Molecular Characteristics of HBV Among Blood Donors in Southern China with Different HBsAg ELISA Results by Two Kits

Abstract

Background: The blood donors tested reactive (R) for hepatitis B surface antigen (HBsAg) but were missed by routine nucleic acid test (NAT), which can be resulted by the infection of hepatitis B virus (HBV) with extremely low viral load. This phenomenon remains a strict threat to blood transfusion in China.Objectives: We aimed to investigate and analyze the molecular characteristics of HBV among blood donors that detected as HBsAg reactive (R) in single ELISA Reagent. Methods: The blood donations were detected as HBsAg R in one ELISA kit, using two kinds of ELISA reagents. These samples with non-reactive (NR) results by NAT were collected and tested for HBsAg by Abbott chemiluminescent microparticle immunoassay (CLIA) with neutralization test, the level of HBsAg were further detected by Roche electrochemiluminescence immunoassay (ECLIA). The viral basic core promoter (BCP) and pre-core (PC) and S regions were also amplified by nested-PCRs. Quantitative real-time PCR (qPCR) for viral load determination and individual donation NAT of Roche reagent were adopted simultaneously. HBsAg was confirmed by the results of CLIA, ECLIA, nested-PCR, qPCR, and ID-NAT.Results: Of 100,252 donations, 38 and 41 were identified as HBsAg reactive only in WanTai and DiaSorin ELISA kit respectively, after blood screening using WanTai and DiaSorin ELISA assays. 79 (0.077%, 79/100,252) blood samples with ELISA R-NR and NAT NR results were enrolled in the study. Of which, 17 (21.5%,17/79) were confirmed as HBsAg positive. Of 14 cases genotyped, genotype B were 78.6% (11/14), C and D was observed in 2 (14.2%, 2/14) and 1 sample (7.1%, 1/14), respectively. Mutations in the S gene such as Y100C, Y103I, G145R, and L175S were found, which can affect the detection of HBsAg. A high-frequency mutation, T1719G (93.3%), was detected in BCP/PC and would reduce the replication of the virus. Conclusion: A small part of blood samples with HBsAg ELISA R-NR and NAT NR results were confirmed as HBV infection, viral nucleic acids were found in most of those samples through the in-house NAT methods. It is necessary to apply more sensitive and specific assays for the detection of HBV infection among blood donors.