Mobility shapes plasmid GC content evolution

Abstract Plasmids are frequently AT-rich relative to their bacterial hosts. Despite this tendency towards lower GC content, plasmid and host chromosome GC content are positively correlated across diverse collections of plasmid–host pairs. However, the evolutionary processes underlying this pattern remain unclear. The classic model of amelioration predicts that horizontally acquired DNA gradually converges on host nucleotide composition, but since plasmids can repeatedly transfer between bacterial hosts, the opportunity for such host-associated evolution may depend on their transmission dynamics. Using 50,936 plasmid–host pairs from a public sequence database, we found that the apparent global correlation between plasmid and host chromosome GC content was largely driven by differences between bacterial species rather than within species. We therefore accounted for plasmid and host population structure when testing how plasmid mobility shaped host-associated compositional evolution. We compared two contrasting regimes: a population of 3,682 Enterobacterales plasmids distributed across diverse host backgrounds, and six long-term host-associated plasmids from a Rhizobium leguminosarum lineage with INSeq-determined gene essentiality data. In the Enterobacterales population, GC content variation was overwhelmingly explained by plasmid lineage rather than host phylogeny, and conjugative plasmids showed greater similarity to their host chromosomes than mobilisable or non-mobilisable plasmids. In the Rhizobium leguminosarum plasmids, synonymous-site composition was more similar to the host chromosome among genes required across multiple host life stages. Together, these results support a model in which plasmid mobility influences the opportunity for host-associated evolutionary processes to alter nucleotide composition.