Intrinsic replication advantage of a Hepatitis B Virus genotype C strain is independent of the basal core promoter A1762T/G1764A mutation

This study aimed to isolate and characterize individual hepatitis B virus (HBV) strains from a patient co-infected with genotype B and C, and to directly compare their intrinsic replication capacities. HBV DNA was isolated from a chronic hepatitis B patient. Genotypes were determined by genotype-specific polymerase chain reaction (PCR), and full-length genomes were cloned. Two clones representing subgenotype B2 (c17) and C1 (c21) were selected, sequenced and transfected into HepG2 cells. Viral replication was assessed via Southern blot and quantitative PCR. The role of the common Adenine (A)1762Thymine (T/)Guanine (G)1764A mutation in the C1 clone was investigated by site-directed mutagenesis. As a result, the genotype C1 clone exhibited significantly higher replication capacity than the B2 clone. Counterintuitively, reverting the A1762T/G1764A mutation in the C1 clone to wild-type further enhanced its replication, indicating this mutation was not the primary driver of its high replicative fitness. In conclusion, HBV genotype C1 possesses a greater inherent replication capacity than B2, independent of the classic basal core promoter (BCP) mutation, providing a virological explanation for its association with severe disease outcomes in co-infection.