20例中国人HCVⅡ/1b型高变区1序列变异的动态观察

Dynamic study of genotype Ⅱ/1b hepatitis C virus hypervariable region 1 sequence diversity in 20 Chinese patients

目的 动态研究中国人群HCVⅡ /1b型包膜蛋白E2 /NS1高变区 1(HVR1)序列变异规律、意义及影响因素。方法 应用逆转录巢式PCR技术从 2 0例HCVⅡ /1b型感染的中国病人血清中扩增了HCV部分包膜区基因片段 (nt144 9～ 15 86 ,HCV J) ,纯化后直接采用双脱氧链末端终止法进行序列分析。结果 中国人群HCVHVR1位于氨基酸 (AA) 384～ 410位 ,有 6个较保守的AA位点 :385位Thr,389、390、40 6位Gly ,40 3位Phe ,40 9位Gln。自然病程组核苷酸 (nt)变异率每年 [0～ 2 0 7(平均0 5 0 ) ]× 10 1个 /位点 ,AA变异率每年 [0～ 6 2 2 (平均 1 2 4) ]× 10 1个 /位点 ,干扰素治疗组nt变异率为每年 [0～ 5 0 3(平均 2 31) ]× 10 1个 /位点 ,AA变异率每年 [0～ 10 7(平均 4 40 ) ]× 10 1个 /位点。干扰素治疗对HVR1区变异有较强的免疫选择作用 ,HVR1变异率高低与慢性肝炎的临床病程有一定联系 ,肝炎活动期变异明显。结论 对HCVHVR1区变异规律及其生物学意义的进一步研究将有助于了解HCV慢性感染机理、制定新的治疗方案及疫苗设计等。

Objective To study the sequence diversity of the hypervariable region 1(HVR1) in the putative envelope protein E2/NS1 of hepatitis C virus (HCV) in Chinese patients and its possible influencing factor. Methods The cDNAs (nt1449 1586, HCV J) derived from sequential plasma of 20 patients infected with genotypeⅡ/1b HCV were amplified, purified and directly sequenced by using RT nested polymerase chain reaction (PCR) and dideoxynucleotide chain termination method. Results The HVR1 was found in amino acid (aa) 384 410 positions of E2 protein, which had six highly conserved amino acids: Thr at position 385, Gly at position 389, 390 and 406, Phe at position 403 and Gln at position 409. The rate of nucleotide (nt) and amino acid sequence variation in natural course was 0 2.07 (mean 0.50) and 0 6.22 (1.24)×10 1 /genome site/year. In the interferon (IFN) treatment group, the rate of nucleotide and amino acid sequence variation was 0 5.05 (2.31) and 0 10.74 (4.40)×10 1 /genome site/year. The rate of sequence variation was also higher in patients with flare up of their ALT levels. Conclusion Our data on sequence diversity of HVR1 should be useful to the understanding of HCV persistent infection, development of new treatment strategy and vaccine design.