Objectives To study the effects of urbanization on physical fitness (PF), we compare PF between urban and rural children from western Kenya. We hypothesize that active rural children are stronger, more flexible, and have greater endurance, and that PF differences are predictive of endurance running performance. Methods We recruited an age‐matched, cross‐sectional sample of participants (55 males, 60 females; 6–17 years) from schools near Eldoret, Kenya. PF and anthropometrics were assessed using the FITNESSGRAM®. General linear mixed models (GLMM) and path analyses tested for age, sex, and activity group differences in PF, as well as the effects of PF variables on mile run time. Results On average, urban participants had greater body mass (36.8 ± 15.9 vs. 31.9 ± 10.9 kg) but were not taller than rural participants (1.4 ± 0.2 vs. 1.4 ± 0.2 cm). Greater urban body mass appears driven by higher body fat (28.2 ± 9.4 vs. 16.8 ± 4.4%), which increased with age in urban but not rural participants. GLMM analyses showed age effects on strength variables (P<0.05) and sex differences in hip flexibility, sit‐ups, and mile run (P<0.05). There were few differences in PF between groups except rural participants had stronger back muscles (18.2 ± 4.5 vs. 14.18 ± 4.3 cm) and faster mile times (6.3 ± 0.7 vs. 7.9 ± 2.0 min). Body composition and abdominal strength were predictive of mile time (P < 0.06), but the path analysis revealed a network of interacting direct and indirect effects that influenced endurance performance. Conclusions Although differences in endurance and body composition are marked between urban and rural groups, strength and flexibility are not always correlated with overall activity levels. Am. J. Hum. Biol. 28:514–523, 2016. © 2015 Wiley Periodicals, Inc.