Four hundred forty-six serum samples from HBsAg-positive chronic hepatitis B patients were collected from five areas in China (eastern coastal city, Shanghai; southwestern inland city, Chengdu; mid-inland city, Wuhan; southern island city, Haikou; and northeastern city, Changchun). Precore stop codon variants (e-minus mutants) were screened using a rapid method of polymerase chain reaction (PCR) amplification of a precore and partial core gene fragment (nucleotides 1785-2172), followed by dot-blot hybridization with specific oligonucleotide probes (M0, and M1 + M2). The sequence of the M0 probe covered the distal precore region of wild-type virus (nucleotides 1887-1908), and the sequences of the M1 and M2 probes were from sequences mutated at nt. 1898, (TGG-->TAG) with or without additional change at nt. 1901. A significantly lower incidence of the precore stop codon was found in anti-HBe-positive serum samples from Haikou (17.6%), whereas in other areas the percentages of this mutation in anti-HBe positive sera ranged from 47.4% to 78.9%. In HBeAg-positive samples, the rate of e-minus mutant in coexistence with wild-type virus was low in specimens from Haikou (9.5%) and Changchun (2.9%) compared to other areas in China. In contrast, coexistence of mutant and wild-type virus was frequently detected in samples from Wuhan (50.0%).