Biology subjects

Botelho, J.

Publications and source records attributed to Botelho, J..

13 recordsLinked to original sources

bioRxiv2026

Rsm-mediated post-translational control of the Pseudomonas putida Type VI Secretion System

The Type VI secretion system (T6SS) is a bacterial nanoweapon that injects toxic effectors into prokaryotic and eukaryotic cells. It is widely found among gram-negative bacteria and provides a significant fitness advantage in interbacterial competition. Pseudomonas putida KT2440 possesses three T6SS clusters (K1-, K2- and K3-T6SS) that combat phytopathogens. This makes this strain a potent biocontrol agent that protects plants from pathogens and can be further enhanced by a better understanding of its T6SS regulation. Although the core components of T6SS are conserved, the elements controlling its regulation differ among bacterial species. T6SS activity is regulated by various factors acting at different levels, from transcription to post-translational modification, to ensure precise control of its activity. Here, we demonstrate the critical importance that the three Rsm proteins, RsmIEA, have in controlling the K1-T6SS structural components and related orphan elements at the post-transcriptional level in Pseudomonas putida. We identified multiple Rsm-binding sites responsible for directly repressing the translation of T6SS proteins (Hcp1 and Hcp5) and their associated effectors (Tke2 and Tke7). Derepression of K1-T6SS mRNA in the rsmIEA mutant led to enhanced translation and expression of the K1-T6SS components and effectors, and critically increased the number of cells in the population with assembled T6SS. This results in a greater capacity to secrete toxins and kill prey cells via the T6SS-dependent mechanism. Finally, we demonstrate the K1-T6SS ability to kill environmental pathogens, including Salmonella enterica and Erwinia amylovora.

microbiology↗

bioRxiv2025

Resistance variation and bacterial interactions shape the adaptation of a genetically diverse bacterial population to antimicrobial treatment

Bacterial infections are often polymicrobial and subject to the evolution of antimicrobial resistance (AMR). Existing knowledge on AMR in such polymicrobial infections usually relies on observational patient data, for which cause-effect relationships are difficult to infer, or on studying interactions between different bacterial species, ignoring the commonly encountered variation within species. Here, we therefore asked how mixed populations with strains from the same species evolve under antibiotic treatment. We used a genetically diverse population of the high-risk human pathogen Pseudomonas aeruginosa, and first identified strain variation in both AMR and pairwise bacteria-bacteria interactions, the latter ranging from beneficial, neutral, to competitive. Using experimental evolution, we subsequently demonstrate that the response to selection by different antibiotic treatments is significantly influenced by AMR strain variation, bacterial interactions, and also spatial population structure. Moreover, de novo AMR evolution was additionally impacted by variation in resistance rates towards the two considered antibiotics. A second evolution experiment emphasized the central role of strain variation and bacterial interactions in determining the evolutionary outcome. We conclude that ecological dynamics in genetically diverse pathogen populations are key for our general understanding of infection characteristics and AMR evolution, and, therefore, deserve particular attention during treatment of polymicrobial infections.

evolutionary biology↗

bioRxiv2024

Timescales and genetic linkage explain the variable impact of defense systems on horizontal gene transfer

Prokaryotes have evolved a wide repertoire of defense systems to prevent invasion by mobile genetic elements (MGE). However, because MGE are vehicles for the exchange of beneficial accessory genes, defense systems could consequently impede rapid adaptation in microbial populations. Here, we study how defense systems impact horizontal gene transfer (HGT) in the short and long terms. By combining comparative genomics and phylogeny-aware statistical methods, we quantified the association between the presence of 7 widespread defense systems and the abundance of MGE in the genomes of 196 bacterial and 1 archaeal species. We also calculated the differences in the rates of gene gain and loss between lineages that possess and lack each defense system. Our results show that the impact of defense systems on HGT is highly taxon- and system-dependent. CRISPR-Cas stands out as the defense system that most often associates with a decrease in the number of MGE and reduced gene acquisition. Timescale analysis reveals that defense systems must persist in a lineage for a relatively long time to exert an appreciable negative impact on HGT. In contrast, at short evolutionary times, defense systems, MGE, and gene gain rates tend to be positively correlated. Based on these results and given the high turnover rates experienced by defense systems, we propose that the inhibitory effect of most defense systems on HGT is masked by recent co-transfer events involving MGE.

microbiology↗

bioRxiv2024

Dynamics of CRISPR-mediated virus-host interactions in the human gut microbiome

Arms races between mobile genetic elements and prokaryotic hosts are major drivers of ecological and evolutionary change in microbial communities. Prokaryotic defense systems such as CRISPR-Cas have the potential to regulate microbiome composition by modifying the interactions among bacteria, plasmids, and phages. Here, we used longitudinal metagenomic data from 130 healthy and diseased individuals to study how the interplay of genetic parasites and CRISPR-Cas immunity reflects on the dynamics and composition of the human gut microbiome. Based on the coordinated study of 80,000 CRISPR-Cas loci and their targets, we show that CRISPR-Cas immunity effectively modulates bacteriophage abundances in the gut. Acquisition of CRISPR-Cas immunity typically leads to a decrease in the abundance of lytic phages, but does not necessarily cause their complete disappearance. Much smaller effects are observed for lysogenic phages and plasmids. Conversely, phage-CRISPR interactions shape bacterial microdiversity by producing weak selective sweeps that benefit immune host lineages. Interestingly, distal (and chronologically older) regions of CRISPR arrays are enriched in spacers that are potentially functional and target crass-like phages and local prophages. This suggests that exposure to reactivated prophages and other endemic viruses is a major selective pressure in the gut microbiome that drives the maintenance of long-lasting immune memory.

microbiology↗

bioRxiv2024

Variation in the response to antibiotics and life-history across the major Pseudomonas aeruginosa clone type (mPact) panel

Pseudomonas aeruginosa is a ubiquitous, opportunistic human pathogen. Since it often expresses multidrug resistance, it is ranked by the World Health Organization among the top 3 high priority pathogens, for which new treatment options are urgently required. An evaluation of new treatments is usually performed experimentally with one of the canonical laboratory strains (e.g., PAO1 or PA14). However, these two strains are unlikely representative of the strains infecting patients, because they have adapted to laboratory conditions and do not capture the enormous genomic diversity of the species. Here, we characterized the major P. aeruginosa clone type (mPact) panel. This panel consists of 20 strains, which reflect the genomic diversity of the species, cover all major clone types, and have both patient and environmental origins. We found significant strain variation in distinct responses towards antibiotics and general growth characteristics. Only few of the measured traits are related, and if so, only for specific antibiotics. Moreover, high levels of resistance were only identified for clinical mPact isolates and could be linked to known AMR (antimicrobial resistance) genes in the sequenced genomes. One strain also produced highly unstable AMR, indicating an evolutionary cost to resistance expression. By linking isolation source, growth, and virulence to life history traits, we further identified specific adaptive strategies for individual mPact strains towards either host processes or degradation pathways. Overall, the mPact panel provides a reasonably sized set of distinct strains, enabling in-depth analysis of new treatment designs or evolutionary dynamics in consideration of the species genomic diversity. ImportanceNew treatment strategies are urgently needed for high risk pathogens such as the opportunistic and often multidrug resistant pathogen Pseudomonas aeruginosa. Here, we characterize the major P. aeruginosa clone type (mPact) panel. It consists of 20 strains with different origins that cover the major clone types of the species as well as its genomic diversity. This mPact panel shows significant variation in (i) resistance against distinct antibiotics, including several last resort antibiotics, (ii) related traits associated with the response to antibiotics, and (iii) general growth characteristics. We further developed a novel approach that integrates information on resistance, growth, virulence, and life-history characteristics, allowing us to demonstrate the presence of distinct adaptive strategies of the strains that focus either on host interaction or resource processing. In conclusion, the mPact panel provides a manageable number of representative strains for this important pathogen for further in-depth analyses of treatment options and evolutionary dynamics.

microbiology↗

bioRxiv2022

Defense systems are pervasive across chromosomally integrated mobile genetic elements and are inversely correlated to virulence and antimicrobial resistance

Mobile genetic elements (MGEs) are key promoters of microbial evolution. These elements can be located extrachromosomally or integrated into the chromosome. Well-known examples of chromosomally integrated MGEs (ciMGEs) are integrative and conjugative/mobilizable elements (ICEs and IMEs), and most studies to date have focused on the biological mechanisms that shape their lifestyle. It is crucial to profile the diversity and understand their distribution across the microbial community, as the number of genome sequences increases exponentially. Herein, I scanned a collection of more than 20000 bacterial and archaeal non-redundant genomes and found over 13000 ciMGEs across multiple phyla, representing a massive increase in the number of ciMGEs available in public databases (<1000). Although ICEs are the most important ciMGEs for the accretion of defense systems, virulence, and antimicrobial resistance (AMR) genes, IMEs outnumbered ICEs. Moreover, defense systems, AMR, and virulence genes were negatively correlated in both ICEs and IMEs. Multiple ciMGEs form heterogeneous communities and challenge inter-phylum barriers. Finally, I observed that the functional landscape of ICEs was populated by uncharacterized proteins. Altogether, this study provides a comprehensive catalog of nucleotide sequences and associated metadata for ciMGEs from 34 phyla across the bacterial and archaeal domains.

microbiology↗

bioRxiv2022

Phylogroup-specific variation shapes pangenome dynamics in Pseudomonas aeruginosa

BackgroundPseudomonas 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. MethodsIn 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. FindingsWe 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. InterpretationAltogether, 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. FundingGerman Science Foundation, Max-Planck Society, Leibniz ScienceCampus Evolutionary Medicine of the Lung, BMBF program Medical Infection Genomics, Kiel Life Science Postdoc Award. Research in contextO_ST_ABSEvidence before this studyC_ST_ABSTo 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 studyTo 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 evidenceThese 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.

microbiology↗

bioRxiv2022

The ESKAPE mobilome contributes to the spread of antimicrobial resistance and CRISPR-mediated conflict between mobile genetic elements

Mobile genetic elements (MGEs) mediate the shuffling of genes among organisms. They contribute to the spread of virulence and antibiotic resistance genes in human pathogens, such as the particularly problematic group of ESKAPE pathogens, including Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter sp. Here, we performed the first systematic analysis of MGEs, including plasmids, prophages, and integrative and conjugative/mobilizable elements (ICEs/IMEs), in the ESKAPE pathogens. We characterized over 1700 complete ESKAPE genomes and found that different MGE types are asymmetrically distributed across these pathogens. While some MGEs are capable of exchanging DNA beyond the genus (and phylum) barrier, most horizontal gene transfer (HGT) events are restricted by phylum or genus. We show that the MGEs proteome is involved in diverse functional processes and distinguish widespread proteins within the ESKAPE context. Moreover, anti-CRISPRs and antimicrobial resistance (AMR) genes are overrepresented in the ESKAPE mobilome. Our results also underscore species-specific trends shaping the number of MGEs, AMR, and virulence genes across pairs of conspecific ESKAPE genomes with and without CRISPR-Cas systems. Finally, we found that CRISPR targets vary according to MGE type: while plasmid and prophage CRISPRs almost exclusively target other plasmids and prophages, respectively, ICEs/IME CRISPRs preferentially target prophages. Overall, our study highlights the general importance of the ESKAPE mobilome in contributing to the spread of AMR and mediating conflict among MGEs.

microbiology↗

bioRxiv2020

ICEs are the main reservoirs of the ciprofloxacin-modifying crpP gene in Pseudomonas aeruginosa

The ciprofloxacin-modifying crpP gene was recently identified in a plasmid isolated from a clinical Pseudomonas aeruginosa clinical isolate. Homologues of this gene were also identified in Escherichia coli, Klebsiella pneumoniae and Acinetobacter baumannii. We set out to explore the mobile genetic elements involved in the acquisition and spread of this gene in publicly available and complete genomes of Pseudomonas. The crpP gene was identified only in P. aeruginosa, in more than half of the complete chromosomes (61.9%, n=133/215) belonging to 52 sequence types, of which the high-risk clone ST111 was the most frequent. We identified 136 crpP-harboring ICEs, with 93.4% belonging to the mating-pair formation G (MPFG) family. The ICEs were integrated at the end of a tRNALys gene and were all flanked by highly conserved 45-bp direct repeats. The core ICEome contains 26 genes (2.2% of all genes), which are present in 99% or more of the crpP-harboring ICEs. The most frequently encoded traits on these ICEs include replication, transcription, intracellular trafficking and cell motility. Our work reveals that ICEs are the main vectors promoting the dissemination of the ciprofloxacin-modifying crpP gene in P. aeruginosa. Author NotesAll supporting data has been provided within the article or through supplementary data files. Supplementary material is available with the online version of this article. Impact StatementA high proportion of Pseudomonas aeruginosa clinical isolates are resistant to ciprofloxacin. Resistance to this antibiotic is often mediated by chromosomal mutations, but recently horizontally transferred genes have been identified. We assessed the repartition of the ciprofloxacin-modifying crpP gene among Pseudomonas genomes and we characterized the mobile elements associated with its acquisition. We found that this gene is prevalent in P. aeruginosa and frequently associated with integrative and conjugative elements (ICEs). Importantly, we also identified highly conserved direct repeats that can be used to accurately delimit crpP-carrying ICEs in P. aeruginosa genomes. Data SummaryAll the bacterial genomes scanned in this study have been deposited previously in the National Center for Biotechnology Information genome database and are listed on the supplementary tables. The newick files used to create the trees in Figures 1 and 4 are deposited on figshare at https://figshare.com/projects/ICEs_are_the_main_reservoirs_of_the_ciprofloxacin-modifying_crpP_gene_in_Pseudomonas_aeruginosa/79308. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=124 SRC="FIGDIR/small/991208v2_fig1.gif" ALT="Figure 1"> View larger version (26K): org.highwire.dtl.DTLVardef@f66f4forg.highwire.dtl.DTLVardef@105b335org.highwire.dtl.DTLVardef@138882eorg.highwire.dtl.DTLVardef@74b9f9_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOFigure 1.C_FLOATNO Approximately-maximum-likelihood phylogenetic tree base on the core genome alignment of 215 complete P. aeruginosa genomes. Branches and labels from crpP-positive hits are colored blue and the branches have twice the standard width. Only bootstrap values from 0.9 to 1 are displayed. C_FIG O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=164 SRC="FIGDIR/small/991208v2_fig4.gif" ALT="Figure 4"> View larger version (19K): org.highwire.dtl.DTLVardef@15dcb4borg.highwire.dtl.DTLVardef@cdba0aorg.highwire.dtl.DTLVardef@70f131org.highwire.dtl.DTLVardef@eaf032_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOFigure 4.C_FLOATNO Approximately-maximum-likelihood phylogenetic tree base on the alignment of the 26 core genes identified in the crpP-harboring ICEs. Genome accession numbers containing more than one crpP-harboring ICE are highlighted in orange. C_FIG

microbiology↗

bioRxiv2019

Integrative and conjugative elements, the neglected drivers of carbapenemase-encoding genes

Carbapenemases inactivate most {beta}-lactam antibiotics, including carbapenems and have been frequently reported among Enterobacteriaceae, Acinetobacter spp. and Pseudomonas spp. Traditionally, the horizontal gene transfer of carbapenemase encoding genes (CEGs) has been linked to plasmids. However, given that integrative and conjugative elements (ICEs) are possibly the most abundant conjugative elements among prokaryotes, we conducted an in-silico analysis to ascertain the likely role of ICEs in the spread of CEGs among all bacterial genomes (n=182,663). We detected 17,520 CEGs, of which 66 were located within putative ICEs among several bacterial species (including clinically-relevant bacteria as Pseudomonas aeruginosa, Klebsiella pneumoniae and Escherichia coli). Most CEGs detected within ICEs belong to the IMP, NDM and SPM metallo-beta-lactamase families, and the serine beta-lactamase KPC and GES families. Different mechanisms were likely responsible for acquisition of these genes. The majority of CEG-bearing ICEs belong to the MPFG, MPFT and MPFF classes and often encode resistance to other antibiotics (e.g., aminoglycosides and fluoroquinolones). This study provides a snapshot of the different CEGs associated with ICEs among available bacterial genomes and sheds light on the underappreciated contribution of ICEs to the spread of carbapenem resistance globally. Author NotesAll supporting data has been provided within the article or through supplementary data files. Supplementary material is available with the online version of this article. Impact StatementCarbapenems are commonly used to treat severe infections in humans. Resistance is often mediated by carbapenemases. These enzymes degrade carbapenems and are frequently present in plasmids. Here, we demonstrate that common carbapenemase-encoding genes (CEGs) found in clinical isolates (e.g. blaKPC, blaGES, blaIMP, blaNDM, blaVIM) can also be located within integrative and conjugative elements (ICEs). CEG-bearing ICEs belong to three mating-pair formation families. These mobile elements may be particularly important in bacteria where plasmids do not seem to play a significant role in the spread of antibiotic resistance genes, as Pseudomonas spp. This study considerably expands the knowledge of the repertoire of CEGs-bearing ICEs among clinically-relevant bacterial pathogens, such as Pseudomonas aeruginosa, Klebsiella pneumoniae and Escherichia coli. Data SummaryAll the bacterial genomes scanned in this study have been deposited previously in the National Center for Biotechnology Information genome database and are listed on the supplementary tables. The extracted 66 ICEs in fasta format and the outputs for the profile HMMs scanned on the 386 putative MGEs identified in this study are deposited on figshare at https://figshare.com/projects/_Comprehensive_genome_data_analysis_establishes_a_triple_whammy_of_carbapenemases_ICEs_and_multiple_clinically-relevant_bacteria/78369.

microbiology↗

bioRxiv2019

Megaplasmids on the rise: combining sequencing approaches to fully resolve a carbapenemase-encoding plasmid in a proposed novel Pseudomonas species

Horizontal transfer of plasmids plays a pivotal role in the dissemination of antibiotic resistance genes and emergence of multidrug-resistant bacteria. Sequencing of plasmids is thus paramount for the success of accurate epidemiological tracking strategies in the hospital setting and routine surveillance. Here, we combine Nanopore and Illumina sequencing to fully assemble a carbapenemase-encoding megaplasmid carried by a clinical isolate belonging to a putative novel Pseudomonas species. FFUP_PS_41 has a multidrug resistance phenotype and was initially identified as Pseudomonas putida, but an average nucleotide identity below the cut-off for species delineation suggests a new species related to the P. putida phylogenetic group. FFUP_PS_41 harbors a 498,516-bp untypable megaplasmid (pJBCL41) with low similarity compared with publicly available plasmids. pJBCL41 contains a full set of genes for self-transmission and genes predicted to be responsible for plasmid replication, partitioning, maintenance and heavy metal resistance. pJBCL41 carries a class 1 integron with the |aacA7|blaVIM-2|aacA4| cassette array (In103) located within a defective Tn402-like transposon that forms part of a 50,273-bp mosaic region bound by 38-bp inverted repeats typical of the Tn3 family and flanked by 5-bp direct repeats. This region is composed of different elements, including additional transposon fragments, five insertion sequences and a Tn3-Derived Inverted-Repeat Miniature Element. The hybrid Nanopore/Illumina approach resulted in contiguous assemblies and allowed us to fully resolve a carbapenemase-encoding megaplasmid from Pseudomonas spp. The identification of novel megaplasmids will shed a new light on the evolutionary effects of gene transfer and the selective forces driving AR.

microbiology↗

bioRxiv2019

A large self-transmissible plasmid from Nigeria confers resistance to multiple antibacterials without a carrying cost

Antimicrobial resistance is rapidly expanding, in a large part due to mobile genetic elements. We screened 94 fecal fluoroquinolone-resistant Escherichia coli isolates from Nigeria for six plasmid-mediated quinolone resistance (PMQR) genes. Sixteen isolates harbored at least one of the PMQR genes and four were positive for aac-6-Ib-cr. In one strain, aac-6-Ib-cr was mapped to a 125 Kb self-transmissible IncFII plasmid, pMB2, which also bears blaCTX-M-15, seven other functional resistance genes and multiple resistance pseudogenes. We hypothesized that pMB2 had been selected by antimicrobials and that its large size would confer a growth disadvantage. However, laboratory strains carrying pMB2 grew at least as fast as isogenic strains lacking the plasmid in both rich and minimal media. We excised a 32 Kb fragment containing the sitABCD and another putative transporter, pefB, a papB homolog, and several open-reading frames of unknown function. The resulting 93 Kb mini-plasmid conferred slower growth rates and lower fitness than wildtype pMB2. Trans-complementing the deletion with the cloned sitABCD genes confirmed that they accounted for the growth advantage conferred by pMB2 in iron-depleted media. The mini-plasmid additionally conferred autoaggregation and was less transmissible and both phenotypes could be complemented with a pefB clone. pMB2 is a large plasmid with a flexible resistance region that contains multiple loci that can account for evolutionary success in the absence of antimicrobials. Ancillary functions conferred by resistance plasmids can mediate their retention and transmissibility, worsening the trajectory for antimicrobial resistance and potentially circumventing efforts to contain resistance through restricted use.

microbiology↗

bioRxiv2018

Carbapenemases on the move: it’s good to be on ICE

The evolution and spread of antibiotic resistance is often mediated by mobile geneticelements. Integrative and conjugative elements (ICEs) are the most abundant conjugativeelements among prokaryotes. However, the contribution of ICEs to horizontal gene transferof antibiotic resistance has been largely unexplored. Here we report that ICEs belonging tomating-pair formation (MPF) classes G and T are highly prevalent among the opportunisticpathogen Pseudomonas aeruginosa, contributing to the spread of carbapenemase-encodinggenes (CEGs). Most CEGs of the MPFG class were encoded within class I integrons, which co-harbour genes conferring resistance to other antibiotics. The majority of the integrons werelocated within Tn3-like and composite transposons. A conserved attachment site could bepredicted for the MPFGclass ICEs. MPFTclass ICEs carried the CEGs within compositetransposons which were not associated with integrons. The data presented here provides aglobal snapshot of the different CEG-harbouring ICEs and sheds light on the underappreciatedcontribution of these elements for the evolution and dissemination of antibiotic resistanceon P. aeruginosa.

microbiology↗