Stress is known to contribute to overall health status. Many individuals in sub-Saharan Africa are believed to be stressed about their employment, income, and health. This study aimed to investigate hair cortisol as a biomarker of chronic stress in settlement communities in Kenya. Hair samples were collected from 108 volunteers from settlement communities in Kenya. An enzyme-linked immunosorbent assay technique was used to measure hair cortisol concentrations. In parallel, a health survey was completed. The mean ± SD for the cortisol concentration in the hair of volunteers from the settlement communities in Naivasha was 639 ± 300 ng/g, which was higher than found for a Caucasian reference group (299 ± 110 ng/g; one-way ANOVA, P = 0.0003). There were no differences in hair cortisol concentrations between members of slum settlements adjacent to large floriculture farms in Naivasha (Karagita, Kamere/Kwa Muhia/DCK, and Kasarani) compared with those well-removed from all floriculture in Mogotio (Mogotio and Westlands/Katorongot). However, hair cortisol concentrations were significantly higher in females, divorced volunteers, those who made below minimum wage, and those who reported feeling unsafe collecting water or using sanitation facilities within these 2 settlement groups. We found no evidence for increased chronic stress (measured by hair cortisol content) between members of slum settlements adjacent to versus distant to large floriculture farms. Cultural and socio-economic conditions that prevail in much of sub-Saharan Africa were found to be factors contributing to chronic stress.