福州地区HBsAg-/HBVDNA+献血者乙肝病毒S区MHR变异特征分析

Analysis of Variation Characteristics of MHR in Region S of HBSAG-/HBVDNA+ among Blood Donors in Fuzhou

目的分析福州地区HBsAg-/HBV DNA+献血者HBV的基因型及主要亲水区(MHR),尤其是其中"a"决定簇区氨基酸的变异情况,为进一步研究隐匿性乙肝病毒感染的发生机制提供参考。方法对收集的2013年7月—2014年10月福州地区献血者的90 874名献血者血液标本进行HBsAg ELISA与单人份核酸检测,对筛查出的HBsAg-/HBV DNA+标本,进行HBV DNA的提取和S区基因的扩增和测序。测序结果进行进化分型,分析MHR中氨基酸序列的变异情况。选取15例前期研究所得的HBsAg+/HBV DNA+标本的测序结果作为对照,比较两类标本在"a"决定簇区蛋白序列变异上的差异。结果 16份测序结果中B基因型15份,C基因型1份。15例标本中13例在MHR内共发生了33次氨基酸的变异,其中25次集中在"a"决定簇区,HBsAg-/HBV DNA+标该组"a"决定簇区的变异率要高于HBsAg+/HBV DNA+标该组。变异的形式主要为单个的氨基酸的点替换,另有7例标本在124位与125位氨基酸间发生了9个碱基的非移码插入,造成该位点间"TNR"3个氨基酸的插入。结论福州地区献血者HBsAg-/HBV DNA+血液标本中HBV的基因型以B型为主,MHR的氨基酸变异主要集中在"a"决定簇区,有本地区的地域特点。研究中新发现的3个氨基酸"TNR"的插入变异,可能影响HBsAg的检出。

Objective This paper tries to analyze the HBV genotypes and amino acids variability in major hydrophilic region（MHR）, especially in ＂a＂ determinants region, among HBsAg-/HBV DNA＋ blood donors in Fuzhou and to provide the reference for the further study of occult hepatitis B virus infection mechanism. Methods Plasma samples of 90 874 cases of blood donors from July 2013 to October 2014 in Fuzhou were screened and tested by HBsAg ELISA and nucleic acid. HBV DNA was extracted and gene of S region was amplified for HBsAg-/HBV DNA＋ samples. Evolutionary typing was conducted for sequencing results, and variations of amino acid sequence in MHR were analyzed. 15 cases of HBsAg＋/HBV DNA in previous study were selected as the control group, the sequence variations of proteins in ＂a＂ determinants region was compared among sub-specimens. Results Among the 16 cases of sequencing results, 15 cases were genotype B and 1 case was genotype C. Among the 15 specimens, a total of 33 variations of amino acids occurred in MHR in 13 specimens, 25 of which concentrated in ＂a＂ determinants region. Variation rate in the ＂a＂ determinants region of specimens in the group of HBsAg-/HBV DNA ＋ specimens was higher than that in the group of HBsAg ＋/HBV DNA ＋ specimens. The main variation was the point substitution of a single amino acid. In addition, in another 7 specimens, non-frameshift insertions of 9 bases occurred between 124 and 125 amino acids,causing the insertion of three amino acids TNR at this site. Conclusion The genotypes of HBV in specimens of blood donors in Fuzhou were mainly type B, and the variation of MHR mainly concentrated in ＂a＂ determinants region, showing regional characteristics of this region. The insertion variation of newly discovered three amino acids TNR may affect the detection of HBsAg.