乙型肝炎病毒P基因变异的检测及其意义

目的采用错配PCR扩增方法进行乙型肝炎病毒(HBV)P基因YMDD变异的检测,并对其临床意义进行初步研究.方法应用分子克隆方法,构建YMDD、YVDD、YIDD变异的阳性质粒,并进行敏感性及特异性试验.对44例经拉米夫定治疗1年、HBVDNA持续阳性的乙型肝炎患者检测YMDD变异情况.结果经PCR方法、酶切鉴定及直接测序鉴定出YMDD、YVDD和YIDD质粒构建成功,且该方法具有较好的敏感性和特异性.44例拉米夫定治疗1年、HBVDNA持续阳性的乙型肝炎患者存在YMDD变异,半年出现变异5例,变异率为11.36%;1年出现变异7例,变异率为15.91%.结论本检测方法简便、实用,抗病毒药物压力是导致HBVYMDD变异的重要因素.

Detection of YMDD mutations in HBV polymerase gene and study of clinical practice

Objective To assess the clinical significance of YMDD mutations.Methods Plasmids of YMDD, YVDD and YIDD were constructed by molecular clones and its sensitivity and specificity were tested.In 44 patients with HBV infection whose HBV DNA were persistently positive after oral lamivudine therapy for 1 year,YMDD mutations were analyzed by mismatch amplify mutation assay.Results PCR,restriction endonuclease method and direct sequence analysis demonstrated that plasmids of YMDD,YVDD and YIDD were constructed successfully.YMDD mutations occurred in all of above patients.There were 5 patients of YMDD mutations (11.36%) and 7 patients of YMDD mutations (15.91%) after therapy for half a year and one year respectively.Conclusion The mismatch amplify mutation assay is simple and practical in detecting YMDD mutations of P gene of HBV.Pressure of antiviral drug is very important for YMDD mutations.