Abstract: A cross-sectional seroepidemiologic study was carried out between 1985 and 1990 in 1,567 heterosexual intravenous drug users who had been seen at the AIDS Regional Reference Center in Palermo, Italy, to evaluate the rate of human immunodeficiency virus type 1 (HIV-1) seroprevalence in this group and its long-term trend. Sixty serum samples collected from drug users in 1980 and 1983, before the founding of the Center (1985), were tested as well. Some demographic and behavioral risk factors were studied in a subgroup of intravenous drug users enrolled in 1985, 1987, and 1990 for their possible association with HIV-1. These factors were also studied in relation to hepatitis B virus infection, since both viruses share the same modes of spread. These drug users had a higher prevalence of markers for hepatitis B virus than of HIV-1 antibodies, and the prevalence rates in sera collected declined over time for both infections. The presence of both antibodies to HIV-1 and markers for hepatitis B virus was independently associated with the age of the drug user, the duration of drug use, and the year of serum collection. Antibodies to HIV-1 were observed more frequently in females than in males. No relation was found between education or employment status and the presence of HIV-1 antibodies or hepatitis B virus markers. Although new HIV-1 infections still occur, the decline in seroprevalence observed at the end of the 1980s might be related to modifications in social behavior among newer drug users, partial exhaustion of the susceptible population, and increasing risk awareness in more experienced users.