Background Chlamydia trachomatis (CT) and Neisseria gonorrhoeae (NG) are extremely common sexually transmitted infections (STIs) that are associated with adverse birth and neonatal outcomes, and the risk of vertical transmission of CT and NG during delivery is high. The majority of CT and NG infections are asymptomatic and missed by the standard of care in most countries (treatment based on symptoms). Thus, it is likely that missed maternal CT and NG infections contribute to preventable adverse health outcomes among women and children globally. This study aims to assess the effectiveness of CT and NG testing for asymptomatic pregnant women to prevent adverse neonatal outcomes, understand the inflammatory response linking CT and NG infections to adverse neonatal outcomes, and conduct an economic analysis of the CT and NG testing intervention. Methods The Maduo (“results” in Setswana) is a prospective, cluster-controlled trial in Gaborone, Botswana to compare a near point-of-care CT and NG testing and treatment intervention implemented in “study clinics” with standard antenatal care (World Health Organization-endorsed “syndromic management” strategy based on signs and symptoms without laboratory confirmation) implemented in “standard of care clinics” among asymptomatic pregnant women. The primary outcome is vertical transmission of CT/NG infection. Secondary outcomes include preterm birth (delivery < 37 completed weeks of gestation) and/or low birth weight (< 2500 g). The trial will also evaluate immunological and inflammatory markers of adverse neonatal outcomes, as well as the costs and cost-effectiveness of the intervention compared with standard care. Discussion The Maduo study will improve our understanding of the effectiveness and cost-effectiveness of CT and NG testing among asymptomatic pregnant women. It will also increase knowledge about the CT/NG-related immune responses that might drive adverse neonatal outcomes. Further, results from this study could encourage expansion of STI testing during antenatal care in low resource settings and improve maternal and neonatal health globally. Trial registration : This trial is registered with ClinicalTrials.gov (Identifier NCT04955717, First posted: July 9, 2021)).