Biology subjects

Torrents, E.

Publications and source records attributed to Torrents, E..

7 recordsLinked to original sources

bioRxiv2025

Burkholderia cenocepacia and Pseudomonas aeruginosa coinfection alters antimicrobial tolerance, infection dynamics and host immune effectiveness

Polymicrobial infections promote the appearance of a network of interactions that can lead to an increase in their antimicrobial tolerance or to the evasion of the host immune system. Pseudomonas aeruginosa and Burkholderia cenocepacia are two multidrug-resistant opportunistic pathogens that significantly influence host health and alter antibiotic responses when coinfected. To characterize host-pathogen dynamics, we examined infection progression, immune responses, bacterial virulence gene expression, global transcriptomic analysis, and antibiotic susceptibility in single and coinfections involving acute and chronic P. aeruginosa strains combined with B. cenocepacia. This work was performed in vivo using G. mellonella larvae as a model. Larval survival and bacterial dissemination were monitored, revealing tissue-specific patterns of infection. Our findings indicated that coinfections increased larval lethality and worsened overall host health. Notably, B. cenocepacia suppressed host melanization and immune responses, while P. aeruginosa triggered a strong immune activation. Coinfection also induced upregulation of virulence genes in both pathogens. This study advances understanding of host-pathogen interactions in polymicrobial infections and highlights the need for improved therapeutic strategies.

microbiology↗

bioRxiv2025

FleQ-Dependent Regulation of the Ribonucleotide Reductase Repressor nrdR in Pseudomonas aeruginosa During Biofilm Growth and Infection

Ribonucleotide reductases (RNRs) are essential enzymes involved in DNA synthesis and repair, catalyzing the conversion of ribonucleotides to deoxyribonucleotides (dNTPs). While all living cells possess at least one RNR encoded in their genome, certain organisms, such as Pseudomonas aeruginosa, encode multiple RNR classes. This multiplicity provides a competitive advantage, allowing these organisms to adapt and colonize different environments. Despite their importance, the mechanisms coordinating the expression of different RNRs in microorganisms with multiple RNR classes remain poorly understood. The transcriptional regulator NrdR has been implicated in controlling the expression of all three RNR classes in P. aeruginosa by binding to conserved NrdR boxes within the promoter regions of the RNR genes. To gain insights into the regulation of the different RNR genes, it is first necessary to understand how nrdR itself is transcriptionally regulated. In this study, we employed a bioinformatics approach to identify potential transcription factors (TF) involved in nrdR regulation. We combined this with promoter-probe vectors nrdR promoter fusions to investigate nrdR transcriptional regulation and identify TFs that modulate its expression in vitro. Our analysis identified four potential TF that could regulate nrdR, and we experimentally confirmed that FleQ is responsible for regulating nrdR expression under aerobic and anaerobic conditions. Furthermore, we explored nrdR regulation under biofilm-forming conditions and in the Galleria mellonella infection model to gain insights into how nrdR might be regulated in vivo. ImportanceThis study reveals a nuanced regulatory mechanism by which the transcription factor FleQ, modulated by intracellular c-di-GMP levels, governs the expression of the essential gene nrdR in Pseudomonas aeruginosa. By demonstrating that FleQ acts as an activator under planktonic and infection conditions and as a repressor during biofilm formation, the findings reveal a dual regulatory role that aligns with the bacteriums transition between acute and chronic infection states. This dynamic control of nrdR, a key repressor of ribonucleotide reductases, links environmental sensing to nucleotide metabolism, offering new insights into how P. aeruginosa adapts to diverse and hostile environments. These results not only deepen our understanding of bacterial gene regulation but also highlight potential targets for disrupting biofilm-associated persistence in clinical settings.

microbiology↗

bioRxiv2025

Structural basis and mechanism of action of NrdR, a bacterial master regulator of ribonucleotide reduction

Ribonucleotide reductases (RNRs) are the essential enzymes responsible for synthesizing dNTPs, the building blocks of DNA. In bacteria, the entire RNR network is controlled by the master regulator NrdR. As a regulator of an essential pathway with no eukaryotic equivalent, NrdR is a promising antimicrobial target. Recent studies have outlined a mechanism of action for NrdR, in which ATP and dATP induce changes in the protein quaternary structure, regulating RNR repression. However, due to a lack of functional studies linking the known structures to their biological roles, the role of NrdR is not yet fully understood. Here, we conducted a comprehensive study of NrdR in Escherichia coli and Pseudomonas aeruginosa. We delimited the NrdR regulon, combining transcriptomics and motif-based sequence analysis. We crystallized NrdR and obtained structural data on nucleotide binding and its relation to the stability of protein-protein interfaces involved in NrdR oligomerization. We examined the quaternary structures of NrdR using SEC-MALS and atomic force microscopy and correlated structure to function using point mutations, EMSAs, and in vitro transcription assays. Overall, our results demonstrate the mechanism used by NrdR to modulate its quaternary structure and activity and provide structural data that pave the way for targeted antimicrobial therapies.

molecular biology↗

bioRxiv2024

Candida albicans enhance Staphylococcus aureus virulence by progressive generation of new phenotypes

Candida albicans and Staphylococcus aureus have been co-isolated from several biofilm- associated diseases, including those related to medical devices. This association confers advantages to both microorganisms, resulting in detrimental effects on the host. To elucidate this phenomenon, the present study investigated colony changes derived from non-physical interactions between C. albicans and S. aureus. We performed proximity assays by confronting colonies of the yeast and the bacteria on agar plates at six different distances for 9 days. We found that colony variants of S. aureus originated progressively after prolonged exposure to C. albicans proximity, specifically in response to pH neutralization of the media by the fungi. The new phenotypes of S. aureus were more virulent in a Galleria mellonella larvae model compared to colonies grown without C. albicans influence. This event was associated with an upregulation of RNAIII and AgrA expression, suggesting a role for -toxin. Our findings indicate that C. albicans enhances S. aureus virulence by inducing the formation of more aggressive colonies. ImportanceFor decades, it has been known that C. albicans increase S. aureus virulence, resulting in a "lethal synergism". However, it was only recently identified that this outcome is driven by the sustained activation of the staphylococcal agr system in response to C. albicans environmental modifications. Our experimental design allowed us to observe individual changes over time caused by the proximity of both microorganisms. As a result, we report for first time that C. albicans exposure induces the generation and favors the growth of S. aureus colony variants with increased expression of virulence factors. Our findings highlight the importance to understanding the intricate connection between environmental responses, virulence and fitness in S. aureus pathogenesis.

microbiology↗

bioRxiv2024

Antimicrobial and antibiofilm activity of human recombinant H1 histones against bacterial infections

Histones possess significant antimicrobial potential, yet their activity against biofilms remains underexplored. Moreover, concerns regarding adverse effects limit their clinical implementation. We investigated the antibacterial efficacy of human recombinant histone H1 subtypes against Pseudomonas aeruginosa PAO1, both planktonic and in biofilms. After the in vitro tests, toxicity and efficacy were assessed in a P. aeruginosa PAO1 infection model using Galleria mellonella larvae. Histones were also evaluated in combination with ciprofloxacin and gentamicin. Our results demonstrate antimicrobial activity against of all three histones against P. aeruginosa PAO1, with H1.0 and H1.4 showing efficacy at lower concentrations. The bactericidal effect was associated with a mechanism of membrane disruption. In vitro studies using static and dynamic models showed that H1.4 had antibiofilm potential by reducing cell biomass. Neither H1.0 nor H1.4 showed toxicity in G. mellonella larvae, and both increased larvae survival when infected with P. aeruginosa PAO1. Although in vitro synergism was observed between ciprofloxacin and H1.0, no improvement over the antibiotic alone was noted in vivo. Differences in antibacterial and antibiofilm activity were attributed to sequence and structural variations among histone subtypes. Moreover, the efficacy of H1.0 and H1.4 was influenced by the presence and strength of the extracellular matrix. These findings suggest histones hold promise for combating acute and chronic infections caused by pathogens such as P. aeruginosa. ImportanceThe constant increase of multidrug-resistant bacteria is a critical global concern. The inefficacy of current therapies to treat bacterial infections is attributed to multiple mechanisms of resistance, including the capacity to form biofilms. Therefore, the identification of novel and safe therapeutic strategies is imperative. This study confirms the antimicrobial potential of three histone H1 subtypes against both Gram-negative and Gram-positive bacteria. Furthermore, histones H1.0 and H1.4 demonstrated in vivo efficacy without associated toxicity in an acute infection model of Pseudomonas aeruginosa PAO1 in Galleria mellonella larvae. The bactericidal effect of these proteins also resulted in reduction in biomass of P. aeruginosa PAO1 biofilms. Given the clinical significance of this opportunistic pathogen, our research provides a comprehensive initial evaluation of the efficacy, toxicity, and mechanism of action of a potential new therapeutic approach against acute and chronic bacterial infections.

microbiology↗

bioRxiv2024

Interplay of Mycobacterium abscessus and Pseudomonas aeruginosa in coinfection: Biofilm Dynamics and Host Immune Response

The incidence of infection by nontuberculous mycobacteria, mainly Mycobacterium abscessus, in patients with cystic fibrosis and other chronic pulmonary illnesses is increasing, translating into an acceleration in the decline of lung function. In most cases, M. abscessus coinfects with Pseudomonas aeruginosa, the most common pathogen in these chronic diseases. However, it is unknown how these two bacterial species interact when coinfecting. This study aims to explore the behavior of both species in three relevant pathogenic settings: dual-species biofilm development using a recently developed method to monitor individual species in dual-species biofilms; coinfection in bronchial epithelial cells using in vitro assays; and in vivo coinfection using the Galleria mellonella model. The results demonstrate the capability of both species to form stable mixed biofilms and to reciprocally inhibit single-biofilm progression. Coinfections in bronchial epithelial cells were correlated with significantly decreased cell viability, while in G. mellonella, coinfections induced lower survival rates than individual infections. Outstandingly, the analysis of the immune response triggered by each bacterium in bronchial epithelial cell assays and G. mellonella larvae revealed that P. aeruginosa induces the overexpression of proinflammatory and melanization cascade responses, respectively. In contrast, M. abscessus and P. aeruginosa coinfection significantly inhibited the immune response in both models, resulting in worse consequences for the host than those generated by single P. aeruginosa infection. Overall, the presence of M. abscessus produces a decline in the immune responses that worsens the infection and compromises the host. ImportanceThe appearance of bacterial infections in the respiratory tract of patients with chronic respiratory diseases suppose a serious and difficult to treat health problem. This complication is exacerbated by the increase resistance against antibiotics generated by pathogenic microorganisms. The most common and virulent pathogenic bacteria reported in the respiratory airway is Pseudomonas aeruginosa. It is a Gram-negative, ubiquitous, and intrinsic resistant to antibiotics bacteria. However, the incidence of a rapidly growing, multi-drug resistant mycobacteria; Mycobacterium abscessus, is growing worldwide. The pulmonary coinfection by both pathogens is directly related with higher rates of morbidity and mortality of patients. The significance of our research is characterizing the behavior of these two pathogens when they coinfects together, exploring the immune response triggered by the host and its impact in the survival. The purpose is enhancing the limited understanding we have of this clinically relevant coinfection to favor the development of new effective treatments.

microbiology↗

bioRxiv2022

Genomic analysis of Mycobacterium brumae sustains its nonpathogenic and immunogenic phenotype

M. brumae is a rapid-growing, non-pathogenic Mycobacterium species, originally isolated from environmental and human samples in Barcelona, Spain. M. brumae is not pathogenic and its in vitro phenotype and immunogenic properties have been well characterized. However, the knowledge of its underlying genetic composition is still incomplete. In this study, we first describe the 4 Mb genome of the M. brumae type strain ATCC 51384T assembling PacBio reads, and second, we assess the low intraspecies variability by comparing the type strain with Illumina reads from three additional strains. M. brumae genome is composed of a circular chromosome with a high GC content of 69.2 % and containing 3,791 CDSs, 97 pseudogenes, one prophage and no CRISPR loci. M. brumae has shown no pathogenic potential in in vivo experiments, and our genomic analysis confirms its phylogenetic position with other non-pathogenic and rapid growing mycobacteria. Accordingly, we determined the absence of virulence related genes, such as ESX-1 locus and most PE/PPE genes, among others. Although immunogenic potential of M. brumae was proved to be as high as Mycobacterium bovis BCG, the only mycobacteria licensed to treat cancer, the genomic content of M. tuberculosis T cell and B cell antigens in M. brumae is considerably lower than those of M. bovis BCG. Overall, this work provides relevant genomic data on one of the species of the mycobacterial genus with high therapeutic potential.

genetics↗