Project description:Plasmid fitness is directed by two orthogonal processes—vertical transfer through cell division and horizontal transfer through conjugation. When considered individually, improvements in either mode of transfer can promote how well a plasmid spreads and persists. Together, however, the metabolic cost of conjugation could create a tradeoff that constrains plasmid evolution. Here we present evidence for the presence, consequences, and molecular basis of a conjugation-growth tradeoff across 40 plasmids derived from clinical E. coli pathogens. We discover that most plasmids operate below a conjugation efficiency threshold for major growth effects, indicating strong natural selection for vertical transfer. Below this threshold, E. coli demonstrates a remarkable growth tolerance to over four orders of magnitude change in conjugation efficiency. This tolerance fades as nutrients become scarce and horizontal transfer attracts a greater share of host resources. Our results provide insight into evolutionary constraints directing plasmid fitness and strategies to combat the spread of antibiotic resistance.
Project description:Plasmids are extrachromosomal genetic elements commonly found in bacteria. Plasmids are known to fuel bacterial evolution through horizontal gene transfer (HGT), but recent analyses indicate that they can also promote intragenomic adaptations. However, the role of plasmids as catalysts of bacterial evolution beyond HGT remains poorly explored. In this study, we investigate the impact of a widespread conjugative plasmid, pOXA-48, on the evolution of various multidrug-resistant clinical enterobacteria. Combining experimental and within-patient evolution analyses, we unveil that plasmid pOXA-48 promotes bacterial evolution through the transposition of plasmid-encoded IS1 elements. Specifically, IS1-mediated gene inactivations expedite the adaptation rate of clinical strains in vitro and foster within-patient adaptation in the gut. We decipher the mechanism underlying the plasmid-mediated surge in IS1 transposition, revealing a negative feedback loop regulated by the genomic copy number of IS1. Given the overrepresentation of IS elements in bacterial plasmids, our findings propose that plasmid-mediated IS transposition represents a crucial mechanism for swift bacterial adaptation.
Project description:Plasmid conjugation is a key facilitator of horizontal gene transfer. Since plasmids often carry antibiotic resistance genes, they are crucial drivers of the world-wide rise of antibiotic resistance among pathogens. In natural, engineered and clinical environments, bacteria often grow in protective biofilms. Therefore, a better understanding of plasmid transfer in biofilms is needed. Our aim was to investigate plasmid transfer in a biofilm adapted wrinkly colony mutant of Xanthomonas retroflexus (XRw) with enhanced matrix production and reduced motility. We found that XRw biofilms had an increased uptake of the broad host-range IncP-1ϵ plasmid pKJK5 compared to the wild type. Proteomics revealed fewer flagellum associated proteins in XRw, suggesting that flagella were responsible for reducing plasmid uptake. This was confirmed by the higher plasmid uptake of non-flagellated ∆fliM mutants of X. retroflexus wild type and wrinkly mutant. Moreover, testing several flagella mutants of Pseudomonas putida suggested that the flagella effect was more general. We identified seven mechanisms with the potential to explain the flagella effect and simulated them in an individual-based model. Two mechanisms could thus be eliminated (increased distances between cells and increased lag times due to flagella). Another mechanism identified as viable in the modelling was eliminated by further experiments. Four additional proposed mechanisms have a reduced probability of plasmid transfer in common. Our findings highlight the important yet complex effects of flagella during bacterial conjugation in biofilms.
Project description:The study aimed to characterize plasmids mediating carbepenem resistance in Klebsiella pneumoniae in Pretoria, South Africa. We analysed 56 K. pneumoniae isolates collected from academic hospital around Pretoria. Based on phenotypic and molecular results of these isolates, 6 representative isolates were chosen for further analysis using long reads sequencing platform. We observed multidrug resistant phenotype in all these isolates, including resistance to aminoglycosides, tetracycline, phenicol, fosfomycin, floroquinolones, and beta-lactams antibiotics. The blaOXA-48/181 and blaNDM-1/7 were manily the plasmid-mediated carbapenemases responsible for carbapenem resistance in the K. pneumoniae isolates in these academic hospitals. These carbapenemase genes were mainly associated with plasmid replicon groups IncF, IncL/M, IncA/C, and IncX3. This study showed plasmid-mediated carbapenemase spread of blaOXA and blaNDM genes mediated by conjugative plasmids in Pretoria hospitals.
Project description:Horizontal transfer of the integrative and conjugative element ICEMlSymR7A converts non symbiotic Mesorhizobium spp. into nitrogen-fixing legume symbionts. Here we discover subpopulations of Mesorhizobium japonicum R7A become epigenetically primed for quorum-sensing (QS) and QS-activated horizontal transfer. Isolated populations in this state termed R7A* maintained these phenotypes in laboratory culture but did not transfer the R7A* state to recipients of ICEMlSymR7A following conjugation. We previously demonstrated ICEMlSymR7A transfer and QS are repressed by the antiactivator QseM in R7A populations and that the adjacently-coded DNA-binding protein QseC can represses qseM transcription. Here RNA-sequencing revealed qseM expression was repressed in R7A* cells and that RNA antisense to qseC was abundant in R7A but not R7A*. Deletion of the antisense-qseC promoter converted cells into an R7A*-like state. An adjacently-coded QseC2 protein bound two operator sites and repressed antisense-qseC transcription. Plasmid overexpression of QseC2 stimulated the R7A* state, which persisted following curing of this plasmid. The epigenetic maintenance of the R7A* state required ICEMlSymR7A-encoded copies of both qseC and qseC2. Therefore QseC and QseC2, together with their DNA-binding sites and overlapping promoters, form a stable epigenetic switch that establishes binary control over qseM transcription and primes a subpopulation of R7A cells for QS and horizontal transfer.
Project description:Horizontal gene transfer via plasmid conjugation is a major driving force in microbial evolution. Transfer of conjugative plasmids is a complex process that needs to be synchronized with the physiological state of the bacterial host. While several host transcription factors are known to control the plasmid-borne transfer control genes, RNA-based regulatory circuits for host-plasmid communication remain unknown. Here, we describe a post-transcriptional mechanism whereby the Hfq-dependent small RNA, RprA, inhibits transfer of pSLT, the virulence plasmid of Salmonella enterica. RprA employs two different seed pairing domains to recognize and activate the mRNAs of both the sigma-factor S and RicI, a cytoplasmic membrane protein. The latter is a hitherto unknown conjugation inhibitor whose transcription requires S. Together, RprA and S constitute a feed-forward loop with AND-gate logic which tightly controls RicI synthesis for selective suppression of plasmid conjugation under membrane stress. This study reports the first sRNA-controlled feed-forward loop based on double target activation and an unexpected function for a core-genome encoded small RNA in controlling extrachromosomal DNA transfer.
Project description:Antibiotic resistance is exacerbated by the exchange of antibiotic resistance genes (ARGs) between microbes from diverse habitats. Plasmids are important ARGs mobile elements and are spread by horizontal gene transfer (HGT). In this study, we demonstrated the presence of multi-resistant plasmids from inhalable particulate matter (PM) and its effect on gene horizontal transfer. Three transferable multi-resistant plasmids were identified from PM in a hospital, using conjugative mating assays and nanopore sequencing. pTAir-3 contained 26 horizontal transfer elements and 10 ARGs. Importantly pTAir-5 harbored carbapenem resistance gene (blaOXA) which shows homology to plasmids from human and pig commensal bacteria, thus indicating that PM is a media for antibiotic resistant plasmid spread. In addition, 125 μg/mL PM2.5 and PM10 significantly increased the conjugative transfer rate by 110% and 30%, respectively, and augmented reactive oxygen species (ROS) levels. Underlying mechanisms were revealed by identifying the upregulated expressional levels of genes related to ROS, SOS, cell membranes, pilus generation, and transposition via genome-wide RNA sequencing. The study highlights the airborne spread of multi-resistant plasmids and the impact of inhalable PM on the horizontal transfer of antibiotic resistance.