Gain and loss of plasmid-borne antibiotic resistance genes are associated with their chromosomal counterparts and the broader plasmid gene repertoire in Enterobacteriaceae

Plasmids are central vehicles for the dissemination of antibiotic resistance genes (ARGs). They are among the most mobile and evolvable genetic elements, with broad host ranges and high rates of gene turnover, making them especially effective in spreading antibiotic resistance across bacterial lineages. Using the phylogeny-aware gene gain and loss model applied to 6,895 Enterobacteriaceae genomes, we quantified four evolutionary processes, gene gain, loss, expansion, and reduction, for plasmid-borne genes. We found that, overall, plasmid-borne ARGs (pARGs) exhibit similar gain and loss rates compared with other plasmid genes, but significantly higher expansion and reduction rates. All four processes were strongly species dependent, with only a minor influence of antibiotic class. Further, pARGs were co-transferred with a broad range of plasmid functional genes, including defense, metabolic, and mobilome modules throughout evolutionary history. Moreover, bacterial clades harboring chromosomal ARGs (cARGs) showed significantly higher acquisition of plasmid-borne resistance than did their sister clades lacking corresponding cARGs. Our results demonstrate that pARGs in Enterobacteriaceae are co-transferred with the broader plasmid gene repertoire on an evolutionary timescale, whereas copy-number dynamics are distinct from plasmids. Moreover, cARGs can serve as markers of lineages predisposed to accumulate plasmid-mediated resistance.