Conjugation drives the horizontal transfer of adaptive traits across prokaryotes. One-fourth of the plasmids encode the functions necessary to conjugate autonomously, the others being eventually mobilizable by conjugation. To understand the evolution of plasmid mobility, we studied plasmid size, gene repertoires, and conjugation-related genes. Plasmid gene repertoires were found to vary rapidly in relation to the evolutionary rate of relaxases, for example, most pairs of plasmids with 95% identical relaxases have fewer than 50% of homologs. Among 249 recent transitions of mobility type, we observed a clear excess of plasmids losing the capacity to conjugate. These transitions are associated with even greater changes in gene repertoires, possibly mediated by transposable elements, including pseudogenization of the conjugation locus, exchange of replicases reducing the problem of incompatibility, and extensive loss of other genes. At the microevolutionary scale of plasmid taxonomy, transitions of mobility type sometimes result in the creation of novel taxonomic units. Interestingly, most transitions from conjugative to mobilizable plasmids seem to be lost in the long term. This suggests a source-sink dynamic, where conjugative plasmids generate nonconjugative plasmids that tend to be poorly adapted and are frequently lost. Still, in some cases, these relaxases seem to have evolved to become efficient at plasmid mobilization in trans, possibly by hijacking multiple conjugative systems. This resulted in specialized relaxases of mobilizable plasmids. In conclusion, the evolution of plasmid mobility is frequent, shapes the patterns of gene flow in bacteria, the dynamics of gene repertoires, and the ecology of plasmids.

In this study, we searched for integrative conjugative elements (ICEs) in chromosomes of 124 strains of Streptococci. These exhaustive analyses revealed 105 putative ICEs and 26 slightly decayed elements that were grouped in 7 distinct families. Comparison of the conjugation modules of ICEs belonging to each family allowed the identification of the conserved proteins. The phylogenetic analysis also revealed that exchanges or acquisition of conjugation and recombination modules are frequent. Integration of these streptococcal ICEs is catalyzed either by site-specific tyrosine integrases, low-specificity tyrosine integrases, site-specific single serine integrases, triplet of site-specific serine integrases or DDE transposases. Analysis of their integration sites led to the detection of 8 new target-genes for streptococcal ICE insertion that have not been identified previously (ftsK, guaA, lysS, mutT, rpmG, rpsI, traG and ybaB/EbfC). It also suggests that all specificities have evolved to minimize the impact of the insertion on the host.

Plasmids can be transferred between cells by conjugation, thereby driving bacterial evolution by horizontal gene transfer. Yet, we ignore the molecular mechanisms of transfer for many plasmids because they lack all protein-coding genes required for conjugation. We solved this conundrum by identifying hundreds of plasmids and chromosomes with conjugative origins of transfer in Escherichia coli and Staphylococcus aureus. These plasmids (pOriT) hijack the relaxases of conjugative or mobilizable elements, but not both. The functional dependencies between pOriT and other plasmids explain their co-occurrence: pOriT are abundant in cells with many plasmids, whereas conjugative plasmids are the most common in the others. We systematically characterized plasmid mobility in relation to conjugation and alternative mechanisms of transfer and can now propose a putative mechanism of transfer for ∼90% of them. In most cases, plasmid mobility seems to involve conjugation. Interestingly, the mechanisms of mobility are important determinants of plasmid-encoded accessory traits, since pOriTs have the highest densities of antimicrobial resistance genes, whereas plasmids lacking putative mechanisms of transfer have the lowest. We illuminate the evolutionary relationships between plasmids and suggest that many pOriT may have arisen by gene deletions in other types of plasmids. These results suggest that most plasmids can be transferred by conjugation. Some plasmids encode complex cellular structures to transfer between bacteria by conjugation, which facilitates the spread of adaptive trains between cells. Yet, half of all plasmids lack the protein-coding genes required for conjugation. Their mobility has remained a mystery that we now contribute to solve by proposing a mechanism of transfer for 9 out of 10 plasmids. We identified the only non-coding sequence required for conjugation, the origin of transfer (oriT), in hundreds of plasmids of Escherichia coli and Staphylococcus aureus. These plasmids might have originated from larger ones and have the highest density of antibiotic resistance genes. Their horizontal transfer depends on complex functional dependencies with other plasmids, which explains their co-existence in bacterial cells.

Microorganisms endure novel challenges for which other microorganisms in other biomes may have already evolved solutions. This is the case of nosocomial bacteria under antibiotic therapy because antibiotics are of ancient natural origin and resistances to them have previously emerged in environmental bacteria. In such cases, the rate of adaptation crucially depends on the acquisition of genes by horizontal transfer of plasmids from distantly related bacteria in different biomes. We hypothesized that such processes should be driven by plasmids among the most mobile and evolvable. We confirmed these predictions by showing that plasmid species encoding antibiotic resistance are very mobile, have broad host ranges, while showing higher rates of homologous recombination and faster turnover of gene repertoires than the other plasmids. These characteristics remain outstanding when we remove resistance plasmids from our dataset, suggesting that antibiotic resistance genes are preferentially acquired and carried by plasmid species that are intrinsically very mobile and plastic. Evolvability and mobility facilitate the transfer of antibiotic resistance, and presumably of other phenotypes, across distant taxonomic groups and biomes. Hence, plasmid species, and possibly those of other mobile genetic elements, have differentiated and predictable roles in the spread of novel traits.

Mobile Genetic Elements (MGEs) are integrated in bacterial genomes and key elements that drive prokaryote genome evolution. Among them are Integrative and Conjugative Elements (ICEs) and Integrative Mobilizable Elements (IMEs) which are important for bacterial fitness since they frequently carry genes participating in important bacterial adaptation phenotypes such as antibiotic resistance, virulence or specialized metabolic pathways. Although ICEs and IMEs are widespread, they are as yet almost never annotated in public bacterial genomes. To address the need of dedicated strategies for the annotation of these elements, we developed ICEscreen, a tool that introduces two new features to detect ICEs and IMEs in Firmicute genomes. First, ICEscreen uses an efficient strategy to detect Signature Proteins of ICEs and IMEs based on a database dedicated to Firmicutes and composed of manually curated proteins and Hidden Markov Models (HMM) profiles. Second, ICEscreen includes a new original algorithm that detects composite structures of ICEs and IMEs that are frequent in genomes of Firmicutes but are currently not resolved by any other tool. We benchmarked ICEscreen on experimentally supported elements and on a public dataset of 246 manually annotated elements including the genomes of 40 Firmicutes and demonstrate its efficiency to detect ICEs and IMEs.