To define the epidemiology of HIV-2 infection, we conducted a case-control study among hospitalized patients at an acute care hospital in Bissau, Guinea-Bissau, a country with endemic HIV-2 infection. Among 128 patients with various diagnoses, 23 (18%) were positive for HIV-2 by ELISA and Western blot. One of these patients was serologically reactive for HIV-1 also, but PCR and viral culture revealed the presence of HIV-2 only. To study risk factors, behaviors, and AIDS knowledge related to the acquisition of HIV infection, 22 HIV-2-seropositive and 21 seronegative hospitalized patients were given a physical examination and administered a questionnaire. Among women, transfusion was associated with HIV-2 infection (OR = 14.4, p = 0.02); among men, sex with a prostitute was the principal risk factor (OR = undefined, p = 0.02). Although 79% of HIV-infected patients and controls had heard of AIDS, only 17% of all study participants and 50% of males reporting sex with prostitutes had used condoms in the previous year. These data suggest that the risk factors for HIV-2 infection are similar to those for HIV-1 and support previous studies showing that HIV-2 is the predominant HIV in Guinea-Bissau. Efforts to decrease transmission of HIV-2 should include screening for HIV-2 in blood for transfusion in endemic areas (now done in Bissau) and education about the risk of sexual transmission.