Objective: To determine the effect of prenatal maternal vitamin D supplementation on infant vitamin D status in a tropical region where vitamin D supplementation is not routine. Design: A prospective observational follow-up of a randomized trial. Setting: Maternal-child health facility in Dhaka, Bangladesh (23°N). Subjects: Infants born to pregnant women (n 160) randomized to receive 875 µg (35 000 IU) cholecalciferol (vitamin D 3 ) per week (VD) or placebo (PL) during the third trimester were followed from birth until 6 months of age (n 115). Infant serum 25-hydroxyvitamin D concentration (25(OH)D) was measured at <1, 2, 4 and 6 months of age. Results: Mean infant 25(OH)D was higher in the VD v. PL group at <1 month of age (mean (SD): 80 (20) nmol/l v. 22 (18) nmol/l; P < 0·001), but the difference was attenuated by 2 months (52 (19) nmol/l v. 40 (23) nmol/l; P = 0·05). Groups were similar at 4 months (P = 0·40) and 6 months (n 72; P = 0·26). In the PL group, mean infant 25(OH)D increased to 78 (95 % CI 67, 88) nmol/l by 6 months of age (n 34). 25(OH)D was higher with infant formula-feeding and higher in summer v. winter. Conclusions: Prenatal third-trimester vitamin D supplementation (875 µg (35 000 IU)/ week) significantly ameliorated infant vitamin D status during the neonatal period when the risk of vitamin D deficiency is greatest. Further research is warranted to determine factors that contribute to the rise in 25(OH)D during the first 6 months of life among breast-fed infants in this setting.