Background: Although habitual physical activity energy expenditure (PAEE) and cardio-respiratory fitness (CRF) are now well-established determinants of metabolic disease, there is scarcity of such data from Africa. The aim of this study was to describe objectively measured PAEE and CRF in different ethnic populations of rural Kenya. Methods: A cross-sectional study was done among 1,099 rural Luo, Kamba, and Maasai of Kenya. Participants were 17-68 years old and 60.9% were women. Individual heart rate (HR) response to a submaximal steptest was used to assess CRF (estimated VO2max). Habitual PAEE was measured with combined accelerometry and HR monitoring, with individual calibration of HR using information from the step test. Results: Men had higher PAEE than women (∼78 vs. ∼67 kJ day-1 kg-1, respectively). CRF was similar in all three populations (∼38 and ∼43 mlO2·kg-1 min-1 in women and men, respectively), while habitual PAEE measures were generally highest in the Maasai and Kamba. About 59% of time was spent sedentary (<1.5 METs), with Maasai women spending significantly less (55%). Both CRF and PAEE were lower in older compared to younger rural Kenyans, a difference which was most pronounced for PAEE in Maasai (-6.0 and -11.9 kJ day-1 kg-1 per 10-year age difference in women and men, respectively) and for CRF in Maasai men (-4.4 mlO2·min-1 kg-1 per 10 years). Adjustment for hemoglobin did not materially change these associations. Conclusion: Physical activity levels among rural Kenyan adults are high, with highest levels observed in the Maasai and Kamba. The Kamba may be most resilient to age-related declines in physical activity.