PROJECT SUMMARY/ABSTRACT – PROJECT 1 Atopic diseases, including atopic dermatitis (AD), food allergy (FA), allergic rhinitis, and asthma, are the most common chronic medical conditions affecting children in the US, and prevention strategies remain largely unsuccessful. While the risk of asthma is reduced in infants with a farming lifestyle, which includes exposure to a diverse microbiome in early life, the microbial mechanisms of protection against AD and FA under this lifestyle are still being investigated. Multiple bacterial metabolites have been also explored for their connection with atopic disease: short-chain fatty acids (SCFA), which can induce the differentiation of anti-inflammatory colonic Tregs, tryptophan metabolism, which modulates epithelial barrier permeability, or bile acids, which regulate T-cell differentiation in the lamina propria. Our published and preliminary results demonstrate that, compared to infants from urban/suburban Rochester (ROC), the traditional agrarian community of Old Order Mennonites (OOM) are protected against atopic diseases and harbor a distinct gut bacterial community, including an enrichment in Bifidobacteria and Clostridia species, suggesting that lifestyle and early life microbiome accelerate immunocompetence and might play a protective role in atopic diseases. The goals of this proposal are to determine how the gut microbiome and its byproducts develop in infants to modulate the risk of atopic diseases, to identify microbial and metabolite biomarkers to screen high-risk infants, and to assess mechanisms associated with protective innate and adaptive immune markers and skin barrier integrity and microbiome by integration of data with Projects 2 and 3. Our central hypothesis is that the microbiome of infants who do not develop AD and FA modulates the bacterial metabolite pool that confers protection or risk of atopic disease. We hypothesize that the gut microbiome of non-atopic urban Rochester infants and OOM infants will be enriched in species with anti-inflammatory properties and genes related to short-chain fatty acid production, lactic acid metabolism, and tryptophan and bile acid metabolism, as well as an accelerated gut microbiome maturation compared with atopic infants. In parallel, we will also assess the fecal metabolome of OOM and ROC infants, and test its association with atopic outcomes: we hypothesize that tryptophan, short-chain fatty acids and bile acids will be differential between atopic and non-atopic infants, and that correlations between microbiome and metabolome will also be differential based on health outcomes. Finally, we will evaluate the longitudinal development of the gut microbiome and metabolome to identify groups of infants with similar changes over time that are associated with atopic disease, as well as with biomarkers from Projects 2 and 3. The rational design of our strategy will identify novel bacterial and metabolite biomarkers that are associated with protection or risk of atopic disease, and which will facilitate the development of microbial interventions in the future.