Hepatitis B virus (HBV) has been classified into seven genotypes, designated A–G. The HBV genotype has a characteristic geographical distribution. The Republic of Uzbekistan is located in the heart of Asia and has been considered to be a region with high endemicity of hepatitis viruses. However, the present distribution of hepatitis virus infection in this region is unknown. The aim of this study was to investigate the distribution of HBV genotypes and to elucidate the validity of two genotyping systems in Uzbekistan. Fifty-four patients with hepatitis B surface antigen were investigated. HBV genotypes were determined by two methods: one based on restriction fragment length polymorphism (RFLP) targeting to S region, and another on enzyme-linked immunosorbent assay (ELISA), using monoclonal antibodies to pre-S2 region. Seven (13%) and 47 (87%) of the 54 subjects were classified into genotypes A and D, respectively. Dual infection of two viral populations of the same genotype was observed in one subject. No significant difference of ALT level (203.3 ± 244.7 vs. 190.6 ± 39.5) and HBeAg (42.9% vs. 42.6%) were found between genotypes A and D. In this study, the validity of the genotyping systems in this region was confirmed. J. Med. Virol. 67:477–483, 2002. © 2002 Wiley-Liss, Inc.