BACKGROUND: In the Netherlands, no guidelines exist for routine sexually transmitted infection (STI) screening of human immunodeficiency virus (HIV)-infected men having sex with men (MSM). We assessed prevalence and factors associated with asymptomatic STI. METHODS: MSM visiting HIV outpatient clinics of academic hospitals were tested for Chlamydia trachomatis (CT), Neisseria gonorrhoeae (NG), syphilis, and hepatitis B and C infection. Prevalence and risk factors were studied using logistic regression. RESULTS: In total, 659 MSM were included between 2007 and 2008. STI were found in 16.0% of patients, mostly anal CT and syphilis. One new hepatitis B and 3 new hepatitis C infections were identified. In multivariate analyses, any STI (syphilis, CT, or NG) was associated with patient's age below 40 years (odds ratio [OR]: 2.5, 95% confidence interval [CI]: 1.3-5.0), having had sex with 2 or more sexual partners (OR 2.1, 95% CI: 1.2-3.5), the use of the same sexual toys with a sexual partner (OR 2.2, 95% CI: 1.0-4.9), and enema use before sex (OR: 2.3, 95% 1.2-4.2). Syphilis was independently associated with fisting with gloves versus no fisting (OR: 4.9, 95% CI: 1.7-13.7) and with rimming (OR: 5.0, 95% CI: 1.7-15.0). CT or NG were associated with age below 45 years (age 40-44 years: OR: 2.4, 95% CI: 1.1-5.3; age <40 years: OR: 2.4, 95% CI: 1.1-5.4), enema use before sex (OR: 2.4, 95% CI: 1.3-4.4) and drug use during sex (OR: 2.4, 95% CI: 1.4-4.0). CONCLUSIONS: High-risk sexual behavior was very common, and 16% of HIV-infected MSM in HIV care had an asymptomatic STI, mostly anal CT and syphilis. Development of STI screening guidelines is recommended.