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4 results on '"Tiffany Bonnefois"'

1. Interactions between Mycoplasma mycoides subsp. mycoides and bovine macrophages under physiological conditions.

Author

Philippe Totté, Tiffany Bonnefois, and Lucia Manso-Silván

Subjects
Medicine, Science
Abstract

We investigated the interactions of unopsonized and opsonized Mycoplasma mycoides subsp. mycoides (Mmm) with bovine macrophages in vitro. Mmm survived and proliferated extracellularly on bovine macrophage cell layers in the absence of Mmm-specific antisera. Bovine complement used at non-bactericidal concentrations did neither have opsonizing effect nor promoted intracellular survival, whereas Mmm-specific antisera substantially increased phagocytosis and Mmm killing. A phagocytosis-independent uptake of Mmm by macrophages occurred at a high multiplicity of infection, also found to induce the production of TNF, and both responses were unaffected by non-bactericidal doses of bovine complement. Bovine complement used at higher doses killed Mmm in cell-free cultures and completely abrogated TNF responses by macrophages. These results provide a framework to identify Mmm antigens involved in interactions with macrophages and targeted by potentially protective antibodies and point towards a pivotal role of complement in the control of inflammatory responses in contagious bovine pleuropneumonia.

Published
2024
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2. Interactions betweenMycoplasma mycoidessubsp.mycoidesand bovine macrophages under physiological conditions

Author

Philippe Totté, Tiffany Bonnefois, and Lucia Manso-Silván

Abstract

We investigated the interactions of unopsonized and opsonizedMycoplasma mycoidessubsp.mycoides(Mmm) with bovine macrophagesin vitro. Mmmsurvived and proliferated extracellularly on bovine macrophage cell layers in the absence ofMmm-specific antisera. Bovine complement used at non-bactericidal concentrations did neither have opsonizing effect nor promoted intracellular survival, whereasMmm-specific antisera substantially increased phagocytosis andMmmkilling. A phagocytosis-independent uptake ofMmmby macrophages occurred at high MOIs, also found to induce the production of TNF, and both responses were unaffected by non-bactericidal doses of bovine complement. Bovine complement used at higher doses killedMmmin cell-free cultures and completely abrogated TNF responses by macrophages. These results provide a framework to identifyMmmantigens involved in interactions with macrophages and targeted by potentially protective antibodies and point towards a pivotal role of complement in the control of inflammatory responses in contagious bovine pleuropneumonia.

Published
2022
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3. Development of fluorescence expression tools to study host-mycoplasma interactions and validation in two distant mycoplasma clades

Author

François Thiaucourt, Tiffany Bonnefois, Marie-Stéphanie Vernerey, Philippe Totté, Valérie Rodrigues, Lucia Manso-Silvan, Carinne Puech, Chantal Ripoll, Manso-Silvan, Lucia, Contrôle des maladies animales exotiques et émergentes (UMR CMAEE), Institut National de la Recherche Agronomique (INRA)-Centre de Coopération Internationale en Recherche Agronomique pour le Développement (Cirad), Biologie et Génétique des Interactions Plante-Parasite (UMR BGPI), Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Centre de Coopération Internationale en Recherche Agronomique pour le Développement (Cirad)-Institut National de la Recherche Agronomique (INRA), Institut des Neurosciences de Montpellier - Déficits sensoriels et moteurs (INM), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Montpellier (UM), Centre de Coopération Internationale en Recherche Agronomique pour le Développement (Cirad)-Institut National de la Recherche Agronomique (INRA)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro), Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro), Centre de Coopération Internationale en Recherche Agronomique pour le Développement (Cirad)-Institut National de la Recherche Agronomique (INRA), Institut des Neurosciences de Montpellier (INM), and ProdInra, Archive Ouverte

Subjects
0301 basic medicine, Biodiversité et Ecologie, mycoplasma, fluorescence, whole cell labelling, host-pathogen interactions, flow cytometry, confocal microscopy, L73 - Maladies des animaux, medicine.disease_cause, Applied Microbiology and Biotechnology, law.invention, law, Haemobartonella, Flow cytometry, Cells, Cultured, Genetics, Phagocytes, biology, medicine.diagnostic_test, cytotoxicité, General Medicine, Recombinant Proteins, Molecular Imaging, 3. Good health, Mycoplasma mycoides, host pathogen systems, Biotechnology, Whole cell labelling, Green Fluorescent Proteins, 030106 microbiology, protéine fluorescente, Mutagenesis (molecular biology technique), Bioengineering, Fluorescence, Microbiology, Biodiversity and Ecology, 03 medical and health sciences, Confocal microscopy, medicine, Animals, fluorescent protein, Mycoplasma Infections, clone, Host-pathogen interactions, interaction hôte pathogène, Reproducibility of Results, Mycoplasma, biology.organism_classification, In vitro, [SDE.BE] Environmental Sciences/Biodiversity and Ecology, Spectrometry, Fluorescence, Cytoplasm, Cattle, U30 - Méthodes de recherche, [SDE.BE]Environmental Sciences/Biodiversity and Ecology, mCherry
Abstract

Fluorescence expression tools for stable and innocuous whole mycoplasma cell labelling have been developed. A Tn4001-derivative mini-transposon affording unmarked, stable mutagenesis in mycoplasmas was modified to allow the constitutive, high-level expression of mCherry, mKO2 and mNeonGreen. These tools were used to introduce the respective fluorescent proteins as chromosomal tags in the phylogenetically distant species Mycoplasma mycoides subsp. mycoides and Mycoplasma bovis. The production, selection and characterisation of fluorescent clones were straightforward and resulted in the unprecedented observation of red and green fluorescent mycoplasma colonies in the two species, with no apparent cytotoxicity. Equivalent fluorescence expression levels were quantified by flow cytometry in both species, suggesting that these tools can be broadly applied in mycoplasmas. A macrophage infection assay was performed to assess the usefulness of mNeonGreen-expressing strains for monitoring mycoplasma infections, and notably cell invasion. The presence of fluorescent mycoplasmas inside live phagocytic cells was detected and quantified by flow cytometry and corroborated by confocal microscopy, which allowed the identification of individual mycoplasmas in the cytoplasm of infected cells. The fluorescence expression tools developed in this study are suitable for host-pathogen interaction studies and offer innumerable perspectives for the functional analysis of mycoplasmas both in vitro and in vivo.

Published
2016
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4. Characterization of the lipid envelope of exosome encapsulated HEV particles protected from the immune response

Author

Jacques Izopet, Martine Dubois, Pierre-Emmanuel Gleizes, Tiffany Bonnefois, Florence Abravanel, Célia Plisson-Chastang, Sabine Chapuy-Regaud, Justine Bertrand-Michel, Benoit Flan, Steve Simoneau, Sébastien Lhomme, Bruno You, Laboratoire de Virologie, Hôpital Purpan [Toulouse], CHU Toulouse [Toulouse]-CHU Toulouse [Toulouse], Laboratoire de biologie moléculaire eucaryote (LBME), Centre National de la Recherche Scientifique (CNRS)-Centre de Biologie Intégrative (CBI), Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Centre National de la Recherche Scientifique (CNRS)-Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Centre National de la Recherche Scientifique (CNRS), Centre de Biologie Intégrative (CBI), Animal, Santé, Territoires, Risques et Ecosystèmes (UMR ASTRE), Centre de Coopération Internationale en Recherche Agronomique pour le Développement (Cirad)-Institut National de la Recherche Agronomique (INRA), Ecole des Ingénieurs de la Ville de Paris, MetaToul-MetaboHUB, Génopole Toulouse Midi-Pyrénées [Auzeville] (GENOTOUL), Institut National des Sciences Appliquées - Toulouse (INSA Toulouse), Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Ecole Nationale Vétérinaire de Toulouse (ENVT), Institut National Polytechnique (Toulouse) (Toulouse INP), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Institut National Polytechnique (Toulouse) (Toulouse INP), Université Fédérale Toulouse Midi-Pyrénées-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)-Institut National des Sciences Appliquées - Toulouse (INSA Toulouse), Université Fédérale Toulouse Midi-Pyrénées-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), Centre de recherche et de Diagnostic sur le Sida [Abidjan, Côte d'Ivoire] (CeDreS), Centre Hospitalier Universitaire de Treichville [Abidjan, Côte d'Ivoire] (CHU de Treichville), LFB Biomédicaments, Centre de Physiopathologie Toulouse Purpan (CPTP), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Contrôle des maladies animales exotiques et émergentes (UMR CMAEE), Institut National de la Recherche Agronomique (INRA)-Centre de Coopération Internationale en Recherche Agronomique pour le Développement (Cirad), Plateforme Metatoul, Institut National de la Santé et de la Recherche Médicale (INSERM), Centre de recherche et de Diagnostic sur le Sida (CeDreS), CHU Treichville, Centre de Physiopathologie Toulouse Purpan ex IFR 30 et IFR 150 (CPTP), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Toulouse III - Paul Sabatier (UT3), and Université Fédérale Toulouse Midi-Pyrénées-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)-Université Toulouse III - Paul Sabatier (UT3)

Subjects
0301 basic medicine, viruses, [SDV]Life Sciences [q-bio], 030106 microbiology, Population, medicine.disease_cause, Exosomes, Biochemistry, Exosome, Microbiology, 03 medical and health sciences, Membrane Lipids, Hepatitis E virus, Viral envelope, Cell-Derived Microparticles, medicine, Humans, education, ComputingMilieux_MISCELLANEOUS, Infectivity, education.field_of_study, biology, virus diseases, RNA virus, General Medicine, Hep G2 Cells, biology.organism_classification, Virology, digestive system diseases, Microvesicles, 3. Good health, Hepatitis E, 030104 developmental biology, biology.protein, Antibody
Abstract

The hepatitis E virus (HEV) is the most common cause of acute hepatitis worldwide. Although HEV is a small, naked RNA virus, HEV particles become associated with lipids in the blood of infected patients and in the supernatant of culture systems. The egress of these particles from cells implies the exocytosis pathway but the question of the role of the resulting HEV RNA containing exosomes and the nature of the lipids they contain has not been fully addressed. We determined the lipid proportions of exosomes from uninfected and HEV-infected cells and their role in HEV spreading. We cultured a suitable HEV strain on HepG2/C3A cells and analyzed the population of exosomes containing HEV RNA using lipidomics methods and electron microscopy. We also quantified HEV infectivity using an infectivity endpoint method based on HEV RNA quantification to calculate the tissue culture infectious dose 50. Exosomes produced by HEV-infected HepG2/C3A cells contained encapsidated HEV RNA. These HEV RNA-containing exosomes were infectious but ten times less than stools. HEV from stools, but not exosome-associated HEV from culture supernatant, was neutralized by anti-HEV antibodies in a dose-dependent manner. HEV infection did not influence the morphology or lipid proportions of the bulk of exosomes. These exosomes contained significantly more cholesterol, phosphatidylserine, sphingomyelin and ceramides than the parent cells, but less phosphoinositides and polyunsaturated fatty acids. Exosomes play a major role in HEV egress but HEV infection does not modify the characteristics of the bulk of exosomes produced by infected cells. PS and cholesterol enriched in these vesicles could then be critical for HEV entry. HEV particles in exosomes are protected from the immune response which could lead to the wide circulation of HEV in its host.

Published
2017
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