PROJECT SUMMARY More than one million HIV exposed uninfected (HEU) children are born annually, with the majority in sub-Saharan Africa. HEU infants are at an increased risk of poor linear growth, infectious morbidity, and mortality compared to their HIV unexposed uninfected peers. Breastfeeding reduces but does not eliminate poor health outcomes in HEU infants, despite improving maternal health with antiretroviral treatment. The biological mechanisms for vulnerabilities in breastfed HEU infants remain unclear. There are significant gaps in our understanding of how HIV infection affects breastmilk composition and the role of breastmilk composition in the development of poor growth among HEU infants. It is biologically plausible that systemic metabolic dysfunction associated with maternal HIV infection may alter metabolite levels in breastmilk and thereby affect infant outcomes. We hypothesize that HIV will alter the maternal plasma and breastmilk metabolome and that these alterations will be associated with the infant gut microbiome and infant growth. We propose to leverage the Tunza Mwana Kenyan birth cohort (NICHD 5R01HD096999) including lactating women living with and without HIV and their infants. Utilizing existing maternal plasma and breastmilk samples from 60 cohort participants (30 with HIV and 30 without HIV) and prospective infant growth monitoring, we will expand this unique cohort performing targeted metabolomics and machine learning to characterize metabolic relationships between the mother-breastmilk-infant triad and to identify metabolomic profiles in plasma and breastmilk associated with the infant gut microbiome composition and growth. Specific Aims are to 1) determine how maternal plasma metabolites correlate with breastmilk metabolites, and how plasma and breastmilk metabolites are associated with HIV in early (3 weeks) and later (6 months) lactation, 2) evaluate how breastmilk metabolites influence infant gut microbiome composition and diversity at 3 weeks and 6 months, 3) identify maternal plasma and breastmilk metabolites and infant gut microbiome compositions associated with growth to 24 months of life. Career and Learning Objectives are to 1) acquire foundational and advanced skills in epidemiology and biostatistics, 2) develop core competency in the conduct and interpretation of multi-omics studies, and 3) strengthen skills in longitudinal cohort studies and clinical-translational trials. By accomplishing these learning objectives and through the achievement of the specific aims in the research plan, the applicant will be well- positioned to attain competitive external funding to implement future studies assessing the maternal- breastmilk-infant triad and nutrition and growth outcomes. Study Impact: This work will identify key elements in breastmilk that are disrupted and lead to profound effects on infant health. New data generated on maternal metabolomic and infant gut microbiome profiles associated with infant growth wi...