We assessed HIV antibody and risk exposures in a cross-sectional sample of 446 fishermen in Sihanouk Ville, a port and fishing area in Cambodia, where high HIV prevalence has been found in sentinel surveillance studies. HIV prevalence was 16.1%, and was highest among unmarried men (17.3%) compared with married (14.6%). Men who stayed in port over one day had a significantly higher prevalence of HIV (31.7%) than those in port for ≤ day (14.6%). Men who had sex exclusively with their wife or girlfriend in the previous three months were significantly less likely to be HIV-infected than those who reported no sexual relations during the same time period (11% vs 23.7%, respectively). HIV infection in Cambodian fishermen was more than double that estimated in the general Khmer population. Although a large-scale condom promotion programme has been implemented in Sihanouk Ville, additional prevention programmes are needed to prevent further spread of HIV.