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Prenatal exposure to organophosphate (OP) pesticides associate with impaired neurodevelopment in humans and animal models. However, much uncertainty exists about the brain structural alterations underlying these associations. The objective of this study was to determine whether maternal OP pesticide metabolite concentrations in urine repeatedly measured during gestation are associated with brain morphology and white matter microstructure in 518 preadolescents aged 9–12 years. Data came from 518 mother–child pairs participating in the Generation R Study, a population-based birth cohort from Rotterdam, the Netherlands. Maternal urine concentrations were determined for 6 dialkylphosphates (DAPs) including 3 dimethyl (DM) and 3 diethyl (DE) alkyl phosphate metabolites, collected at early, mid, and late pregnancy. At child's age 9–12 years, magnetic resonance imaging was performed to obtain T1-weighted images for brain volumes and surface-based cortical thickness and cortical surface area, and diffusion tensor imaging was used to measure white matter microstructure through fractional anisotropy (FA) and mean diffusivity (MD). Linear regression models were fit for the averaged prenatal exposure across pregnancy. DM and DE metabolite concentrations were not associated with brain volumes, cortical thickness, and cortical surface area. However, a 10-fold increase in averaged DM metabolite concentrations across pregnancy was associated with lower FA (B = −1.00, 95%CI = −1.80, −0.20) and higher MD (B = 0.13, 95%CI = 0.04, 0.21). Similar associations were observed for DE concentrations. This study provides the first evidence that OP pesticides may alter normal white matter microstructure in children, which could have consequences for normal neurodevelopment. No associations were observed with structural brain morphology, including brain volumes, cortical thickness, and cortical surface area. • Little is known about the effect of prenatal OP pesticide exposure on offspring brain morphology and white matter. • Maternal urinary concentrations of 6 DAP metabolites were measured during early, mid, and late pregnancy. • MRI was performed to obtain brain volumes, cortical thickness, and cortical surface area in 518 preadolescents. • DTI was used to measure white matter microstructure through fractional anisotropy (FA) and mean diffusivity (MD). • Prenatal exposure to OP pesticides was predictive of lower FA and higher MD, indicating atypical white matter microstructure. [ABSTRACT FROM AUTHOR]