Objectives: In preparation for a phase III HIV vaccine trial, we report the results of men who have sex with men (MSM) cohort (‘Project0 Rio’). Methods: HIV-negative MSM were followed 6-monthly over a 3-year period. Results: High seroprevalence for sexually transmitted disease (HIV, syphilis and hepatitis B seropositivities of 24%, 29%, a,nd 37%, respectively) was seen in the 1165 potential volunteers, and this seroprevalence showed significant differences according to HIV serologic status. Among the 647 HIV-negative cohort participants, HIV incidence rate (IR) was 3.33 (95% CI 1.93-4.67) per 100 men-years, with 21 newly acquired HIV infections during 7572 men-months of observation. IR differences were observed in four categories: (1) non-commercial sex workers and non-transvestites who had protected sex prior to study entry (IR 1.6 per 100 men-years); (2) non-commercial sex workers and non-transvestites who had unprotected sex prior to study entry (IR 2.7 per 100 men-years); (3) commercial sex workers (IR 3.5 per 100 men-years); and (4) transvestite groups (IR 16.8 per 100 men-years). Unprotected penetrative anal sex was frequent (66%) in the 6 months before study entry, and when we take into consideration the fact that bisexual men engaged in unprotected vaginal intercourse with their female partners, the risk behavior in this group escalates to 73%. HIV seroconverters had high risk behavior prior to study entry (76%), and all but one admitted to having unprotected penetrative sex prior to infection. Conclusions: MSM in this study had a high rate of unprotected penetrative sexual practices, which caused a significant HIV incidence rate, and, with improved study adherence, this study site could be used for future vaccine trials.