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Introduction: Only 18% of high school girls compared with 37% of boys are reported to meet national physical activity guidelines, but much less is known about sex differences in fitness levels during the elementary and middle school years. This study aimed to describe sex differences in longitudinal fitness outcomes after participation in Fit2PlayTM, a park-based afterschool health and wellness program targeting high risk youth ages 6-to-14 years old.Hypothesis: It was hypothesized that over time boys would have more improvement in (1) cardiorespiratory fitness via the Progressive Aerobic Cardiovascular Endurance Run (PACER) and 400 meter run test; (2) strength via 1-minute timed push-ups and sit-ups; and (3) flexibility via the sit-and-reach test, versus girls.Methods: Youth who participated in Fit2PlayTM for either 1, 2 or 3 school years between 2010-2016 (N=2129, mean age 9.1 years, 52% Hispanic, 48% non-Hispanic black, 54% male) participated in a fitness battery at the beginning and end of the school year(s). Effects of length of Fit2PlayTM participation on fitness outcomes were assessed via 3-level repeated measures analysis stratified by sex and adjusted for child sociodemographics, weight category, area poverty, and year.Results: Adjusted models showed significant improvements for both boys and girls in the PACER and 400 meter run tests (p<0.001 for both), with greater magnitude of effects and dose-response trends for girls after up to 3 years of participation in Fit2PlayTM. Specifically, from baseline to 1, 2 and 3 years of program participation, girls demonstrated 8% (95% CI: 0.87, 0.97), 14% (95% CI: 0.77, 0.96), and 23% (95% CI: 0.65, 0.92) mean improvement in 400 meter run times versus 9% (95% CI: 0.86, 0.96), 9% (95% CI: 0.82, 1.01), and 17% (95% CI: 0.70, 0.98) for boys, respectively (p<0.001 for all). Significant strength improvements were found for both boys and girls for push-ups (p<.01 for both, though 4%, 11% and 12% higher magnitude of effects in girls vs. boys for 1, 2 and 3 years of participation, respectively), and only girls for sit-ups (p<.001).Conclusions: Park-based afterschool physical activity programs have the potential to improve fitness performance in all youth, and particularly girls. Future research should further examine sex differences in the effects of park and other community based programs to ultimately reduce sex disparities in youth fitness, particularly in light of the current youth obesity epidemic that continues to challenge our nation.