OBJECTIVE This study was performed to determine the prevalence and distribution of hepatitis C virus (HCV) genotypes in Myanmar. METHODS A total of 1333 peripheral blood samples were collected from four different border cities of Myanmar. The anti-HCV antibody-positive serum samples were identified. HCV was genotyped by reverse transcriptase polymerase chain reaction, direct DNA sequencing and phylogenetic analysis on the partial core genome. RESULTS The overall prevalence of HCV infection was 11.6% (154/1333). Regionally, it was 13.5% (47/349) in the north-eastern city, 12.8% (64/501) in the north-western city, 4.2% (16/380) in the southern city and 26.2% (27/103) in the western city. HCV was genotyped in 145/154 (94.2%) samples. Genotype 6 was the most prevalent genotype in this study (71/145, 49%), followed by genotype 3 (57/145, 39.3%), genotype 1 (16/145, 11%), and genotype 2 (1/145, 0.7%). Genotype 6 was mostly found in the northern cities and genotype 3 in the southern and western cities of Myanmar. Multiple HCV genotypes/subtypes were successfully characterized as 1a, 1b, 2a, 3a, 3b, 6m, 6n, and a new 6 subtype. Among them, subtype 6n was the most predominant subtype (38.6%), followed by subtype 3b (29.7%), 3a (9.6%), 6m (9%), 1b (6.9%), 1a (4.1%), new 6 subtype (1.4%) and 2a (0.7%). Subtype 6n was more widely distributed in the northern cities whereas subtype 3b was more common in the western city. The newly discovered genotype 6 subtype was from the northern cities. CONCLUSIONS The results indicate there are regional differences of HCV genotype distribution in Myanmar. There is a distinct geographic variation from other South-East Asian countries in terms of the existence of the new genotype 6 subtype.