We investigated for the first time the subtype distribution, prevalence of multiple HIV-1 infections, sexual networks, and partnership histories in a cohort of women engaged in high-risk sexual behavior such as female sex workers (FSWs) and women employed in entertainment facilities. Viral RNA was extracted from blood samples collected from 324 HIV-1-positive women; the gp-41 and pol-IN genes were directly sequenced. Women found to have closely related viruses and those with recombinant viruses were further analyzed in the pol-IN gene by clonal sequencing to determine HIV-1 multiple infections. Individual partnership histories were used to provide information on when sex work was undertaken and where. Subtyping in both gp-41 and pol-IN was successfully done in 210/324 (64.8%) women. Subtype distribution in these two genes was 54.3% (n=114) A/A, 2.9% (n=6) C/C, 24.3% (n=51) D/D, 11.9% (n=25) A/D, 4.8% (n=10) D/A, 0.5% (n=1) C/A, 1.0% (n=2) B/A, and 0.5% (n=1) B/D. Sexual networks were identified in six pairs and one triplet of women with closely related subtype A viruses. Partnership histories showed that women having phylogenetically similar viruses had worked in the same localities. Five cases of multiple infections were confirmed: four dual infections and one triple infection. In this first molecular epidemiology study among FSWs in Kampala, subtype A was the predominant subtype. About 9% of a subgroup had multiple infections. Partnership histories and multiple infections observed in this population suggest sexual mixing of the FSWs and their clients confirming their high-risk characteristics.