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目的研究慢性乙型肝炎患者在核苷(酸)类药物(NAs)规范化治疗前耐药突变情况。方法在我国北方地区4家医院收集148例慢性乙型肝炎患者在NAs规范化治疗前的血清,提取血清中HBV DNA,用巢式聚合酶链反应-直接测序法获得HBV全长RT区序列,用生物信息学技术筛查该区内11个经典耐药突变位点(rt169、rt181、rt184、rt194、rt202、rt204、rt236、rt250、rt80、rt173和rt180)并鉴定患者HBV基因型。采用SPSS 13.0软件进行统计分析。计量资料采用x±s表示,计量资料的比较采用t检验,计数资料的比较采用χ2检验。P<0.05为差异有统计学意义。结果 148例慢性乙型肝炎患者中,有142例血清HBV RT区在11个经典耐药突变位点上均为野生型氨基酸;6例患者检出经典耐药突变,分别为rtM204I2例,rtA181V、rtA181T、rtL180M+rtM204I、rtL80V+rtL180M+rtM204I各1例,其在半年以前均有不规范NAs治疗史。148例患者基因型分布为:B基因型占4.1%(6/148),C基因型占95.3%(141/148),D基因型占0.6%(1/148)。结论依据PCR产物直接测序结果,本研究中慢性乙型肝炎患者抗病毒治疗基线时体内HBV以野生株为优势株,检出NAs耐药突变株的患者均有既往不规律抗病毒治疗史。基线耐药检测有助于NAs个体化治疗方案的优化。
Objective To study the mutations of drug resistance in patients with chronic hepatitis B before the standardized treatment of nucleoside (acid) drugs (NAs). Methods Serum samples of 148 patients with chronic hepatitis B before standardized treatment of NAs were collected from 4 hospitals in the northern part of China. HBV DNA in serum was extracted, and full length RT region of HBV was obtained by nested polymerase chain reaction - direct sequencing Bioinformatics techniques were used to screen 11 classical resistance mutations (rt169, rt181, rt184, rt194, rt202, rt204, rt236, rt250, rt80, rt173 and rt180) in this region and identify HBV genotypes. SPSS 13.0 software for statistical analysis. Measurement data using x ± s said measurement data comparison using t test, count data comparison using χ2 test. P <0.05 for the difference was statistically significant. Results Of the 148 patients with chronic hepatitis B, 142 cases of serum HBV RT region were wild-type amino acids at 11 classical drug-resistant mutation sites. Six patients were detected with classical drug resistance mutations, including rtM204I2 cases, rtA181V, rtA181T, rtL180M + rtM204I, rtL80V + rtL180M + rtM204I in 1 case, which had non-standard NAs treatment history of six months ago. The genotypes of 148 patients were 4.1% (6/148) in B genotype, 95.3% (141/148) in C genotype and 0.6% (1/148) in D genotype. Conclusions According to the direct sequencing results of PCR products, in this study, in the study of patients with chronic hepatitis B, the dominant strain of HBV was wild-type strain at the baseline of anti-virus treatment. The patients with detected NAs-resistant mutant strains all had the history of previous irregular antiviral therapy. Baseline drug resistance testing can be used to optimize NAs individualized regimens.