Biology subjects

Ploy, M.-C.

Publications and source records attributed to Ploy, M.-C..

9 recordsLinked to original sources

bioRxiv2025

The bacterial SOS response promotes the expression of the transposase encoded by ISCR mobile genetic elements

Insertion sequences (IS) are widely involved in bacterial genomic plasticity by disrupting, adding, moving genomic sequences, or by activating or extinguishing gene expression. A specific family of IS, ISCR (for insertion sequence of Common Region), is thought to be involved in the dissemination of antibiotic resistance genes (ARG). While some ISCR members are commonly found in bacteria isolated in clinical settings and can contribute to downstream ARG expression, the mechanisms regulating the ISCR-encoded transposases expression have remained uncharacterized. Here, we investigated the expression of the transposase gene of ISCR1, ISCR2 and ISCR8, and its regulation in Escherichia coli. Using in silico analyses and in vitro experiments, we showed that expression levels were extremely low, as observed for most IS transposases. We further demonstrated the direct role of DNA damages and the key SOS response repressor, LexA, in controlling the activity of transposase promoter. These results provide evidence that the mobility of at least some ISCR elements may be promoted upon bacterial exposure to antibiotics inducing the SOS response. IMPORTANCEMobile genetic elements are the most prevalent cause of antibiotic resistance emergence. Among these mobile elements, insertion sequences (IS) are well known to allow the dissemination of antibiotic resistance genes through the action of their tranposase. Here, we studied the regulation of the transposase expression in a specific family of IS, the ISCR family, some members of which are known to be involved in antibiotic resistance. Characterizing the regulation of transposase expression is an important starting point for understanding how these IS can contribute, through their movement, to the spread and expression of antibiotic resistance. This knowledge is necessary if we are to hope to prevent this spread one day.

microbiology↗

bioRxiv2025

Dual regulatory role of IS91-encoded Orf121 in IS91 transposition

Prokaryotic insertion sequences (IS) are pivotal in the propagation of bacterial multidrug resistance, with IS91 notably linked to virulence and antibiotic resistance genes. However, the precise mechanism by which IS91 contributes to gene dissemination remains elusive. Unique among its family, IS91 features a small open reading frame (orf) upstream of the tnpA transposase gene, potentially encoding a 121-amino acid protein, orf121, which may be translationally coupled with tnpA. Using a genetic system based on the mating-out assay in Escherichia coli, we explored the role of orf121 in the in vivo transposition of IS91. Our findings indicate that the overlap between orf121 and tnpA is crucial for tnpA transcription, with both being primarily transcribed as bicistronic mRNAs from the Porf121 promoter. Additionally, the expression of orf121 (whether in cis or trans) significantly reduces the frequency of IS91 transposition and the rate of one-ended transposition. Furthermore, only the single-stranded DNA circles of IS91 intermediates can integrate into new target sequences, and Orf121 negatively influences this insertion step. In summary, orf121 acts as a negative regulator of transposition while ensuring the expression of tnpA, thus providing insights into the complex mechanisms underlying IS91-mediated gene dissemination and its potential role in antibiotic resistance propagation.

molecular biology↗

bioRxiv2023

Hospital and urban wastewaters shape the structure and active resistome of environmental biofilms

BackgroundDemonstration of the transfer, dynamics, and regulation of antibiotic resistance genes (ARGs) and mobile genetic elements (MGEs) in a complex environmental matrix is yet experimentally challenging, with many essential open questions such as how and where transfer and dissemination of ARGs happens in nature. The extent and conditions of MGEs transfer that carry ARGs is still largely unexplored in natural environments and microbial communities. Biofilms are structures that include high density multi-species bacterial communities embedded in self-produced extracellular polymeric substances (EPS) constituting a matrix that facilitates gene transfer and where bacteria exhibit high tolerance to stress and to antibiotics. In this study we implemented a sampling and analysis approach that allows phenotypic and genomic analyses of in situ and reconstituted in vitro hospital and urban wastewater (WW) biofilms. To assess the potential of hospital and urban WW biofilms to efficiently disseminate ARGs in the WW system, we explored the EPS within the biofilm matrix and assessed the expression of the resistome (ARGs) and mobilome (MGEs) by metatranscriptomics. ResultsWe first showed that a) the composition of EPS differs depending on their growth environment (in situ and in vitro) and their sampling origin (hospital vs urban WW) and that b) a low amount of ciprofloxacin impacted the composition of the EPS. Next, the metatranscriptomic approach showed that a) expression of ARGs and MGEs increase upon adding a low amount of ciprofloxacin for biofilms from hospital WW but not for those from urban WW and b) that expression of specific plasmids that carry individual or multiple ARGs varies depending on the WW origins of the biofilms. When the same plasmids were expressed in both, urban and hospital WW biofilms, they carried and expressed different ARGs. ConclusionWe show that hospital and urban wastewaters shape the structure and active resistome of environmental biofilms, and we confirmed that hospital WW is an important hot spot for the dissemination and selection of AMR. The different responses to antibiotic pressure in hospital vs urban biofilms, coupled with differences in biofilm structure helps delineate distinct characteristics of hospital and urban WW biofilms highlighting the relationships between the resistome and its expression in environmental biofilms and their surrounding ecosystems.

microbiology↗

bioRxiv2021

Modelling the impact of urban and hospital wastewaters eco-exposomes on the antibiotic-resistance dynamics.

Antibiotic-resistance emergence and selection have become major public health issues globally. The presence of antibiotic resistant bacteria (ARB) in natural and anthroposophical environments threatens to compromise the sustainability of care in human and animal populations. This study was undertaken to develop a simple model formalizing the selective impact of antibiotics and pollutants on the dynamics of bacterial resistance in water and use the model to analyze longitudinal spatiotemporal data collected in hospital and urban wastewaters. Longitudinal-sampling data were collected between 2012 and 2015 in four different locations in Haute-Savoie, France: hospital and urban wastewaters, before and after water-treatment plants. Concentration in three different types of compounds: 1) heavy metals 2) antibiotics and 3) surfactants; and abundance of 88 individual genes and mobile genetic elements, mostly conferring resistance to antibiotics, were simultaneously collected. A simple hypothesis-driven model describing the weekly ARB dynamics was proposed to fit available data by assuming normalized gene abundance to be proportional to ARB populations in water. Compounds impacts on the dynamics of 17 genes found in multiple sites were estimated. We found that while mercury and vancomycin had relevant effects on ARB dynamics, respectively positively affecting the dynamics of 10 and 12 identified genes, surfactants antagonistically affected genes dynamics (identified for three genes). This simple model enables analyzing the relationship between resistance-gene persistence in aquatic environments and specific compounds inherent to human activities. Applying our model to longitudinal data, we identified compounds that act as co-selectors for antibiotic resistance. HighlightsO_LIWe analyzed longitudinal wastewater resistance genes and environmental data C_LIO_LIWe developed a simple hypothesis-driven model to assess resistance selection C_LIO_LIMercury and vancomycin were key drivers of antibiotic resistance in wastewater C_LI

microbiology↗

bioRxiv2021

Seasonal resistome diversity and dissemination of WHO priority antibiotic-resistant pathogens in Lebanese estuaries

Anthropogenic activities are demonstrated to be the key drivers of antimicrobial resistance (AMR) dissemination in the environment. Lebanese rivers that lead to the Mediterranean Sea were sampled at estuaries sites, under high anthropogenic pressure, in spring 2017 and winter 2018 to study seasonal variation of antimicrobial-resistant bacteria (ARBs) and antimicrobial resistance genes (ARGs). Methods: A combined approach using culture techniques and high throughput qPCR identified hotspots for antimicrobial resistance and anthropogenic pressure in particular locations along the Lebanese coast. Results: Multi-resistant Gram-negative (Enterobacterales and Pseudomonas spp) and Gram-positive bacterial pathogens were isolated. A high abundance of certain ARGs (vanB, blaBIC-1, blaGES, tetM, and mcr-1) was detected in 5 Lebanese estuaries. The relative abundance of ARGs was highest in winter and areas with high anthropogenic activities and population growth with an influx of refugees. Conclusion: Qualitative analysis of ARB and the analysis of the Lebanese estuaries resistome revealed critical levels of contamination with pathogenic bacteria and provided significant information about the spread of ARGs in anthropogenically impacted estuaries.

microbiology↗

bioRxiv2021

Microniches in biofilm depth are hot-spots for antibiotic resistance acquisition in response to in situ stress

Class 1 integrons play a major role in antibiotic resistance dissemination among Gram-negative bacteria. They are genetic platforms able to capture, exchange and express antibiotic resistance gene cassettes. The integron integrase, whose expression is regulated by the bacterial SOS response, is the key element of the integron catalyzing insertion/excision/shuffling of gene cassettes. We previously demonstrated that the basal level of integrase expression and in consequence, its activity, is increased via the starvation-induced stringent response in the biofilm population. However, biofilms are heterogeneous environments where bacteria are under various physiological states. Here we thus analyzed at the bacterial level, the SOS response and integrase expression within the biofilm, using confocal microscopy and flow cytometry. We showed that in the absence of exogenous stress, only a small number of bacteria (~ 1%) located in the depth of the biofilm induce the SOS-response leading to a high level of integrase expression, through both a stringent response-dependent and -independent manner. Our results thus indicate that few bacteria located in microniches of the biofilm depth undergo sufficient endogenous stress to promote the acquisition of antibiotic resistance, forming a reservoir of bacteria ready to rapidly resist antibiotic treatments.

microbiology↗

bioRxiv2021

A qnr-plasmid allows aminoglycosides to induce SOS in Escherichia coli

The plasmid-mediated quinolone resistance (PMQR) genes have been shown to promote high-level bacterial resistance to fluoroquinolone antibiotics, potentially leading to clinical treatment failures. In Escherichia coli, sub-inhibitory concentrations (sub-MIC) of the widely used fluoroquinolones are known to induce the SOS response. Interestingly, the expression of several PMQR qnr genes is controlled by the SOS master regulator. During the characterization of a small qnrD-plasmid carried in E. coli, we observed that the aminoglycosides become able to induce the SOS response in this species, thus leading to the transcription of qnrD. We found that induction of the SOS response is due to nitric oxide (NO) accumulation in presence of sub-MIC of aminoglycosides. We demonstrated that the NO accumulation is driven by two plasmid genes, ORF3 and ORF4, whose products act at two levels. ORF3 encode a FAD-binding oxidoreductase which helps NO synthesis, while ORF4 code for an FNR-type transcription factor, related to an O2-responsive regulator of hmp expression, able to repress the Hmp-mediated NO detoxification pathway of E. coli. Thus, this discovery, that other major classes of antibiotics may induce the SOS response could have worthwhile implications for antibiotic stewardship efforts in preventing the emergence of resistance.

microbiology↗

bioRxiv2019

Classification of hospital and urban wastewater resistome and microbiota over time and their relationship to the eco-exposome.

Wastewaters (WW) are important sources for the dissemination of antimicrobial resistance (AMR) into the environment. Hospital WW (HWW) contain higher loads of micro-pollutants and AMR markers than urban WW (UWW). Little is known about the long-term dynamics of H and U WW and the impact of their joined treatment on the general burden of AMR. Here, we characterized the resistome, microbiota and eco-exposome signature of 126 H and U WW samples treated separately for three years, and then mixed, over one year. Multi-variate analysis and machine learning revealed a robust signature for each WW with no significant variation over time before mixing, and once mixed, both WW closely resembled U signatures. We demonstrated a significant impact of pharmaceuticals and surfactants on the resistome and microbiota of H and U WW. Our results present considerable targets for AMR related risk assessment of WW.

microbiology↗

bioRxiv2018

Novel Staphylococcal Cassette Chromosome composite island (SCC-CI) with a new subtype of SCCmecVI cassette found in ST5 MRSA in France

An emergent kanamycin-susceptible ST5-MRSA lineage has been identified in France. Whole genome sequencing revealed a 40 kb SCC composite island with a mosaic structure including 3 SCC elements: a {Psi}SCCcop/ars, a SCCLim88A with a ccrC recombinase, and a novel subtype of SCCmec type VI (VIb). This mosaic structure suggests a high recombination rate of SCC elements from distinct staphylococci species.

microbiology↗