Among injection drug users (IDUs), those at highest risk for HIV infection include Latinos, young women, and young men who have sex with men (homosexual men). We examined how HIV infection prevalence is affected by gender and sexual orientation among young Latino IDUs in New York City. We used baseline data from a cohort study of young (18-30 years) IDUs in Harlem, New York City, conducted from 1997 through 1999. Participants were asked about drug use and sexual behaviors, and blood was taken for HIV, hepatitis B, and hepatitis C viral antibody testing. Of 156 participants who self-identified as Latino, 145 (94%) were Puerto Rican. Overall, 101 (65%) were heterosexual men, 11 (7%) were men who have sex with men (MSM), 32 (20%) were heterosexual women, and 12 (8%) were women who have sex with women (WSW). Of the whole cohort, 17 (11%) were HIV positive. HIV infection rates were higher among WSW (42%, p < 0.05), heterosexual women (16%, p < 0.05), and homosexual men (18%, p = 0.09) than heterosexual men (5%). Compared with heterosexual men, homosexual men were significantly (p < 0.05) more likely to have received money or drugs for sex (64% versus 33%), and WSW were significantly more likely to have had unprotected sex with an IDU 5 years or more older (50% versus 16%). Multivariate analysis showed being a WSW (adjusted odds ratio [AOR] = 8.68, 95% confidence interval [CI] 1.78-42.26) and having unprotected sex with an older IDU (AOR = 7.01, 95% CI 2.23-21.96) to be associated with HIV infection. Sexual transmission may account for many HIV infections among young Latino IDUs. The high prevalence of HIV infection among WSW may, in part, be due to their having unprotected sex with older men, but studies with larger sample sizes are needed to confirm this.