Publications by authors named "Jean-Paul Latgé"

223 Publications

Comparative host transcriptome in response to pathogenic fungi identifies common and species-specific transcriptional antifungal host response pathways.

Comput Struct Biotechnol J 2021 26;19:647-663. Epub 2020 Dec 26.

Department of Internal Medicine and Radboudumc Center for Infectious Diseases (RCI), Radboud University Medical Center, Nijmegen, The Netherlands.

Candidiasis, aspergillosis, and mucormycosis cause the majority of nosocomial fungal infections in immunocompromised patients. Using an unbiased transcriptional profiling in PBMCs exposed to the fungal species causing these infections, we found a core host response in healthy individuals that may govern effective fungal clearance: it consists of 156 transcripts, involving canonical and non-canonical immune pathways. Systematic investigation of key steps in antifungal host defense revealed fungal-specific signatures. As previously demonstrated, induced type I and Type II interferon-related pathways. In contrast, central pattern recognition receptor, reactive oxygen species production, and host glycolytic pathways were down-regulated in response to , which was associated with an ER-stress response. was identified to be uniquely regulated by and to control cytokine release in response to this fungus. In conclusion, our data reveals the transcriptional profiles induced by , and describes both the common and specific antifungal host responses that could be exploited for novel therapeutic strategies.
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http://dx.doi.org/10.1016/j.csbj.2020.12.036DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7817431PMC
December 2020

Aspergillus fumigatus tryptophan metabolic route differently affects host immunity.

Cell Rep 2021 Jan;34(4):108673

Department of Medicine and Surgery, University of Perugia, 06132 Perugia, Italy.

Indoleamine 2,3-dioxygenases (IDOs) degrade l-tryptophan to kynurenines and drive the de novo synthesis of nicotinamide adenine dinucleotide. Unsurprisingly, various invertebrates, vertebrates, and even fungi produce IDO. In mammals, IDO1 also serves as a homeostatic regulator, modulating immune response to infection via local tryptophan deprivation, active catabolite production, and non-enzymatic cell signaling. Whether fungal Idos have pleiotropic functions that impact on host-fungal physiology is unclear. Here, we show that Aspergillus fumigatus possesses three ido genes that are expressed under conditions of hypoxia or tryptophan abundance. Loss of these genes results in increased fungal pathogenicity and inflammation in a mouse model of aspergillosis, driven by an alternative tryptophan degradation pathway to indole derivatives and the host aryl hydrocarbon receptor. Fungal tryptophan metabolic pathways thus cooperate with the host xenobiotic response to shape host-microbe interactions in local tissue microenvironments.
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http://dx.doi.org/10.1016/j.celrep.2020.108673DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7844877PMC
January 2021

, One Uninucleate Species with Disparate Offspring.

J Fungi (Basel) 2021 Jan 6;7(1). Epub 2021 Jan 6.

Unité des Aspergillus, Institut Pasteur, 75015 Paris, France.

Establishment of a fungal infection due to relies on the efficient germination of the airborne conidia once they penetrate the respiratory tract. However, the features of conidial germination have been poorly explored and understood in this fungal species as well as in other species of filamentous fungi. We show here that the germination of is asynchronous. If the nutritional environment and extensive gene deletions can modify the germination parameters for , the asynchrony is maintained in all germinative conditions tested. Even though the causes for this asynchrony of conidial germination remain unknown, asynchrony is essential for the completion of the biological cycle of this filamentous fungus.
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http://dx.doi.org/10.3390/jof7010030DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7825634PMC
January 2021

Macrophages: Checking Toxicity of Fungal Metabolites in the Colon.

Trends Endocrinol Metab 2021 02 17;32(2):63-65. Epub 2020 Dec 17.

Institute of Molecular Biology and Biotechnology - Foundation for Research and Technology HELLAS (IMBB-FORTH), University of Crete, Heraklion, Crete, Greece. Electronic address:

It is well known that the intestine absorbs nutrients, electrolytes, and water. Chikina et al. recently demonstrated that it is also able to sense, recognize, and block the absorption of toxins through a very sophisticated interactive cellular cooperation between novel subpopulations of macrophages and epithelial cells.
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http://dx.doi.org/10.1016/j.tem.2020.11.012DOI Listing
February 2021

Galactosaminogalactan activates the inflammasome to provide host protection.

Nature 2020 Dec 2;588(7839):688-692. Epub 2020 Dec 2.

Department of Immunology, St Jude Children's Research Hospital, Memphis, TN, USA.

Inflammasomes are important sentinels of innate immune defence that are activated in response to diverse stimuli, including pathogen-associated molecular patterns (PAMPs). Activation of the inflammasome provides host defence against aspergillosis, which is a major health concern for patients who are immunocompromised. However, the Aspergillus fumigatus PAMPs that are responsible for inflammasome activation are not known. Here we show that the polysaccharide galactosaminogalactan (GAG) of A. fumigatus is a PAMP that activates the NLRP3 inflammasome. The binding of GAG to ribosomal proteins inhibited cellular translation machinery, and thus activated the NLRP3 inflammasome. The galactosamine moiety bound to ribosomal proteins and blocked cellular translation, which triggered activation of the NLRP3 inflammasome. In mice, a GAG-deficient Aspergillus mutant (Δgt4c) did not elicit protective activation of the inflammasome, and this strain exhibited enhanced virulence. Moreover, administration of GAG protected mice from colitis induced by dextran sulfate sodium in an inflammasome-dependent manner. Thus, ribosomes connect the sensing of this fungal PAMP to the activation of an innate immune response.
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http://dx.doi.org/10.1038/s41586-020-2996-zDOI Listing
December 2020

Galactomannan Produced by : An Update on the Structure, Biosynthesis and Biological Functions of an Emblematic Fungal Biomarker.

J Fungi (Basel) 2020 Nov 12;6(4). Epub 2020 Nov 12.

Institute of Molecular Biology and Biotechnology FORTH and School of Medicine, University of Crete Heraklion, 71003 Crete, Greece.

The galactomannan (GM) that is produced by the human fungal pathogen is an emblematic biomarker in medical mycology. The GM is composed of two monosaccharides: mannose and galactofuranose. The furanic configuration of galactose residues, absent in mammals, is responsible for the antigenicity of the GM and has favoured the development of ELISA tests to diagnose aspergillosis in immunocompromised patients. The GM that is produced by is a unique fungal polysaccharide containing a tetramannoside repeat unit and having three different forms: (i) membrane bound through a glycosylphosphatidylinositol (GPI)-anchor, (ii) covalently linked to β-1,3-glucans in the cell wall, or (iii) released in the culture medium as a free polymer. Recent studies have revealed the crucial role of the GM during vegetative and polarized fungal growth. This review highlights these recent data on its biosynthetic pathway and its biological functions during the saprophytic and pathogenic life of this opportunistic human fungal pathogen.
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http://dx.doi.org/10.3390/jof6040283DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7712326PMC
November 2020

The Role of RodA-Conserved Cysteine Residues in the Conidial Surface Organization.

J Fungi (Basel) 2020 Aug 26;6(3). Epub 2020 Aug 26.

Molecular Mycology Unit, Institut Pasteur, CNRS-UMR-2000, 10098 Paris, France.

Immune inertness of conidia is attributed to its surface rodlet-layer made up of RodAp, characterized by eight conserved cysteine residues forming four disulfide bonds. Earlier, we showed that the conserved cysteine residue point (ccrp mutations result in conidia devoid of the rodlet layer. Here, we extended our study comparing the surface organization and immunoreactivity of conidia carrying ccrp-mutations with the deletion mutant (∆). Western blot analysis using anti-RodAp antibodies indicated the absence of RodAp in the cytoplasm of ccrpmutant conidia. Immunolabeling revealed differential reactivity to conidial surface glucans, the ccrp-mutant conidia preferentially binding to α-(1,3)-glucan, ∆ conidia selectively bound to β-(1,3)-glucan; the parental strain conidia showed negative labeling. However, permeability of ccrp-mutants and ∆ was similar to the parental strain conidia. Proteomic analyses of the conidial surface exposed proteins of the ccrp-mutants showed more similarities with the parental strain, but were significantly different from the ∆. Ccrp-mutant conidia were less immunostimulatory compared to ∆ conidia. Our data suggest that (i) the conserved cysteine residues are essential for the trafficking of RodAp and the organization of the rodlet layer on the conidial surface, and (ii) targeted point mutation could be an alternative approach to study the role of fungal cell-wall genes in host-fungal interaction.
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http://dx.doi.org/10.3390/jof6030151DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7558875PMC
August 2020

Characterization of Extracellular Vesicles Produced by Aspergillus fumigatus Protoplasts.

mSphere 2020 08 12;5(4). Epub 2020 Aug 12.

Instituto de Microbiologia Paulo de Góes (IMPG), Universidade Federal do Rio de Janeiro, Rio de Janeiro, Brazil

Extracellular vesicles (EVs) are membranous compartments produced by yeast and mycelial forms of several fungal species. One of the difficulties in perceiving the role of EVs during the fungal life, and particularly in cell wall biogenesis, is caused by the presence of a thick cell wall. One alternative to have better access to these vesicles is to use protoplasts. This approach has been investigated here with , one of the most common opportunistic fungal pathogens worldwide. Analysis of regenerating protoplasts by scanning electron microscopy and fluorescence microscopy indicated the occurrence of outer membrane projections in association with surface components and the release of particles with properties resembling those of fungal EVs. EVs in culture supernatants were characterized by transmission electron microscopy and nanoparticle tracking analysis. Proteomic and glycome analysis of EVs revealed the presence of a complex array of enzymes related to lipid/sugar metabolism, pathogenic processes, and cell wall biosynthesis. Our data indicate that (i) EV production is a common feature of different morphological stages of this major fungal pathogen and (ii) protoplastic EVs are promising tools for undertaking studies of vesicle functions in fungal cells. Fungal cells use extracellular vesicles (EVs) to export biologically active molecules to the extracellular space. In this study, we used protoplasts of , a major fungal pathogen, as a model to evaluate the role of EV production in cell wall biogenesis. Our results demonstrated that wall-less exports plasma membrane-derived EVs containing a complex combination of proteins and glycans. Our report is the first to characterize fungal EVs in the absence of a cell wall. Our results suggest that protoplasts represent a promising model for functional studies of fungal vesicles.
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http://dx.doi.org/10.1128/mSphere.00476-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7426166PMC
August 2020

RNA-dependent sterol aspartylation in fungi.

Proc Natl Acad Sci U S A 2020 06 15;117(26):14948-14957. Epub 2020 Jun 15.

Université de Strasbourg, CNRS, Génétique Moléculaire, Génomique, Microbiologie, UMR 7156, 67084 Strasbourg Cedex, France;

Diverting aminoacyl-transfer RNAs (tRNAs) from protein synthesis is a well-known process used by a wide range of bacteria to aminoacylate membrane constituents. By tRNA-dependently adding amino acids to glycerolipids, bacteria change their cell surface properties, which intensifies antimicrobial drug resistance, pathogenicity, and virulence. No equivalent aminoacylated lipids have been uncovered in any eukaryotic species thus far, suggesting that tRNA-dependent lipid remodeling is a process restricted to prokaryotes. We report here the discovery of ergosteryl-3β--l-aspartate (Erg-Asp), a conjugated sterol that is produced by the tRNA-dependent addition of aspartate to the 3β-OH group of ergosterol, the major sterol found in fungal membranes. In fact, Erg-Asp exists in the majority of "higher" fungi, including species of biotechnological interest, and, more importantly, in human pathogens like We show that a bifunctional enzyme, ergosteryl-3β--l-aspartate synthase (ErdS), is responsible for Erg-Asp synthesis. ErdS corresponds to a unique fusion of an aspartyl-tRNA synthetase-that produces aspartyl-tRNA (Asp-tRNA)-and of a , which actually transfers aspartate from Asp-tRNA onto ergosterol. We also uncovered that removal of the Asp modifier from Erg-Asp is catalyzed by a second enzyme, ErdH, that is a genuine Erg-Asp hydrolase participating in the turnover of the conjugated sterol in vivo. Phylogenomics highlights that the entire Erg-Asp synthesis/degradation pathway is conserved across "higher" fungi. Given the central roles of sterols and conjugated sterols in fungi, we propose that this tRNA-dependent ergosterol modification and homeostasis system might have broader implications in membrane remodeling, trafficking, antimicrobial resistance, or pathogenicity.
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http://dx.doi.org/10.1073/pnas.2003266117DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7334510PMC
June 2020

Functional Coupling between the Unfolded Protein Response and Endoplasmic Reticulum/Golgi Ca-ATPases Promotes Stress Tolerance, Cell Wall Biosynthesis, and Virulence of Aspergillus fumigatus.

mBio 2020 06 2;11(3). Epub 2020 Jun 2.

Department of Pathology & Laboratory Medicine, University of Cincinnati College of Medicine, Cincinnati, Ohio, USA

Many species of pathogenic fungi deploy the unfolded protein response (UPR) to expand the folding capacity of the endoplasmic reticulum (ER) in proportion to the demand for virulence-related proteins that traffic through the secretory pathway. Although Ca plays a pivotal role in ER function, the mechanism by which transcriptional upregulation of the protein folding machinery is coordinated with Ca homeostasis is incompletely understood. In this study, we investigated the link between the UPR and genes encoding P-type Ca-ATPases in the human-pathogenic mold We demonstrate that acute ER stress increases transcription of the gene, encoding a member of the sarco/endoplasmic reticulum Ca-ATPase (SERCA) family, as well as that of , encoding a secretory pathway Ca-ATPase (SPCA) in the Golgi membrane. Loss of the UPR transcription factor HacA prevented the induction of and transcription during ER stress, defining these ER/Golgi Ca pumps as novel downstream targets of this pathway. While deletion of alone caused no major deficiencies, a Δ/Δ mutant displayed a severe polarity defect, was hypersensitive to ER stress, and showed attenuated virulence. In addition, cell wall analyses revealed a striking reduction in mannose levels in the absence of both Ca pumps. The Δ mutant was hypersensitive to agents that block calcineurin-dependent signaling, consistent with a functional coupling between the UPR and Ca homeostasis. Together, these findings demonstrate that the UPR integrates the need for increased levels of chaperone and folding enzymes with an influx of Ca into the secretory pathway to support fungal growth, stress adaptation, and pathogenicity. The UPR is an intracellular signal transduction pathway that maintains homeostasis of the ER. The pathway is also tightly linked to the expression of virulence-related traits in diverse species of human-pathogenic and plant-pathogenic fungal species, including the predominant mold pathogen infecting humans, Despite advances in the understanding of UPR signaling, the linkages and networks that are governed by this pathway are not well defined. In this study, we revealed that the UPR is a major driving force for stimulating Ca influx at the ER and Golgi membranes and that the coupling between the UPR and Ca import is important for virulence, cell wall biosynthesis, and resistance to antifungal compounds that inhibit Ca signaling.
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http://dx.doi.org/10.1128/mBio.01060-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7267887PMC
June 2020

GPI Anchored Proteins in Aspergillus fumigatus and Cell Wall Morphogenesis.

Curr Top Microbiol Immunol 2020 ;425:167-186

Aspergillus Unit, Institut Pasteur, 25 Rue Du Docteur Roux, 75015, Paris, France.

Glycosylphosphatidylinositol (GPI) anchored proteins are a class of proteins attached to the extracellular leaflet of the plasma membrane via a post-translational modification, the glycolipid anchor. GPI anchored proteins are expressed in all eukaryotes, from fungi to plants and animals. They display very diverse functions ranging from enzymatic activity, signaling, cell adhesion, cell wall metabolism, and immune response. In this review, we investigated for the first time an exhaustive list of all the GPI anchored proteins present in the Aspergillus fumigatus genome. An A. fumigatus mutant library of all the genes that encode in silico identified GPI anchored proteins has been constructed and the phenotypic analysis of all these mutants has been characterized including their growth, conidial viability or morphology, adhesion and the ability to form biofilms. We showed the presence of different fungal categories of GPI anchored proteins in the A. fumigatus genome associated to their role in cell wall remodeling, adhesion, and biofilm formation.
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http://dx.doi.org/10.1007/82_2020_207DOI Listing
September 2020

Revisiting Old Questions and New Approaches to Investigate the Fungal Cell Wall Construction.

Curr Top Microbiol Immunol 2020 ;425:331-369

Institute of Molecular Biology and Biotechnology of the Foundation for Research and Technology Hellas (IMBB-FORTH), Heraklion, Greece.

The beginning of our understanding of the cell wall construction came from the work of talented biochemists in the 70-80's. Then came the era of sequencing. Paradoxically, the accumulation of fungal genomes complicated rather than solved the mystery of cell wall construction, by revealing the involvement of a much higher number of proteins than originally thought. The situation has become even more complicated since it is now recognized that the cell wall is an organelle whose composition continuously evolves with the changes in the environment or with the age of the fungal cell. The use of new and sophisticated technologies to observe cell wall construction at an almost atomic scale should improve our knowledge of the cell wall construction. This essay will present some of the major and still unresolved questions to understand the fungal cell wall biosynthesis and some of these exciting futurist approaches.
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http://dx.doi.org/10.1007/82_2020_209DOI Listing
September 2020

Cell Wall Composition Heterogeneity between Single Cells in Aspergillus fumigatus Leads to Heterogeneous Behavior during Antifungal Treatment and Phagocytosis.

mBio 2020 05 12;11(3). Epub 2020 May 12.

Manchester Fungal Infection Group (MFIG), Infection, Immunity and Respiratory Medicine, The University of Manchester, Manchester, United Kingdom.

can cause a variety of lung diseases in immunocompromised patients, including life-threatening invasive aspergillosis. There are only three main classes of antifungal drugs currently used to treat aspergillosis, and antifungal resistance is increasing. Experimental results in fungal biology research are usually obtained as average measurements across whole populations while ignoring what is happening at the single cell level. In this study, we show that conidia with the same genetic background in the same cell population at a similar developmental stage show heterogeneity in their cell wall labeling at the single cell level. We present a rigorous statistical method, newly applied to quantify the level of cell heterogeneity, which allows for direct comparison of the heterogeneity observed between treatments. We show the extent of cell wall labeling heterogeneity in dormant conidia and how the level of heterogeneity changes during germination. The degree of heterogeneity is influenced by deletions of cell wall synthesizing genes and environmental conditions, including medium composition, method of inoculation, age of conidia, and the presence of antifungals. This heterogeneity results in subpopulations of germinating conidia with heterogeneous fitness to the antifungal caspofungin, which targets cell wall synthesis and heterogeneous sensitivity of dormant conidia to phagocytosis by macrophages. The fungus can cause invasive lung diseases in immunocompromised patients resulting in high mortality. Treatment using antifungal compounds is often unsuccessful. Average population measurements hide what is happening at the individual cell level. We set out to test what impact individual differences between the cell walls of fungal conidia have on their behavior. We show that a population of cells having the same genetic background gives rise to subpopulations of cells that exhibit distinct behavior (phenotypic heterogeneity). This cell heterogeneity is dependent on the strain type, gene deletions, cell age, and environmental conditions. By looking at the individual cell level, we discovered subpopulations of cells that show differential fitness during antifungal treatment and uptake by immune cells.
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http://dx.doi.org/10.1128/mBio.03015-19DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7218287PMC
May 2020

Phagosomal removal of fungal melanin reprograms macrophage metabolism to promote antifungal immunity.

Nat Commun 2020 05 8;11(1):2282. Epub 2020 May 8.

Life and Health Sciences Research Institute (ICVS), School of Medicine, University of Minho, 4710-057, Braga, Portugal.

In response to infection, macrophages adapt their metabolism rapidly to enhance glycolysis and fuel specialized antimicrobial effector functions. Here we show that fungal melanin is an essential molecule required for the metabolic rewiring of macrophages during infection with the fungal pathogen Aspergillus fumigatus. Using pharmacological and genetic tools, we reveal a molecular link between calcium sequestration by melanin inside the phagosome and induction of glycolysis required for efficient innate immune responses. By remodeling the intracellular calcium machinery and impairing signaling via calmodulin, melanin drives an immunometabolic signaling axis towards glycolysis with activation of hypoxia-inducible factor 1 subunit alpha (HIF-1α) and phagosomal recruitment of mammalian target of rapamycin (mTOR). These data demonstrate a pivotal mechanism in the immunometabolic regulation of macrophages during fungal infection and highlight the metabolic repurposing of immune cells as a potential therapeutic strategy.
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http://dx.doi.org/10.1038/s41467-020-16120-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7210971PMC
May 2020

What Are the Functions of Chitin Deacetylases in ?

Front Cell Infect Microbiol 2020 6;10:28. Epub 2020 Feb 6.

Aspergillus Unit, Pasteur Institut, Paris, France.

Deacetylation of chitin by chitin deacetylases (Cda) results in the formation of chitosan. Chitosan, a polymer of β1,4 linked glucosamine, plays multiple roles in the function of the fungal cell wall, including virulence and evasion of host immune responses. In this study, the roles of chitosan and putative s in cell wall structure and virulence of were investigated. Low levels of chitosan were found in the conidial and cell wall of . Seven putative genes were identified, disrupted and the phenotype of the single mutants and the septuple mutants were investigated. No alterations in fungal cell wall chitosan levels, changes in fungal growth or alterations in virulence were detected in the single or septuple Δ mutant strains. Collectively, these results suggest that chitosan is a minority component of the cell wall, and that the seven candidate Cda proteins do not play major roles in fungal cell wall synthesis or virulence. However, Cda2 is involved in conidiation, suggesting that this enzyme may play a role in N-acetyl-glucosamine metabolism.
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http://dx.doi.org/10.3389/fcimb.2020.00028DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7016196PMC
February 2020

Infection in Humans With STAT3-Deficiency Is Associated With Defective Interferon-Gamma and Th17 Responses.

Front Immunol 2020 28;11:38. Epub 2020 Jan 28.

Unité des Aspergillus, Institut Pasteur, Paris, France.

In humans, loss-of-function mutation in the gene is frequently associated with susceptibility to bacterial as well as fungal infections including aspergillosis, although its pathogenesis remains largely unknown. In the present study, we investigated the immune responses obtained after stimulation with in STAT3-deficient patients. conidial killing efficiencies of both monocytes and neutrophils isolated from whole blood samples of STAT3-deficient patients were not different compared to those of healthy controls. After stimulation with conidia, lower concentrations of adaptive cytokines (IFN-γ, IL-17 and IL-22) were secreted by peripheral blood mononuclear cells from STAT3-deficient patients compared to those from healthy controls. Moreover, the frequency of IFN-γ and IL-17 producing CD4+ T cells was lower in STAT3-deficient patients vs. healthy controls. Among the STAT3-deficient patients, those with aspergillosis showed further lower secretion of IFN-γ upon stimulation of their PBMCs with conidia compared to the patients without aspergillosis. Together, our study indicated that STAT3-deficiency leads to a defective adaptive immune response against infection, particularly with a lower IFN-γ and IL-17 responses in those with aspergillosis, suggesting potential therapeutic benefit of recombinant IFN-γ in STAT3-deficient patients with aspergillosis.
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http://dx.doi.org/10.3389/fimmu.2020.00038DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6997434PMC
February 2021

Calcineurin A Is Essential in the Regulation of Asexual Development, Stress Responses and Pathogenesis in .

Front Microbiol 2019 21;10:3094. Epub 2020 Jan 21.

Department of Dermatology and Venereology, The First Affiliated Hospital of Guangxi Medical University, Nanning, China.

is a common cause of infection in immunocompromised patients in Southeast Asia and Southern China. The pathogenicity of depends on the ability of the fungus to survive the cytotoxic processes of the host immune system and grow inside host macrophages. These mechanisms that allow to survive macrophage-induced death are poorly understood. In this study, we examined the role of a calcineurin homolog () from during growth, morphogenesis and infection. Deletion of the gene in resulted in a strain with significant defects in conidiation, germination, morphogenesis, cell wall integrity, and resistance to various stressors. The Δ mutant showed a lower minimal inhibitory concentration (MIC) against caspofungin (16 μg/ml to 2 μg/ml) and micafungin (from 32 μg/ml to 4 μg/ml) compared with the wild-type. These results suggest that targeting calcineurin in combination with echinocandin treatment may be effective for life-threatening systemic infection. Importantly, the mutant was incapable of adapting to the macrophage environment and displayed virulence defects in a mouse model of invasive talaromycosis. For the first time, a role has been shown for in the morphology and pathogenicity of a dimorphic pathogenic filamentous fungus.
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http://dx.doi.org/10.3389/fmicb.2019.03094DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6985273PMC
January 2020

The negative cofactor 2 complex is a key regulator of drug resistance in Aspergillus fumigatus.

Nat Commun 2020 01 22;11(1):427. Epub 2020 Jan 22.

Manchester Fungal Infection Group, Division of Infection, Immunity and Respiratory Medicine, Faculty of Biology, Medicine and Health, University of Manchester, CTF Building, 46 Grafton Street, Manchester, M13 9NT, UK.

The frequency of antifungal resistance, particularly to the azole class of ergosterol biosynthetic inhibitors, is a growing global health problem. Survival rates for those infected with resistant isolates are exceptionally low. Beyond modification of the drug target, our understanding of the molecular basis of azole resistance in the fungal pathogen Aspergillus fumigatus is limited. We reasoned that clinically relevant antifungal resistance could derive from transcriptional rewiring, promoting drug resistance without concomitant reductions in pathogenicity. Here we report a genome-wide annotation of transcriptional regulators in A. fumigatus and construction of a library of 484 transcription factor null mutants. We identify 12 regulators that have a demonstrable role in itraconazole susceptibility and show that loss of the negative cofactor 2 complex leads to resistance, not only to the azoles but also the salvage therapeutics amphotericin B and terbinafine without significantly affecting pathogenicity.
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http://dx.doi.org/10.1038/s41467-019-14191-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7194077PMC
January 2020

Potential of Chemically Synthesized Oligosaccharides To Define the Carbohydrate Moieties of the Fungal Cell Wall Responsible for the Human Immune Response, Using Aspergillus fumigatus Galactomannan as a Model.

mSphere 2020 01 8;5(1). Epub 2020 Jan 8.

Laboratory of Glycoconjugate Chemistry, N. D. Zelinsky Institute of Organic Chemistry, Russian Academy of Sciences, Moscow, Russia

Methodologies to identify epitopes or ligands of the fungal cell wall polysaccharides influencing the immune response of human pathogens have to date been imperfect. Using the galactomannan (GM) of as a model, we have shown that synthetic oligosaccharides of distinct structures representing key fragments of cell wall polysaccharides are the most precise tools to study the serological and immunomodulatory properties of a fungal polysaccharide.
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http://dx.doi.org/10.1128/mSphere.00688-19DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6952192PMC
January 2020

Biotinylated Oligo-α-(1 → 4)-d-galactosamines and Their N-Acetylated Derivatives: α-Stereoselective Synthesis and Immunology Application.

J Am Chem Soc 2020 01 10;142(3):1175-1179. Epub 2020 Jan 10.

N.D. Zelinsky Institute of Organic Chemistry , Russian Academy of Sciences , Leninsky Prospect 47 , 119991 Moscow , Russian Federation.

Using 3--benzoyl-4,6--di--butylsilylidene-2-azido-2-deoxy-selenogalactoside, biotinylated oligo-α-(1 → 4)-d-galactosamines comprising from two to six GalN units were prepared for the first time together with their N-acetylated derivatives. The combination of blocking groups used herein provided stereocontrol for the α-stereospecific glycosylation, to show also high efficiency of phenyl 2-azido-2-deoxy-selenogalactosides as glycosyl donors. The obtained glycoconjugates are related to fragments of exopolysaccharide galactosaminogalactan (GG) found in , which is the most important airborne human fungal pathogen in industrialized countries. The synthesized glycoconjugates were arrayed on streptavidin-coated plates and used to investigate the GG epitopes recognized by mouse monoclonal antibodies against GG and by human antibodies in the sera of patients with aspergillosis. The obtained data showed that the oligo-α-(1 → 4)-d-galactosamines and their N-acetylated derivatives allowed the first precise analysis of the specificity of the antibody responses to this extremely complex fungal polysaccharide.
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http://dx.doi.org/10.1021/jacs.9b11703DOI Listing
January 2020

Genome Sequences of Two Strains of the Food Spoilage Mold Aspergillus fischeri.

Microbiol Resour Announc 2019 Dec 12;8(50). Epub 2019 Dec 12.

Department of Food Science, University of Massachusetts, Amherst, Massachusetts, USA

is a common food spoilage fungus and a close relative of the opportunistic human pathogen Here, we sequenced the genomes of two isolates of to build resources for comparative genomics and to aid in differentiation between subspecies.
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http://dx.doi.org/10.1128/MRA.01328-19DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6908800PMC
December 2019

Definition of the Anti-inflammatory Oligosaccharides Derived From the Galactosaminogalactan (GAG) From .

Front Cell Infect Microbiol 2019 6;9:365. Epub 2019 Nov 6.

Unité des Aspergillus, Institut Pasteur, Paris, France.

Galactosaminogalactan (GAG) is an insoluble aminosugar polymer produced by and has anti-inflammatory properties. Here, the minimum glycosidic sequences required for the induction of IL-1Ra by peripheral blood mononuclear cells (PBMCs) was investigated. Using chemical degradation of native GAG to isolate soluble oligomers, we have found that the de--acetylation of galactosamine residues and the size of oligomer are critical for the immune response. A minimal oligomer size of 20 galactosamine residues is required for the anti-inflammatory response but the presence of galactose residues is not necessary. In a Dextran sulfate induced colitis mouse model, a fraction of de--acetylated oligomers of 13 < dp < 20 rescue inflammatory damage like the native GAG polymer in an IL-1Ra dependent pathway. Our results demonstrate the therapeutic suitability of water-soluble GAG oligosaccharides in IL-1 mediated hyper-inflammatory diseases and suggest that α-1,4-galactosamine oligomers chemically synthesized could represent new anti-inflammatory glycodrugs.
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http://dx.doi.org/10.3389/fcimb.2019.00365DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6851199PMC
July 2020

Aspergillus fumigatus and Aspergillosis in 2019.

Clin Microbiol Rev 2019 12 13;33(1). Epub 2019 Nov 13.

School of Medicine, University of Crete, Heraklion, Crete, Greece.

is a saprotrophic fungus; its primary habitat is the soil. In its ecological niche, the fungus has learned how to adapt and proliferate in hostile environments. This capacity has helped the fungus to resist and survive against human host defenses and, further, to be responsible for one of the most devastating lung infections in terms of morbidity and mortality. In this review, we will provide (i) a description of the biological cycle of ; (ii) a historical perspective of the spectrum of aspergillus disease and the current epidemiological status of these infections; (iii) an analysis of the modes of immune response against in immunocompetent and immunocompromised patients; (iv) an understanding of the pathways responsible for fungal virulence and their host molecular targets, with a specific focus on the cell wall; (v) the current status of the diagnosis of different clinical syndromes; and (vi) an overview of the available antifungal armamentarium and the therapeutic strategies in the clinical context. In addition, the emergence of new concepts, such as nutritional immunity and the integration and rewiring of multiple fungal metabolic activities occurring during lung invasion, has helped us to redefine the opportunistic pathogenesis of .
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http://dx.doi.org/10.1128/CMR.00140-18DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6860006PMC
December 2019

-Derived Volatile Sulfur Compounds Promote Distal Growth and a Synergistic Pathogen-Pathogen Interaction That Increases Pathogenicity in Co-infection.

Front Microbiol 2019 9;10:2311. Epub 2019 Oct 9.

Manchester Fungal Infection Group, Division of Infection, Immunity and Respiratory Medicine, Faculty of Biology, Medicine and Health, School of Biological Sciences, University of Manchester, Manchester Academic Health Science Centre, Manchester, United Kingdom.

Pathogen-pathogen interactions in polymicrobial infections are known to directly impact, often to worsen, disease outcomes. For example, co-infection with and , respectively the most common bacterial and fungal pathogens isolated from cystic fibrosis (CF) airways, leads to a worsened prognosis. Recent studies of microbial cross-talk demonstrated that -derived volatile sulfur compounds (VSCs) can promote growth . However, the mechanistic basis of such cross-talk and its physiological relevance during co-infection remains unknown. In this study we combine genetic approaches and GC-MS-mediated volatile analysis to show that assimilates VSCs via cysteine (CysB)- or homocysteine (CysD)-synthase. This process is essential for utilization of VSCs as sulfur sources, since -derived VSCs trigger growth of wild-type, but not of a ΔΔ mutant, on sulfur-limiting media. produces VSCs when infecting and co-infection with in this model results in a synergistic increase in mortality and of fungal and bacterial burdens. Interestingly, the increment in mortality is much greater with the wild-type than with the ΔΔ mutant. Therefore, ability to assimilate derived VSCs significantly triggers a synergistic association that increases the pathobiology of infection. Finally, we show that can promote fungal growth when growing on substrates that resemble the lung environment, which suggests that this volatile based synergism is likely to occur during co-infection of the human respiratory airways.
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http://dx.doi.org/10.3389/fmicb.2019.02311DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6794476PMC
October 2019

Aspergillus fumigatus exoβ(1-3)glucanases family GH55 are essential for conidial cell wall morphogenesis.

Cell Microbiol 2019 12 30;21(12):e13102. Epub 2019 Aug 30.

Aspergillus Unit, Institut Pasteur, Paris, France.

The cell wall of Aspergillus fumigatus is predominantly composed of polysaccharides. The central fibrillar core of the cell wall is composed of a branched β(1-3)glucan, to which the chitin and the galactomannan are covalently bound. Softening of the cell wall is an essential event during fungal morphogenesis, wherein rigid cell wall structures are cleaved by glycosyl hydrolases. In this study, we characterised the role of the glycosyl hydrolase GH55 members in A. fumigatus fungal morphogenesis. We showed that deletion of the six genes of the GH55 family stopped conidial cell wall maturation at the beginning of the development process, leading to abrogation of conidial separation: the shape of conidia became ovoid, and germination was delayed. In conclusion, the reorganisation and structuring of the conidial cell wall mediated by members of the GH55 family is essential for their maturation, normal dissemination, and germination.
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http://dx.doi.org/10.1111/cmi.13102DOI Listing
December 2019

The Glycosylphosphatidylinositol-Anchored Family Is Essential for the Insertion of Galactomannan into the β-(1,3)-Glucan-Chitin Core of the Cell Wall of Aspergillus fumigatus.

mSphere 2019 07 31;4(4). Epub 2019 Jul 31.

Unité des Aspergillus, Institut Pasteur, Paris, France

The fungal cell wall is a complex and dynamic entity essential for the development of fungi. It is composed mainly of polysaccharides that are synthetized by protein complexes. At the cell wall level, enzyme activities are involved in postsynthesis polysaccharide modifications such as cleavage, elongation, branching, and cross-linking. Glycosylphosphatidylinositol (GPI)-anchored proteins have been shown to participate in cell wall biosynthesis and specifically in polysaccharide remodeling. Among these proteins, the DFG family plays an essential role in controlling polar growth in yeast. In the filamentous fungus and opportunistic human pathogen , the gene family contains seven orthologous genes among which only six are expressed under growth conditions. Deletions of single genes revealed that plays the most important morphogenetic role in this gene family. A sextuple-deletion mutant resulting from the deletion of all expressed genes did not contain galactomannan in the cell wall and has severe growth defects. This study has shown that DFG members are absolutely necessary for the insertion of galactomannan into the cell wall of and that the proper cell wall localization of the galactomannan is essential for correct fungal morphogenesis in The fungal cell wall is a complex and dynamic entity essential for the development of fungi. It is composed mainly of polysaccharides that are synthetized by protein complexes. Enzymes involved in postsynthesis polysaccharide modifications, such as cleavage, elongation, branching, and cross-linking, are essential for fungal life. Here, we investigated in the role of the members of the Dfg family, one of the 4 GPI-anchored protein families common to yeast and molds involved in cell wall remodeling. Molecular and biochemical approaches showed that DFG members are required for filamentous growth, conidiation, and cell wall organization and are essential for the life of this fungal pathogen.
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http://dx.doi.org/10.1128/mSphere.00397-19DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6669337PMC
July 2019

Interactions between and Pulmonary Bacteria: Current State of the Field, New Data, and Future Perspective.

J Fungi (Basel) 2019 Jun 12;5(2). Epub 2019 Jun 12.

Aspergillus Unit, Institut Pasteur, 75015 Paris, France.

and are central fungal and bacterial members of the pulmonary microbiota. The interactions between and have only just begun to be explored. A balance between inhibitory and stimulatory effects on fungal growth was observed in mixed cultures. Negative interactions have been seen for homoserine-lactones, pyoverdine and pyochelin resulting from iron starvation and intracellular inhibitory reactive oxidant production. In contrast, several types of positive interactions were recognized. Dirhamnolipids resulted in the production of a thick fungal cell wall, allowing the fungus to resist stress. Phenazines and pyochelin favor iron uptake for the fungus. is able to use bacterial volatiles to promote its growth. The immune response is also differentially regulated by co-infections.
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http://dx.doi.org/10.3390/jof5020048DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6617096PMC
June 2019

Chitinases Play a Key Role in Stipe Cell Wall Extension in the Mushroom .

Appl Environ Microbiol 2019 08 18;85(15). Epub 2019 Jul 18.

Jiangsu Key Laboratory for Microbes and Microbial Functional Genomics, Jiangsu Engineering and Technology Research Center for Industrialization of Microbial Resources, College of Life Science, Nanjing Normal University, Nanjing, People's Republic of China

The elongation growth of the mushroom stipe is a characteristic but not well-understood morphogenetic event of basidiomycetes. We found that extending native stipe cell walls of were associated with the release of -acetylglucosamine and chitinbiose and with chitinase activity. Two chitinases among all detected chitinases from , ChiE1 and ChiIII, reconstituted heat-inactivated stipe wall extension and released -acetylglucosamine and chitinbiose. Interestingly, both ChiE1 and ChiIII hydrolyze insoluble crystalline chitin powder, while other chitinases do not, suggesting that crystalline chitin components of the stipe cell wall are the target of action for ChiE1 and ChiIII. ChiE1- or ChiIII-reconstituted heat-inactivated stipe walls showed maximal extension activity at pH 4.5, consistent with the optimal pH for native stipe wall extension ; ChiE1- or ChiIII-reconstituted heat-inactivated stipe wall extension activities were associated with stipe elongation growth regions; and the combination of ChiE1 and ChiIII showed a synergism to reconstitute heat-inactivated stipe wall extension at a low action concentration. Field emission scanning electron microscopy (FESEM) images showed that the inner surface of acid-induced extended native stipe cell walls and ChiE1- or ChiIII-reconstituted extended heat-inactivated stipe cell walls exhibited a partially broken parallel microfibril architecture; however, these broken transversely arranged microfibrils were not observed in the unextended stipe cell walls that were induced by neutral pH buffer or heat inactivation. Double knockdown of ChiE1 and ChiIII resulted in the reduction of stipe elongation, mycelium growth, and heat-sensitive cell wall extension of native stipes. These results indicate a chitinase-hydrolyzing mechanism for stipe cell wall extension. A remarkable feature in the development of basidiomycete fruiting bodies is stipe elongation growth that results primarily from manifold cell elongation. Some scientists have suggested that stipe elongation is the result of enzymatic hydrolysis of cell wall polysaccharides, while other scientists have proposed the possibility that stipe elongation results from nonhydrolytic disruption of the hydrogen bonds between cell wall polysaccharides. Here, we show direct evidence for a chitinase-hydrolyzing mechanism of stipe cell wall elongation in the model mushroom that is different from the expansin nonhydrolysis mechanism of plant cell wall extension. We presumed that in the growing stipe cell walls, parallel chitin microfibrils are tethered by β-1,6-branched β-1,3-glucans, and that the breaking of the tether by chitinases leads to separation of these microfibrils to increase their spacing for insertion of new synthesized chitin and β-1,3-glucans under turgor pressure .
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http://dx.doi.org/10.1128/AEM.00532-19DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6643254PMC
August 2019

β-Glucan Grafted Microcapsule, a Tool for Studying the Immunomodulatory Effect of Microbial Cell Wall Polysaccharides.

Bioconjug Chem 2019 06 7;30(6):1788-1797. Epub 2019 Jun 7.

Unité des Aspergillus , Institut Pasteur , Paris 75015 , France.

β-(1,3)-Glucan is one of the antigenic components of the bacterial as well as fungal cell wall. We designed microcapsules (MCs) ligated with β-(1,3)-glucan, to study its immunomodulatory effect. The MCs were obtained by interfacial polycondensation between diacyl chloride (sebacoyl chloride and terephtaloyl chloride) and diethylenetriamine in organic and aqueous phases, respectively. Planar films were first designed to optimize monomer compositions and to examine the kinetics of film formation. MCs with aqueous fluorescent core were then obtained upon controlled emulsification-polycondensation reactions using optimized monomer compositions and adding fluorescein into the aqueous phase. The selected MC-formulation was grafted with Curdlan, a linear β-(1,3)-glucan from  Agrobacterium species or branched β-(1,3)-glucan isolated from the cell wall of Aspergillus fumigatus. These β-(1,3)-glucan grafted MCs were phagocytosed by human monocyte-derived macrophages, and stimulated cytokine secretion. Moreover, the blocking of dectin-1, a β-(1,3)-glucan recognizing receptor, did not completely inhibit the phagocytosis of these β-(1,3)-glucan grafted MCs, suggesting the involvement of other receptors in the recognition and uptake of β-(1,3)-glucan. Overall, grafted MCs are a useful tool for the study of the mechanism of phagocytosis and immunomodulatory effect of the microbial polysaccharides.
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http://dx.doi.org/10.1021/acs.bioconjchem.9b00304DOI Listing
June 2019