Biology subjects

Janet-Maitre, M.

Publications and source records attributed to Janet-Maitre, M..

9 recordsLinked to original sources

bioRxiv2026

Experimental evolution reveals temporally opposing roles for Csu pili during Acinetobacter baumannii respiratory infection

Multidrug-resistant (MDR) Acinetobacter baumannii is an opportunistic pathogen responsible for a wide range of infections, including ventilator-associated pneumonia, bloodstream infections, and urinary tract infections. Most studies investigating A. baumannii pulmonary pathogenesis rely on acute pneumonia models that use high bacterial inocula and do not recapitulate the prolonged infections observed in some patients. Recently, our laboratory developed a murine model of chronic pulmonary infection that enabled the identification of virulence factors specifically required for long-term lung persistence. To investigate how A. baumannii adapts during chronic pulmonary infection, we performed an experimental evolution study using this model. This approach revealed an unexpected role for the archaic chaperone-usher (Csu) pili in A. baumannii pathogenesis. At early time points, Csu expression reduced bacterial burdens in both acute and chronic pneumonia models, an effect associated with enhanced host inflammatory responses. In contrast, at later stages of chronic infection, Csu promoted bacterial persistence, potentially by facilitating adhesion to pulmonary epithelial cells. These opposing effects were conserved across two distinct A. baumannii clinical isolates. We also found that Csu contributes to bacterial colonization in a catheter-associated urinary tract infection (CAUTI) model in a strain-dependent manner. Together, these findings reveal an infection-stage-dependent role for Csu in A. baumannii pathogenesis, demonstrating that a bacterial surface structure can impair early colonization while promoting long-term persistence.

microbiology↗

bioRxiv2026

An Expanded Lpt Pathway Couples Surface Lipoprotein Export to Outer Membrane Vesicle Biogenesis in Bacteroides

The ecological dominance of Bacteroidota in the human gut microbiota derives from their exceptional capacity to metabolize diverse dietary and host-derived glycans. This ability relies on surface-exposed lipoproteins (SLPs), which are displayed on the bacterial surface and selectively packaged into outer membrane vesicles (OMVs). Although SLP export and OMV biogenesis are tightly coupled, the mechanism linking these processes has remained unknown. Here, we identify LptZ, a previously uncharacterized protein, as a critical link between SLP surface translocation and OMV biogenesis. Cryo-electron tomography of the lptZ mutant revealed chains of vesicles tethered to the outer membrane, offering a rare glimpse into an arrested stage of OMV formation in which vesicle fission is impaired. Proteomic analysis further showed that, in the absence of LptZ, a subset of SLPs fails to reach the cell surface and instead accumulates at the inner membrane. Unexpectedly, LptZ associates with the lipopolysaccharide transport (Lpt) machinery and repurposes it for SLP translocation. Crosslinking experiments revealed selective associations between multiple SLPs and Lpt proteins, providing mechanistic evidence for the direct involvement of the Lpt machinery in SLP trafficking. We further show that the Bacteroides Lpt machinery contains previously unrecognized components, revealing an unanticipated level of specialization relative to canonical Lpt systems. Together, our findings uncover a previously unknown protein export pathway, expand the functional role of the Lpt machinery, and establish a direct mechanistic link between SLP translocation and OMV biogenesis in Bacteroidota.

microbiology↗

bioRxiv2026

Elongasome Dysfunction Triggers Dependence on MepM-Mediated Peptidoglycan Recycling

The bacterial peptidoglycan (PG) layer is responsible for maintaining cell shape and protecting the cell against both external stress and internal turgor pressure. In rod-shaped bacteria, PG synthesis is orchestrated by two major multi-protein complexes: the elongasome and the divisome, which drive lateral and septal cell wall synthesis, respectively. The membrane protein MreD plays a central role in regulating the elongasome activity. Here, we leveraged a viable Pseudomonas aeruginosa transposon mutant, with reduced mreD expression, to dissect cellular adaptations to impaired elongasome function. A genome-wide synthetic lethality screen revealed that cell division and PG recycling pathways become essential in the context of an impaired elongasome machinery. Moreover, Tn-seq analysis identified the endopeptidase MepM as essential in the mreD downregulated background, whereas its overexpression promoted cell elongation in the parental strain, supporting a conserved role in lateral wall biogenesis. We demonstrate that MepM sustains viability during elongasome impairment by maintaining an active MurU-dependent PG recycling pathway. Together, our findings uncover a functional coupling between PG hydrolysis, recycling, and synthesis. We also identify MepM and PG recycling as critical determinants of bacterial survival when elongasome activity is compromised, positioning these pathways as attractive targets for the development of next-generation or combinatory antibacterial therapies.

microbiology↗

bioRxiv2025

Intracellular Acinetobacter baumannii Acts as a Reservoir in Lung Infection via a 'Persist and Resist' Strategy

Although considered primarily extracellular, Acinetobacter baumannii can survive and replicate within macrophages in vitro. Intracellular bacteria are often protected from the host immune system and antibiotic treatment, potentially leading to chronic or recurrent infections. To investigate the role of intracellular A. baumannii during infection, we transferred infected bronchoalveolar lavage fluid (BALF), containing an intracellular bacterial population, to naive immunocompromised mice. This BALF transfer resulted in A. baumannii lung infection, suggesting that intracellular bacteria may serve as a reservoir for recurrent lung infections. Using dual-proteomics, we characterized the A. baumannii-macrophages interactions. Infected macrophages exhibit an inflammatory and Type I interferon response, marked by increased ACOD1 (IRG1) protein levels. Intracellular A. baumannii upregulates proteins involved in evading nutritional immunity, stress response, surface modification, and metabolic adaptation. These findings suggest that A. baumannii employs a multifactorial strategy to survive and replicate within macrophages, which may contribute to recurrent and chronic infections, and compromise treatment efficacy. TeaserIntracellular A. baumannii can act as a reservoir in lung infection

microbiology↗

bioRxiv2025

Strain-Specific Variation in the Complement Resistome of Pseudomonas aeruginosa

Bloodstream infections caused by Pseudomonas aeruginosa are associated with the highest mortality rates. The complement system, key component of the innate immune response, is mostly responsible for P. aeruginosa elimination in human blood. Nevertheless, the plasma sensitivity of P. aeruginosa strains varies, ranging from highly sensitive, persistent to fully resistant. Whereas most studies rely on model strains, given the species high genomic and phenotypic diversity, the interaction between P. aeruginosa and the complement system may be highly complex. In this work, we characterized the plasma resistome of P. aeruginosa in three strains displaying variable plasma sensitivity using Tn-seq. The gain-of-function screen performed on the sensitive strain PA14 highlighted numerous bacterial factors impacting plasma resistance, including members of the RetS-LadS/Gac/Rsm regulatory system and outer membrane porins. Tn-seq in plasma-resistant strains CHA and YIK, suggested that each strain utilizes a specific and limited set of proteins to evade complement-dependent killing. Despite this strain-specific response, we identified common strategies across all strains, including the production of exopolysaccharides (EPSs), the presence of surface appendages and modification in the O-specific antigen. We evidence Ssg and Crc as common factors contributing to plasma resistance. However, whereas mutants lacking ssg and/or crc in plasma had reduced survival, a subpopulation of these mutants persisted over prolonged exposure to human plasma. Overall, this work provides valuable insights into the complex interplay between P. aeruginosa and the complement system in the context of bloodstream infections and raises concerns for the development of therapies targeting individual virulence factors.

microbiology↗

bioRxiv2024

A chronic murine model of pulmonary Acinetobacter baumannii infection enabling the investigation of late virulence factors, long-term antibiotic treatments, and polymicrobial infections

Acinetobacter baumannii can cause prolonged infections that disproportionately affect immunocompromised populations. Our understanding of A. baumannii respiratory pathogenesis relies on an acute murine infection model with limited clinical relevance that employs an unnaturally high number of bacteria and requires the assessment of bacterial load at 24-36 hours post-infection. Here, we demonstrate that low intranasal inoculums in immunocompromised mice with a tlr4 mutation leads to reduced inflammation, allowing for persistent infections lasting at least 3 weeks. Using this "chronic infection model," we determined the adhesin InvL is an imperative virulence factor required during later stages of infection, despite being dispensable in the early phase. We also demonstrate that the chronic model enables the distinction between antibiotics that, although initially reduce bacterial burden, either lead to complete clearance or result in the formation of bacterial persisters. To illustrate how our model can be applied to study polymicrobial infections, we inoculated mice with an active A. baumannii infection with Staphylococcus aureus or Klebsiella pneumoniae. We found that S. aureus exacerbates the infection, while K. pneumoniae enhances A. baumannii clearance. In all, the chronic model overcomes some limitations of the acute pulmonary model, expanding our capabilities to study of A. baumannii pathogenesis and lays the groundwork for the development of similar models for other important opportunistic pathogens.

microbiology↗

bioRxiv2023

Pseudomonas aeruginosa MipA-MipB envelope proteins act as new sensors of polymyxin

Due to the rising incidence of antibiotic resistant infections, the last-line antibiotics polymyxins have resurged in the clinics in parallel with new bacterial strategies of escape. The Gram-negative opportunistic pathogen Pseudomonas aeruginosa develops resistance to colistin/polymyxin by distinct molecular mechanisms, mostly through modification of the lipid A component of the LPS by proteins encoded within the arnBCDATEF-ugd (arn) operon. In this work, we characterized a polymyxin-induced operon, named mipBA, notably present in P. aeruginosa strains devoid of the arn operon. We showed that mipBA is activated by ParR/ParS two-component regulatory system in response to polymyxin. MipA and MipB localize to bacterial outer membrane and form a complex in vitro. Structural modeling revealed that the lipoprotein MipB adopts a {beta}-lactamase fold with two additional C-terminal domains,while MipA folds as an outer-membrane {beta}-barrel, harboring an internal negatively charged channel, able to host a polymyxin molecule. Nano differential scanning fluorimetry (DSF) showed that polymyxin stabilized MipA protein in vitro. Mass spectrometry-based quantitative proteomics on whole bacterial membranes demonstrated that the {Delta}mipBA mutant synthesized less MexXY-OprA proteins in response to polymyxin compared to the wild-type strain, as a consequence of impaired transcriptional activation of the mex operon. We propose MipA/MipB to act as membrane (co)sensors working in concert to activate ParS histidine kinase and help the bacterium to cope with polymyxin-mediated envelope stress through synthesis of the efflux pomp, MexXY-OprA.

microbiology↗

bioRxiv2022

Colistin resistance mutations in phoQ sensitize Klebsiella pneumoniae to IgM-mediated complement killing

The Gram-negative bacterium Klebsiella pneumoniae is notorious for a strong increase of infections with antibiotic resistant strains. To treat infections with antibiotic resistant K. pneumoniae, clinicians increasingly need to use the last resort antibiotic colistin. K. pneumoniae can develop colistin resistance by modifying its membranes. During infection the membranes of Gram-negative bacteria are also targeted by the human immune system via the complement system. Gram-negative bacteria have an outer and inner membrane separated by a thin cell wall. Activation of the complement system leads to the formation of the membrane attack complex (MAC), a pore that inserts into the outer membrane, and ultimately leads to lysis of the bacterium. As both colistin and the MAC interact with the outer membrane of Gram-negative bacteria, we wondered if developing colistin resistance influences MAC-mediated killing of K. pneumoniae. Using clinical isolates that developed colistin resistance, we found that the strain Kp209_CSTR became more sensitive to MAC-mediated killing compared to the wild-type strain. MAC-mediated membrane permeabilization of Kp209_CSTR required antibody dependent activation of the classical complement pathway. Strikingly, Kp209_CSTR was bound by IgM in human serum that did not recognise the wild-type strain. Depletion of Kp209_CSTR-specific antibodies from serum prevented MAC-mediated membrane permeabilization, which was restored by adding back IgM. Genomic sequence comparison revealed that Kp209_CSTR has a deletion in the phoQ gene. RNAseq analysis suggested that this mutation locks PhoQ in a constitutively active state. These results indicate that PhoQ activation in Kp209_CSTR leads to both colistin resistance and sensitivity to MAC-mediated killing. Together, our results show that developing colistin resistance can sensitize K. pneumoniae to killing by the immune system.

microbiology↗

bioRxiv2021

Molecular features underlying Pseudomonas aeruginosa persistence in human plasma

Pseudomonas aeruginosa, an opportunistic Gram-negative pathogen, is a leading cause of bacteremia with a high mortality rate. We recently reported that P. aeruginosa forms a persister-like sub-population of evaders in human plasma and blood. However, the molecular mechanisms underlying the formation of evaders remained unknown. Here, using a gain-of-function genetic screen, we examined the molecular determinants of P. aeruginosa persistence in plasma. We found that, among other factors, ATP and biotin availability greatly influence bacterial survival in plasma; mutants in pur and bio genes display higher tolerance and persistence, respectively. Electron microscopy combined with energy-dispersive X-ray spectroscopy (EDX) revealed the formation of polyphosphate granules upon incubation in plasma in several clinical strains, implying the bacterial response to a low-energy stress signal. Indeed, mutants with transposon insertions in ppk genes were eliminated in the plasma. Analysis of several steps of the complement cascade and exposure to an outer-membrane-impermeable drug, nisin, suggested that the mutants impede membrane attack complex (MAC) activity per se. Through this study, we shed light on P. aeruginosa response to the plasma conditions and discovered the multifactorial origin of bacterial resilience to MAC that provides a comprehensive picture of the complex interplay between P. aeruginosa and the human complement system. Author summaryPersistence of bacterial pathogens is a main cause of treatment failure and establishment of chronic bacterial infection. Despite innate immune responses, some bacteria may persist in human blood and plasma. Here we used a genome-wide screen to investigate the molecular determinants influencing Pseudomonas aeruginosa persistence in human plasma facing the complement system. Alongside a multifactorial strategy, we found intracellular levels of ATP and biotin to significantly influence bacterial capacity to deal with membrane attack complex (MAC)-dependent killing. These results underline the need to understand the complex interplay between bacterial pathogens and the human immune system when seeking to develop efficient antibacterial strategies.

microbiology↗