HBsAg ELISA-/NAT+的HBV血清学模式和病毒变异分析

Analysis of HBV serological pattern and virus variation in HBsAg ELISA-/NAT+

目的结合乙型肝炎病毒核酸(HBV DNA)的检测结果,分析血清标本血清学模拟及DNA序列,了解HBV DNA以及HBsAg血型模式之间的关系。方法选自该院54例血液标本作为本研究对象;对标本行HBV DNA提取、PCR扩增处理、PCR产物纯化、DNA测序、HBV基因分型和序列分析,并运用生物信息学软件对测序结果进行分析处理,对比分析基因突变情况。结果 PCR扩增结果,54例标本中40例表现为非常显著的PCR扩增不佳,14例标本PCR产物电泳均能够观察到1 400bp特异性条带;22例为HBsAg ELISA阳性,14例为隐匿性HBV阳性,PCR扩增阳性行基因型检测,B型基因在HBsAg ELISA中占81.82%,C型基因在隐匿性HBV阳性中比例为78.57%,差异具有统计学意义(P<0.05);14株隐匿性HBV基因中,6例在S区区域内发生基因序列点突变,其中2例在1个碱基点出现突变,另3例在2个位点的碱基点出现突变;3例HBV基因C型感染者出现基因点突变,2例B基因型感染者出现基因点突变;5例标本在9个位点部位的"a"决定族内碱基点出现突变,A-C的突变较多。结论血液筛查中,检测显示为HBsAg的阴性合格血液,仍可能有隐匿性乙肝感染和窗口期漏检,其病毒株主要为C型,且有变异株,B型病毒株相对较少,但突变株相对较高,可能逃避现有筛查试剂。

Objective To investigate the relationship between HBV DNA and HBsAg blood group model by analyzing the serological and DNA sequences of serum samples.Methods Blood samples from 54 patients were selected from this hospital,collecting blood samples;on the HBV DNA extraction,PCR amplification,purification of PCR products,DNA sequencing,HBV gene analysis and sequence,and the use of bioinformatics software.The results were analyzed and compared the gene mutation.Results The results of PCR amplification,54 specimens in 40 cases showed significant PCR was poor,14 specimens were able to observe the electrophoresis of PCR amplified 1 400 bp specific band;22cases were HBsAg ELISA positive,14 cases of occult HBV positive,PCR positive amplification for genotype,B genotype accounted for in 81.82% HBsAg ELISA,C gene in occult HBV positive accounted for 78.57%（P0.05）;occult HBV gene of 14 strains,6cases in the S area of the area occurred in the gene sequence of point mutations,including 2cases in 1base point mutations,the other 3cases of base point in 2sites the emergence of mutation;3cases of HBV genotype C infected gene point mutation,2cases of genotype B infected gene point mutation;5cases were in the 9site of the ＂a＂ family decided to base mutation,A-C mutation.Conclusion Blood screening,detection showed HBsAg negative blood qualified,may still have occult HBV infection and missed the window period,were mainly C type,B type and mutant strain is relatively less,but the mutant is relatively high,may escape the existing screening reagent.