Biology subjects

Rossez, Y.

Publications and source records attributed to Rossez, Y..

10 recordsLinked to original sources

bioRxiv2026

Delayed onset and heterogeneous collective organization characterize twitching motility in Acinetobacter baumannii

Type IV pili (T4P) mediate twitching motility and contribute to surface colonization, biofilm formation, and host interactions in Acinetobacter baumannii. However, the prevalence, dynamics, and diversity of twitching motility across A. baumannii populations remain poorly understood. Here, we compared twitching motility in a collection of 35 A. baumannii strains originating from clinical, environmental, and animal sources, using Pseudomonas aeruginosa PAO1 as a reference. Standardization of assay conditions revealed a strong influence of agar composition on twitching motility, with Eiken agar supporting the most robust surface translocation. Under these conditions, 14 of 35 A. baumannii isolates exhibited detectable twitching motility. Time-lapse microscopy revealed major differences between A. baumannii and P. aeruginosa. Whereas PAO1 initiated twitching within minutes after inoculation and formed characteristic multicellular rafts, motile A. baumannii strains displayed a prolonged non-motile phase before movement initiation and exhibited distinct patterns of collective organization. Two major expansion phenotypes were identified, termed Homogeneous Front (HF) and Raft-Like Front (RLF), together with Early-Onset Motility (EOM) and Delayed-Onset Motility (DOM) subgroups. Quantitative analyses further revealed substantial variation in speed, directional persistence, and migration dynamics among strains. Because a majority of isolates were non-motile, we investigated the contribution of the minor pilin FimT. Although deletion of fimT abolished twitching motility and specific substitutions modulated motility efficiency, sequence variation in FimT alone could not account for the observed phenotypic diversity. Collectively, these findings reveal extensive heterogeneity in T4P-mediated surface motility in A. baumannii and identify delayed twitching activation and distinct collective migration strategies as key features of surface colonization in this species. HighlightsO_LIPopulation analysis reveals extensive twitching diversity in A. baumannii C_LIO_LIDelayed motility onset distinguishes A. baumannii from P. aeruginosa C_LIO_LIHomogeneous and raft-like fronts define distinct migration strategies C_LIO_LIFimT contributes to motility but not all non-motile phenotypes C_LI

microbiology↗

bioRxiv2025

How flagella glycosylation of the phytopathogenic bacteria Pseudomonas amygdali pv. tabaci 6605 affect transport and deposition in saturated sandy porous media

To mitigate bacterial contamination in underground farmland, a comprehensive understanding of the transport and adhesion mechanisms of phytopathogenic bacteria in porous media is crucial for safeguarding soil and groundwater. This study aims to elucidate the effects of Pseudomonas amygdali pv. tabaci 6605 flagella (Wild type, {Delta}fliC strain) and its glycosylation ({Delta}fgt1 and {Delta}fgt2 strains) on bacterial transport and deposition in sandy porous media through a combination of experimental observations and numerical simulations. Flagella play a key role in bacterial transport and deposition dynamics through its surface properties. Its intrinsic hydrophobicity enhances bacterial adhesion and promotes deposition onto sandy grains, while simultaneously limiting transport through the porous medium. However, glycosylation of flagellin introduces hydrophilic glycans, which counteract this effect by increasing the overall hydrophilicity of the bacterial surface. As a result, glycosylated flagella facilitate bacterial mobility and improves recovery in the effluent, while reducing retention within the sand matrix. These findings highlight the critical influence of flagellas biochemical modifications on bacterial behavior in porous environments. They provide valuable insights for understanding and managing microbial contamination in subsurface systems. ImportanceThis work, conducted using homogeneous laboratory sand, could be extended to other types of abiotic media found in natural environments, such as clay, heterogeneous sands, and soils. Our study highlights the impact of flagellar glycosylation on bacterial behavior, an essential factor for assessing the risk posed by phytopathogenic bacteria in agricultural settings and for developing effective soil bioremediation strategies. Moreover, this study provided valuable insights into the mechanisms governing bacterial transport and deposition at the macroscopic (column) scale under dynamic flow conditions. Investigating unsaturated flow conditions, which better approximate real field scenarios, may further our understanding of bacterial interactions at air/solid/water interfaces. Future research should explore bacterial movement across different spatial scales. In particular, pore-scale experiments can provide direct evidence of processes such as attachment and motility. This could significantly enhance our understanding of microbial dynamics in complex environments.

microbiology↗

bioRxiv2024

Biochemical characterization of the Escherichia coli surfaceome: A focus on type I fimbriae and flagella.

The Escherichia coli surfaceome consists mainly of the large surface organelles expressed by the organism to navigate and interact with the surrounding environment. The current study focuses on type I fimbriae and flagella. These large polymeric surface organelles are composed of hundreds to thousands of subunits, with their large size often preventing them from being studied in their native form. Recent studies are accumulating which demonstrate the glycosylation of surface proteins or virulence factors in pathogens, including E. coli. Using biochemical and glycobiological techniques, including biotin-hydrazide labelling of glycans and chemical and glycosidase treatments, we demonstrate i) the presence of a well-defined and chemically resistant FimA oligomer in several strains of pathogenic and non-pathogenic E. coli, ii) the major subunit of type I fimbriae, FimA, in pathogenic and laboratory strains is recognized by concanavalin A, iii) standard methods to remove N-glycans (PNGase F) or a broad-specificity mannosidase fail to remove the glycan structure, despite the treatments resulting in altered migration in SDS-PAGE, iv) PNGase F treatment results in a novel 32 kDa band recognized by anti-FliC antiserum. While the exact identity of the glycan(s) and their site of attachment currently elude detection by conventional glycomics/glycoproteomics, the current findings highlight a potential additional layer of complexity of the surface (glyco)proteome of the commensal or adhesive and invasive E. coli strains studied.

microbiology↗

bioRxiv2023

Unraveling the Interplay of Temperature Adaptation, Lipidomics, and Environmental Factors in Acinetobacter baumannii Clinical Strains.

Acinetobacter baumannii has gained prominence due to its heightened antibiotic resistance and adaptability within healthcare settings. Unlike other Acinetobacter species, A. baumannii predominantly thrives within healthcare environments, where its persistence is underscored by physiological adaptations, including homeoviscous adaptation that modifies glycerophospholipids (GPL) to enhance membrane flexibility. The bacteriums substantial genetic diversity highlights the paramount importance of prudent strain selection for research involving drug resistance and virulence. This study investigates the lipid composition of six clinical A. baumannii strains, incorporating the highly virulent model strain AB5075 with multiple antibiotic resistances. Our objective is to scrutinize the adaptations of glycerophospholipids (GPL) and glycerolipids (GL) within these isolated strains, each characterized by unique antibiotic resistance profiles, under variable temperature conditions mimicking environmental and physiological scenarios. The strains differential performance in motilities and biofilm formation across varying temperatures reveals intriguing patterns. Notably, the study uncovers a consistent elevation in palmitoleic acid (C16:1) content in five of the six strains at 18{degrees}C. Utilizing LC-HRMS2 analysis, we elucidate shifts in GPL and GL compositions as temperatures oscillate between 18{degrees}C and 37{degrees}C for all strains. Exploration of lipid subspecies further exposes disparities in PE and PG lipids containing C16:1 and oleic acid (C18:1). This investigation not only provides insights into the physiological attributes and survival strategies of A. baumannii but also deepens our comprehension of its adaptive responses to temperature changes. By unraveling the dynamics of lipid composition and fatty acid profiles, this study enriches our understanding of the bacteriums ecological fitness and behavior in diverse environments. IMPORTANCEAcinetobacter baumannii, a bacterium known for its resistance to antibiotics, is a concern in healthcare settings. This study focused on understanding how this bacterium adapts to different temperatures and how its lipid composition changes. Lipids are like the building blocks of its cell membranes. By studying these changes, scientists can gain insights into how the bacterium survives and behaves in various environments. This knowledge helps us better understand its ability to cause infections and resist treatments. The studys findings contribute to our broader understanding of how Acinetobacter baumannii functions, which is important for developing strategies to combat its impact on patient health.

microbiology↗

bioRxiv2023

Bridges instead of boats? The Mla system of diderm Firmicute Veillonella parvula reveals an ancestral transenvelope core of phospholipid trafficking

Despite extensive characterisation of envelope biogenesis systems in diderm bacteria, glycerophospholipid (GPL) trafficking remains poorly understood, and has only been studied in a handful of model species. Within the Proteobacteria, the maintenance of lipid asymmetry (Mla) system facilitates retrograde GPL trafficking via six proteins, MlaA-F. GPLs are extracted from the outer leaflet of the outer membrane by the lipoprotein MlaA which associates with porin trimers, then shipped through the periplasmic space by the chaperone MlaC, which finally delivers GPLs to the inner membrane complex formed by MlaBDEF. Here, we investigate GPL trafficking in Veillonella parvula, a diderm member of the Firmicutes which encodes an Mla system devoid of MlaA and MlaC. V. parvula {Delta}mla mutants display phenotypes characteristic of disrupted lipid asymmetry such as hypervesiculation and detergent hypersensitivity, and lipid content analysis from outer membrane vesicles reveals an enrichment for the major lipid component phosphatidylethanolamine. Interestingly, suppressor analysis identifies mutations in tamB that rescue detergent hypersensitivity and hypervesiculation of {Delta}mla strains, supporting the involvement of these two systems in antagonistic GPL trafficking functions across diverse bacterial lineages. A combination of structural modeling and subcellular localisation assays shows that MlaDVp is longer than in classical diderm models and forms a transenvelope bridge, encoding both an inner membrane-localised MCE domain and an outer membrane {beta}-barrel. These results strongly suggest that V. parvula possesses a minimal Mla system for GPL trafficking, replacing the need for chaperones and outer membrane lipoproteins by directly connecting the two membranes. Finally, phylogenomic analysis indicates that this MlaEFD self-contained architecture is widely distributed in diderm bacteria and most likely represents the ancestral functional core of the Mla system, which subsequently increased in complexity in Proteobacteria and closely related phyla following the emergence of MlaABC. Our work broadens the diversity of current models of GPL trafficking in diderm bacteria, challenging the paradigm set by classical models and shedding light on the evolution of a crucial system in the biogenesis and maintenance of the bacterial outer membrane.

microbiology↗

bioRxiv2022

A bioorthogonal chemistry approach to detect the K1 polysialic acid capsule in Escherichia coli

Most Escherichia coli strains associated with neonatal meningitis express the K1 capsule, a sialic acid polysaccharide that is directly related to their pathogenicity. Metabolic oligosaccharide engineering (MOE) has mostly been developed in eukaryotes, but has also been successfully applied to the study of several oligosaccharides or polysaccharides constitutive of the bacterial cell wall. However, bacterial capsules are seldom targeted despite their important role as virulence factors, and the K1 polysialic acid (PSA) antigen that shields bacteria from the immune system still remains untackled. Herein, we report a fluorescence microplate assay that allows the fast and facile detection of K1 capsules with an approach that combines MOE and bioorthogonal chemistry. We exploit the incorporation of synthetic analogues of N-acetylmannosamine or N-acetylneuraminic acid, metabolic precursors of PSA, and copper-catalysed azide-alkyne cycloaddition (CuAAC) as the click chemistry reaction to specifically label the modified K1 antigen with a fluorophore. The method was optimized, validated by capsule purification and fluorescence microscopy, and applied to the detection of whole encapsulated bacteria in a miniaturized assay. We observe that analogues of ManNAc are readily incorporated into the capsule while those of Neu5Ac are less efficiently metabolized, which provides useful information regarding the capsule biosynthetic pathways and the promiscuity of the enzymes involved. Moreover, this microplate assay is transferable to screening approaches and may provide a platform to identify novel capsule-targeted antibiotics that would circumvent resistance issues.

microbiology↗

bioRxiv2021

The role of L-arabinose metabolism for Escherichia coli O157:H7 in edible plants

Arabinose is a major plant aldopentose in the form of arabinans complexed in cell wall polysaccharides or glycoproteins (AGP), but comparatively rare as a monosaccharide. L-arabinose is an important bacterial metabolite, accessed by pectolytic microorganisms such as Pectobacterium atrosepticum via pectin and hemicellulose degrading enzymes. However, not all plant-associated microbes encode cell wall degrading enzymes, yet can metabolise L-arabinose, raising questions about their use of and access to the glycan in plants. Therefore, we examined L-arabinose metabolism in the food-borne pathogen Escherichia coli O157:H7 (isolate Sakai) during its colonisation of plants. L-arabinose metabolism (araBA) and transport (araF) genes were activated at 18 {degrees}C in vitro by L-arabinose and expressed over prolonged periods in planta. Although deletion of araBAD did not impact the colonisation ability of E. coli O157:H7 (Sakai) on plants, araA was induced on exposure to spinach cell wall polysaccharides. Furthermore, debranched and arabinan oligosaccharides induced ara metabolism gene expression in vitro, and stimulated modest proliferation, while immobilised pectin did not. Thus, E. coli O157:H7 (Sakai) can utilise pectin/AGP- derived L-arabinose as a metabolite, but differs fundamentally in ara gene organisation, transport and regulation from the related pectinolytic species P. atrosepticum, reflective of distinct plant- associated lifestyles.

microbiology↗

bioRxiv2020

The impact of plasma membrane lipid composition on flagella-mediated adhesion of enterohemorrhagic Escherichia coli

Enterohemorrhagic Escherichia coli (EHEC) O157:H7 is a major cause of foodborne gastrointestinal illness. The adhesion of EHEC on host tissues is the first step enabling bacterial colonization. Adhesins like fimbriae and flagella mediate this mechanism. Here, we studied the interaction of the bacterial flagellum with the host cell’s plasma membrane using Giant Unilamellar Vesicles (GUVs) as a biologically relevant model. Cultured cell lines contain many different molecular components including proteins and glycoproteins. In contrast, with GUVs we can characterize the bacterial mode of interaction solely with a defined lipid part of the cell membrane. Bacterial adhesion on GUVs was dependent on the presence of the flagellar filament and its motility. By testing different phospholipid head groups, the nature of the fatty acid chains or the liposome curvature, we found that lipid packing is a key parameter to enable bacterial adhesion. Using HT-29 cells grown in the presence of polyunsaturated fatty acid (α-linolenic acid) or saturated fatty acid (palmitic acid), we found that α-linolenic acid reduced adhesion of wild type EHEC but not of a non-flagellated mutant. Finally, our results reveal that the presence of flagella is advantageous for the bacteria to bind to lipid rafts. We speculate that polyunsaturated fatty acids prevent flagellar adhesion on membrane bilayers and play a clear role for optimal host colonization. Flagella-mediated adhesion to plasma membranes has broad implications to host-pathogen interactions.Importance Bacterial adhesion is a crucial step to allow bacteria to colonize their hosts, invade tissues and form biofilm. Enterohemorrhagic E. coli O157:H7 is a human pathogen and the causative agent of diarrhea and hemorrhagic colitis. Here, we use biomimetic membrane models and cell lines to decipher the impact of lipid content of the plasma membrane on enterohemorrhagic E. coli flagella-mediated adhesion. Our findings provide evidence that polyunsaturated fatty acid (α-linolenic acid) inhibits E. coli flagella adhesion to the plasma membrane in a mechanism separate from its antimicrobial and anti-inflammatory functions. In addition, we confirm that cholesterol-enriched lipid microdomains, often called lipid rafts are important in bacterial adhesion. These findings significantly strengthen plasma membrane adhesion via bacterial flagella in an important human pathogen. This mechanism represents a promising target for the development of novel anti-adhesion therapies.Competing Interest StatementThe authors have declared no competing interest.View Full Text

microbiology↗

bioRxiv2020

Bacterial flagella disrupt host cell membranes and interact with cytoskeletal components

Bacterial flagella have many established roles beyond swimming motility. Despite clear evidence of flagella-dependent adherence, the specificity of ligands and mechanisms of binding are still debated. In this study, the molecular basis of E. coli O157:H7 and S. Typhimurium flagella binding to epithelial cell cultures was investigated. Flagella interactions with host cell surfaces were intimate and crossed cellular boundaries as demarcated by actin and membrane labelling. SEM revealed flagella disappearing into cellular surfaces and TEM of S. Typhiumurium indicated host membrane deformation and disruption in proximity to flagella. Motor mutants of E. coli O157:H7 and S. Typhimurium caused reduced haemolysis compared to wild-type, indicating that membrane disruption was in part due to flagella rotation. Flagella from E. coli O157 (H7), EPEC O127 (H6), and S. Typhimurium (P1 & P2 flagella) were shown to bind to purified intracellular components of the actin cytoskeleton and directly increase in vitro actin polymerisation rates.

microbiology↗

bioRxiv2019

Flagella methylation promotes bacterial adhesion and host cell invasion

The flagellum is the motility device of many bacteria and the long external filament is made of several thousand copies of a single protein, flagellin. While posttranslational modifications of flagellin are common among bacterial pathogens, the role of lysine methylation remained unknown. Here, we show that both flagellins of Salmonella enterica, FliC and FljB, are methylated at surface-exposed lysine residues. A Salmonella mutant deficient in flagellin methylation was outcompeted for gut colonization in a gastroenteritis mouse model. In support, methylation of flagellin promoted invasion of epithelial cells in vitro. Lysine methylation increased the surface hydrophobicity of flagellin and enhanced flagella-dependent adhesion of Salmonella to phosphatidylcholine vesicles and epithelial cells. In summary, posttranslational flagellin methylation constitutes a novel mechanism how flagellated bacteria facilitate adhesion to hydrophobic host cell surfaces and thereby contributes to efficient gut colonization and successful infection of the host.

microbiology↗