Background: Adiposity, low aerobic fitness and low levels of activity are all associated with clustered cardiovascular disease risk in children and their high prevalence represents a major public health concern. Objective: The aim of this study is to investigate the relationship of objectively measured physical activity (PA) with motor skills (agility and balance), aerobic fitness and %body fat in young children. Design: This study is a cross-sectional and longitudinal analyses using mixed linear models. Longitudinal data were adjusted for baseline outcome parameters. Subjects: In all, 217 healthy preschool children (age 4-6 years, 48% boys) participated in this study. Measurements: PA (accelerometers), agility (obstacle course), dynamic balance (balance beam), aerobic fitness (20-m shuttle run) and %body fat (bioelectric impedance) at baseline and 9 months later. Results: PA was positively associated with both motor skills and aerobic fitness at baseline as well as with their longitudinal changes. Specifically, only vigorous, but not total or moderate PA, was related to changes in aerobic fitness. Higher PA was associated with less %body fat at baseline, but not with its change. Conversely, baseline motor skills, aerobic fitness or %body fat were not related to changes in PA. Conclusion: In young children, baseline PA was associated with improvements in motor skills and in aerobic fitness, an important determinant of cardiovascular risk.