Contributors: MICrobiologie de l'ALImentation au Service de la Santé (MICALIS); AgroParisTech-Université Paris-Saclay-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE); Plateforme technologique Nanoimagerie / Nanoimaging Technological Platform; Institut Pasteur Paris (IP)-Université Paris Cité (UPCité); Plate-forme de bioimagerie ultrastructurale - Ultrastructural BioImaging Core Facility; Mathématiques et Informatique Appliquées du Génome à l'Environnement Jouy-En-Josas (MaIAGE); Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE); Infectiologie et Santé Publique (ISP); Université de Tours (UT)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE); This work was supported by grants from the Agence Nationale de la Recherche (ANR) to HB (PERMALI: ANR-20-CE35-0001-01) and AP (THOR: ANR-20-CE15-0008-01); from the Université Paris-Saclay to HB (DEPISTALIS, AAP Poc in labs 2019); from the INRAE’s MICA department and Micalis Institute to AP (AAP Micalis 2023); and from the European Research Council, under the Horizon 2020 research and innovation program, to RCL (agreement ID: 772178). FC was supported by postdoctoral grants from INRAE MICA department and ANR (ANR-20-PAMR-0011). AB received funding from the European Union’s Horizon 2020 research and innovation program, under the Marie Skłodowska-Curie Actions grant (agreement ID: 101030628).; ANR-20-CE35-0001,PERMALI,Marqueurs de la persistance intracellulaire de Listeria monocytogenes(2020); ANR-20-CE15-0008,THOR,Le ciblage de l'ARN de l'hôte par des effecteurs bacteriens secrétés(2020); ANR-20-PAMR-0011,TherAEPI,Thérapies épigénétiques pour esquiver la résistance(2020); European Project: 772178,BACTIN; European Project: 101030628,H2020-MSCA-IF-2020 ,10.3030/101030628,FCS_BACSUB(2021)
نبذة مختصرة : Bacteria can respond to environmental stresses by entering a dormant state, called viable but non-culturable (VBNC) state, in which they no longer grow in routine culture media. VBNC pathogens pose thus a significant risk for human and animal health as they are not detected by standard growth-based techniques and can “wake up” back into a vegetative and virulent state. Although hundreds of species were reported to become VBNC in response to different stresses, the molecular mechanisms governing this phenotypic switch remain largely elusive. Here, we characterized the VBNC state transition process in the Gram-positive pathogen Listeria monocytogenes in response to nutritional deprivation. By combining fluorescence microscopy, cryo-electron tomography and analytical biochemistry, we found that starvation in mineral water drives L. monocytogenes into a VBNC state via a mechanism of cell wall (CW) shedding that generates osmotically stable CW-deficient (CWD) coccoid forms. This phenomenon occurs in multiple L. monocytogenes strains and in other Listeria species, suggesting it may be a stress-adapting process transversal to the Listeria genus. Transcriptomic and gene-targeted approaches revealed the stress response regulator SigB and the autolysin NamA as major moderators of CW loss and VBNC state transition. Finally, we show that this CWD dormant state is transient as VBNC Listeria revert back to a walled, vegetative and virulent state after passage in embryonated eggs. Our findings provide unprecedented detail on the mechanisms governing the transition to a VBNC state, and reveal that dormant CWD bacterial forms can naturally arise in aquatic environments without osmotic stabilization. This may represent an alternative strategy for bacterial survival in oligotrophic conditions, which can potentially generate public health-threatening reservoirs of undetectable pathogens.
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