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Hepatitis B virus replication is very sensitive to lamivudine. A single amino acid change in human immunodeficiency virus reverse transcriptase is responsible for high-level resistance to this compound. Duck hepatitis B virus mutants were created bearing the analogous amino acid change in the duck hepatitis B virus polymerase. Viral DNA production was reduced 92% for the wild-type virus at 2 micrograms of lamivudine per ml, while the mutants required 40 micrograms of lamivudine per ml to inhibit replication by greater than 80%.