Abstract Objectives Streptococcus agalactiae constitutes an important cause of neonatal infections in sub-Saharan Africa. Sulfadoxine/pyrimethamine—the current intermittent preventive treatment of malaria in pregnancy (IPTp)—has proven in vitro activity against group B Streptococcus (GBS). Because of specific drug resistance to sulfadoxine/pyrimethamine, mefloquine—an antimalarial without in vitro activity against GBS—was evaluated as a potential alternative. This study assessed the potential of sulfadoxine/pyrimethamine-IPTp to reduce the prevalence of GBS colonization in pregnant women in Gabon when compared with the inactive control mefloquine-IPTp. Methods Pregnant women participating in a randomized controlled clinical trial evaluating mefloquine-IPTp versus sulfadoxine/pyrimethamine-IPTp were invited to participate and recto-vaginal swabs were collected at delivery for detection of GBS colonization. Prevalence of recto-vaginal GBS colonization was compared between IPTp regimens and risk factor and birth outcome analyses were computed. Results Among 549 participants, 106 were positive for GBS colonization at delivery (19%; 95% CI = 16%–23%). Prevalence of maternal GBS colonization showed no significant difference between the two IPTp regimens (mefloquine-IPTp: 67 of 366 women = 18%; 95% CI = 14%–22%; sulfadoxine/pyrimethamine-IPTp: 39 of 183 women = 21%; 95% CI = 15%–27%). Risk factor analysis for GBS colonization demonstrated a significant association with illiteracy (adjusted OR = 2.03; 95% CI = 1.25–3.30). GBS colonization had no impact on birth outcome, anaemia at delivery, gestational age and birth weight. Conclusions Sulfadoxine/pyrimethamine did not reduce colonization rates when used as the IPTp drug during pregnancy. Illiteracy was associated with GBS colonization.