The replication of two HIV-1 variants (>4.0% divergent in the env gene) was observed during primary infection of a frequent plasma donor. Phylogenetic analysis indicated that both HIV-1 variants likely originated from the same source. Heteroduplex tracking analysis of the env V3-V5 region indicated that one of these variant emerged in the plasma at the time of seroconversion, 15 days after the initial detection of HIV-1 RNA. Sequencing of the entire protein-coding region of plasma viruses from Days 2, 22, and 31 showed possible regions of recombination in the pol locus occurring within the first month of infection. The very rapid fluctuations of HIV-1 variant frequencies and their recombination during primary infection may reflect changes in their relative fitness in the face of developing immunological responses.