Hepatitis B virus (HBV) mutants with mutations in the YMDD motif of viral DNA polymerase/reverse transcriptase are described in patients infected with HBV who have not received lamivudine therapy, but their pathogenetic potential is not clear. These mutants were detected by the polymerase chain reaction with peptide nucleic acid clamping in pretreatment sera from two patients who later received lamivudine. One patient developed acute exacerbation with hepatic encephalopathy and received lamivudine along with plasma exchange, which were effective on his illness. YIDD mutants were detected in all three pretreatment sera and both posttreatment sera from him. HBV DNA clones from pretreatment and posttreatment sera, however, did not have the same amino acid sequence. In the other patient who developed severe breakthrough hepatitis after receiving lamivudine, YIDD mutants were detected in two pretreatment and two posttreatment sera. When amino acid sequences of HBV DNA clones with the YIDD mutation were compared before and after he received lamivudine, however, they were not in accord. Hence, YIDD mutants in both patients with chronic hepatitis B before treatment were not selected by lamivudine after they had been placed on it. Numerous amino acid conversions were detected in HBV DNA clones with YIDD mutations, and some of them created stop codons in the overlapping S gene sequence. In Conclusion, HBV mutants with mutations in the YMDD motif in patients before treatment would not be selected by lamivudine or induce breakthrough hepatitis, and some of these would not be replication-competent due to stop codons in the S gene. J. Med. Virol. 74:361–366, 2004. © 2004 Wiley-Liss, Inc.