PROJECT SUMMARY This data science project proposal seeks to utilize innovative methods in untargeted metabolomics and microbiome analysis to study the effect of maternal antibiotic exposure in breastfeeding infants. When breastfeeding mothers are taking antibiotics for common infections, the infant's microbiome and their metabolism may be affected. This could have long-term consequences on the infant's overall health because of the important role of the microbiome in health and disease. It can be difficult to monitor drug effects in infants because of their low blood volume and the invasiveness of blood draws. We have been able to non-invasively monitor drugs in skin using skin swabs. We think this may be an important new way to monitor drugs in infants. The three areas to be investigated in this project are: Aim 1: Monitor infant antibiotic exposure levels through breast milk by monitoring the skin metabolome using non-invasive swabs. We will test skin from breast milk fed infants whose mothers were given antibiotics, breast milk fed infants whose mothers were not given antibiotics, and infants who are not breast fed whose mothers were given antibiotics from Project 1. Untargeted metabolomics and molecular networking will be used to detect antibiotics and their metabolites in skin. Aim 2: Determine the impact of antibiotic exposure through breast milk on the infant microbiome, metabolome and metabolic transformation capacity. Infant fecal samples obtained from Project 1 will be analyzed for changes in the microbiome due to antibiotics from the mother. We will compare how the four different antibiotics given in the clinical study affect the infant microbiome and metabolome. Aim 3: Elucidate the gut metabolic transformations in infant mice from direct or indirect (via maternal treatment or breastfeeding) exposure to ampicillin and investigate if host cytochrome P450 (CYP) activities are altered. Mouse samples obtained from Project 2 will be analyzed using new data science applications to determine the effect of ampicillin through breast milk on the infant's metabolism throughout the intestine and the liver including the cytochrome P450 drug metabolizing enzymes (Cyp). We will also have the targeted analysis of ampicillin in breast milk from our Pharmacometrics and Analytical Chemistry Core and the breast milk analysis from the Milk Analytics Core to complement our untargeted approaches. We will also assess the effect of ampicillin on Cyp activity in the mouse intestine and liver.