贵州地区慢性乙型肝炎病毒感染者病毒基因亚型研究

A study of hepatitis B virus subtypes in patients chronically infected with genotype B or C of hepatitis Bvirus in Guizhou DING

目的调查贵州省B、C基因型慢性HBV感染者的病毒基因亚型。方法PCR扩增HBV P区长309 bp的基因片段,扩增产物分别经限制性内切酶NciⅠ、VspⅠ、BstEⅡ酶切,琼脂糖凝胶电泳,根据酶切图谱多态性,用限制性片段长度多态性分析（PCR-RFLP）检测HBV C基因亚型。直接测序确定B基因亚型、对178例用S基因限制性片段长度多态性鉴定为B、C基因型的不同临床类型慢性HBV感染者进行亚型分析。结果84例C基因型HBV感染者中,27例（32.14%）为C1亚型、56例（66.67%）为C2亚型,1例为C1、C2亚型混合感染。94例B基因型HBV感染者中,93例（98.94%）为Ba、1例为Bj亚型感染。从无症状乙型肝炎表面抗原携带者、CHB到肝硬化/肝癌,C1亚型在各组中的比例逐渐降低,分别为60.00%、30.65%和16.67%;而C2亚型在各对应组中的分布逐渐增高,分别为40.00%、67.74%和83.33%。结论贵州地区B、C基因型慢性HBV感染者中,以Ba、C2亚型为主。C1、C2亚型在疾病中的分布有一定差异。PCR-RFLP分析HBV C1、C2亚型,方法简便、特异性强,适合较大样本分析,可用于流行病学调查。

Objectives To investigate hepatitis B virus （HBV） subtypes in patients chronically infected with genotype B or C of hepatitis B virus in Guizhou and to study the relationship between the subtypes and the progression of their liver diseases. Method Using PCR, 309 bp gene fragments in the HBV p region were amplified. The products of PCR were digested by NciⅠ,VspⅠ,BstEⅡ and subjected to agarose gel electrophoresis. The subtypes of C 1 and C2 were detected by restriction fragment length polymorphism （RFLP）. B subtype was determined by direct sequencing of PCR product. One hundred seventy-eight patients with genotype B or C HBV infection in Guizhou, including 50 asymptomatic carriers （ASC）, 100 chronic hepatitis （CHB）, 14 liver cirrhosis （LC）, and 14 hepatocellular carcinoma （HCC） patients were examined. The relationship between HBV C subtypes and the progression of their liver diseases was studied by analyzing the HBeAg positivity, HBV DNA loads and ALT levels of the patients. Results Of 84 patients with HBV genotype C, 27 （32.14%） and 56 （66.67%） were subtype C1 and C2, respectively. In 94 genotype B, 93 （98.94%） were subtype Ba and only one was subtype Bj. Subtype C1 showed a trend of gradual decrease from ASC and CHB to LC/HCC groups. In contrast, subtype C2 showed a gradual increase （trend） in the same order. The HBeAg positivity was significantly lower in subtype C1 than that in subtype C2 （P 〈 0.05）. The ALT levels and HBV DNA loads were higher in patients with subtype C2 than those in subtype C1, however no statistical significance was found in these primes （t = 0.95, 0.79, P 〉 0.05）. Conclusion Subtype Ba is major and subtype C2 is more common in Guizhou. The distribution of subtype C1 and C2 are different in various stages of liver disease. The PCR-RFLP method is simple and accurate and can be used in a large-scale survey.