The aim of the study was to establish the seroprevalence of herpes simplex virus type 2 (HSV-2) among sexually active adults from East Croatia and to evaluate its correlates with demographic and behavioural risk factors. This cross-sectional study was conducted from the 1st June until the 30th September 2005 at the Department of Transfusion Medicine, Osijek University Hospital Center. The study included 423 blood donors composed of 366 (86.5%) men and 57 (13.5%) women. Demographic data and data on the risky sexual behaviour during the lifetime were collected with usage of an anonymous questionnaire consisting of 15 questions. Western blot testing revealed that 3.3% (14/423) subjects were positive for the presence of IgG antibodies to herpes simplex virus 2, i.e. 10 (2.7%) men and 4 (7.0%) women. A higher HSV-2 seroprevalence was associated with female sex, older age, marital status (with single subjects been under the greater risk), irregular condom use on sexual intercourses with new partners and with higher number (five or more) of sexual partners during lifetime. Among determined risk factors in the study population, only the connection between the HSV-2 positivity and the older age was statistically significant. The study has showed that the demographic rather than the behavioural risk factors are associated with higher prevalence of HSV-2 antibodies in the observed population.