Accompanying the transition from early to middle childhood are substantial changes in children’s educational and recreational circumstances. These changes may affect physical activity levels. This study examined levels of physical activity and sedentary behaviours in kindergarten (age range 5–6 years) and grade 2 (age range 7–8 years). Participants were 96 kindergarten children recruited in the 2010–2011 and 2011–2012 school years and 94 grade 2 children recruited in the 2012–2013 and 2013–2014 school years. A sub-cohort of children was tracked longitudinally from kindergarten to grade 2. Accelerometers were used to measure physical activity and sedentary behaviour. Independent t-tests revealed that children in grade 2 spent significantly less time engaged in physical activity compared to those in kindergarten (292 min/day compared with 354 min/day) and more in sedentary behaviours (443 min/day compared with 368 min/day). For the longitudinal sample, the pattern was similar. Repeated measures ANOVA revealed a significant decrease in physical activity levels (364 min/day to 292 min/day) and a significant increase in sedentary behaviour (368 min/day to 435 min/day) over time. There is a critical need to invest in strategies to maintain higher levels of physical activity across the primary years and reduce sedentary time, since these behaviours affect health outcomes.