Publications by authors named "Mark S Gresnigt"

60 Publications

Human albumin enhances the pathogenic potential of Candida glabrata on vaginal epithelial cells.

PLoS Pathog 2021 10 28;17(10):e1010037. Epub 2021 Oct 28.

Department of Microbial Pathogenicity Mechanisms, Leibniz Institute for Natural Product Research and Infection Biology, Hans Knoell Institute, Jena, Germany.

The opportunistic pathogen Candida glabrata is the second most frequent causative agent of vulvovaginal candidiasis (VVC), a disease that affects 70-75% of women at least once during their life. However, C. glabrata is almost avirulent in mice and normally incapable of inflicting damage to vaginal epithelial cells in vitro. We thus proposed that host factors present in vivo may influence C. glabrata pathogenicity. We, therefore, analyzed the impact of albumin, one of the most abundant proteins of the vaginal fluid. The presence of human, but not murine, albumin dramatically increased the potential of C. glabrata to damage vaginal epithelial cells. This effect depended on macropinocytosis-mediated epithelial uptake of albumin and subsequent proteolytic processing. The enhanced pathogenicity of C. glabrata can be explained by a combination of beneficial effects for the fungus, which includes an increased access to iron, accelerated growth, and increased adhesion. Screening of C. glabrata deletion mutants revealed that Hap5, a key regulator of iron homeostasis, is essential for the albumin-augmented damage potential. The albumin-augmented pathogenicity was reversed by the addition of iron chelators and a similar increase in pathogenicity was shown by increasing the iron availability, confirming a key role of iron. Accelerated growth not only led to higher cell numbers, but also to increased fungal metabolic activity and oxidative stress resistance. Finally, the albumin-driven enhanced damage potential was associated with the expression of distinct C. glabrata virulence genes. Transcriptional responses of the epithelial cells suggested an unfolded protein response (UPR) and ER-stress responses combined with glucose starvation induced by fast growing C. glabrata cells as potential mechanisms by which cytotoxicity is mediated.Collectively, we demonstrate that albumin augments the pathogenic potential of C. glabrata during interaction with vaginal epithelial cells. This suggests a role for albumin as a key player in the pathogenesis of VVC.
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http://dx.doi.org/10.1371/journal.ppat.1010037DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8577789PMC
October 2021

Candida albicans-induced leukotriene biosynthesis in neutrophils is restricted to the hyphal morphology.

FASEB J 2021 10;35(10):e21820

Department of Pharmaceutical/Medicinal Chemistry, Institute of Pharmacy, Greifswald University, Greifswald, Germany.

Neutrophils are the most abundant leukocytes in circulation playing a key role in acute inflammation during microbial infections. Phagocytosis, one of the crucial defence mechanisms of neutrophils against pathogens, is amplified by chemotactic leukotriene (LT)B , which is biosynthesized via 5-lipoxygenase (5-LOX). However, extensive liberation of LTB can be destructive by over-intensifying the inflammatory process. While enzymatic biosynthesis of LTB is well characterized, less is known about molecular mechanisms that activate 5-LOX and lead to LTB formation during host-pathogen interactions. Here, we investigated the ability of the common opportunistic fungal pathogen Candida albicans to induce LTB formation in neutrophils, and elucidated pathogen-mediated drivers and cellular processes that activate this pathway. We revealed that C. albicans-induced LTB biosynthesis requires both the morphological transition from yeast cells to hyphae and the expression of hyphae-associated genes, as exclusively viable hyphae or yeast-locked mutant cells expressing hyphae-associated genes stimulated 5-LOX by [Ca ] mobilization and p38 MAPK activation. LTB biosynthesis was orchestrated by synergistic activation of dectin-1 and Toll-like receptor 2, and corresponding signaling via SYK and MYD88, respectively. Conclusively, we report hyphae-specific induction of LTB biosynthesis in human neutrophils. This highlights an expanding role of neutrophils during inflammatory processes in the response to C. albicans infections.
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http://dx.doi.org/10.1096/fj.202100516RRDOI Listing
October 2021

Candida albicans colonization of the gastrointestinal tract: A double-edged sword.

PLoS Pathog 2021 07 22;17(7):e1009710. Epub 2021 Jul 22.

Departamento de Microbiología y Parasitología-IRYCIS, Facultad de Farmacia, Universidad Complutense de Madrid, Madrid, Spain.

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http://dx.doi.org/10.1371/journal.ppat.1009710DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8297749PMC
July 2021

Uncoupling of IL-6 signaling and LC3-associated phagocytosis drives immunoparalysis during sepsis.

Cell Host Microbe 2021 Aug 1;29(8):1277-1293.e6. Epub 2021 Jul 1.

Laboratory of Clinical Microbiology and Microbial Pathogenesis, School of Medicine, University of Crete, Voutes, 71110 Heraklion, Crete, Greece; Institute of Molecular Biology and Biotechnology, Foundation for Research and Technology, 71300 Heraklion, Crete, Greece. Electronic address:

Immune deactivation of phagocytes is a central event in the pathogenesis of sepsis. Herein, we identify a master regulatory role of IL-6 signaling on LC3-associated phagocytosis (LAP) and reveal that uncoupling of these two processes during sepsis induces immunoparalysis in monocytes/macrophages. In particular, we demonstrate that activation of LAP by the human fungal pathogen Aspergillus fumigatus depends on ERK1/2-mediated phosphorylation of p47phox subunit of NADPH oxidase. Physiologically, autocrine IL-6/JAK2/Ninein axis orchestrates microtubule organization and dynamics regulating ERK recruitment to the phagosome and LC3 phagosome (LAPosome) formation. In sepsis, loss of IL-6 signaling specifically abrogates microtubule-mediated trafficking of ERK, leading to defective activation of LAP and impaired killing of bacterial and fungal pathogens by monocytes/macrophages, which can be selectively restored by IL-6 supplementation. Our work uncovers a molecular pathway linking IL-6 signaling with LAP and provides insight into the mechanisms underlying immunoparalysis in sepsis.
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http://dx.doi.org/10.1016/j.chom.2021.06.002DOI Listing
August 2021

Albumin Neutralizes Hydrophobic Toxins and Modulates Pathogenicity.

mBio 2021 06 22;12(3):e0053121. Epub 2021 Jun 22.

Junior Research Group Adaptive Pathogenicity Strategies, Leibniz Institute for Natural Product Research and Infection Biology, Hans Knöll Institute, Jena, Germany.

Albumin is abundant in serum but is also excreted at mucosal surfaces and enters tissues when inflammation increases vascular permeability. Host-associated opportunistic pathogens encounter albumin during commensalism and when causing infections. Considering the ubiquitous presence of albumin, we investigated its role in the pathogenesis of infections with the model human fungal pathogen, Candida albicans. Albumin was introduced in various models that mimic different stages of systemic or mucosal candidiasis, where it reduced the ability of C. albicans to damage host cells. The amphipathic toxin candidalysin mediates necrotic host cell damage induced by C. albicans. Using cellular and biophysical assays, we determined that albumin functions by neutralizing candidalysin through hydrophobic interactions. We discovered that albumin, similarly, can neutralize a variety of fungal (α-amanitin), bacterial (streptolysin O and staurosporin), and insect (melittin) hydrophobic toxins. These data suggest albumin as a defense mechanism against toxins, which can play a role in the pathogenesis of microbial infections. Albumin is the most abundant serum protein in humans. During inflammation, serum albumin levels decrease drastically, and low albumin levels are associated with poor patient outcome. Thus, albumin may have specific functions during infection. Here, we describe the ability of albumin to neutralize hydrophobic microbial toxins. We show that albumin can protect against damage induced by the pathogenic yeast C. albicans by neutralizing its cytolytic toxin candidalysin. These findings suggest that albumin is a toxin-neutralizing protein that may play a role during infections with toxin-producing microorganisms.
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http://dx.doi.org/10.1128/mBio.00531-21DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8262992PMC
June 2021

Neuraminidase and SIGLEC15 modulate the host defense against pulmonary aspergillosis.

Cell Rep Med 2021 May 18;2(5):100289. Epub 2021 May 18.

Department of Internal Medicine, Radboud University Medical Center, Nijmegen, the Netherlands.

Influenza-associated pulmonary aspergillosis (IAPA) has been reported increasingly since the advent of use of neuraminidase (NA) inhibitors following the 2009 influenza pandemic. We hypothesize that blocking host NA modulates the immune response against . We demonstrate that NA influences the host response against and that oseltamivir increases the susceptibility of mice to pulmonary aspergillosis. Oseltamivir impairs the mouse splenocyte and human peripheral blood mononuclear cell (PBMC) killing capacity of , and adding NA restores this defect in PBMCs. Furthermore, the sialic acid-binding receptor is upregulated in PBMCs stimulated with . Silencing of decrease PBMC killing of . We provide evidence that host NA activity and sialic acid recognition are important for anti- defense. NA inhibitors might predispose individuals with severe influenza to invasive aspergillosis. These data shed light on the pathogenesis of invasive fungal infections and may identify potential therapeutic targets.
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http://dx.doi.org/10.1016/j.xcrm.2021.100289DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8149467PMC
May 2021

Data of common and species-specific transcriptional host responses to pathogenic fungi.

Data Brief 2021 Apr 4;35:106928. Epub 2021 Mar 4.

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

Using a comparative RNA-Sequencing based transcriptional profiling approach, responses of primary human peripheral blood mononuclear cells (PBMCs) to common human pathogenic fungi have been characterized (Bruno et al. Computational and Structural Biology Journal). Primary human PBMCs were stimulated in vitro with the fungi , and after which RNA was isolated and sequenced. From raw sequencing reads differential expressed genes in response to the different fungi where calculated by comparison with unstimulated cells. By overlapping differentially expressed genes in response to the pathogenic fungi , and a dataset was generated that encompasses a common response to these three distinct fungi as well as species-specific responses. Here we present datasets on these common and species-specific responses that complement the original study (Bruno et al. Computational and Structural Biology Journal). These data serve to facilitate further fundamental research on the immune response to opportunistic pathogenic fungi such as , and .
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http://dx.doi.org/10.1016/j.dib.2021.106928DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8039545PMC
April 2021

Candida pathogens induce protective mitochondria-associated type I interferon signalling and a damage-driven response in vaginal epithelial cells.

Nat Microbiol 2021 05 22;6(5):643-657. Epub 2021 Mar 22.

Department of Microbial Pathogenicity Mechanisms, Leibniz Institute for Natural Product Research and Infection Biology, Hans Knoell Institute, Jena, Germany.

Vaginal candidiasis is an extremely common disease predominantly caused by four phylogenetically diverse species: Candida albicans; Candida glabrata; Candida parapsilosis; and Candida tropicalis. Using a time course infection model of vaginal epithelial cells and dual RNA sequencing, we show that these species exhibit distinct pathogenicity patterns, which are defined by highly species-specific transcriptional profiles during infection of vaginal epithelial cells. In contrast, host cells exhibit a homogeneous response to all species at the early stages of infection, which is characterized by sublethal mitochondrial signalling inducing a protective type I interferon response. At the later stages, the transcriptional response of the host diverges in a species-dependent manner. This divergence is primarily driven by the extent of epithelial damage elicited by species-specific mechanisms, such as secretion of the toxin candidalysin by C. albicans. Our results uncover a dynamic, biphasic response of vaginal epithelial cells to Candida species, which is characterized by protective mitochondria-associated type I interferon signalling and a species-specific damage-driven response.
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http://dx.doi.org/10.1038/s41564-021-00875-2DOI Listing
May 2021

In vitro infection models to study fungal-host interactions.

FEMS Microbiol Rev 2021 Sep;45(5)

Department of Microbial Pathogenicity Mechanisms, Leibniz Institute for Natural Product Research and Infection Biology-Hans Knoell Institute, Beutenbergstrasse 11a, 07745, Jena, Germany.

Fungal infections (mycoses) affect over a billion people per year. Approximately, two million of these infections are life-threatening, especially for patients with a compromised immune system. Fungi of the genera Aspergillus, Candida, Histoplasma and Cryptococcus are opportunistic pathogens that contribute to a substantial number of mycoses. To optimize the diagnosis and treatment of mycoses, we need to understand the complex fungal-host interplay during pathogenesis, the fungal attributes causing virulence and how the host resists infection via immunological defenses. In vitro models can be used to mimic fungal infections of various tissues and organs and the corresponding immune responses at near-physiological conditions. Furthermore, models can include fungal interactions with the host-microbiota to mimic the in vivo situation on skin and mucosal surfaces. This article reviews currently used in vitro models of fungal infections ranging from cell monolayers to microfluidic 3D organ-on-chip (OOC) platforms. We also discuss how OOC models can expand the toolbox for investigating interactions of fungi and their human hosts in the future.
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http://dx.doi.org/10.1093/femsre/fuab005DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8498566PMC
September 2021

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

The impact of the Fungus-Host-Microbiota interplay upon Candida albicans infections: current knowledge and new perspectives.

FEMS Microbiol Rev 2021 05;45(3)

NEXBIOME Therapeutics, 22 allée Alan Turing, 63000 Clermont-Ferrand, France.

Candida albicans is a major fungal pathogen of humans. It exists as a commensal in the oral cavity, gut or genital tract of most individuals, constrained by the local microbiota, epithelial barriers and immune defences. Their perturbation can lead to fungal outgrowth and the development of mucosal infections such as oropharyngeal or vulvovaginal candidiasis, and patients with compromised immunity are susceptible to life-threatening systemic infections. The importance of the interplay between fungus, host and microbiota in driving the transition from C. albicans commensalism to pathogenicity is widely appreciated. However, the complexity of these interactions, and the significant impact of fungal, host and microbiota variability upon disease severity and outcome, are less well understood. Therefore, we summarise the features of the fungus that promote infection, and how genetic variation between clinical isolates influences pathogenicity. We discuss antifungal immunity, how this differs between mucosae, and how individual variation influences a person's susceptibility to infection. Also, we describe factors that influence the composition of gut, oral and vaginal microbiotas, and how these affect fungal colonisation and antifungal immunity. We argue that a detailed understanding of these variables, which underlie fungal-host-microbiota interactions, will present opportunities for directed antifungal therapies that benefit vulnerable patients.
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http://dx.doi.org/10.1093/femsre/fuaa060DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8100220PMC
May 2021

Human recombinant interleukin-38 suppresses inflammation in mouse models of local and systemic disease.

Cytokine 2021 01 28;137:155334. Epub 2020 Oct 28.

Department of Medicine, University of Colorado Denver, Aurora, CO, USA; Department of Internal Medicine, Radboud University Medical Center, Nijmegen, The Netherlands. Electronic address:

Interleukin (IL)-38 belongs to the IL-1 family and is part of the IL-36 subfamily due to its binding to the IL-36 Receptor (IL-1R6). In the current study, we assessed the anti-inflammatory properties of IL-38 in murine models of arthritis and systemic inflammation. First, the anti-inflammatory properties of mouse and human IL-38 precursors were compared to forms with a truncated N-terminus. In mouse bone marrow derived dendritic cells (BMDC), human and mouse IL-38 precursors with a truncation of the two N-terminal amino acids (3-152) suppressed LPS-induced IL-6. Recombinant human IL-38 (3-152) was further investigated for its immunomodulatory potential using four murine models of inflammatory disease: streptococcal cell wall (SCW)-induced arthritis, monosodium urate (MSU) crystal-induced arthritis, MSU crystal-induced peritonitis, and systemic endotoxemia. In each of these models IL-38 significantly reduced inflammation. In SCW and MSU crystal-induced arthritis, joint swelling, inflammatory cell influx, and synovial levels of IL-1β, IL-6, and KC were reduced by 50% or greater. These suppressive properties of IL-38 in SCW-induced arthritis were independent of the anti-inflammatory co-receptor IL-1R8, as IL-38 reduced arthritis equally in IL-1R8 deficient and WT mice. In MSU crystal-induced peritonitis, IL-38 reduced hypothermia, while plasma IL-6 and KC and peritoneal KC levels were reduced by 65-70%. In the LPS endotoxemia model, IL-38 pretreatment reduced systemic IL-6, TNFα and KC. Furthermore, in ex vivo cultured bone marrow, LPS-induced IL-6, TNFα and KC were reduced by 75-90%. Overall, IL-38 exhibits broad anti-inflammatory properties in models of systemic and local inflammation and therefore may be an effective cytokine therapy.
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http://dx.doi.org/10.1016/j.cyto.2020.155334DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7725974PMC
January 2021

The gut, the bad and the harmless: Candida albicans as a commensal and opportunistic pathogen in the intestine.

Curr Opin Microbiol 2020 08 27;56:7-15. Epub 2020 Jun 27.

Department of Microbial Pathogenicity Mechanisms, Leibniz Institute for Natural Product Research and Infection Biology - Hans-Knoell-Institute, Beutenbergstraße 11a, 07745 Jena, Germany; Institute of Microbiology, Faculty of Biological Sciences, Friedrich Schiller University, 07743 Jena, Germany. Electronic address:

Candida albicans is a regular member of the intestinal microbiota in the majority of the human population. This underscores C. albicans' adaptation to life in the intestine without inducing competitive interactions with other microbes, or immune responses detrimental to its survival. However, specific conditions such as a dysbalanced microbiome, a suppression of the immune system, and an impaired intestinal barrier can predispose for invasive, mostly nosocomial, C. albicans infections. Colonization of the intestine and translocation through the intestinal barrier are fundamental aspects of the processes preceding life-threatening systemic candidiasis. Insights into C. albicans' commensal lifestyle and translocation can thus help us to understand how patients develop candidiasis, and may provide leads for therapeutic strategies aimed at preventing infection. In this review, we discuss the commensal lifestyle of C. albicans in the intestine, the role of morphology for commensalism, the influence of diet, and the interactions with bacteria of the microbiota.
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http://dx.doi.org/10.1016/j.mib.2020.05.006DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7744392PMC
August 2020

I want to break free - macrophage strategies to recognize and kill Candida albicans, and fungal counter-strategies to escape.

Curr Opin Microbiol 2020 12 27;58:15-23. Epub 2020 Jun 27.

Junior Research Group Adaptive Pathogenicity Strategies, Leibniz Institute for Natural Product Research and Infection Biology, Hans Knöll Institute (HKI), Adolf-Reichwein-Straße 23, 07745, Jena, Germany. Electronic address:

Candida albicans is a major cause of fungal nosocomial infections. Host defense against disseminated infections caused by this yeast strongly relies on myeloid cells of the innate immune system. Recently, several breakthroughs have been made that significantly improved our understanding of the role of macrophages during candidiasis and how C. albicans and macrophages interact. Resident tissue macrophages and macrophages derived from monocytes that infiltrate infected tissues are essential for the initiation of the antifungal immune response, as well as elimination of C. albicans from the bloodstream and infected organs. These cells engulf and try to eliminate the invading fungi through specialized mechanisms. Concurrently, C. albicans tries to survive the stresses imposed by the macrophage, acquires nutrients, and can break free from their captive environment. This review focuses on the most recent insights into the strategies of macrophages to eliminate C. albicans and the fungal counterstrategies to overcome these threats.
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http://dx.doi.org/10.1016/j.mib.2020.05.007DOI Listing
December 2020

Keeping commensal: how lactobacilli antagonize pathogenicity of in an gut model.

Dis Model Mech 2019 09 12;12(9). Epub 2019 Sep 12.

Department of Microbial Pathogenicity Mechanisms, Leibniz Institute for Natural Product Research and Infection Biology-Hans-Knoell-Institute, Beutenbergstraße 11A, 07745 Jena, Germany

The intestine is the primary reservoir of that can cause systemic infections in immunocompromised patients. In this reservoir, the fungus exists as a harmless commensal. However, antibiotic treatment can disturb the bacterial microbiota, facilitating fungal overgrowth and favoring pathogenicity. The current gut models that are used to study the pathogenesis of investigate the state in which behaves as a pathogen rather than as a commensal. We present a novel gut model in which the fungal pathogenicity is reduced to a minimum by increasing the biological complexity. In this model, enterocytes represent the epithelial barrier and goblet cells limit adhesion and invasion. Significant protection against induced necrotic damage was achieved by the introduction of a microbiota of antagonistic lactobacilli. We demonstrated a time-, dose- and species-dependent protective effect against -induced cytotoxicity. This required bacterial growth, which relied on the presence of host cells, but was not dependent on the competition for adhesion sites. reduced hyphal elongation, a key virulence attribute. Furthermore, bacterial-driven shedding of hyphae from the epithelial surface, associated with apoptotic epithelial cells, was identified as a main and novel mechanism of damage protection. However, host cell apoptosis was not the driving mechanism behind shedding. Collectively, we established an gut model that can be used to experimentally dissect commensal-like interactions of with a bacterial microbiota and the host epithelial barrier. We also discovered fungal shedding as a novel mechanism by which bacteria contribute to the protection of epithelial surfaces.This article has an associated First Person interview with the joint first authors of the paper.
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http://dx.doi.org/10.1242/dmm.039719DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6765188PMC
September 2019

A three-dimensional immunocompetent intestine-on-chip model as in vitro platform for functional and microbial interaction studies.

Biomaterials 2019 11 2;220:119396. Epub 2019 Aug 2.

Center for Sepsis Control and Care, Jena University Hospital, Friedrich-Schiller-University of Jena, Jena, Germany; Institute of Biochemistry II, Jena University Hospital, Jena, Germany. Electronic address:

Alterations of the microbial composition in the gut and the concomitant dysregulation of the mucosal immune response are associated with the pathogenesis of opportunistic infections, chronic inflammation, and inflammatory bowel disease. To create a platform for the investigation of the underlying mechanisms, we established a three-dimensional microphysiological model of the human intestine. This model resembles organotypic microanatomical structures and includes tissue resident innate immune cells exhibiting features of mucosal macrophages and dendritic cells. The model displays the physiological immune tolerance of the intestinal lumen to microbial-associated molecular patterns and can, therefore, be colonised with living microorganisms. Functional studies on microbial interaction between probiotic Lactobacillus rhamnosus and the opportunistic pathogen Candida albicans show that pre-colonization of the intestinal lumen of the model by L. rhamnosus reduces C. albicans-induced tissue damage, lowers its translocation, and limits fungal burden. We demonstrate that microbial interactions can be efficiently investigated using the in vitro model creating a more physiological and immunocompetent microenvironment. The intestinal model allows a detailed characterisation of the immune response, microbial pathogenicity mechanisms, and quantification of cellular dysfunction attributed to alterations in the microbial composition.
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http://dx.doi.org/10.1016/j.biomaterials.2019.119396DOI Listing
November 2019

A Genome-Wide Functional Genomics Approach Identifies Susceptibility Pathways to Fungal Bloodstream Infection in Humans.

J Infect Dis 2019 07;220(5):862-872

Department of Internal Medicine and Radboud Center for Infectious Diseases, Radboud University Medical Center, Nijmegen, the Netherlands.

Background: Candidemia, one of the most common causes of fungal bloodstream infection, leads to mortality rates up to 40% in affected patients. Understanding genetic mechanisms for differential susceptibility to candidemia may aid in designing host-directed therapies.

Methods: We performed the first genome-wide association study on candidemia, and we integrated these data with variants that affect cytokines in different cellular systems stimulated with Candida albicans.

Results: We observed strong association between candidemia and a variant, rs8028958, that significantly affects the expression levels of PLA2G4B in blood. We found that up to 35% of the susceptibility loci affect in vitro cytokine production in response to Candida. Furthermore, potential causal genes located within these loci are enriched for lipid and arachidonic acid metabolism. Using an independent cohort, we also showed that the numbers of risk alleles at these loci are negatively correlated with reactive oxygen species and interleukin-6 levels in response to Candida. Finally, there was a significant correlation between susceptibility and allelic scores based on 16 independent candidemia-associated single-nucleotide polymorphisms that affect monocyte-derived cytokines, but not with T cell-derived cytokines.

Conclusions: Our results prioritize the disturbed lipid homeostasis and oxidative stress as potential mechanisms that affect monocyte-derived cytokines to influence susceptibility to candidemia.
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http://dx.doi.org/10.1093/infdis/jiz206DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6667794PMC
July 2019

Frontline Science: Endotoxin-induced immunotolerance is associated with loss of monocyte metabolic plasticity and reduction of oxidative burst.

J Leukoc Biol 2019 07 6;106(1):11-25. Epub 2019 Jun 6.

Department of Internal Medicine, Radboud University Medical Center, Nijmegen, the Netherlands.

Secondary infections are a major complication of sepsis and associated with a compromised immune state, called sepsis-induced immunoparalysis. Molecular mechanisms causing immunoparalysis remain unclear; however, changes in cellular metabolism of leukocytes have been linked to immunoparalysis. We investigated the relation of metabolic changes to antimicrobial monocyte functions in endotoxin-induced immunotolerance, as a model for sepsis-induced immunoparalysis. In this study, immunotolerance was induced in healthy males by intravenous endotoxin (2 ng/kg, derived from Escherichia coli O:113) administration. Before and after induction of immunotolerance, circulating CD14 monocytes were isolated and assessed for antimicrobial functions, including cytokine production, oxidative burst, and microbial (Candida albicans) killing capacity, as well metabolic responses to ex vivo stimulation. Next, the effects of altered cellular metabolism on monocyte functions were validated in vitro. Ex vivo lipopolysaccharide stimulation induced an extensive rewiring of metabolism in naive monocytes. In contrast, endotoxin-induced immunotolerant monocytes showed no metabolic plasticity, as they were unable to adapt their metabolism or mount cytokine and oxidative responses. Validation experiments showed that modulation of metabolic pathways, affected by immunotolerance, influenced monocyte cytokine production, oxidative burst, and microbial (C. albicans) killing in naive monocytes. Collectively, these data demonstrate that immunotolerant monocytes are characterized by a loss of metabolic plasticity and these metabolic defects impact antimicrobial monocyte immune functions. Further, these findings support that the changed cellular metabolism of immunotolerant monocytes might reveal novel therapeutic targets to reverse sepsis-induced immunoparalysis.
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http://dx.doi.org/10.1002/JLB.5HI0119-018RDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6852552PMC
July 2019

The fungal peptide toxin Candidalysin activates the NLRP3 inflammasome and causes cytolysis in mononuclear phagocytes.

Nat Commun 2018 10 15;9(1):4260. Epub 2018 Oct 15.

Department of Microbial Pathogenicity Mechanisms, Leibniz Institute for Natural Product Research and Infection Biology - Hans Knoell Institute, Beutenbergstrasse 11a, Jena, 07745, Germany.

Clearance of invading microbes requires phagocytes of the innate immune system. However, successful pathogens have evolved sophisticated strategies to evade immune killing. The opportunistic human fungal pathogen Candida albicans is efficiently phagocytosed by macrophages, but causes inflammasome activation, host cytolysis, and escapes after hypha formation. Previous studies suggest that macrophage lysis by C. albicans results from early inflammasome-dependent cell death (pyroptosis), late damage due to glucose depletion and membrane piercing by growing hyphae. Here we show that Candidalysin, a cytolytic peptide toxin encoded by the hypha-associated gene ECE1, is both a central trigger for NLRP3 inflammasome-dependent caspase-1 activation via potassium efflux and a key driver of inflammasome-independent cytolysis of macrophages and dendritic cells upon infection with C. albicans. This suggests that Candidalysin-induced cell damage is a third mechanism of C. albicans-mediated mononuclear phagocyte cell death in addition to damage caused by pyroptosis and the growth of glucose-consuming hyphae.
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http://dx.doi.org/10.1038/s41467-018-06607-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6189146PMC
October 2018

The Itaconate Pathway Is a Central Regulatory Node Linking Innate Immune Tolerance and Trained Immunity.

Cell Metab 2019 01 4;29(1):211-220.e5. Epub 2018 Oct 4.

Department of Internal Medicine (463) and Radboud Center for Infectious Diseases (RCI), Radboud University Nijmegen Medical Centre, Geert Grooteplein 8, Nijmegen 6500 HB, the Netherlands; Department for Genomics & Immunoregulation, Life and Medical Sciences Institute (LIMES), University of Bonn, Bonn, Germany; Human Genomics Laboratory, Craiova University of Medicine and Pharmacy, Craiova, Romania.

Sepsis involves simultaneous hyperactivation of the immune system and immune paralysis, leading to both organ dysfunction and increased susceptibility to secondary infections. Acute activation of myeloid cells induced itaconate synthesis, which subsequently mediated innate immune tolerance in human monocytes. In contrast, induction of trained immunity by β-glucan counteracted tolerance induced in a model of human endotoxemia by inhibiting the expression of immune-responsive gene 1 (IRG1), the enzyme that controls itaconate synthesis. β-Glucan also increased the expression of succinate dehydrogenase (SDH), contributing to the integrity of the TCA cycle and leading to an enhanced innate immune response after secondary stimulation. The role of itaconate was further validated by IRG1 and SDH polymorphisms that modulate induction of tolerance and trained immunity in human monocytes. These data demonstrate the importance of the IRG1-itaconate-SDH axis in the development of immune tolerance and training and highlight the potential of β-glucan-induced trained immunity to revert immunoparalysis.
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http://dx.doi.org/10.1016/j.cmet.2018.09.003DOI Listing
January 2019

Adjunctive interferon-γ immunotherapy in a pediatric case of Aspergillus terreus infection.

Eur J Clin Microbiol Infect Dis 2018 Oct 19;37(10):1915-1922. Epub 2018 Jul 19.

Department of Pediatric Infectious Diseases and Immunology, Radboudumc Amalia Children's Hospital, Radboud University Medical Center, Nijmegen, the Netherlands.

Aspergillus terreus causes invasive aspergillosis (IA) in immunocompromised patients. Treatment is complicated by intrinsic resistance to amphotericin B and thereby contributing to a high mortality. Therefore, we conducted in vitro studies to investigate the effectivity of adjunctive recombinant interferon-γ immunotherapy. We describe a pediatric patient with A. terreus IA who received adjunctive recombinant interferon-γ (rIFNγ) immunotherapy. In vitro studies were conducted to investigate the capacity of rIFNγ to improve antifungal host defense in terms of fungal killing ability and the release of pro-inflammatory cytokines in cells of the patient as well as healthy controls. An 8-year-old female pediatric patient with leukemia developed A. terreus IA. She clinically deteriorated and had high serum galactomannan levels despite broad antifungal therapy. Therefore, adjunctive immune stimulatory therapy with rIFNγ was initiated. After 3 weeks of treatment, galactomannan levels decreased and the patient clinically showed improvement. Addition of rIFNγ boosted the capacity of monocytes of healthy volunteers to mount TNFα and IL-1β cytokine responses to Escherichia coli LPS, and increased TNFα response to both A. terreus and Aspergillus fumigatus. Monocytes isolated from the patient's blood demonstrated a similar augmented cytokine induction in response to rIFNγ. In addition, rIFNγ increased the capacity of monocytes from healthy volunteers as well as monocytes from the patient to kill A. terreus spores. Adjuvant immunotherapy with rIFNγ might be a promising additional treatment strategy that could be used to improve outcome in patients with refractory invasive A. terreus infections or other resistant invasive Aspergillus infections.
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http://dx.doi.org/10.1007/s10096-018-3325-4DOI Listing
October 2018

Microbiological and immunological characteristics of a lethal pulmonary infection in a non-neutropenic patient.

Med Mycol Case Rep 2018 Sep 7;21:4-7. Epub 2018 Mar 7.

Department of Medical Microbiology, Radboud University Medical Center, Nijmegen, The Netherlands.

Invasive pulmonary aspergillosis is increasingly described in non-neutropenic patients, such as patients with COPD receiving corticosteroids and the critically ill. Here, we present a case of a lethal pulmonary infection in a COPD patient. Immunological tests showed an impaired innate and adaptive immune response to . A history of COPD, unresponsiveness to antibiotics and especially a suggestive CT-scan should trigger the clinician to consider diseases caused by
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http://dx.doi.org/10.1016/j.mmcr.2018.03.002DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6026298PMC
September 2018

Genetic deficiency of NOD2 confers resistance to invasive aspergillosis.

Nat Commun 2018 07 6;9(1):2636. Epub 2018 Jul 6.

Department of Experimental Internal Medicine and Radboud Center for Infectious Diseases (RCI), Radboud University Medical Center, Geert Grooteplein zuid 8, 6525GA, Nijmegen, The Netherlands.

Invasive aspergillosis (IA) is a severe infection that can occur in severely immunocompromised patients. Efficient immune recognition of Aspergillus is crucial to protect against infection, and previous studies suggested a role for NOD2 in this process. However, thorough investigation of the impact of NOD2 on susceptibility to aspergillosis is lacking. Common genetic variations in NOD2 has been associated with Crohn's disease and here we investigated the influence of these  genetic variations on the anti-Aspergillus host response. A NOD2 polymorphism reduced the risk of IA after hematopoietic stem-cell transplantation. Mechanistically, absence of NOD2 in monocytes and macrophages increases phagocytosis leading to enhanced fungal killing, conversely, NOD2 activation reduces the antifungal potential of these cells. Crucially, Nod2 deficiency results in resistance to Aspergillus infection in an in vivo model of pulmonary aspergillosis. Collectively, our data demonstrate that genetic deficiency of NOD2 plays a protective role during Aspergillus infection.
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http://dx.doi.org/10.1038/s41467-018-04912-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6035256PMC
July 2018

The Multifaceted Role of T-Helper Responses in Host Defense against Aspergillus fumigatus.

J Fungi (Basel) 2017 Oct 4;3(4). Epub 2017 Oct 4.

Department of Experimental Internal Medicine and Radboud Center for Infectious diseases (RCI), Radboud University Medical Center, Geert Grooteplein Zuid 10, 6525 GA Nijmegen, The Netherlands.

The ubiquitous opportunistic fungal pathogen rarely causes infections in immunocompetent individuals. A healthy functional innate immune system plays a crucial role in preventing -infection. This pivotal role for the innate immune system makes it a main research focus in studying the pathogenesis of aspergillosis. Although sometimes overshadowed by the innate immune response, the adaptive immune response, and in particular T-helper responses, also represents a key player in host defense against . Virtually all T-helper subsets have been described to play a role during aspergillosis, with the Th1 response being crucial for fungal clearance. However; morbidity and mortality of aspergillosis can also be partly attributed to detrimental immune responses resulting from adaptive immune activation. Th2 responses benefit fungal persistence; and are the foundation of allergic forms of aspergillosis. The Th17 response has two sides; although crucial for granulocyte recruitment, it can be involved in detrimental immunopathology. Regulatory T-cells, the endogenous regulators of inflammatory responses, play a key role in controlling detrimental inflammatory responses during aspergillosis. The current knowledge of the adaptive immune response against is summarized in this review. A better understanding on how T-helper responses facilitate clearance of -infection and control inflammation can be the fundamental basis for understanding the pathogenesis of aspergillosis and for the development of novel host-directed therapies.
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http://dx.doi.org/10.3390/jof3040055DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5753157PMC
October 2017

The Absence of NOD1 Enhances Killing of Through Modulation of Dectin-1 Expression.

Front Immunol 2017 13;8:1777. Epub 2017 Dec 13.

Laboratory for Experimental Internal Medicine, Department of Internal Medicine, Radboud University Medical Center, Nijmegen, Netherlands.

One of the major life-threatening infections for which severely immunocompromised patients are at risk is invasive aspergillosis (IA). Despite the current treatment options, the increasing antifungal resistance and poor outcome highlight the need for novel therapeutic strategies to improve outcome of patients with IA. In the current study, we investigated whether and how the intracellular pattern recognition receptor NOD1 is involved in host defense against . When exploring the role of NOD1 in an experimental mouse model, we found that mice were protected against IA and demonstrated reduced fungal outgrowth in the lungs. We found that macrophages derived from bone marrow of mice were more efficiently inducing reactive oxygen species and cytokines in response to . Most strikingly, these cells were highly potent in killing compared with wild-type cells. In line, human macrophages in which NOD1 was silenced demonstrated augmented killing and NOD1 stimulation decreased fungal killing. The differentially altered killing capacity of NOD1 silencing versus NOD1 activation was associated with alterations in dectin-1 expression, with activation of NOD1 reducing dectin-1 expression. Furthermore, we were able to demonstrate that mice have elevated dectin-1 expression in the lung and bone marrow, and silencing of gene expression in human macrophages increases dectin-1 expression. The enhanced dectin-1 expression may be the mechanism of enhanced fungal killing of cells and human cells in which NOD1 was silenced, since blockade of dectin-1 reversed the augmented killing in these cells. Collectively, our data demonstrate that NOD1 receptor plays an inhibitory role in the host defense against . This provides a rationale to develop novel immunotherapeutic strategies for treatment of aspergillosis that target the NOD1 receptor, to enhance the efficiency of host immune cells to clear the infection by increasing fungal killing and cytokine responses.
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http://dx.doi.org/10.3389/fimmu.2017.01777DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5733348PMC
December 2017

Differential Kinetics of Aspergillus nidulans and Aspergillus fumigatus Phagocytosis.

J Innate Immun 2018 16;10(2):145-160. Epub 2017 Dec 16.

Medical Research Council Centre for Medical Mycology at the University of Aberdeen, Aberdeen Fungal Group, Institute of Medical Sciences, University of Aberdeen, Aberdeen, UK.

Invasive aspergillosis mainly occurs in immunocompromised patients and is commonly caused by Aspergillus fumigatus, while A.nidulans is rarely the causative agent. However, in chronic granulomatous disease (CGD) patients, A. nidulans is a frequent cause of invasive aspergillosis and is associated with higher mortality. Immune recognition of A. nidulans was compared to A. fumigatus to offer an insight into why A. nidulans infections are prevalent in CGD. Live cell imaging with J774A.1 macrophage-like cells and LC3-GFP-mCherry bone marrow-derived macrophages (BMDMs) revealed that phagocytosis of A. nidulans was slower compared to A. fumigatus. This difference could be attributed to slower migration of J774A.1 cells and a lower percentage of migrating BMDMs. In addition, delayed phagosome acidification and LC3-associated phagocytosis was observed with A. nidulans. Cytokine and oxidative burst measurements in human peripheral blood mononuclear cells revealed a lower oxidative burst upon challenge with A. nidulans. In contrast, A. nidulans induced significantly higher concentrations of cytokines. Collectively, our data demonstrate that A. nidulans is phagocytosed and processed at a slower rate compared to A. fumigatus, resulting in reduced fungal killing and increased germination of conidia. This slower rate of A. nidulans clearance may be permissive for overgrowth within certain immune settings.
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http://dx.doi.org/10.1159/000484562DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6757152PMC
September 2019

Rewiring monocyte glucose metabolism via C-type lectin signaling protects against disseminated candidiasis.

PLoS Pathog 2017 Sep 18;13(9):e1006632. Epub 2017 Sep 18.

Department of Internal Medicine and Radboud Center for Infectious diseases (RCI), Radboud University Nijmegen Medical Centre, Geert Grooteplein 8, Nijmegen, the Netherlands.

Monocytes are innate immune cells that play a pivotal role in antifungal immunity, but little is known regarding the cellular metabolic events that regulate their function during infection. Using complementary transcriptomic and immunological studies in human primary monocytes, we show that activation of monocytes by Candida albicans yeast and hyphae was accompanied by metabolic rewiring induced through C-type lectin-signaling pathways. We describe that the innate immune responses against Candida yeast are energy-demanding processes that lead to the mobilization of intracellular metabolite pools and require induction of glucose metabolism, oxidative phosphorylation and glutaminolysis, while responses to hyphae primarily rely on glycolysis. Experimental models of systemic candidiasis models validated a central role for glucose metabolism in anti-Candida immunity, as the impairment of glycolysis led to increased susceptibility in mice. Collectively, these data highlight the importance of understanding the complex network of metabolic responses triggered during infections, and unveil new potential targets for therapeutic approaches against fungal diseases.
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http://dx.doi.org/10.1371/journal.ppat.1006632DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5619837PMC
September 2017

Aspergillus fumigatus morphology and dynamic host interactions.

Nat Rev Microbiol 2017 Nov 18;15(11):661-674. Epub 2017 Sep 18.

Unité des Aspergillus, Institut Pasteur, 25-28 Rue du Dr Roux, 75015 Paris, France.

Aspergillus fumigatus is an environmental filamentous fungus that can cause life-threatening disease in immunocompromised individuals. The interactions between A. fumigatus and the host environment are dynamic and complex. The host immune system needs to recognize the distinct morphological forms of A. fumigatus to control fungal growth and prevent tissue invasion, whereas the fungus requires nutrients and needs to adapt to the hostile environment by escaping immune recognition and counteracting host responses. Understanding these highly dynamic interactions is necessary to fully understand the pathogenesis of aspergillosis and to facilitate the design of new therapeutics to overcome the morbidity and mortality caused by A. fumigatus. In this Review, we describe how A. fumigatus adapts to environmental change, the mechanisms of host defence, and our current knowledge of the interplay between the host immune response and the fungus.
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http://dx.doi.org/10.1038/nrmicro.2017.90DOI Listing
November 2017

Toll-like receptor 2 induced cytotoxic T-lymphocyte-associated protein 4 regulates Aspergillus-induced regulatory T-cells with pro-inflammatory characteristics.

Sci Rep 2017 09 13;7(1):11500. Epub 2017 Sep 13.

Department of Experimental Internal Medicine and Radboud Center for Infectious diseases (RCI), Radboud University Medical Center, Geert Grooteplein zuid 8, 6525GA, Nijmegen, The Netherlands.

Patients with cystic fibrosis, chronic obstructive pulmonary disease, severe asthma, pre-existing pulmonary lesions, and severely immunocompromised patients are susceptible to develop infections with the opportunistic pathogenic fungus Aspergillus fumigatus, called aspergillosis. Infections in these patients are associated with persistent pro-inflammatory T-helper (T)2 and T17 responses. Regulatory T-cells, natural suppressor cells of the immune system, control pro-inflammatory T-cell responses, but can also contribute to disease by shifting to a pro-inflammatory T17-like phenotype. Such a shift could play an important role in the detrimental immunopathology that is seen in aspergillosis. Our study demonstrates that Aspergillus fumigatus induces regulatory T-cells with a T17-like phenotype. We also demonstrate that these regulatory T-cells with a pro-inflammatory T17-like phenotype can be reprogrammed to their "classical" anti-inflammatory phenotype by activating Toll-like receptor 2 (TLR2), which regulates the induction of cytotoxic T-lymphocyte-associated protein 4 (CTLA4). Similarly, soluble CTLA4 could reverse the pro-inflammatory phenotype of Aspergillus-induced regulatory T-cells. In conclusion, our results suggest a role for regulatory T-cells with a pro-inflammatory T17-like phenotype in Aspergillus-associated immunopathology, and identifies key players, i.e. TLR2 and CTLA4, involved in this mechanism.
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http://dx.doi.org/10.1038/s41598-017-11738-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5597613PMC
September 2017
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