Biology subjects

LeibundGut-Landmann, S.

Publications and source records attributed to LeibundGut-Landmann, S..

17 recordsLinked to original sources

bioRxiv2026

Starve, stress, and mate: Activation of the CCAAT-Binding complex overrides sterility of mating-type heterozygous Candida albicans

Sexual reproduction in the human fungal pathogen Candida albicans is typically restricted by a strict genetic lock: the a1-2 repressor present in standard mating-type heterozygous (MTLa/) diploid strains, which actively represses mating competency. While severe environmental pressures are known to influence fungal development, the molecular mechanisms that allow cells to override this genetic sterility block remain poorly understood. Here, we identify the highly conserved CCAAT-binding complex (CBC) as the critical developmental hub that integrates environmental stress to unlock mating in heterozygous diploids. By combining proteomic mapping (ChIP-SICAP) and transcriptomics (RNA-Seq) to define the regulatory network of the core CBC subunit Hap5, we demonstrate that CBC hyperactivation completely overrides the a1-2 repressor. This activation triggers a robust white-to-opaque phenotypic switch and initiates a mating-competence program in MTLa/ strains genetically locked in a sterile state. Crucially, this developmental transition is antagonized by the iron-regulatory partner Hap43, but is strictly dependent on physical interaction at the chromatin level with Cap1, the master regulator of the oxidative stress response. Our findings reveal that the CBC and Cap1 couple metabolic starvation (glucose and iron limitation) and oxidative stress to a core reproductive program, providing a mechanistic basis for how C. albicans bypasses genetic constraints on sexual reproduction to generate genetic diversity under hostile host conditions.

microbiology↗

bioRxiv2026

Host skin lipids trigger MAT-dependent mating, pathogenic hyphal growth, and parasexual reproduction of Malassezia furfur

The lipophilic yeast genus Malassezia dominates the human skin mycobiome and, while typically commensal, is associated with skin disorders in which hyphal cells are frequently observed. However, the mechanisms underlying hyphal differentiation and its contribution to pathogenesis remain poorly understood. Here, we show that hyphal growth in Malassezia furfur requires a complete mating-type system and is promoted by noncanonical reproductive interactions. Interspecific mating generates stable diploid hybrids with evidence of recombination, whereas intraspecific mating produces cell-fusion products that undergo recombination and haploidization, consistent with a parasexual cycle. Functional analyses reveal that the pheromone/receptor (P/R) locus governs cell recognition and fusion, while the homeodomain (HD) locus is required for hyphal development despite having lost canonical mating-type determination. Leveraging in vitro and in vivo skin models, we show that hyphal cells invade the epidermis more efficiently and elicit stronger inflammatory responses than yeast cells. Together, these findings link parasexual-like reproduction, mating-type loci function, and morphogenesis in Malassezia to pathogenic interactions with the host.

microbiology↗

bioRxiv2026

Fourier transform infrared spectroscopy reveals high intraspecies diversity of Malassezia pachydermatis in dogs with atopic dermatitis

1Canine atopic dermatitis (CAD) is a chronic inflammatory skin condition sometimes associated with microbial dysbiosis, including alterations in colonization by the lipophilic yeast Malassezia pachydermatis. This study investigated the population diversity of M. pachydermatis in the ear canals of healthy and CAD-affected dogs using Fourier-transform infrared (FTIR) spectroscopy and whole genome sequencing (WGS). Among 60 dogs, M. pachydermatis prevalence was significantly higher in CAD cases than in healthy controls. FTIR spectroscopy revealed greater strain heterogeneity in CAD-affected dogs, often with distinct genotypes in each ear, while healthy dogs exhibited more homogeneous populations. Using a previously developed FTIR-based artificial neural network classifier, we assigned strains to three phylogroups. Strains from phylogroups I and III were significantly enriched in CAD-affected dogs, while phylogroup II was most prevalent overall and the dominant phylogroup in healthy controls. This suggests that CAD-associated inflammation may favor specific M. pachydermatis phylogroups and sub-clusters within phylogroups, shaping colonization dynamics. FTIR-based typing showed full concordance with WGS across 35 sequenced isolates, recapitulating relationships among phylogenetically related isolates and their similar phenotypic profiles. Overall, our findings reveal strain-level shifts in M. pachydermatis populations associated with CAD and establish FTIR spectroscopy as a rapid, cost-effective tool for large-scale epidemiological studies.

microbiology↗

bioRxiv2025

Clec7a-mediated regulation of Killer-like Lectin Receptor expression controls T cell immunity

Clec7a is a C-type lectin receptor (CLR) originally defined for its non-redundant role in anti-fungal immunity. Subsequent work has broadened this view, implicating Clec7a in host defense against diverse pathogens and in the pathogenesis of cancer, autoimmunity, neuroinflammation, and developmental disorders. How a single innate receptor orchestrates such wide-ranging outcomes remains unresolved. We previously demonstrated that dendritic cell (DC)-expressed Clec7a is required for protective anti-fungal immunity in the gastrointestinal tract through regulation of fungus-specific CD4 T cell responses. Here, we show that Clec7a controls the expression of multiple C-type lectins in DCs, including a cluster of killer lectin-like receptors (KLRs). Notably, we reveal that these KLRs directly regulate DC function and control CD4 T cell responses. These findings define a novel Clec7a-KLR axis that integrates innate and adaptive immunity, highlighting a regulatory pathway with broad relevance for immune homeostasis, inflammation, and host defense.

immunology↗

bioRxiv2025

KLRE1 shapes antifungal immunity by tuning dendritic cell and T cell activation through distinct heterodimeric partners

Dendritic cells (DCs) instruct adaptive immunity by integrating inflammatory cues to regulate co-signalling molecules and T cell activation. We recently discovered that the pattern recognition receptor Dectin-1 (Clec7a) controls expression of a network of killer lectin-like receptors (KLRs), including KLRI1 and KLRI2, which regulate adaptive immune responses. Here, we identify KLRE1 as a key regulator of DC phenotype and function, acting through distinct heterodimeric interactions with KLRI1 and KLRI2. KLRE1 is expressed by tissue-resident and bone marrow-derived DCs and is dynamically regulated by inflammatory signals, including fungal stimuli. Mechanistically, KLRE1:KLRI1 heterodimers promote DC activation, increasing CD40, CD80, CD86, and MHC-II, whereas KLRE1:KLRI2 heterodimers restrain activation. Differential regulation of KLRI1 and KLRI2 by inflammatory signals shapes heterodimer availability and immune outcomes under distinct conditions. Functionally, KLRE1 deficiency enhances DC activation and T cell responses in vitro and in vivo during Candida albicans infection. Strikingly, KLRI1- and KLRI2-deficient mice showed opposite survival outcomes during systemic candidiasis, despite similar fungal burdens, implicating KLRE1 heterodimers in disease tolerance rather than pathogen clearance. Protection in KLRI1-deficient mice was associated with a more balanced T cell response, whereas susceptibility to infection associated with KLRI2 deficiency resulted from increased T cell activation and migratory potential, leading to systemic inflammation and renal dysfunction. Thus, KLRE1 heterodimers fine-tune DC-driven T cell immunity, balancing protection and immunopathology during fungal infection.

immunology↗

bioRxiv2025

Fourier transform infrared spectroscopy enables rapid species discrimination across Malassezia and strain-level typing in M. pachydermatis

1Malassezia pachydermatis is a zoophilic yeast found on the skin and in the outer ear canal of many mammals. It normally maintains a commensal lifestyle but can cause dermatitis and otitis in predisposed hosts, particularly in atopic dogs. M. pachydermatis is genetically diverse, with strains clustering into at least three phylogroups based on molecular typing, a pattern we now confirm through whole-genome sequencing (WGS). Accurate species and strain-level identification is essential for understanding its epidemiology, pathogenic potential, and response to treatment. In this study, we established Fourier Transform Infrared (FTIR) spectroscopy as a rapid, cost-effective method for distinguishing M. pachydermatis from other Malassezia species, including M. globosa, M. furfur, M. restricta, and M. sympodialis. Within M. pachydermatis, FTIR spectroscopy resolved even closely related strains with high accuracy producing clusters congruent with WGS-based phylogeny. The incorporation of an Artificial Neural Network classifier further enhanced the discriminatory power, enabling robust and automated strain assignment. These findings demonstrate the potential of FTIR spectroscopy as a practical tool for large-scale epidemiological surveillance of M. pachydermatis and for clinical and veterinary applications where strain-level identification could inform treatment and management of Malassezia-associated diseases. ImportanceMalassezia pachydermatis is a yeast that commonly inhabits the skin and ear canals of mammals but can cause dermatitis and otitis in predisposed hosts, especially dogs with allergies. This species displays substantial genetic diversity, with strains falling into distinct phylogroups that may differ in their biology and clinical significance. Determining these differences has typically required advanced molecular or genomic methods, which can be costly and time-consuming. In this study, we demonstrate that Fourier transform infrared spectroscopy can rapidly and accurately distinguish M. pachydermatis from other Malassezia species and resolve genetic groups within the species in a way that reflects whole-genome relationships. This capability offers a practical tool for investigating the epidemiology and inter-/intraspecies diversity of M. pachydermatis and for guiding targeted management of Malassezia-associated diseases in both veterinary and, potentially, human medicine.

microbiology↗

bioRxiv2025

Genome-guided manipulation of regulators of morphogenesis in a C. albicans strain with low virulence is not sufficient to trigger a high-virulence phenotype

As a member of the microbiome, C. albicans colonizes the oral cavity and other mucosal surfaces of the human body. While commensalism is tightly controlled by the host immune system, the fungal determinants enabling the fungus to colonize the host mucosa without causing tissue damage and inflammation remain less clear. In search of genetic determinants that may underly the commensal lifestyle of the low-virulent C. albicans isolate 101, we identified a small sequence duplication in one of the BRG1 allele resulting in a truncated loss-of-function allele (BRG1TRUNC). Replacing BRG1TRUNC by the full-length allele (BRG1FL) resulted in a modest increase in filamentation, but did not alter the phenotype of the fungus in the oral mucosa of experimentally colonized mice. Spontaneous outgrowth of highly virulent variants of the 101 strain with the BRG1TRUNC alleles replaced by a full-length allele identified the Cyr1-cAMP-PKA signalling pathway as a modulator of fungal virulence. Although the Glu-to-Lys mutation in CYR1 (CYR1E1541K), which converts Cyr1 into a hyperactive adenylate cyclase, greatly increased filamentation, the expression of hyphae-associated genes, and host cell damage when tested in vitro, it was insufficient to increase virulence of C. albicans strain 101 in the oral mucosa in vivo, irrespective of the BRG1 status. Together, this highlights that the low-virulent nature of strain 101 is firmly anchored and cannot be overcome by manipulating BRG1 and CYR1, two genes with known roles in C. albicans virulence. IMPORTANCEDuring homeostasis, the fungus Candida albicans establishes mutualistic interactions with its human host. It can however also adopt a pathogenic state and cause infections with diverse clinical manifestions that pose a significant challenge for diagnosis and therapy. Understanding the fungal determinants that underlie C. albicans colonization under steady state conditions may thus provide new avenues for modulating the fungus-host interaction in candidiasis patients to restore homeostasis. Here, we report gene variants that distinguish high-virulent from low-virulent C. albicans. Gene-exchange mutants provided evidence for the contribution of a BRG1 loss-of-function allele and a CYR1 gain-of-function mutation toward fungal pathogenicity. However, in vivo in an experimental model of C. albicans oral colonization, none of these gene variants individually or in combination were sufficient to change the virulence profile of the fungus. These findings indicate that C. albicans mucosal colonization is regulated by a complex gene network rather than by single genetic determinants.

microbiology↗

bioRxiv2025

The skin commensal yeast Malassezia promotes tissue homeostasis via the aryl hydrocarbon receptor.

As an abundant fungal colonizer of human skin, Malassezia has long been associated with pathological skin conditions, yet its role in skin homeostasis remain poorly understood. Here, we demonstrate that Malassezia furfur plays an active role in maintaining epidermal integrity by producing tryptophan-derived metabolites that activate the aryl hydrocarbon receptor (AhR), a key regulator of keratinocyte differentiation and inflammation. Using a fungal mutant defective in indole production, we show that M. furfur-derived AhR activation is required to restore barrier function and control inflammation in diseased skin. AhR-deficient mice fail to benefit from M. furfur-mediated barrier protection, underscoring the importance of microbial-derived AhR agonists in skin physiology. These findings establish a previously unrecognized mutualistic role for Malassezia in epidermal homeostasis, challenging its perception as solely a pathogenic fungus and expanding our understanding of the skin microbiotas influence on barrier function and immune regulation. KEY FINDINGSO_LIMalassezia-derived indoles reprogram epidermal gene expression to enhance keratinocyte function. C_LIO_LIAhR activation by Malassezia restores skin barrier integrity and reduces inflammation. C_LIO_LIMalassezia Sul1-dependent tryptophan metabolism is essential for the production of AhR agonists. C_LIO_LIThe barrier protective effects of Malassezia are mediated specifically through keratinocyte intrinsic AhR signaling. C_LI

microbiology↗

bioRxiv2024

The oxidative stress response-related peroxiredoxin Tsa1b of Candida auris functions as a virulence factor that promotes infection

The difficulty of accurately identifying Candida auris and the high resistance rates presented have increased the concern in the healthcare setting. Due to this, the aim of this study was to analyse the fungal response to oxidative stress. To achieve this goal, gene and protein expression were examined using qPCR and two-dimensional electrophoresis, respectively, peroxiredoxin Tsa1b being discovered to be overexpressed under oxidative stress. Besides, its antigenicity was also confirmed by western blotting. Subsequently, the significance of Tsa1b was next investigated by creating and characterizing the C. auris {Delta}TSA1B and C. auris {Delta}TSA1B::TSA1B strains using CRISPR-Cas9. The findings demonstrated that the{Delta} TSA1B strain was more susceptible to oxidative and cell wall stressors than the wild-type strain, which was consistent with an increase in the cell wall {beta}-glucan amounts when grown in the presence of oxidative stress. Furthermore, the{Delta} TSA1B strain was also more vulnerable to the presence of dendritic cells and bone marrow-derived macrophages. Finally, in vivo infections performed in Galleria mellonella and mice showed a slower progression of the disease in those animals infected with the mutant strain. In conclusion, the peroxiredoxin Tsa1b has been identified as an important protein for the C. auris response to oxidative stress and as a virulence factor, allowing for a more thorough knowledge of the pathobiology of this yeast. This study points out the potential that this protein may have for the development of new diagnostic and therapeutic approaches. HIGHLIGHTSO_LISeveral metabolic proteins are implicated in C. auris response to oxidative stress C_LIO_LIC. auris response to oxidative stress is influenced by the Tsa1b peroxiredoxin C_LIO_LILack of Tsa1b generates more susceptibility to stresses and an altered cell wall C_LIO_LIC. auris Tsa1b is involved in the fungal interaction with host immune cells C_LIO_LIThe Tsa1b of C. auris contributes to the progression of the infection in vivo C_LI

microbiology↗

bioRxiv2024

Card9 and MyD88 differentially regulate Th17 immunity to the commensal yeast Malassezia in the murine skin

The fungal community of the skin microbiome is dominated by a single genus, Malassezia. Besides its symbiotic lifestyle at the host interface, this commensal yeast has also been associated with diverse inflammatory skin diseases in humans and pet animals. Stable colonization is maintained by antifungal type 17 immunity. The mechanisms driving Th17 responses to Malassezia remain, however, unclear. Here, we show that the C-type lectin receptors Mincle, Dectin-1, and Dectin-2 recognize conserved patterns in the cell wall of Malassezia and induce dendritic cell activation in vitro, while only Dectin-2 is required for Th17 activation during experimental skin colonization in vivo. In contrast, Toll-like receptor recognition was redundant in this context. Instead, inflammatory IL-1 family cytokines signaling via MyD88 were also implicated in Th17 activation in a T cell-intrinsic manner. Taken together, we characterized the pathways contributing to protective immunity against the most abundant member of the skin mycobiome. This knowledge contributes to the understanding of barrier immunity and its regulation by commensals and is relevant considering how aberrant immune responses are associated with severe skin pathologies.

immunology↗

bioRxiv2024

Malassezia responds to environmental pH signals through the conserved Rim/Pal pathway

During mammalian colonization and infection, microorganisms must be able to rapidly sense and adapt to changing environmental conditions including alterations in extracellular pH. The fungus-specific Rim/Pal signaling pathway is one process that supports microbial adaptation to alkaline pH. This cascading series of interacting proteins terminates in the proteolytic activation of the highly conserved Rim101/PacC protein, a transcription factor that mediates microbial responses that favor survival in neutral/alkaline pH growth conditions, including many mammalian tissues. We identified the putative Rim pathway proteins Rim101 and Rra1 in the human skin colonizing fungus Malassezia sympodialis. Gene deletion by transconjugation and homologous recombination revealed that Rim101 and Rra1 are required for M. sympodialis growth at higher pH. Additionally, comparative transcriptional analysis of the mutant strains compared to wild-type suggested mechanisms for fungal adaptation to alkaline conditions. These pH-sensing signaling proteins are required for optimal growth in a murine model of atopic dermatitis, a pathological condition associated with increased skin pH. Together these data elucidate both conserved and phylum-specific features of microbial adaptation to extracellular stresses. ImportanceThe ability to adapt to host pH has been previously associated with microbial virulence in several pathogenic fungal species. Here we demonstrate that a fungal-specific alkaline response pathway is conserved in the human skin commensal fungus Malassezia sympodialis (Ms). This pathway is characterized by the pH-dependent activation of the Rim101/PacC transcription factor that controls cell surface adaptations to changing environmental conditions. By disrupting genes encoding two predicted components of this pathway, we demonstrated that the Rim/Pal pathway is conserved in this fungal species as a facilitator of alkaline pH growth. Moreover, targeted gene mutation and comparative transcriptional analysis supports the role of the Ms Rra1 protein as a cell surface pH sensor conserved within the basidiomycete fungi, a group including plant and human pathogens. Using an animal model of atopic dermatitis, we demonstrate the importance of Ms Rim/Pal signaling in this common inflammatory condition characterized by increased skin pH.

microbiology↗

bioRxiv2023

γδ T cells respond directly and selectively to the skin commensal yeast Malassezia for IL-17-dependent fungal control.

Stable microbial colonization of the skin depends on tight control by the host immune system. The lipid-dependent yeast Malassezia typically colonizes skin as a harmless commensal and is subject to host type 17 immunosurveillance, but this fungus has also been associated with diverse skin pathologies in both humans and animals. Using a murine model of Malassezia exposure, we show that V{gamma}4+ dermal {gamma}{delta} T cells expand rapidly and are the major source of IL-17A mediating fungal control in colonized skin. A pool of memory-like Malassezia-responsive V{gamma}4+ T cells persisted in the skin, were enriched in draining lymph nodes even after fungal clearance, and were protective upon fungal re-exposure up to several weeks later. Induction of {gamma}{delta}T17 immunity depended on IL-23 and IL-1 family cytokine signalling, whereas Toll-like and C-type lectin receptors were dispensable. Furthermore, V{gamma}4+ T cells from Malassezia-exposed hosts were able to respond directly and selectively to Malassezia-derived ligands, independently of antigen-presenting host cells. Reactivity of human {gamma}{delta} T cells against Malassezia spp. confirmed the relevance of this fungus-specific response across different host species. The fungal moieties detected were shared across diverse species of the Malassezia genus, but not conserved in other Basidiomycota or Ascomycota. These data provide novel mechanistic insight into the induction and maintenance of type 17 immunosurveillance of skin commensal colonization that has significant implications for cutaneous health. AUTHOR SUMMARYMalassezia is the most abundant fungus living on our skin and is usually harmless, but this microbe has also been shown to play a role in pathological conditions such as eczema and dermatitis. Here, we investigated how a population of V{gamma}4+ {gamma}{delta} T cells protect mouse skin against fungal overgrowth. While generally considered part of the innate immune system, we found that {gamma}{delta} T cells were maintained long after Malassezia was cleared from the skin of experimentally-infected animals. These cells displayed memory-like features and were highly efficient at fighting the fungus after a secondary challenge. We observed that classic fungal pattern recognition receptors were not involved, and that antigen-presenting cells were not required to generate Malassezia-protective {gamma}{delta} T cells. Finally, we confirmed that {gamma}{delta} T cells can recognise Malassezia, both in mice and human hosts. The fungal structure that triggers these responses was highly specific to Malassezia and was not conserved among other fungi. Our results highlight for the first time an important role for {gamma}{delta} T cells in preventing uncontrolled growth of the most abundant skin fungus. These findings have implications for several Malassezia-associated pathologies including eczema, dermatitis, and potentially even cancer.

immunology↗

bioRxiv2023

Epidermal barrier dysregulation in atopic skin predisposes for excessive growth of the allergy-associated yeast Malassezia

The skin barrier is vital for protection against environmental threats including insults caused by skin-resident microbes. Dysregulation of the barrier is a hallmark of atopic dermatitis (AD) and ichthyosis, with variable consequences for host immune control of colonizing commensals and opportunistic pathogens. While sensitisation to Malassezia, the most abundant commensal fungus of the skin, is common in AD, its relevance for pathogenesis remains unclear. Here we show that in barrier-disrupted skin, Malassezia acquires enhanced fitness. This is not a consequence of the dysregulated allergic immune status characteristic for AD but is rather explained by structural and metabolic changes in the cutaneous niche that provide increased accessibility and a favourable lipid profile, to which the lipid-dependent yeast adapts for enhanced nutrient assimilation. These findings reveal fundamental insights into the role of the mycobiota in the pathogenesis of common skin barrier disorders.

microbiology↗

bioRxiv2021

Candida albicans commensalism in the oral mucosa is favoured by limited virulence and metabolic adaptation .

As part of the human microbiota, the fungus Candida albicans colonizes the oral cavity and other mucosal surfaces of the human body. Commensalism is tightly controlled by complex interactions of the fungus and the host to preclude fungal elimination but also fungal overgrowth and invasion, which can result in disease. As such, defects in antifungal T cell immunity render individuals susceptible to oral thrush due to interrupted immunosurveillance of the oral mucosa. The factors that promote commensalism and ensure persistence of C. albicans in a fully immunocompetent host remain less clear. Using an experimental model of C. albicans oral colonization in mice we explored fungal determinants of commensalism in the oral cavity. Transcript profiling of the oral isolate 101 in the murine tongue tissue revealed a characteristic metabolic profile tailored to the nutrient poor conditions in the stratum corneum of the epithelium where the fungus resides. Metabolic adaptation of isolate 101 was also reflected in enhanced nutrient acquisition when grown on oral mucosa substrates. Persistent colonization of the oral mucosa by C. albicans also correlated inversely with the capacity of the fungus to induce epithelial cell damage and to elicit an inflammatory response. Here we show that these immune evasive properties of isolate 101 are explained by a strong attenuation of a number of virulence genes, including those linked to filamentation. De-repression of the hyphal program by deletion or conditional repression of NRG1 abolished the commensal behaviour of isolate 101, thereby establishing a central role of this factor in the commensal lifestyle of C. albicans in the oral niche of the host. AUTHOR SUMMARYThe oral microbiota represents an important part of the human microbiota and includes several hundreds to several thousands of bacterial and fungal species. One of the most prominent fungus colonizing the oral cavity is the yeast Candida albicans. While the presence of C. albicans usually remains unnoticed, the fungus can under certain circumstances cause lesions on the lining of the mouth referred to as oral thrush or contribute to other common oral diseases such as caries. Maintaining C. albicans commensalism in the oral mucosa is therefore of utmost importance for oral health and overall wellbeing. While overt fungal growth and disease is limited by immunosurveillance mechanisms during homeostasis, C. albicans strives to survive and evades elimination from the host. Here, we show that while commensalism in the oral cavity is characterized by a restricted fungal virulence and hyphal program, enforcing filamentation in a commensal isolate is sufficient for driving pathogenicity and fungus-induced inflammation in the oral mucosa thwarting persistent colonization. Our results further support a critical role for specialized nutrient acquisition allowing the fungus to thrive in the nutrient poor environment of the squamous epithelium. Together, this work revealed key determinants of C. albicans commensalism in the oral niche.

microbiology↗

bioRxiv2020

Horizontal gene transfer in the human and skin commensal Malassezia: a bacterially-derived flavohemoglobin is required for NO resistance and host interaction

The skin of humans and animals is colonized by commensal and pathogenic fungi and bacteria that share this ecological niche and have established microbial interactions. Malassezia are the most abundant fungal skin inhabitant of warm-blooded animals, and have been implicated in skin diseases and systemic disorders, including Crohns disease and pancreatic cancer. Flavohemoglobin is a key enzyme involved in microbial nitrosative stress resistance and nitric oxide degradation. Comparative genomics and phylogenetic analyses within the Malassezia genus revealed that flavohemoglobin-encoding genes were acquired through independent horizontal gene transfer events from different donor bacteria that are part of the mammalian microbiome. Through targeted gene deletion and functional complementation in M. sympodialis, we demonstrated that bacterially-derived flavohemoglobins are cytoplasmic proteins required for nitric oxide detoxification and nitrosative stress resistance under aerobic conditions. RNAseq analysis revealed that endogenous accumulation of nitric oxide resulted in upregulation of genes involved in stress response, and downregulation of the MalaS7 allergen-encoding genes. Solution of the high-resolution X-ray crystal structure of Malassezia flavohemoglobin revealed features conserved with both bacterial and fungal flavohemoglobins. In vivo pathogenesis is independent of Malassezia flavohemoglobin. Lastly, we identified additional 30 genus- and species-specific horizontal gene transfer candidates that might have contributed to the evolution of this genus as the most common inhabitants of animal skin. Significance statementMalassezia species are the main fungal components of the mammalian skin microbiome and are associated with a number of skin disorders. Recently, Malassezia has also been found in association with Crohns Disease and with pancreatic cancer. The elucidation of the molecular bases of skin adaptation by Malassezia is critical to understand its role as commensal and pathogen. In this study we employed evolutionary, molecular, biochemical, and structural analyses to demonstrate that the bacterially-derived flavohemoglobins acquired by Malassezia through horizontal gene transfer resulted in a gain of function critical for nitric oxide detoxification and resistance to nitrosative stress. Our study underscores horizontal gene transfer as an important force modulating Malassezia evolution and niche adaptation.

microbiology↗

bioRxiv2019

A novel mycovirus evokes transcriptional rewiring in Malassezia and provokes host inflammation and an immunological response

Mycoviruses infect fungi, and while most persist asymptomatically, there are examples of mycoviruses having both beneficial and detrimental effects on their host. Virus-infected Saccharomyces and Ustilago strains exhibit a killer phenotype conferring a growth advantage over uninfected strains and other competing yeast species, whereas hypovirus-infected Cryphonectria parasitica displays defects in growth, sporulation, and virulence. In this study we identify a dsRNA mycovirus in five Malassezia species. Sequence analysis reveals it to be a totivirus with two dsRNA segments: a larger 4.5 kb segment with genes encoding components for viral replication and maintenance, and a smaller 1.4 kb segment encoding a novel protein. Furthermore, RNA-seq of virus-infected versus virus-cured Malassezia sympodialis revealed an upregulation of dozens of ribosomal components in the cell, suggesting the virus modifies the transcriptional and translational landscapes of the cell. Given that Malassezia is the most abundant fungus on human skin, we assessed the impact of the mycovirus in a murine epicutaneous infection model. Although infection with virus-infected strains was not associated with an increased inflammatory response, we did observe enhanced skin colonization in one of two virus-infected M. sympodialis strains. Noteworthy, interferon-{beta} expression was significantly upregulated in bone marrow-derived macrophages when challenged with virus-infected, compared to virus-cured M. sympodialis, suggesting that the presence of the virus can induce an immunological response. Although many recent studies have illuminated how widespread mycoviruses are, there are relatively few in-depth studies about their impact on disease caused by the host fungus. We describe here a novel mycovirus in Malassezia and its possible implications in pathogenicity. ImportanceMalassezia species represent the most common fungal inhabitant of the mammalian skin microbiome, and are natural skin commensal flora. However, these fungi are also associated with a variety of clinical skin disorders. Recent studies have reported associations of Malassezia with Crohns disease and pancreatic cancer, further implicating this fungal genus in inflammatory and neoplastic disease states. Because M. sympodialis has lost genes involved in RNAi, we hypothesized Malassezia could harbor dsRNA mycoviruses. Indeed, we identified a novel mycovirus of the totivirus family in several Malassezia species, and characterized the MsMV1 mycovirus of M. sympodialis. We found conditions that lead to curing of the virus, and analyzed isogenic virus-infected/virus-cured strains to determine MsMV1 genetic and pathogenic impacts. MsMV1 induces a strong overexpression of transcription factors and ribosomal genes, while downregulating cellular metabolism. Moreover, MsMV1 induced a significantly higher level of interferon-{beta} expression in cultured macrophages. This study sheds light on the mechanisms of pathogenicity of Malassezia, focusing on a previously unidentified novel mycovirus.

microbiology↗

bioRxiv2018

The fungal ligand chitin directly binds and signals inflammation dependent on oligomer size and TLR2

Chitin is a highly abundant polysaccharide and linked to fungal infection and asthma. Unfortunately, its polymeric structure has hampered the identification of immune receptors directly binding chitin and signaling immune activation and inflammation, because purity, molecular structure and molarity are not well definable for a polymer typically extracted from biomass. Therefore, by using defined chitin (N-acetyl-glucosamine) oligomers, we identified six subunit long chitin chains as the smallest immunologically active motif and the innate immune receptor Toll-like receptor (TLR) 2 as the primary fungal chitin receptor on human and murine immune cells. Chitin oligomers directly bound TLR2 with nanomolar affinity and showed both overlapping and distinct signaling outcomes compared to known mycobacterial TLR2 ligands. Conversely, chitin oligomers shorter than 6 subunits were inactive or showed antagonistic effects on chitin/TLR2-mediated signaling, hinting to a size-dependent sensing/activation system unexpectedly conserved in plants and humans. Since blocking the chitin-TLR2 interaction effectively prevented chitin-mediated inflammation in vitro and in vivo, our study highlights the chitin TLR2 interaction as a potential target for developing novel therapies in chitin-related pathologies and fungal disease.

immunology↗