To investigate whether Epstein-Barr virus (EBV) type 2 infection is highly prevalent among homosexual men, the prevalence of EBV type 2 was studied among homosexual and heterosexual white men who were at high and low risk for sexually transmitted diseases; these data were correlated with sexual behavior. The prevalence of EBV type 2 among homosexual men was significantly higher than it was among heterosexual men (39% vs. 6%). Among high-risk heterosexual men, prevalence was significantly higher than it was among low-risk heterosexual men (15% vs. 0). In univariate analyses, EBV type 2 infection in homosexual men was significantly associated with human immunodeficiency virus (HIV) seropositivity, increased numbers of intercourse partners, non-Dutch nationality, and human herpesvirus 8 seropositivity. In multivariate analyses, an independent association with EBV type 2 was observed only for HIV seropositivity and number of sex partners. These data support the conclusion that EBV type 2 infection is more prevalent among white homosexual men and is caused by sexual transmission.