戊型肝炎病毒3种核酸检测方法的比较

Comparison of three nucleic acid detection methods for hepatitis E virus

目的比较戊型肝炎病毒(hepatitis E virus,HEV)3种核酸检测方法的敏感性及特异性。方法将HEV基因4型代表株的开放阅读框(open reading frame,ORF)2区部分基因序列克隆至pUC57载体,合成质粒DNA,使用A、B两种实时定量方法检测该质粒,比较最低检出限;使用其他标本做特异性检测。使用3种方法平行检测急性戊肝病例血清标本,比较检测结果。结果A、B方法对质粒DNA的最低检出限均可达35拷贝数/反应,特异度均为100%;A、B和C方法对急性戊肝病例血清标本HEV RNA检出阳性率分别为47.8%(43/90)、43.3%(39/90)和41.1%(37/90),一致率为88.9%(80/90),3种方法的检出率之间无统计学差异(χ^2=0.8414,P=0.6566)。A方法检测Ct值≤34.6或者B方法检测Ct值≤35.6的血清标本,C方法扩增成功率为100%。结论A方法的敏感性和特异性均较高,C方法为灵敏的基因检测方法。

Objective To compare the performance of three nucleic acid detection methods for hepatitis E virus.Methods The open reading frame(ORF)2 gene sequence of HEV genotype 4 representative strain was cloned into pUC57 vector.Plasmid DNA was detected by two real-time quantitative method A and B,and the detection limits were compared.Other samples were used for specificity detection.Serum specimens of acute hepatitis E patients were detected by three method,and the results were compared.Results The lowest detection limit of plasmid DNA by A and B method can both reach 35 copies/reaction,with specificity of 100%.The HEV RNA positive rate of serum samples from acute hepatitis E patients by A,B and C method was 47.8%(43/90),43.3%(39/90)and 41.1%(37/90),with the concordance rate of 88.9%(80/90).There was no statistically significant difference among the three method(χ^2=0.8414,P=0.6566).Serum specimens with Ct values below 34.6 detected by method A,or below 35.6 detected by method B,the success rate of amplification by method C was 100%.Conclusions Method A has both higher sensitivity and specificity,and method C is a sensitive gene detection method.