Background Guidelines recommend three-site (urine, anal, pharynx) three-monthly (3X3 screening) screening for Neisseria gonorrhoeae (NG) and Chlamydia trachomatis (CT) in men who have sex with men (MSM) taking HIV pre-exposure prophylaxis (PrEP). We present the first randomized controlled trial to compare the effect of screening versus non-screening for NG/CT on the incidence of these infections in MSM taking PrEP. Methods A multicenter, randomized, controlled trial of 3X3 screening for NG/CT versus non-screening was conducted among MSM taking PrEP in five HIV reference centers in Belgium. Participants attended the PrEP clinics quarterly for 12 months. NG/CT was tested at each visit in both arms, but results were not provided to the non-screening arm. The primary outcome was the incidence rate (IR) of NG/CT infections in each arm, assessed in the per-protocol population. Non-inferiority of the non-screening arm was proven if the upper limit of the 95% confidence interval of the IR ratio (IRR) was lower than 1.25. The trial protocol was registered at clinicaltrials.gov (NCT04269434). Findings Between September 2020 and June 2021, 508 subjects were randomized to the 3X3 screening arm and 506 to the non-screening arm. The overall IR of NG/CT was 0.155 cases/100 person-days (95%CI 0.128-0.186) in the 3x3 screening arm and 0.205 (95%CI 0.171-0.246) in the non-screening arm. The IR was significantly higher in the non-screening arm (IRR 1.318, 95%CI 1.068-1.627). Participants in the non-screening arm had a higher incidence of CT infections and symptomatic CT infections. There were no significant differences in NG infections. Participants in the non-screening arm consumed significantly less antimicrobials. No serious adverse events were reported. Interpretation We failed to show that non-screening for NG/CT is non-inferior to 3-site 3-monthly screening in MSM taking PrEP in Belgium. However, screening was associated with higher antibiotic consumption and had no effect on the incidence of NG. Therefore, our findings do not provide strong support for screening for NG/CT in this population.