Project Summary Horizontal gene transfer (HGT) is a driver of rapid evolution and adaptation in microbial communities. HGT spreads genes among and between bacterial species and is relevant to human health as this process disseminates antibiotic resistance genes, genes involved in toxin production, and other virulence traits. Transduction, the transfer of genetic material by bacterial viruses (phages) or virus-like particles (VLPs), is widespread among bacteria and plays an essential role in the evolution of bacterial pathogens. Despite the importance of transduction to human health, our understanding of the taxonomic range, frequency, and mechanisms of transduction between commensal and pathogenic bacteria are limited. Currently, the study of transduction in microbial communities relies on the reconstruction of historical transduction events that shaped extant microbial genomes. These reconstructions do not measure ongoing transduction. To address this challenge, we developed a novel method termed transductomics, which allows us to measure near to “real time” transduction in microbial communities and identifies the DNA sequence transduced to new bacterial recipients. Transductomics uses shotgun metagenomic DNA sequencing to identify and characterize DNA originating from microbial cells that is carried in phages or VLPs. This technique has the power to identify transducing viruses within microbiomes and can be used to determine the spread of virulence and antibiotic resistance traits that contribute to pathogen success. Recent work, including our own, has shown that phages and VLPs are abundant in the intestine. This indicates that phages and/or VLPs capable of transducing DNA to and from bacteria are significant are understudied players in the spread of genetics traits that can promote bacterial pathogenesis. Although our transductomics method readily identifies both active and potential transduction occurring within diverse bacterial communities, it lacks automation, is low throughput, and labor intensive. The main goals of this project are to establish high-throughput methods with refined resolution that allow for the measurement and identification of transduction within bacterial pathogens and microbiomes and to examine how transduction spreads virulence traits within the microbiota. Identifying transducing events within the microbiota and among established pathogens will facilitate the development of precision therapies aimed at curtailing the spread of virulence and antibiotic resistance genes among bacteria, which can limit pathogenesis.