Biology subjects

San Millan, A.

Publications and source records attributed to San Millan, A..

At least 19 recordsLinked to original sources

bioRxiv2026

Genomic Characterization of the RyC collection: 50 Multidrug Resistant Clinical Isolates of Escherichia coli and Klebsiella spp.

Antimicrobial resistance (AMR) is a major global public health threat, and Enterobacterales producing extended-spectrum {beta}-lactamases (ESBLs) represent some of the most common and concerning pathogens in clinical settings. Importantly, the dissemination of these resistance mechanisms is largely driven by mobile genetic elements (MGEs), particularly plasmids. Advancing our understanding of AMR evolution through experimentation requires moving beyond domesticated laboratory strains and towards clinically relevant isolates. However, despite the abundance of genomic data in public repositories, there is a lack of well-characterised clinical collections available for experimental work. Here, we characterise the RyC collection, which includes 50 multidrug-resistant, ESBL-producing Escherichia coli and Klebsiella spp. strains isolated from the gut microbiota of hospitalised patients at Hospital Universitario Ramon y Cajal (Madrid, Spain). We generated high-quality genome assemblies for all strains using a combination of short- and long-read sequencing technologies. From these data, we performed a comprehensive characterisation of the pangenome, mobilome, resistome and defensome of the collection. We present the RyC collection as a robust and experimentally tractable resource to study AMR evolution and MGEs dynamics in clinically relevant bacterial backgrounds. Impact statementAntimicrobial resistance (AMR) is a growing global health threat driven by the rapid dissemination of resistance genes among clinically relevant bacteria. A major challenge in studying AMR evolution is the reliance on domesticated laboratory strains, which poorly represent the complexity of pathogens circulating in hospitals. Here, we introduce the RyC collection, a set of well-characterised, multidrug-resistant Enterobacterales isolates obtained from hospitalised patients. By combining high-quality genome sequencing with detailed analyses of their gene content and mobile genetic elements (MGEs), this collection provides a realistic and experimentally tractable system to study how resistance evolves and spreads. The RyC collection will facilitate research on AMR dynamics, plasmid biology and host-MGEs interactions, ultimately contributing to the development of more effective strategies to combat antibiotic-resistant infections.

microbiology↗

bioRxiv2026

Fitness effects of antimicrobial resistance genes in changing environments

The evolutionary success of antimicrobial resistance (AMR) genes is generally seen as a trade-off between their function in the presence and their cost in the absence of antibiotics. Mobile integrons are genetic elements that recruit and disseminate dozens of AMR genes among Gram-negative pathogens. Here, we have measured the fitness effects of 136 integron genes conferring resistance against several antibiotic families. We have found a significant proportion having neutral and positive effects in the absence of antibiotics. We confirmed this using a mouse model, where we also observed cases of changes in the sign of fitness effects. This led us to unveil that oxygen availability modulates the cost of AMR genes. Using a stochastic model, we show that fluctuating aerobic/anaerobic conditions can rescue AMR genes in the absence of selective pressure. Here we provide a comprehensive analysis of the cost of AMR at the gene level challenging the traditional fitness-resistance trade-off hypothesis.

microbiology↗

bioRxiv2026

Coupling metabolic enhancement to plasmid spread enables programmable antimicrobial control

The rise of multidrug-resistant pathogens underscores the need for precise antimicrobial strategies that extend beyond conventional antibiotics. Conjugation-based approaches offer a powerful yet underexploited means of delivering targeted genetic interventions directly within microbial communities. In this work, we combined selective killing modules with rationally optimized conjugative vectors to target antibiotic-resistant pathogens and clinically relevant antimicrobial resistance plasmids. First, we engineered and validated toxin-intein modules, programmable cassettes that restrict toxic activity to highly specific regulatory contexts. Specifically, we developed and validated modules targeting Shigella spp., Salmonella enterica, and bacteria carrying the resistance plasmid pOXA-48, demonstrating a tunable system capable of selective activity at both the species and strain levels. To identify the most effective delivery platform, we compared mobilizable and conjugative systems and found that, in vitro, conjugative plasmids consistently outperformed mobilizable ones by approximately one order of magnitude. To further optimize delivery, we streamlined the broad-host-range plasmid RP4 and enhanced its functionality by incorporating either the metabolic fos locus, which confers a fitness advantage to cells carrying the delivery vehicle; a type IV pilus operon that promotes mating-pair stabilization and enables efficient conjugation in liquid environments; or both features combined. Using these engineered RP4 derivatives, we integrated the toxin-intein module targeting pOXA-48 and evaluated its performance in complex microbial communities. In this setting, the RP4 variant carrying both the fos locus and the type IV pilus operon effectively blocked the spread of pOXA-48. Together, this work advances the use of conjugative plasmids as robust and programmable platforms to combat antibiotic resistance and enable microbiome engineering. Beyond introducing highly specific antimicrobial modules and a new generation of optimized conjugative vectors, our results identify ecological competitiveness and plasmid transfer dynamics as critical determinants of the success of such interventions.

microbiology↗

bioRxiv2026

Post-transfer instability, not initial transfer, limits dissemination of IncI1-blaCTX-M-1 plasmids between chicken and human Escherichia coli.

Plasmids are key vectors in the dissemination of antibiotic resistance genes. Within Escherichia coli, IncI1 plasmids carrying blaCTX-M-1 significantly contribute to extended-spectrum beta-lactamase (ESBL) resistance. To investigate their zoonotic dissemination potential, we compared the conjugative transfer rates, post-transfer stability, and short-term evolution of two IncI1-blaCTX-M-1 plasmids transferred from their original chicken E. coli host to a panel of chicken and human E. coli. In vitro conjugation assays revealed that transfer rates were not affected by recipient host origin or temperature and were primarily governed by recipient genotype. However, post-transfer plasmid loss rates were host-dependent, with four-fold higher loss rates in human than chicken-derived transconjugants. The total number of mutations accumulated post-acquisition was determined by the identity of the specific plasmid rather than the host origin, with each plasmid following a distinct evolutionary trajectory involving convergent parallel deletions. Specifically, one trajectory involved the loss of genes associated with plasmid stability, observed more frequently in human than in chicken strains, while the other involved the loss of part of the conjugation machinery. Our findings show that, although these plasmids transfer easily between bacteria from chicken to human hosts, they are less stable in their new host environment, which may restrict their overall spread. A comprehensive One Health risk assessment must consider these critical downstream dynamics of plasmid stability and host-specific evolution.

microbiology↗

bioRxiv2025

Plasmids promote antimicrobial resistance through Insertion Sequence-mediated gene inactivation

Antimicrobial Resistance (AMR) is a major threat to public health. Plasmids are mobile genetic elements that can rapidly spread across bacterial populations, promoting the dissemination of AMR genes in clinical bacteria. In addition, plasmids are enriched in insertion sequences (IS), which are small transposable elements able to translocate between genetic locations. Importantly, IS transpositions commonly lead to gene inactivation, which can in turn promote AMR (e.g. through the modification of the antibiotic target). In this study, we combined experimental, bioinformatic and computational approaches to investigate the role of plasmids as catalysts of AMR through IS-mediated gene inactivation. Our results revealed that plasmid pOXA-48, which encodes two IS1 elements, increases the rate of resistance acquisition to multiple antibiotics in clinical strains of Klebsiella pneumoniae through IS1-mediated gene disruption. Moreover, a large screen of genome databases confirmed that the inactivation of genes through plasmid-encoded IS elements is an extended mechanism of AMR evolution. Finally, both our experiments and computational model revealed that conjugative plasmids can promote this route of AMR acquisition while invading complex bacterial communities. Overall, our study reveals that conjugative plasmids fuel AMR not only through the dissemination of resistance genes, but also through IS-mediated gene inactivation, promoting the evolution of multidrug resistance in bacteria.

microbiology↗

bioRxiv2025

The unique role of nucS-mediated non-canonical mismatch repair in Mycobacterium tuberculosis resistance evolution

DNA surveillance mechanisms play a vital role in maintaining genome stability and minimizing mutation rates. One such mechanism, post-replicative mismatch repair (MMR), corrects replication errors that escape DNA polymerase proofreading activity. In most bacteria and eukaryotes, MMR is orchestrated by MutS and MutL proteins. However, certain archaeal and actinobacterial species, including the major human pathogen Mycobacterium tuberculosis, lack these components. Instead, they rely on the nuclease EndoMS/NucS, a structurally distinct enzyme that governs a non-canonical MMR pathway. Given that M. tuberculosis acquires drug resistance exclusively through chromosomal mutations, understanding mutation rate regulation in this pathogen is critical. Nevertheless, despite its anticipated significance, the role of NucS in drug resistance evolution remains largely unexplored in this organism. This study investigates NucS function in M. tuberculosis and uncovers a unique resistance dynamic distinct from other Actinobacteria. While nucS deletion alters the mutational spectrum, it minimally affects the emergence of rifampicin-, isoniazid-, and ethambutol-resistant mutations, in stark contrast to its role in other Actinobacteria. We demonstrated that this atypical behaviour is not attributable to the presence of a single NucS polymorphism, R144S, in the NucS sequence of the M. tuberculosis reference strain H37Rv, which differs from the NucS consensus sequence. Constructing and analysing an H37Rv variant possessing the NucS consensus sequence revealed a subtly altered mutational spectrum but unchanged mutation rates. Notably, database analysis of the R144S polymorphism in clinical isolates revealed its prevalence and significant association with ethambutol resistance. These findings challenge the established view that nucS serves as a genome stability guardian that minimizes mutation rates in M. tuberculosis, suggesting additional mismatch repair mechanisms beyond NucS or a highly efficient replication system in this pathogen.

microbiology↗

bioRxiv2025

Dissecting pOXA-48 fitness effects in clinical enterobacteria using plasmid-wide CRISPRi screens

Conjugative plasmids are the main vehicle for the spread of antimicrobial resistance (AMR) genes in clinical bacteria. AMR plasmids allow bacteria to survive antibiotic treatments, but they also produce physiological alterations in their hosts that commonly translate into fitness costs. Despite the key role of plasmid-associated fitness effects in AMR evolution, their origin and molecular bases remain poorly understood. In this study, we introduce plasmid-wide CRISPR interference (CRISPRi) screens as a tool to dissect plasmid-associated fitness effects. We designed and performed CRISPRi screens targeting the globally distributed carbapenem resistance plasmid pOXA-48 in 13 different multidrug resistant clinical enterobacteria. Our results revealed that pOXA-48 gene-level effects are conserved across clinical strains, and exposed the key role of the carbapenemase-encoding gene, blaOXA-48, as the main responsible for pOXA-48 fitness costs. Moreover, our results highlighted the relevance of postsegregational killing systems in pOXA-48 vertical transmission, and uncovered new genes implicated in pOXA-48 stability. This study sheds new light on the biology and evolution of carbapenem resistant enterobacteria and endorses CRISPRi screens as a powerful method for studying plasmid-mediated AMR.

microbiology↗

bioRxiv2024

Plasmid-chromosome transcriptional crosstalk in multidrug resistant clinical enterobacteria

Conjugative plasmids promote the dissemination and evolution of antimicrobial resistance in bacterial pathogens. However, plasmid acquisition can produce physiological alterations in the bacterial host, leading to potential fitness costs that determine the clinical success of bacteria-plasmid associations. In this study, we used a transcriptomic approach to characterize the interactions between a globally disseminated carbapenem resistance plasmid, pOXA-48, and a diverse collection of multidrug resistant clinical enterobacteria. Although pOXA-48 produced mostly strain-specific transcriptional alterations, it also led to the common overexpression of a small chromosomal operon present in Klebsiella spp. and Citrobacter freundii strains. This operon included two genes coding for a pirin and an isochorismatase family proteins (pfp and ifp), and showed evidence of horizontal mobilization across Proteobacteria species. Combining genetic engineering, transcriptomics, and CRISPRi gene silencing, we showed that a pOXA-48-encoded LysR regulator is responsible for the plasmid-chromosome crosstalk. Crucially, the operon overexpression produced a fitness benefit in a pOXA-48-carrying K. pneumoniae clinical strain, suggesting that this crosstalk promotes the dissemination of carbapenem resistance in clinical settings.

microbiology↗

bioRxiv2024

Anaerobiosis modulates the performance of antimicrobial resistance genes in Enterobacteriaceae

Bacteria must face and adapt to a variety of physicochemical conditions in the environment and during infection. A key condition is the concentration of dissolved oxygen, proportional to the partial pressure of oxygen (PO2), which is extremely variable among environmental biogeographical areas and also compartments of the human and animal body. Here, we sought to understand if the phenotype of resistance determinants commonly found in Enterobacterales can be influenced by oxygen pressure. To do so, we have compared the MIC in aerobic and anaerobic conditions of isogenic Escherichia coli strains containing 136 different resistance genes against 9 antibiotic families. Our results show a complex landscape of changes in the performance of resistance genes in anaerobiosis. Certain changes are especially relevant for their intensity and the importance of the antibiotic family, like the large decreases in resistance observed against ertapenem and fosfomycin among blaVIM {beta}-lactamases and certain fos genes, respectively; however, the blaOXA-48 {beta}-lactamase from the clinically relevant pOXA-48 plasmid conferred 4-fold higher ertapenem resistance in anaerobiosis. Strong changes in resistance patterns in anaerobiosis were also conserved in Klebsiella pneumoniae. Our results suggest that anaerobiosis is a relevant aspect that can affect the action and selective power of antibiotics for specific AMRs in different environments.

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↗

bioRxiv2023

IDENTIFICATION OF PROMOTER ACTIVITY IN GENE-LESS CASSETTES FROM VIBRIONACEAE SUPERINTEGRONS

Integrons are genetic platforms that acquire new genes encoded in integron cassettes (ICs), building arrays of adaptive functions. ICs generally encode promoterless genes, whose expression relies on the platform-associated Pc promoter, with the cassette array functioning as an operon-like structure regulated by the distance to the Pc. This is relevant in large sedentary chromosomal integrons (SCIs) carrying hundreds of ICs, like those in Vibrio species. We selected 29 gene-less cassettes in four Vibrio SCIs, and explored whether their function could be related to the transcription regulation of adjacent ICs. We show that most gene-less cassettes have promoter activity on the sense strand, enhancing the expression of downstream cassettes. Additionally, we identified the transcription start sites of gene-less ICs through 5-RACE. Accordingly, we found that most of the superintegron in Vibrio cholerae is not silent. These promoter cassettes can trigger the expression of a silent dfrB9 cassette downstream, increasing trimethoprim resistance >512-fold in V. cholerae and Escherichia coli. Furthermore, one cassette with an antisense promoter can reduce trimethoprim resistance when cloned downstream. Our findings highlight the regulatory role of gene-less cassettes in the expression of adjacent cassettes, emphasizing their significance in SCIs and their clinical importance if captured by mobile integrons.

microbiology↗

bioRxiv2023

Chromosomal Integrons are Genetically and Functionally Isolated Units of Genomes

Integrons are genetic elements that increase the evolvability of bacteria by capturing new genes and stockpiling them in arrays. Sedentary chromosomal integrons (SCIs), can be massive and highly stabilized structures encoding hundreds of genes, whose function remains generally unknown. SCIs have co-evolved with the host for aeons and are highly intertwined with their physiology from a mechanistic point of view. But, paradoxically, other aspects, like their variable content and location within the genome, suggest a high genetic and functional independence. In this work, we have explored the connection of SCIs to their host genome using as a model the Superintegron (SI), a 179-cassette long SCI in the genome of Vibrio cholerae N16961. We have relocated and deleted the SI using SeqDelTA, a novel method that allows to counteract the strong stabilization conferred by toxin-antitoxin systems within the array. We have characterized in depth the impact in V. choleraes physiology, measuring fitness, chromosome replication dynamics, persistence, transcriptomics, phenomics and virulence. The deletion of the SI did not produce detectable effects in any condition, proving that -despite millions of years of co-evolution-, SCIs are genetically and functionally isolated units of genomes.

microbiology↗

bioRxiv2023

The distribution of fitness effects of plasmid pOXA-48 in clinical enterobacteria

Antimicrobial resistance (AMR) in bacteria is a major public health problem. The main route for AMR acquisition in clinically important bacteria is the horizontal transfer of plasmids carrying resistance genes. AMR plasmids allow bacteria to survive antibiotics, but they also entail physiological alterations in the host cell. Multiple studies over the last years indicate that these alterations can translate into a fitness cost when antibiotics are absent. However, due to technical limitations, most of these studies are based on analysing new associations between plasmids and bacteria generated in vitro, and we know very little about the effects of plasmids in their native bacterial hosts. In this study, we used a CRISPR-Cas9-tool to selectively cure plasmids from clinical enterobacteria to overcome this limitation. Using this approach, we were able to study the fitness effects of the carbapenem resistance plasmid pOXA-48 in 35 pOXA-48-carrying isolates recovered from hospitalised patients. Our results revealed that pOXA-48 produces variable effects across the collection of wild type enterobacterial strains naturally carrying the plasmid, ranging from fitness costs to fitness benefits. Importantly, the plasmid was only associated with a significant fitness reduction in 4 out of 35 clones, and produced no significant changes in fitness in the great majority of isolates. Our results suggest that plasmids produce neutral fitness effects in most native bacterial hosts, helping to explain the great prevalence of plasmids in natural microbial communities.

microbiology↗

bioRxiv2023

Differences in vertical and horizontal transmission dynamics shape plasmid distribution in clinical enterobacteria

Conjugative plasmids play a key role in the dissemination of antimicrobial resistance (AMR) genes across bacterial pathogens. AMR plasmids are widespread in clinical settings, but their distribution is not random, and certain associations between plasmids and bacterial clones are particularly successful. For example, the globally spread carbapenem resistance plasmid pOXA-48 can use a wide range of enterobacterial species as hosts, but it is usually associated with a small number of specific Klebsiella pneumoniae clones. These successful associations represent an important threat for hospitalized patients. However, knowledge remains limited about the factors determining AMR plasmid distribution in clinically relevant bacteria. Here, we combined in vitro and in vivo experimental approaches to analyze pOXA-48-associated AMR levels and conjugation dynamics in a collection of wild type enterobacterial strains isolated from hospitalized patients. Our results reveal significant variability in these traits across different bacterial hosts, with Klebsiella spp. strains showing higher pOXA-48-mediated AMR and conjugation frequencies than Escherichia coli strains. Using experimentally determined parameters, we developed a simple mathematical model to interrogate the contribution of AMR levels and conjugation permissiveness to plasmid distribution in bacterial communities. The simulations revealed that a small subset of clones, combining high AMR levels and conjugation permissiveness, play a critical role in stabilizing the plasmid in different polyclonal microbial communities. These results help to explain the preferential association of plasmid pOXA-48 with K. pneumoniae clones in clinical settings. More generally, our study reveals that species- and strain-specific variability in plasmid-associated phenotypes shape AMR evolution in clinically relevant bacterial communities. Significance statementConjugative plasmids disseminate AMR genes across bacterial pathogens. Understanding the rules governing plasmid dynamics in bacterial communities is therefore crucial to controlling the global AMR crisis. In this study, we analyzed the dynamics of an AMR plasmid of great clinical relevance, pOXA-48, in a collection of wild type bacteria recovered from hospitalized patients. We reported a high degree of variability in two key plasmid-associated phenotypes, AMR level and conjugation ability, across the collection of clinical bacteria. Using simulations based on the experimental results, we studied how successful associations between AMR plasmids and clinical strains can arise in bacterial communities. Our results revealed that accounting for variability in plasmid-associated phenotypes help to understand the evolution of AMR in clinical settings.

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↗

bioRxiv2021

Fitness effects of CTX-M-15-encoding IncF plasmids on their native Escherichia coli ST131 H30Rx hosts

ObjectivesThe objective of this study was to investigate effects of large CTX-M-15-encoding IncF plasmids on the fitness of their native E. coli ST131 H30Rx hosts in order to understand possible plasmid-host coevolution. MethodsWe selected five E. coli ST131 H30Rx strains of diverse origin, each carrying a multireplicon IncF plasmid encoding the gene blaCTX-M-15. The plasmid was eliminated from each isolate by displacement using an incompatible plasmid vector pMDP5_cureEC958. Whole-genome sequencing (WGS) was performed to obtain complete chromosome and plasmid sequences of wild-type isolates and to detect chromosomal mutations in plasmid-free strains. Competition assays were conducted to determine the relative fitness of plasmid-free clones compared to the corresponding wild-type isolates. ResultsWe were able to successfully eliminate the IncF plasmids from all of the wild-type strains using the curing vector pMDP5_cureEC958. The chromosomes of plasmid-free clones contained zero to six point mutations. Plasmid-free strains of three isolates showed no significant difference in relative fitness compared to the corresponding plasmid-free strains. In the two remaining isolates, the plasmids produced a small but significant fitness cost. ConclusionWe conclude that IncF plasmids produce moderate fitness effects in their E. coli ST131 H30Rx hosts. This fitness compatibility is likely to promote the maintenance of antibiotic resistance in this worrisome E. coli lineage.

evolutionary biology↗