The aim of this study was to characterize updated HIV subtypes in Yunnan to determine their origins and distribution within the population. RT-PCR of both the gag and env genes were sequenced from Yunnan province inhabitants newly diagnosed with HIV-1. Sequence data from 290 samples were used for statistical analysis of subtype distribution and phylogenetic tree construction. Distribution data were adjusted to account for different geographical distributions of HIV-1 subtypes in the population. Phylogenetic analysis revealed six HIV-1 subtypes in Yunnan, including eight types of unique recombination forms (URFs). The most prevalent subtypes in this province, CRF07_BC (18·9%), CRF08_BC (39·1%), CRF01_AE (22·4%), and URFs (subtype C, 5·9% and subtype B, 4·5%), were all recombinants. We found significant differences in the distribution of these HIV-1 subtypes not only geographically, but also between various ethnic groups and with respect to transmission routes. Our findings indicate a complex population of HIV-1 subtypes, URFs, and recombinant subtypes in Yunnan province. This diversity could make the prevention and control of HIV infection in Yunnan more difficult due to the possibility of virus recombination or infection by multiple subtypes.