Phylogroup-specific variation shapes the clustering of antimicrobial resistance genes and defence systems across regions of genome plasticity

Background Pseudomonas aeruginosa is an opportunistic pathogen consisting of three phylogroups (hereafter named A, B, and C) of unevenly distributed size. Here, we assessed phylogroup-specific evolutionary dynamics in a collection of P. aeruginosa genomes. Methods In this genomic analysis, using phylogenomic and comparative genomic analyses, we generated 18 hybrid assemblies from a phylogenetically diverse collection of clinical and environmental P. aeruginosa isolates, and contextualised this information with 1991 publicly available genomes of the same species. We explored to what extent antimicrobial resistance (AMR) genes, defence systems, and virulence genes vary in their distribution across regions of genome plasticity (RGPs) and “masked” (RGP-free) genomes, and to what extent this variation differs among the phylogroups. Findings We found that members of phylogroup B possess larger genomes, contribute a comparatively larger number of pangenome families, and show lower abundance of CRISPR-Cas systems. Furthermore, AMR and defence systems are pervasive in RGPs and integrative and conjugative/mobilizable elements (ICEs/IMEs) from phylogroups A and B, and the abundance of these cargo genes is often significantly correlated. Moreover, inter- and intra-phylogroup interactions occur at the accessory genome level, suggesting frequent recombination events. Finally, we provide here a panel of diverse P. aeruginosa strains to be used as reference for functional analyses. Interpretation Altogether, our results highlight distinct pangenome characteristics of the P. aeruginosa phylogroups, which are possibly influenced by variation in the abundance of CRISPR-Cas systems and that are shaped by the differential distribution of other defence systems and AMR genes. Funding German Science Foundation, Max-Planck Society, Leibniz ScienceCampus Evolutionary Medicine of the Lung, BMBF program Medical Infection Genomics, Kiel Life Science Postdoc Award. Research in context Evidence before this study To date, pangenome studies exploring the epidemiology and evolution dynamics of bacterial pathogens have been limited due to the use of gene frequencies across whole species dataset without accounting for biased sampling or the population structure of the genomes in the dataset. We searched PubMed without language restrictions for articles published before September 1, 2021, that investigated the phylogroup-specific evolutionary dynamics across bacterial species. In this literature search we used the search terms “pangenome” and “phylogroup” or “uneven”, which returned 14 results. Of these, only one study used a population structure-aware approach to explore pangenome dynamics in a bacterial species consisting of multiple phylogroups with unevenly distributed members. Added value of this study To our knowledge, this study is the first to assess phylogroup-specific evolutionary dynamics in a collection of genomes belonging to the nosocomial pathogen P. aeruginosa. Using a refined approach that challenges traditional pangenome analyses, we found specific signatures for each of the three phylogroups, and we demonstrate that members of phylogroup B contribute a comparatively larger number of pangenome families, have larger genomes, and have a lower prevalence of CRISPR-Cas systems. Additionally, we observed that antibiotic resistance and defence systems are pervasive in regions of genome plasticity and integrative and conjugative/mobilizable elements from phylogroups A and B, and that antibiotic resistance and defence systems are often significantly correlated in these mobile genetic elements. Implications of all the available evidence These results indicate that biases inherent to traditional pangenome approaches can obscure the real distribution of important cargo genes in a bacterial species with a complex population structure. Furthermore, our findings pave the way to new pangenome approaches that are currently under-explored in comparative genomics and, crucially, shed a new light on the role that integrative and conjugative/mobilizable elements may play in protecting the host against foreign DNA.