Biology subjects

Rodriguez-Beltran, J.

Publications and source records attributed to Rodriguez-Beltran, J..

12 recordsLinked to original sources

bioRxiv2026

Multireplicon plasmids emerge under predictable rules and drive the spread of antimicrobial resistance across bacterial hosts

Plasmids are DNA molecules that replicate independently of the bacterial chromosome and are typically associated with the spread of antimicrobial resistance (AMR) and virulence determinants, among other relevant traits. Fusion events between plasmids generate larger, complex backbones that carry two or more replication systems, known as multireplicon plasmids. Despite decades of study, we are still far from understanding how multireplicon plasmids arise, persist, and shape the evolution of AMR. Here, we analyzed 24,000 non-redundant plasmids across bacterial genera and found that more than 30% of them encoded multiple replicons. Compared to single-replicon plasmids, multireplicon plasmids were larger, were enriched in genes encoding antimicrobial, metal, and biocide resistance as well as virulence factors, and showed higher mobility and a broader host range. We also found that multireplicon assembly is not random. Some replicon pairs repeatedly merge into stable multireplicon plasmids, while other pairs rarely fuse even when they commonly coexist intracellularly. We also show that replicon pairs tend to be localized either in close proximity to one another or on opposite poles of the plasmid. We further highlight that multireplicon plasmids can be broadly classified into two groups: long-term coevolving replicon pairs and transient associations that lack a shared evolutionary history. Finally, we reveal the molecular mechanisms underlying multireplicon formation and highlight the role of insertion sequences in their formation and maintenance. Together, our work sheds light on the abundance, gene content, evolutionary patterns, and formation dynamics of multireplicon plasmids and pinpoints their relevance to bacterial evolution and human health.

microbiology↗

bioRxiv2025

Plasmids promote bacterial evolution through a copy number-driven increase in mutation rate

Plasmids are autonomously replicating DNA molecules that stably coexist with chromosomes in bacterial cells. These genetic elements drive horizontal gene transfer and play a fundamental role in bacterial ecology and evolution. Theory suggests that plasmids might evolve faster than chromosomes, as the mutation rate per gene should proportionally increase with plasmid copy number. However, the segregation of plasmid copies to daughter cells is random, introducing an additional layer of genetic drift, known as segregational drift, that might delay plasmid evolution. The interplay between plasmid mutational supply and segregational drift determines the evolutionary rate of plasmid-encoded genes, yet the relative contribution of these opposite forces in plasmid evolution remains unclear. Here, we took a classical population genetics framework to devise a mathematical approximation that predicts the fate of plasmid mutations in bacterial populations. We then validate these predictions by integrating computational, experimental, and bioinformatic approaches. Our findings show that plasmid mutation rates scale logarithmically with copy number: while increasing copy number elevates the mutation rate, the effect diminishes at higher copy numbers, where additional copies yield only marginal increases. Nonetheless, the supply of new mutations consistently surpasses the impact of segregational drift across all copy number levels. These results underscore plasmids as powerful platforms for bacterial evolvability and help explain their remarkable prevalence across microbial phylogeny.

microbiology↗

bioRxiv2024

Universal rules govern plasmid copy number

Plasmids -autonomously replicating DNA molecules- exhibit a broad range of replication and mobility strategies, genetic repertoires, host ranges, sizes, and copies per cell. However, the determinants of plasmid copy number (PCN) remain poorly understood. Here, we use extensive DNA sequencing data to analyse the copy number of thousands of diverse bacterial plasmids in a comprehensive manner. We find that PCN is highly variable, spanning nearly three orders of magnitude, and that it is intrinsically robust against changes in genomic context. We further show that PCN variability is tightly associated with plasmid lifestyles, and propose the concept of replicon dominance to explain interactions in widespread multi-replicon plasmids. Finally, we uncover a universal scaling law that links copy number and plasmid size across bacterial species, indicating that pervasive constraints modulate the PCN-size trade-off.

microbiology↗

bioRxiv2024

Plasmid-encoded insertion sequences promote rapid adaptation in clinical enterobacteria

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 insertion sequence 1 (IS1) elements. Specifically, IS1-mediated gene inactivations expedite the adaptation rate of clinical strains in vitro and foster within-patient adaptation in the gut microbiota. 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.

microbiology↗

bioRxiv2023

β-lactamase expression induces collateral sensitivity in Escherichia coli

Major antibiotic groups are losing effectiveness due to the uncontrollable spread of antimicrobial resistance (AMR) genes. Among these, {beta}-lactam resistance genes -encoding {beta}-lactamases- stand as the most common resistance mechanism in Enterobacterales due to their frequent association with mobile genetic elements. In this context, novel approaches that counter mobile AMR are urgently needed. Collateral sensitivity (CS) occurs when the acquisition of resistance to one antibiotic increases susceptibility to another antibiotic and can be exploited to selectively eliminate AMR. However, most CS networks described so far emerge as a consequence of chromosomal mutations and cannot be leveraged to tackle mobile AMR. Here, we dissected the CS response elicited by the acquisition of a prevalent antibiotic resistance plasmid to reveal that the expression of the {beta}-lactamase blaOXA-48 induces CS to colistin and azithromycin. We next showed that expression of other clinically relevant mobile {beta}-lactamases produces similar CS responses in multiple, phylogenetically unrelated E. coli strains. Finally, by combining experiments with surveillance data comprising thousands of antibiotic susceptibility tests, we showed that {beta}-lactamase-induced CS is pervasive within Enterobacterales. These results highlight that the physiological side-effects of {beta}-lactamases can be leveraged therapeutically, paving the way for the rational design of specific therapies to block mobile AMR or at least counteract their effects.

microbiology↗

bioRxiv2022

Antibiotic heteroresistance generated by multi-copy plasmids

Heteroresistance - in which a clonal bacterial population contains a cell subpopulation with higher resistance to antibiotics than the main population - is a growing clinical problem that complicates susceptibility determination and threatens therapeutic success. Despite the high prevalence of heteroresistance in clinical settings, the underlying genetic mechanisms that stably maintain heterogeneous bacterial populations are poorly understood. Using fluorescence microscopy, single-cell microfluidics, and quantitative image analysis, we show that random replication and segregation of multicopy plasmids produce populations of bacterium Escherichia coli MG1655 in which cells with low-and high-plasmid copy numbers stably co-exist. By combining stochastic simulations of a computational model with high-throughput single-cell measurements of blaTEM-1 expression, we show that copy number variability confers the bacterial population with transient resistance to a lethal concentration of a {beta} -lactam antibiotic. Moreover, this surviving, high plasmid copy minority is capable of regenerating a heterogeneous bacterial population with low and high plasmid copy numbers through segregational instability, rapidly alleviating the fitness burden of carrying large numbers of plasmids. Our results provide further support for the tenet that plasmids are more than simple vehicles for horizontal transmission of genetic information between cells, as they can also drive bacterial adaptation in dynamic environments by providing a platform for rapid amplification and attenuation of gene copy number that can accelerate the rate of resistance adaptation and can lead to treatment failure.

microbiology↗

bioRxiv2022

Within-patient evolution of plasmid-mediated antimicrobial resistance

Antibiotic resistance (AMR) in bacteria is a major threat to public health, and one of the key elements in the spread and evolution of AMR in clinical pathogens is the transfer of conjugative plasmids. The drivers of AMR evolution have been extensively studied in vitro, but the evolution of plasmid-mediated AMR in vivo remains poorly explored. Here, we tracked the evolution of the clinically-relevant plasmid pOXA-48, which confers resistance to the last-resort antibiotics carbapenems, in a large collection of enterobacterial clones isolated from the gut of hospitalised patients. Combining genomic and experimental approaches, we first characterized plasmid diversity and the genotypic and phenotypic effects of multiple plasmid mutations on a common genetic background. Second, using cutting-edge genomic editing in wild-type multidrug resistant enterobacteria, we dissected three cases of within-patient plasmid-mediated AMR evolution. Our results revealed, for the first time, compensatory evolution of plasmid-associated fitness cost, as well as the evolution of enhanced plasmid-mediated AMR, in bacteria evolving within the gut of hospitalised patients. Crucially, we observed that the evolution of plasmid-mediated AMR in vivo involves a pivotal trade-off between resistance levels and bacterial fitness. This study highlights the need to develop new evolution-informed approaches to tackle plasmid-mediated AMR dissemination.

evolutionary biology↗

bioRxiv2020

The distribution of plasmid fitness effects explains plasmid persistence in bacterial communities

Introductory paragraphPlasmid persistence in bacterial populations is strongly influenced by the fitness effects associated with plasmid carriage. However, plasmid fitness effects in wild-type bacterial hosts remain largely unexplored. In this study, we determined the distribution of fitness effects (DFE) for the major antibiotic resistance plasmid pOXA-48 in wild-type, ecologically compatible enterobacterial isolates from the human gut microbiota. Our results show that although pOXA-48 produced an overall reduction in bacterial fitness, the DFE was dominated by quasi-neutral effects, and beneficial effects were observed in several isolates. Incorporating these data into a simple population dynamics model revealed a new set of conditions for plasmid stability in bacterial communities, with plasmid persistence increasing with bacterial diversity and becoming less dependent on conjugation. Moreover, genomic results showed a link between plasmid fitness effects and bacterial phylogeny, helping to explain pOXA-48 epidemiology. Our results provide a simple and general explanation for plasmid persistence in natural bacterial communities.

microbiology↗

bioRxiv2020

Collateral Sensitivity Associated with Antibiotic Resistance Plasmids

Collateral sensitivity (CS) is a promising alternative approach to counteract the rising problem of antibiotic resistance (ABR). CS occurs when the acquisition of resistance to one antibiotic produces increased susceptibility to a second antibiotic. For CS to be widely applicable in clinical practice, it would need to be effective against the different resistance mechanisms available to bacteria. Recent studies have focused on CS strategies designed against ABR mediated by chromosomal mutations. However, one of the main drivers of ABR in clinically relevant bacteria is the horizontal transfer of ABR genes mediated by plasmids. Here, we report the first analysis of CS associated with the acquisition of complete ABR plasmids, including the clinically important carbapenem-resistance conjugative plasmid pOXA-48. In addition, we describe the conservation of CS in clinical E. coli isolates and its application to the selective elimination of plasmid-carrying bacteria. Our results provide new insights that establish the basis for developing CS-informed treatment strategies to combat plasmid-mediated ABR.

microbiology↗

bioRxiv2020

Dissemination routes of the carbapenem resistance plasmid pOXA-48 in a hospital setting

Introductory paragraphInfections caused by carbapenemase-producing enterobacteria (CPE) are a major concern in clinical settings worldwide. Two fundamentally different processes shape the epidemiology of CPE in hospitals: the dissemination of CPE clones from patient to patient (between-patient transfer), and the transfer of carbapenemase-encoding plasmids between enterobacteria in the gut microbiota of individual patients (within-patient transfer). The relative contribution of each process to the overall dissemination of carbapenem resistance in hospitals remains poorly understood. Here, we used mechanistic models combining epidemiological data from more than 9,000 patients with whole genome sequence information from 250 enterobacteria clones to characterise the dissemination routes of the carbapenemase-encoding plasmid pOXA-48 in a hospital setting over a two-year period. Our results revealed frequent between-patient transmission of high-risk pOXA-48-carrying clones, mostly of Klebsiella pneumoniae and sporadically Escherichia coli. The results also identified pOXA-48 dissemination hotspots within the hospital, such as specific wards and individual rooms within wards. Using high-resolution plasmid sequence analysis, we uncovered the pervasive within-patient transfer of pOXA-48, suggesting that horizontal plasmid transfer occurs in the gut of virtually every colonised patient. The complex and multifaceted epidemiological scenario exposed by this study provides new insights for the development of intervention strategies to control the in-hospital spread of CPE.

microbiology↗

bioRxiv2019

Genetic dominance governs the evolution and spread of mobile genetic elements in bacteria

Mobile genetic elements (MGEs), such as plasmids, promote bacterial evolution through horizontal gene transfer (HGT). However, the rules governing the repertoire of traits encoded on MGEs remain unclear. In this study, we uncovered the central role of genetic dominance shaping genetic cargo in MGEs, using antibiotic resistance as a model system. MGEs are typically present in more than one copy per host bacterium and, as a consequence, genetic dominance favors the fixation of dominant mutations over recessive ones. Moreover, genetic dominance also determines the phenotypic effects of horizontally acquired MGE-encoded genes, silencing recessive alleles if the recipient bacterium already carries a wild-type copy of the gene. The combination of these two effects governs the catalogue of genes encoded on MGEs, dictating bacterial evolution through HGT.

microbiology↗

bioRxiv2017

Clay-induced DNA double-strand breaks underlay genetic diversity, antibiotic resistance and molecular basis of asbestosis

Some natural clays and synthetic nanofibres present in the environment have a severe impact on human health. After several decades of research, the molecular mechanism of how asbestos induce cancers is not well understood. Different fibres, including asbestos, can penetrate the membrane and introduce DNA in both, bacterial and eukaryotic cells. Incubating Escherichia coli with sepiolite, a clayey material, and asbestos under friction forces, both fibres cause double-strand breaks in bacteria. Since antibiotics and clays are used together in animal husbandry, the mutagenic effect of these fibres might constitute a pathway to antibiotic resistance due to the friction provided by peristalsis of the gut from farm animals in addition to the previously proposed horizontal gene transfer. Moreover, we raise the possibility that the same mechanism could generate bacteria diversity in natural scenarios with a role in the evolution of species. Finally, we provide a new model on how asbestos may promote mutagenesis and cancer based on the observed mechanical genotoxicity.

evolutionary biology↗