Title: The genomics and consequences of gut bacterial adaptation to high-fat diets Project Summary: The gut microbial communities of humans and other mammals respond rapidly to changes in host diet, and diet-induced changes in the microbiota can feedback to shape host health outcomes. However, little is known about how individual microbial species within the gut adapt to changes in host diet. The central hypothesis underlying the proposed work is that high-fat diets drive adaptive evolutionary responses in gut bacterial genomes that in turn affect host metabolic traits. Guided by preliminary data generated by our lab, we propose that 1) specific genomic loci underlying gut bacterial adaptation to high-fat diets can be discovered using evolve and re-sequence experiments leveraging genome-resolved metagenomics approaches; 2) plasmid transmission and proliferation promoted by high-fat diets can be quantified through high-throughput culturing and low-cost isolate genome sequencing; 3) fitness effects conferred by gut bacterial genome evolution induced by high-fat diets can be tested through microbiota- competition experiments in germ-free mice; and 4) the effects on host metabolic traits of strain variation selected by high-fat host diets can be measured through transplantation into germ-free mice of defined microbial cocktails evolved under high-fat or low-fat host diets. In Aims 1 and 2, we will use an existing panel of wild-derived inbred mouse lines, each of which harbors a distinct wild-derived gut microbiota, to conduct metagenomic evolve and re-sequence experiments with highly replicated sets of sublines maintained for multiple generations on either high-fat or low-fat diets. Analyses of metagenome-assembled genomes will be used to identify population-genomic signatures of gut bacterial adaptation to the diets (Aim 1), and analyses of bacterial isolate genomes will be used to link diet-associated plasmids with their bacterial hosts (Aim 2). In Aim 3, we will use approaches recently developed and employed by our lab to conduct microbiota-competition experiments in germ-free mice fed either high-fat or low-fat diets. These experiments will enable quantification of the degree to which high-fat or low-fat diets select for the bacterial genomic changes identified in Aims 1 and 2. In Aim 4, we will directly test in germ-free mice how strain adaptation to high-fat host diets affects proximal host traits, such as gut gene expression and immune responses, as well as higher level metabolic traits, including adiposity. These experiments will uncover the adaptive landscape experienced by gut bacteria in the context of a high-fat host diet and determine the consequences of diet-induced bacterial evolution for host metabolic health.