Characterization of occult hepatitis B virus from blood donors carrying genotype A2 or genotype D strains

Background/Aims Nucleic acid testing (NAT) for hepatitis B virus (HBV) DNA in blood donations identified occult HBV infection (OBI) as a potential threat to blood safety. Methods A collaborative study was undertaken to explore the molecular basis of OBIs prevalent in Europe in relation to clinical and serological data. Results Ninety-one percent of 77 donor samples of European origin HBV DNA positive but HBV surface antigen (HBsAg) negative were confirmed. Viral load ranged between unquantifiable and 5640 IU/mL (median 25 IU/mL). Fifty-two strains were genotyped (14 HBVA2 and 38 HBVD). Compared to HBsAg+ samples, genotype D was significantly more frequent than genotype A2 in OBIs from Poland or Italy (P < 0.04). Amino acid substitutions were concentrated in the immunologically active parts of the Pre-S/S proteins (P < 0.0001) affecting both cellular CD8 T-cell epitopes and B-cell neutralizing Major Hydrophilic Region epitopes. Substitutions were more frequent in OBIs than in HBsAg+ strains of both genotype D (P < 0.001) and A2 (P < 0.01), in OBIs of genotype D than A2 in the ‘a’ region (P < 0.001) but not cellular epitopes, and in anti-HBs+ than anti-HBs− OBIs (P < 0.001). Conclusions Results support the hypothesis that humoral and cellular immune pressure on the HBV envelope proteins are major mechanisms generating OBI. Nucleic acid testing (NAT) for hepatitis B virus (HBV) DNA in blood donations identified occult HBV infection (OBI) as a potential threat to blood safety. A collaborative study was undertaken to explore the molecular basis of OBIs prevalent in Europe in relation to clinical and serological data. Ninety-one percent of 77 donor samples of European origin HBV DNA positive but HBV surface antigen (HBsAg) negative were confirmed. Viral load ranged between unquantifiable and 5640 IU/mL (median 25 IU/mL). Fifty-two strains were genotyped (14 HBVA2 and 38 HBVD). Compared to HBsAg+ samples, genotype D was significantly more frequent than genotype A2 in OBIs from Poland or Italy (P < 0.04). Amino acid substitutions were concentrated in the immunologically active parts of the Pre-S/S proteins (P < 0.0001) affecting both cellular CD8 T-cell epitopes and B-cell neutralizing Major Hydrophilic Region epitopes. Substitutions were more frequent in OBIs than in HBsAg+ strains of both genotype D (P < 0.001) and A2 (P < 0.01), in OBIs of genotype D than A2 in the ‘a’ region (P < 0.001) but not cellular epitopes, and in anti-HBs+ than anti-HBs− OBIs (P < 0.001). Results support the hypothesis that humoral and cellular immune pressure on the HBV envelope proteins are major mechanisms generating OBI.