PROJECT SUMMARY Late chronotype has been associated with gut dysbiosis and increased cardiometabolic (CM) risk, yet the causal mechanisms are unknown. Existing studies are limited by contradictory findings, small sample sizes, imprecise measurements of sleep and CM risk, as well as lack of detailed measures of dietary intake and other environmental exposures/mediators. The objective of this study is to compare sleep patterns, gut microbiota, lifestyle behaviors, and risk for CM disease in individuals from 5 distinct African-origin populations. The second objective is to humanize mice with gut microbiota from study participants identified by chronotype (early vs. late) in each of the 5 cohorts and challenge the mice to high fat feeding in order to confirm the transferability of the late chronotype impact on gut dysbiosis. The proposed study will investigate sleep timing associations with the gut microbiota and CM risk, including elevated waist circumference, elevated fasting glucose, hypertriglyceridemia, elevated blood pressure, and low HDL, in existing cohorts with significant lifestyle diversity (i.e., diet and physical activity), who have been followed prospectively since 2010. The associations identified will guide fecal microbiota transplant experiments in mice using stool from participants identified by chronotype, and exposed to a 20-week high fat diet challenge to confirm the associations between sleep timing, gut microbiota and CM risk factors. Participants are currently enrolled in METS-Microbiome (R01-DK111848), an ongoing prospective cohort study leveraging an existing cohort of five diverse, well-defined populations from the Modeling the Epidemiologic Transition Study (METS, R01-DK080763). METS is comprised of a cohort of 2,500 adults, living in 5 distinctly different environments: Ghana, South Africa, Jamaica, Seychelles and the US. Our preliminary data from the METS cohorts suggest that gut microbiota diversity is negatively related to CM risk and sleep disruption. In addition to yearly health measurements, including anthropometrics, blood pressure and CM risk measures, we propose to measure sleep timing using actigraphy in 1000 participants (N=200 from each site) currently enrolled in METS-Microbiome. We will use a causal mediation analysis to identify the direct and indirect effects of sleep timing on the gut microbiota. We will thus capitalize upon existing, extensively described cohorts of adults from geographically dispersed populations, resulting in significant variation in environmental covariates. The proposed study will substantially advance our understanding of sleep timing associations with the gut microbiota and CM risk, which is critical given modern 24/7 “on-demand” societies requiring both night and early morning work hours.