Biology subjects

Grenier, F.

Publications and source records attributed to Grenier, F..

11 recordsLinked to original sources

bioRxiv2026

Insights Into The Early Diversification of blaNDM-1 Genetic Contexts Following its Emergence in Acinetobacter species and Spread to Enterobacterales

Since its emergence, the New Delhi metallo-{beta}-lactamase (NDM) has become a major global antimicrobial resistance threat, enabling bacterial hosts to resist most {beta}-lactam antibiotics, including carbapenems, and spreading widely across bacterial species and geographic regions. Believed to have originated in Acinetobacter spp., blaNDM subsequently spread to Enterobacterales through mobile genetic elements, despite barriers to lateral gene transfer between these phylogenetically distant taxa. To investigate the genetic diversity and mechanisms underlying this interspecies transfer and subsequent global expansion, we sequenced and characterized 13 blaNDM-1-positive Gram-negative bacterial isolates collected in India in 2007, among the earliest known isolates harboring blaNDM-1. Clonal diversity was observed among both Escherichia coli and Klebsiella pneumoniae isolates. Comprehensive genomic analysis revealed that blaNDM-1 was consistently embedded within a conserved multidrug resistance region, often associated with mobile genetic elements involved in lateral gene transfer. Notably, the insertion sequence ISAba14 was identified in 6 of the 13 sequences, suggesting a potential role in the early dissemination of blaNDM-1 among diverse bacterial hosts. These findings contribute to our understanding of the evolutionary origins and early genetic context of blaNDM-1, highlighting the importance of continued genomic surveillance and functional studies to better understand the persistence and dissemination of carbapenem resistance.

molecular biology↗

bioRxiv2026

ARG-PASS v2.0 expands global resistome surveillance to reveal previously unrecognized antibiotic resistance genes reaching the human gut

Antimicrobial resistance (AMR) is a significant global public health threat, and antibiotic resistance genes (ARGs) that bacteria depend on to withstand treatment are its molecular basis. How ARGs move from environmental reservoirs into the human microbiome, and eventually into human pathogens, remains one of the most important unresolved questions in AMR research. Surveillance has largely targeted ARGs resembling those already clinically characterised. The environmental reservoir remains poorly described and the routes connecting it to human microbiomes are inferred rather than observed. Describing that reservoir requires computational methods able to recognise ARGs that share little similarity with known enzymes, yet retain accuracy after experimental validation to enable scalable applications. Dietary exposure through unprocessed ferments offers a tractable setting in which such a route can be observed directly. Here we introduce ARG-PASS v2.0 (ARG-prediction by PAirwise Sequence versus Structure v2.0) and first apply it to a lake metagenome from Kolkata, India, where 10 of 11 tested candidates across diverse ARG classes were functionally confirmed after expression in Escherichia coli. Given their clinical relevance, we then predicted 918 class B1 metallo-beta-lactamases (MBLs) from MGnify and the PLSDB, with most diverging from characterised enzymes, but as widely distributed. A number of globally widespread candidates track their hosts distribution, with some recovered only from human-associated biomes, yet most span a wider ecological range. Phylogenetic inference places clinically relevant class B1 MBLs within marine diversity and suggests a marine reservoir. Cross-referencing against the Unified Human Gastrointestinal Protein catalogue identified a carbapenemase, an ANT, and class A beta-lactamases from Bacillus sonorensis and Bacillus licheniformis, detected in South Korean fermented soybean paste (doenjang) as well as gut isolates of populations consuming these foods. Together, these results point to a largely chromosomal reservoir of undescribed ARG diversity that reaches the human gut through both resident and generalist hosts. Once in the human microbiome, treatment may be compromised directly if the host is an opportunistic pathogen, or after transfer to a clinical pathogen.

microbiology↗

bioRxiv2026

Optimization of DNA Transformation in Mesoplasma florum and Identification of a Candidate Recipient Strain for Genome Transplantation

Genome transplantation is a key technology for synthetic genomics, enabling entire genomes to be transferred into recipient cells. Despite its importance, genome transplantation remains confined to a small number of wall-less Mollicute species and is often characterized by low efficiencies, limiting the development and testing of synthetic genomes. Because genome transplantation relies on the same polyethylene glycol mediated DNA delivery process used for plasmid transformation, improving DNA uptake is an important step toward more efficient transplantation systems. Here, we systematically optimized polyethylene glycol-mediated transformation in Mesoplasma florum, a fast-growing, non-pathogenic Mollicute with a reduced genome that represents an attractive chassis for synthetic genomics. We evaluated 20 parameters spanning cell physiology, DNA preparation, membrane conditioning, and recovery conditions. DNA topology emerged as the strongest determinant of transformation efficiency, with highly compacted DNA preparations producing up to three orders of magnitude more transformants than conventional plasmid preparations. Growth phase and polyethylene glycol concentration also strongly influenced transformation outcomes. To identify potential recipients for future genome transplantation experiments, we further screened ten strains belonging to the Mesoplasma lineage. Transformation efficiencies varied widely among strains, and Mesoplasma entomophilum W17 emerged as a particularly promising candidate. Moreover, W17 supported replication of plasmids carrying the M. florum oriC, suggesting compatibility between the replication systems of the two species. Together, these results establish an improved transformation workflow for M. florum, identify DNA topology as a major determinant of DNA uptake, and reveal a new candidate recipient for future genome transplantation studies. These advances provide practical tools and biological insights for synthetic genome engineering in Mollicutes.

synthetic biology↗

bioRxiv2026

Historical isolates provide insights into the evolution of Acinetobacter baumannii international clone 2 and its resistome

Acinetobacter baumannii remains a pervasive nosocomial pathogen, with international clone 2 (IC2) being the most successful lineage. IC2 is often discussed as a singular entity, and the genomic factors behind its success over other lineages are poorly characterised. This study aimed to track genomic changes occurring around the emergence of IC2 and further determine the within-clone population structure. A set of historical A. baumannii isolates from the 1970s to early 2000s were long-read sequenced and assembled into high-quality chromosomes. These were then used to supplement publicly available A. baumannii genomes, producing a dataset of 1,281 chromosomes derived from isolates collected across six continents. A recombination-free phylogeny was produced and combined with antimicrobial resistance gene (ARG) presence/absence data. Analysis showed that IC2 began to emerge as the dominant lineage around 2005, which coincides with the acquisition of oxa23 and AbGRI3. Further, IC2 can be split into at least four distinct groups. Groups 1, 2, and 3 represent incremental evolution of the A. baumannii chromosome over time, while group 4 represents a separate evolutionary path distinct from the main IC2 lineage, which has recently been isolated at higher frequency. Significance statementCharacterising the evolution and population structure of antimicrobial-resistant bacterial pathogens is of key importance both for epidemiological monitoring and developing approaches to combat infections and their spread. Here, we use a unique collection of historical isolates to show that the major international clone in the WHO priority pathogen Acinetobacter baumannii is comprised of different, independent groups with individual antimicrobial resistance gene compliments. This highlights new areas for research to understand the clinical impact of the different groups and the drivers underlying the evolution and spread of epidemic clones of antimicrobial-resistance bacterial pathogens.

microbiology↗

bioRxiv2025

Conserved protein sequence-structure signatures identify emerging antibiotic resistance genes from the human microbiome

Bacteria exhibiting antimicrobial resistance (AMR) is a problem that has grown to become a significant public health challenge worldwide. Antibiotic resistance genes (ARGs), determinants of AMR, mostly emerge from non-clinical settings. Identifying previously undetected ARGs in the human microbiome which confer resistance to clinical concentrations of antibiotics is a crucial component of addressing AMR, yet can be hindered by their low homology to existing ARGs. Here, we attempt to address this by focussing on functionally important protein regions. ARG-PASS (ARG-PAirwise Sequence vs Structure) represents a novel protein function prediction method which leverages a one-class support vector machine trained on pairwise primary and tertiary distributions of structurally conserved regions of proteins encoded by ARGs. ARG-PASS was applied to six reference strains of the Human Microbiome Project. Nine candidates were selected for experimental verification and all were functionally confirmed when expressed in E.coli, belonging to ARG classes: APH, dfr, class B and C {beta}-lactamases, and penicillin binding proteins. We also used ARG-PASS directly on protein structures within the AlphaFold database and predicted a phnP gene (metallo-{beta}-lactamase fold), which is highly divergent from existing {beta}-lactamases and had activity against ampicillin. In total, 80% of the tested genes confer resistance at CLSI resistant breakpoints and the remainder represent pre-resistance genes, with activity but not at clinically relevant minimum inhibitory concentrations. We suggest pre-resistance genes may preferentially evolve into clinically relevant resistance determinants. ARG-PASS represents a novel and precise method of identifying previously uncharacterized ARGs from DNA databases, contributing to resistance surveillance and antibiotic stewardship.

microbiology↗

bioRxiv2023

Emergence of Acinetobacter baumannii International Clone 10 predominantly found in the Middle East

Acinetobacter baumannii is a globally distributed human pathogen. Infections caused by carbapenem-resistant isolates of A. baumannii (CRAB) are of great concern, as treatment options are very limited. Despite having among the highest rates reported worldwide, there exists limited genomic data from CRAB strains isolated in the Middle East. Here we report epidemiological, phenotypic, and genome sequencing data (short reads and long reads) on a set of 60 A. baumannii isolates belonging to Sequence Type ST158 (Pasteur MLST scheme). They represent a novel international clone (IC), designated IC10, with limited geographic spread beyond the Middle East. Specific antibiotic-resistance genes associated with this clone were identified and data on the plasmid content associated with this lineage are presented.

microbiology↗

bioRxiv2023

Earliest observation of the tetracycline destructase tet(X3)

Tigecycline is an antibiotic of last resort for infections with carbapenem-resistant Acinetobacter baumannii. Plasmids harboring variants of the tetracycline destructase tet(X) promote rising tigecycline resistance rates. We report the earliest observation of tet(X3) in a clinical strain predating tigecyclines commercialization, suggesting selective pressures other than tigecycline contributed to its emergence.

microbiology↗

bioRxiv2023

Adaptative Laboratory Evolution reveals biofilm regulating genes as key players in B. subtilis root colonization

Root-associated microorganisms play an important role in plant health, such as plant growth-promoting rhizobacteria from the Bacillus and Pseudomonas genera. Although bacterial consortia including these two genera would represent a promising avenue to efficient biofertilizer formulation, we observed that B. subtilis root colonization is decreased by the presence of P. fluorescens and P. protegens. To determine if B. subtilis can adapt to the inhibitory effect of Pseudomonas on roots, we conducted adaptative laboratory evolution experiments with B. subtilis in mono-association or co-cultured with P. fluorescens on tomato plant roots. Evolved isolates with various colony morphology and stronger colonization capacity of both tomato plant and A. thaliana roots emerged rapidly from the two evolution experiments. Certain evolved isolates had also a better fitness on root in presence of other Pseudomonas species. Whole genome sequencing revealed that single nucleotide polymorphism (SNPs) in negative biofilm regulator genes ywcC or sinR were found in all independent lineages, suggesting their involvement in enhanced root colonization. These findings provide insights into the molecular mechanisms underlying B. subtilis adaptation to root colonization and highlight the potential of directed evolution to enhance beneficial traits of PGPRs.

microbiology↗

bioRxiv2022

Molecular epidemiology of carbapenemase-producing Acinetobacter spp. from Israel, 2001-2006: earliest report of blaNDM predating the oldest known blaNDM-positive strains

BackgroundCarbapenem-resistant Acinetobacter baumannii (CRAb) is a WHO priority 1 critical pathogen. Despite early emergence of elevated CRAb rates in Israel, limited molecular data from this location are available. We searched for carbapenemases among 198 clinical Acinetobacter spp. from Israel between 2001 and 2006. MethodsStrains from 3 archives underwent whole-genome sequencing (Illumina NovaSeq on all, MinION on a subset) and computational analyses: assembly (Unicycler), annotation (prokka), identification (Kraken, rpoB similarity), search for carbapenemases (ResFinder, BLDB curation). FindingsA. baumannii (Ab) represented 179 (90{middle dot}4%) Acinetobacter spp. Eighty-four Ab (46{middle dot}9%) carried a carbapenemase: 38 (45{middle dot}2%) blaOXA-72 (blaOXA-24-like); 28 (33{middle dot}3%) blaOXA-23-like (20 blaOXA-23 and 8 blaOXA-225); 18 (21{middle dot}5%) blaOXA-58 (16 from 2001-2). Carbapenemase rates increased yearly from 2002 (32%) to 2006 (67%). Eight species of non-baumannii Acinetobacter (NbA) accounted for 19 isolates (9{middle dot}6%). Two of three A. junii contained blaOXA-58, one of which, Ajun-H1-3, isolated in January 2004, also possessed blaNDM-1. The pNDM-Ajun-H1-3 plasmid matched numerous NDM-positive plasmids reported from 2005 onwards in Acinetobacter spp. as well as Enterobacterales. InterpretationWe assessed carbapenemase diversity among Acinetobacter spp. in Israel from 2001-2006. Findings in Ab predate observations elsewhere: rapidly rising carbapenemase rates, driven by blaOXA-23-like and blaOXA-24-like genes replacing blaOXA-58. Among NbA, an A. junii isolated in 2004 carried blaNDM-1, making it the earliest NDM-positive isolate reported to date, preceding those from 2005 in India. Further research into blaNDMs emergence is warranted, in order to shed light on the evolution and spread of this and other antibiotic-resistance genes. FundingCentre de recherche Charles-Le Moyne; Department of Microbiology and Infectious Diseases, Faculty of Medicine and Health Sciences, Universite de Sherbrooke; Fonds de recherche du Quebec - Sante; New Frontiers in Research Fund Grant NFRFE-2019-00444; CIFAR-Azrieli Global Scholars Program.

microbiology↗

bioRxiv2021

PixR, a novel activator of conjugative transfer of IncX4 resistance plasmids, mitigates the fitness cost of mcr-1 carriage in Escherichia coli

The emergence of the plasmid-borne colistin resistance gene mcr-1 threats public health. IncX4-type plasmids are one of the most epidemiologically successful vehicles for spreading mcr-1 worldwide. Since MCR-1 is known for imposing a fitness cost to its host bacterium, the successful spread of mcr-1-bearing plasmids might be linked to high conjugation frequency, which would enhance the maintenance of the plasmid in the host without antibiotic selection. However, the mechanism of IncX4 plasmids conjugation remains unclear. In this study, we used high-density transposon mutagenesis to identify factors required for IncX4 plasmid transfer and 18 genes were identified, including five with annotations unrelated to conjugation. The Cappable-seq and RNA-seq analysis confirmed that a novel transcriptional regulator gene, pixR, directly regulates the transfer of IncX4 plasmids by binding the promoter of 13 essential transfer genes to increase their transcription. Plasmid invasion and co-culture competition assays revealed that pixR is essential for the spread and persistence of mcr-1-bearing IncX4 plasmids in bacterial populations, and effective conjugation is crucial for alleviating the fitness cost exerted by mcr-1 carriage. The existence of the IncX4-specific pixR gene increases plasmid transmissibility while promoting the invasion and persistence of mcr-1-bearing plasmids in bacterial populations, which helps explain their global prevalence. IMPORTANCEThe spread of clinical important antibiotic resistance genes is frequently related to some epidemic plasmids. However, the underlying molecular mechanisms contributing to the successful spread of these epidemic plasmids remains unclear. The significant of our research indicated that efficient conjugation could promote the invasion and persistence of plasmids within a bacterial population, resulting in the successful dissemination of epidemic plasmids in nature. Our data also highlight the importance of developing plasmid conjugation inhibitors to solve the antibiotic resistance crisis.

microbiology↗

bioRxiv2019

Sen1 is required for RNA Polymerase III transcription termination in an R-loop independent manner.

R-loop disassembly by the human helicase Senataxin contributes to genome stability and to proper transcription termination at a subset of RNA polymerase II genes. Whether Senataxin-mediated R-loop disassembly also contributes to transcription termination at other classes of genes has remained unclear. Here we show in fission yeast that SenataxinSen1 promotes efficient termination of RNA Polymerase III (RNAP3) transcription in vivo. In the absence of SenataxinSen1, RNAP3 accumulates downstream of the primary terminator at RNAP3-transcribed genes and produces long exosome-sensitive 3-extended transcripts. Importantly, neither of these defects was affected by the removal of R-loops. The finding that SenataxinSen1 acts as an ancillary factor for RNAP3 transcription termination in vivo challenges the pre-existing view that RNAP3 terminates transcription autonomously. We propose that Senataxin is a cofactor for transcription termination that has been co-opted by different RNA polymerases in the course of evolution.

molecular biology↗