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Background: Besides vaccine escape or immune escape hepatitis B virus (HBV) mutants, naturally occurring and drug-induced mutations have been reported in the surface gene (S-gene) of HBV. Aim: To investigate the frequency and profile of naturally occurring S-gene mutants and the influence of long-term lamivudine therapy in patients with chronic hepatitis B (CHB). Materials and Methods: 57 patients with histologically proven CHB, on lamivudine 100 mg/day for more than 24 months, were included. Viral DNA was extracted at baseline and from on-therapy serum samples. The region encoding the complete major hydrophilic region (MHR) and flanking regions (nucleotides 425–840) of major S-gene that overlapped with the viral polymerase was PCR amplified and sequenced. End-of-therapy response (ETR) was assessed. Results: Two (3.5%) patients had naturally occurring HBV mutants, sP127S …