HBsAg反应性/核酸筛查无反应性献血者乙肝感染多方法确认研究

Analysis of NAT non-reactive results implicated in HBsAg ELISA reactive blood donors by multi-assays

目的了解深圳市HBsAg ELISA反应性核酸筛查无反应性献血者乙肝感染状况。方法收集2019~2020年HBsAg ELISA反应性NAT无反应性标本,应用全自动时间分辨荧光分析法(TRFIA)和罗氏电化学发光免疫法(ECLIA)及中和试验检测HBsAg;分别采用商用罗氏MPX和Ultrio Elite单人份检测HBV DNA,经2.5 mL大容量提取病毒核酸,采用实时定量PCR检测病毒载量,巢式PCR扩增BCP/PC区和S区基因;同时检测HBV血清学标志物,对可疑结果标本进行追踪检测。综合多方法分析检测结果。结果67602份血样中有73(0.11%)份HBsAg ELISA反应性NAT无反应性血液标本参与本研究。TRFIA、ECLIA、中和试验及5种核酸检测方法确证HBsAg阳性标本15例(20.5%,15/73),另外2例(2.7%,2/73)标本经追踪检测后确认;17例确证HBsAg+标本病毒载量为(未检出~378)IU/mL,中位数为10.1IU/mL;HBsAg浓度与HBV DNA载量间相关性较弱(R^(2)=0.3944)。结论不同HBsAg检测方法只能检测出大部分乙肝病毒感染标本,血液筛查必须选取灵敏度高的多种方法联合检测以确保血液安全。对于结果不一致标本可进行随访,并增加乙肝血清学标志物检测辅助判断。

Objective To investigate NAT non-reactive results implicated in HBsAg ELISA reactive voluntary blood donors in Shenzhen.Methods HBsAg ELISA+but NAT-blood samples were collected,and HBsAg was further retested by TRFIA,Roche ECLIA and neutralization test.HBV DNA of individual donation was detected by commercial Roche MPX and Uultrio Elite,and virus nucleic acid was extracted via 2.5 mL.Molecular characterizations of HBsAg+/NAT-samples were determined by quantitative polymerase chain reaction(qPCR)and nested PCR amplifification of the precore and core promoter regions and HBsAg(S)region.HBV serological markers were detected,and the samples with suspicious results were followed up and detected by multi-assay.Results Among 67602 samples,73(0.11%)HBsAg ELISA+and NAT-blood samples were enrolled in the study.15(20.5%,15/73)were confirmed HBsAg+by TRFIA,ECLI and five alternative DNA assays,and the other 2(2.7%,2/73)were further identified as HBsAg+by follow-up study.In 17 confirmed HBsAg+samples,the viral loads ranged undetectable to 378 IU/mL,with the median of 10.1 IU/mL.Weak correlation was found between HBsAg and HBV DNA load(R^(2)=0.3944).Conclusion Some Hepatitis B virus infected blood samples may miss even with different HBsAg assays.Multi-assays with high sensitivity should be combined for blood screening to ensure blood safety..The inconsistent results should be followed up and further tested for hepatitis B serological markers to assist the confirmation.