Biology subjects

Costas, C.

Publications and source records attributed to Costas, C..

7 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↗

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

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-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↗

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↗

bioRxiv2018

Differences in firing efficiency, chromatin and transcription underlie the developmental plasticity of Arabidopsis originome

Eukaryotic genome replication depends on thousands of DNA replication origins (ORIs) that constitute the originome. A major challenge is to learn ORI biology in multicellular organisms in the context of growing organs to understand their developmental plasticity. We have determined the originome and chromatin landscape of Arabidopsis thaliana at two stages of postembryonic development. ORIs associate with multiple chromatin signatures including TSS but also regulatory regions and heterochromatin, where ORIs colocalize with retrotransposons. In addition, quantitative analysis of ORI activity led us to conclude that strong ORIs have high GC content and clusters of GGN trinucleotides. Development primarily influences ORI firing strength rather than ORI location. ORIs that preferentially fire at early developmental stages colocalize with GC-rich heterochromatin whereas at later stages with transcribed genes, perhaps as a consequence of changes in chromatin features associated with developmental processes. Our study provides the originome of an organism at the postembryo stage that should allow us to study ORI biology in response to development, environment and mutations with a quantitative approach. In a wider scope, the computational strategies developed here can be transferred to other eukaryotic systems.

plant biology↗

bioRxiv2016

Retrotransposons are specified as DNA replication origins in the gene-poor regions of Arabidopsis heterochromatin

Genomic stability depends on faithful genome replication. This is achieved by the concerted activity of thousands of DNA replication origins (ORIs) scattered throughout the genome. In spite of multiple efforts, the DNA and chromatin features that determine ORI specification are not presently known. We have generated a high-resolution genome-wide map of ORIs in cultured Arabidopsis thaliana cells that rendered a collection of 3230 ORIs. In this study we focused on defining the features associated with ORIs in heterochromatin. We found that while ORIs tend to colocalize with genes in euchromatic gene-rich regions, they frequently colocalize with transposable elements (TEs) in pericentromeric gene-poor domains. Interestingly, ORIs in TEs associate almost exclusively with retrotransposons, in particular, of the Gypsy family. ORI activity in retrotransposons occurs independently of TE expression and while maintaining high levels of H3K9me2 and H3K27me1, typical marks of repressed heterochromatin. ORI-TEs largely colocalize with chromatin signatures defining GC-rich heterochromatin. Importantly, TEs with active ORIs contain a local GC content higher than the TEs lacking them. Our results lead us to conclude that ORI colocalization with TEs is largely limited to retrotransposons, which are defined by their transposition mechanisms based on transcription, and they occur in a specific chromatin landscape. Our detailed analysis of ORIs responsible for heterochromatin replication has also implications on the mechanisms of ORI specification in other multicellular organisms in which retrotransposons are major components of heterochromatin as well as of the entire genome.

genomics↗