In 2002, an HIV surveillance study was performed among more than 5500 individuals representing the general population of urban and rural districts in Burundi. In this report, we genetically characterized a subset of the HIV-1-positive samples identified during this survey, including all the HIV-positive samples from Bujumbura, the capital city, and samples from one semiurban and one rural district. One hundred and nineteen samples were genetically characterized in the V3-V5 region of the env gene and/or in the protease and reverse transcriptase region of the pol gene. Phylogenetic analysis of 101 env/pol sequences revealed that the HIV-1 epidemic in Burundi was driven by subtype C (81.2%), followed by subtype A (7.9 %) and polC/envA recombinants (5.9%). One major mutation associated with resistance to antiretroviral drugs (ARVs) in the pol gene, as defined by the International AIDS Society Resistance Testing-USA panel, was observed in one individual, but many minor resistance-associated mutations were also present in the majority of the samples.