1896G(χ2=6.958,P=0.030)的突变与HBV-DNA的病毒载量存在一定相关性。总之,福建省慢性HBV感染者在HBV不同基因功能区域均存在不同程度的变异,一些与HBeAg表达情况、HBV-DNA载量、疫苗免疫逃避及肝细胞癌发生具有相关联的变异位点已经出现,BCP区A1752G位点的高频率出现应值得关注,对于这些变异位点的患者应加强监测。

【关键词】 乙型肝炎病毒(HBV);基因变异;基本核心启动子(BCP);前C区;

【DOI】

【基金资助】 国家科技重大专项(项目号:2012ZX10002001-002-002),题目:艾滋病和病毒性肝炎等重大传染病防治; 国家科技重大专项(项目号:2017ZX10103008-004),题目:浙江及周边省传染病病原谱流行规律研究——五大症候群病原谱监测;

Genetic Mutations of the Hepatitis B Virus Genome S and BCP/Pre-C Regions of the Hepatitis B Virus (HBV) Genome in Chronic HBV Infection in Fujian Province of China

LI Dong 1 YANG Xiuhui 1,2 CHEN Zhifei 1 ZHANG Suhan 1 ZHENG Ningxuan 1 PAN Weiyi 1,2 ZHOU Yong 1,2 ZHENG Kuicheng 1,2

(1.Key Laboratory of Zoonosis Research of Fujian Province, Fujian Center for Disease Control & Prevention , Fuzhou, China 350001)
(2.Teaching Base of Public Health School of Fujian Medical University , Fuzhou, China 350001)
【Novelty】Based on the previous study selected the sequence of HBV gene S area, basic core promoter (BCP) and the Pre- C mutation as a key discussion,More than previous articles is one of the gene function research.This paper discuss three functional areas, and found that these mutations may cause virus antigen expression, vaccine escape, patients condition change, etc., so should strengthen the monitoring in patients with these mutations.

【Abstract】To investigate the molecular characteristics of the hepatitis B virus (HBV) genome in chronic HBV-infected patients in Fujian Province of China, serum samples were collected from patients with chronic HBV infection. HBV DNA was extracted for amplification of surface genes, basic core promoter (BCP) genes and precore (Pre-C) genes amplification. Polymerase chain reaction (PCR) products were sequenced and compared with standard sequences in GenBank. Sequencher, MEGA5 and Bioedit were used for mutation analyses of the HBV genome. We obtained 82 complete sequences of HBV. In all sequences, 56 were B genotype and 26 were C genotype. Gene sequencing revealed that these HBV gene sequences in S (23.2%), BCP (61.0%) and Pre-C (29.3%) regions had a different degree of variation. The frequency of a determinant amino acid (aa) variant was 45. 8%. Those mutation sites might be correlated with severe liver diseases and immune escape, such as the sites of aa126, aa129 and aa145. Analyses revealed that site G1896A (19.5%), G1764A (11.0%) and A1762T (9.8%) remained the major mutations in the BCP/Pre-C region. Simultaneously, the high mutation rate of A1752G (25.6%) in BCP merited attention. G1764A (χ2 = 6.498, P = 0.013), A1896G (10.444, 0.001) and A1762T/G1764A) mutations were more likely to occur in HBeAg–negative samples. A1846T (χ2 = 11.882, P = 0.003), A1762T (6.561, 0.038) and A1896G (6.958, 0.030) mutations had some relevances with HBV DNA load. In conclusion, people with chronic HBV infection in Fujian Province of China had different degrees of variation in HBV gene-function areas. Some of these mutations were associated with HBeAg expression, HBV DNA load, immune evasion of vaccines, and occurrence of hepatocellular carcinoma. Patients with these mutations need strengthened monitoring.

【Keywords】 Hepatitis B virus (HBV) ; Genetic mutations; Basic core promoter (BCP) ; Pre-C region;

【DOI】

【Funds】 National Science and Technology Major Project of China (No. 2012ZX10002001-002-002) ; National Science and Technology Major Project of China (No. 2017ZX10103008-004) ;

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This Article

ISSN:1000-8721

CN: 11-1865/R

Vol 34, No. 04, Pages 491-497

June 2018

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Abstract

  • Materials and methods
  • Results
  • Discussion
  • References