Plasmid overlap and evolution between Enterobacterales isolates from bloodstream infections and non-human compartments

Horizontal gene transfer in Enterobacterales allows mobile genetic elements to move between strains, species, and genera. In particular, the movement of plasmids is a known route for rapid dissemination of antimicrobial resistance (AMR) genes. However, it is difficult to establish to what extent plasmids are shared between Enterobacterales causing human infections and those from non-human sources, such as livestock or the environment. While some previous studies have found only limited evidence for genetic overlap, these have often been limited in size, restricted to drug-resistant isolates, and have used fragmented genome assemblies. Here, we report a collection of geographically and temporally restricted isolates from human bloodstream infections (BSI), environmental soils, livestock (cattle, pigs, poultry, sheep), wastewater (influent, effluent), and rivers. Isolates were all collected between 2008-2018 from sampling sites <60km apart. The combined dataset contains 1,458 complete Enterobacterales genomes, including 3,697 circularised plasmids of which one third were unclassifiable. We find eight groups of near-identical plasmids seen in both human BSI and non-human isolates, of which two are conjugative F-type plasmids carrying AMR genes. We cluster plasmids based on alignment-free distances and find that 73/247 (30%) plasmid clusters contain plasmids from both human BSI and non-human isolates. Pangenome-style analyses of the 69 most prevalent clusters (1,832/3,697 plasmids) reveals sets of shared core genes alongside accessory gene repertoires. Core-gene phylogenies suggest an intertwined ecology where well-conserved putative plasmid ‘backbones’ carry diverse accessory functions, potentially linked to niche adaptation. Furthermore, we show that closely related human and non-human plasmids are frequently found across distantly related bacterial hosts. Our findings underline the importance of diverse sampling in ‘One Health’ approaches for AMR management. ### Competing Interest Statement The authors have declared no competing interest.