Duck hepatitis B virus polymerase gene mutants associated with resistance to lamivudine have a decreased replication capacity in vitro and in vivo

Background/Aims: Hepatitis B virus mutants of the polymerase gene are frequently selected during lamivudine therapy for chronic hepatitis B. To study the biology of these mutants, we analyzed their replication capacity in the duck hepatitis B virus (DHBV) infection. Methods: The B and C domain polymerase mutants corresponding to the clinical isolates were engineered by site directed mutagenesis in the DHBV genome in different expression vectors. Results: The study of the enzymatic activity of the mutated viral polymerase polypeptides analyzed in a cell free system demonstrated a lower priming activity and a decreased capacity of elongation of viral minus strand DNA that was consistent with the lower replication capacity of these mutants in transfected leghorn male hepatoma cells compared to wild type genome. These mutants had a lower replication capacity in primary hepatocytes and in in vivo transfected ducklings. Although resistant to lamivudine, these mutants remained sensitive to PMEA. Conclusion: YMDD mutants of the DHBV reverse transcriptase have a decreased replication capacity both in vitro and in vivo, and are not cross-resistant to PMEA. These results may be important to design new antiviral strategies to combat the replication of the lamivudine resistant viral strains.