Background: Determine the relationship between obesogenic characteristics of childcare and child adiposity in tribally-affiliated centers in Oklahoma. Methods: The two-day Environment and Policy Assessment and Observation (EPAO) included a total environment (TE), nutrition (N), and physical activity (PA) score and took place in 11 centers across Oklahoma. Eighty-two preschool children (3-5 years) participated. Child height and weight were measured and overweight status (≥ 85th percentile for age and sex) was determined. Regression models, fit using Generalized Estimating Equations methodology to account for clustering by center were used and adjusted for center characteristics. Results: Participants were 3.8 (0.8) years old, 55% male, 67% American Indian (AI) and 38% overweight. A healthier TE and PA was associated with a reduced odds of overweight, which remained significant after adjusting for some center characteristics, but not all. A healthier TE, N, and PA was associated with lower BMI percentile, which remained significant after some center-level adjustments, but not all. Lower sedentary opportunity and sedentary time were no longer associated with reduced odds of overweight following adjustment. Lower opportunity for high sugar and high fat foods and minutes of active play were associated with reduced odds of overweight in some adjusted models. Conclusions: Collectively unadjusted and adjusted models demonstrate that some aspects of a healthier childcare center environment are associated with reduced odds of overweight and lower BMI percentile in preschool children attending tribally-affiliated childcare in Oklahoma. Future research should examine the association of childcare and health behaviors and further explore the role of potential confounders. (author abstract)