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MBio May 2018Microbial secondary metabolites, including isocyanide moieties, have been extensively mined for their repertoire of bioactive properties. Although the first naturally...
Microbial secondary metabolites, including isocyanide moieties, have been extensively mined for their repertoire of bioactive properties. Although the first naturally occurring isocyanide (xanthocillin) was isolated from the fungus over half a century ago, the biosynthetic origins of fungal isocyanides remain unknown. Here we report the identification of a family of isocyanide synthases (ICSs) from the opportunistic human pathogen Comparative metabolomics of overexpression or knockout mutants of ICS candidate genes led to the discovery of a fungal biosynthetic gene cluster (BGC) that produces xanthocillin (). Detailed analysis of xanthocillin biosynthesis in revealed several previously undescribed compounds produced by the BGC, including two novel members of the melanocin family of compounds. We found both the BGC and a second ICS-containing cluster, named the copper-responsive metabolite () BGC, to be transcriptionally responsive to external copper levels and further demonstrated that production of metabolites from the BGC is increased during copper starvation. The BGC includes a novel type of fungus-specific ICS-nonribosomal peptide synthase (NRPS) hybrid enzyme, CrmA. This family of ICS-NRPS hybrid enzymes is highly enriched in fungal pathogens of humans, insects, and plants. Phylogenetic assessment of all ICSs spanning the tree of life shows not only high prevalence throughout the fungal kingdom but also distribution in species not previously known to harbor BGCs, indicating an untapped resource of fungal secondary metabolism. Fungal ICSs are an untapped resource in fungal natural product research. Their isocyanide products have been implicated in plant and insect pathogenesis due to their ability to coordinate transition metals and disable host metalloenzymes. The discovery of a novel isocyanide-producing family of hybrid ICS-NRPS enzymes enriched in medically and agriculturally important fungal pathogens may reveal mechanisms underlying pathogenicity and afford opportunities to discover additional families of isocyanides. Furthermore, the identification of noncanonical ICS BGCs will enable refinement of BGC prediction algorithms to expand on the secondary metabolic potential of fungal and bacterial species. The identification of genes related to ICS BGCs in fungal species not previously known for secondary metabolite-producing capabilities (e.g., spp.) contributes to our understanding of the evolution of BGC in fungi.
Topics: Aspergillus fumigatus; Biosynthetic Pathways; Butadienes; Cyanides; Fungal Proteins; Multigene Family; Peptide Synthases; Phenols; Phylogeny
PubMed: 29844112
DOI: 10.1128/mBio.00785-18 -
Proceedings. Biological Sciences Feb 2019Aspergillus fumigatus causes a range of diseases in humans, some of which are characterized by fungal persistence. Aspergillus fumigatus, being a generalist saprotroph,...
Aspergillus fumigatus causes a range of diseases in humans, some of which are characterized by fungal persistence. Aspergillus fumigatus, being a generalist saprotroph, may initially establish lung colonization due to its physiological versatility and subsequently adapt through genetic changes to the human lung environment and antifungal treatments. Human lung-adapted genotypes can arise by spontaneous mutation and/or recombination and subsequent selection of the fittest genotypes. Sexual and asexual spores are considered crucial contributors to the genetic diversity and adaptive potential of aspergilli by recombination and mutation supply, respectively. However, in certain Aspergillus diseases, such as cystic fibrosis and chronic pulmonary aspergillosis, A. fumigatus may not sporulate but persist as a network of fungal mycelium. During azole therapy, such mycelia may develop patient-acquired resistance and become heterokaryotic by mutations in one of the nuclei. We investigated the relevance of heterokaryosis for azole-resistance development in A. fumigatus. We found evidence for heterokaryosis of A. fumigatus in patients with chronic Aspergillus diseases. Mycelium from patient-tissue biopsies segregated different homokaryons, from which heterokaryons could be reconstructed. Whereas all variant homokaryons recovered from the same patient were capable of forming a heterokaryon, those from different patients were heterokaryon-incompatible. We furthermore compared heterokaryons and heterozygous diploids constructed from environmental isolates with different levels of azole resistance. When exposed to azole, the heterokaryons revealed remarkable shifts in their nuclear ratio, and the resistance level of heterokaryons exceeded that of the corresponding heterozygous diploids.
Topics: Adaptation, Biological; Antifungal Agents; Aspergillus fumigatus; Azoles; Drug Resistance, Fungal; Genetic Variation
PubMed: 30963936
DOI: 10.1098/rspb.2018.2886 -
Microbiology (Reading, England) Aug 2018Aspergillus fumigatus is a saprotrophic fungus that continuously disseminates spores (conidia) into the environment. It is also the most common and opportunistic aerial...
Aspergillus fumigatus is a saprotrophic fungus that continuously disseminates spores (conidia) into the environment. It is also the most common and opportunistic aerial fungal pathogen, causing allergic and chronic lung pathologies including the fatal invasive aspergillosis in immunocompromised patients. The pathobiology of aspergillosis is complex and depends on the competence of the host immune system. Moreover, A. fumigatus has become a model to study unique features of fungi. This includes the fungal cell wall, which not only acts as a rigid skeleton for protection against hostile environments but also plays significant roles during infection by manipulating the host immune response.
Topics: Aspergillus fumigatus; Genome, Fungal; Mycelium; Phylogeny; Pulmonary Aspergillosis; Spores, Fungal
PubMed: 30066670
DOI: 10.1099/mic.0.000651 -
Frontiers in Cellular and Infection... 2019Dendritic cells (DCs) are antigen presenting cells which serve as a passage between the innate and the acquired immunity. Aspergillosis is a major lethal condition in...
Dendritic cells (DCs) are antigen presenting cells which serve as a passage between the innate and the acquired immunity. Aspergillosis is a major lethal condition in immunocompromised patients caused by the adaptable saprophytic fungus . The healthy human immune system is capable to ward off infections however immune-deficient patients are highly vulnerable to invasive aspergillosis. can persist during infection due to its ability to survive the immune response of human DCs. Therefore, the study of the metabolism specific to the context of infection may allow us to gain insight into the adaptation strategies of both the pathogen and the immune cells. We established a metabolic model of central metabolism during infection of DCs and calculated the metabolic pathway (elementary modes; EMs). Transcriptome data were used to identify pathways activated when is challenged with DCs. In particular, amino acid metabolic pathways, alternative carbon metabolic pathways and stress regulating enzymes were found to be active. Metabolic flux modeling identified further active enzymes such as alcohol dehydrogenase, inositol oxygenase and GTP cyclohydrolase participating in different stress responses in . These were further validated by qRT-PCR from RNA extracted under these different conditions. For DCs, we outlined the activation of metabolic pathways in response to the confrontation with . We found the fatty acid metabolism plays a crucial role, along with other metabolic changes. The gene expression data and their analysis illuminate additional regulatory pathways activated in the DCs apart from interleukin regulation. In particular, Toll-like receptor signaling, NOD-like receptor signaling and RIG-I-like receptor signaling were active pathways. Moreover, we identified subnetworks and several novel key regulators such as UBC, EGFR, and CUL3 of DCs to be activated in response to . In conclusion, we analyze the metabolic and regulatory responses of and DCs when confronted with each other.
Topics: Aspergillus fumigatus; Cytokines; Dendritic Cells; Gene Expression; Host-Pathogen Interactions; Humans; Interleukins; Metabolic Networks and Pathways; NLR Proteins; Signal Transduction; Toll-Like Receptors; Transcriptome
PubMed: 31192161
DOI: 10.3389/fcimb.2019.00168 -
Philosophical Transactions of the Royal... Dec 2016Aspergillus fungi are the cause of an array of diseases affecting humans, animals and plants. The triazole antifungal agents itraconazole, voriconazole, isavuconazole... (Review)
Review
Aspergillus fungi are the cause of an array of diseases affecting humans, animals and plants. The triazole antifungal agents itraconazole, voriconazole, isavuconazole and posaconazole are treatment options against diseases caused by Aspergillus However, resistance to azoles has recently emerged as a new therapeutic challenge in six continents. Although de novo azole resistance occurs occasionally in patients during azole therapy, the main burden is the aquisition of resistance through the environment. In this setting, the evolution of resistance is attributed to the widespread use of azole-based fungicides. Although ubiquitously distributed, A. fumigatus is not a phytopathogen. However, agricultural fungicides deployed against plant pathogenic moulds such as Fusarium, Mycospaerella and A. flavus also show activity against A. fumigatus in the environment and exposure of non-target fungi is inevitable. Further, similarity in molecule structure between azole fungicides and antifungal drugs results in cross-resistance of A. fumigatus to medical azoles. Clinical studies have shown that two-thirds of patients with azole-resistant infections had no previous history of azole therapy and high mortality rates between 50% and 100% are reported in azole-resistant invasive aspergillosis. The resistance phenotype is associated with key mutations in the cyp51A gene, including TR/L98H, TR and TR/Y121F/T289A resistance mechanisms. Early detection of resistance is of paramount importance and if demonstrated, either with susceptibility testing or through molecular analysis, azole monotherapy should be avoided. Liposomal amphotericin B or a combination of voriconazole and an echinocandin are recomended for azole-resistant aspergillosis.This article is part of the themed issue 'Tackling emerging fungal threats to animal health, food security and ecosystem resilience'.
Topics: Antifungal Agents; Aspergillosis; Aspergillus fumigatus; Azoles; Drug Resistance, Fungal; Fungicides, Industrial; Phenotype
PubMed: 28080986
DOI: 10.1098/rstb.2015.0460 -
MSphere May 2020is an opportunistic and allergenic pathogenic fungus, responsible for fungal infections in humans. infections are usually treated with polyenes, azoles, or...
is an opportunistic and allergenic pathogenic fungus, responsible for fungal infections in humans. infections are usually treated with polyenes, azoles, or echinocandins. Echinocandins, such as caspofungin, can inhibit the biosynthesis of the β-1,3-glucan polysaccharide, affecting the integrity of the cell wall and leading to fungal death. In some strains, caspofungin treatment at high concentrations induces an increase of fungal growth, a phenomenon called the aspofungin aradoxical ffect (CPE). Here, we analyze the proteome and phosphoproteome of the wild-type strain and of mitogen-activated protein kinase (MAPK) and null mutant strains during CPE (2 μg/ml caspofungin for 1 h). The wild-type proteome showed 75 proteins and 814 phosphopeptides (corresponding to 520 proteins) altered in abundance in response to caspofungin treatment. The Δ (Δ caspofungin/wild-type caspofungin) and Δ (Δ caspofungin/wild-type caspofungin) strains displayed 626 proteins and 1,236 phosphopeptides (corresponding to 703 proteins) and 101 proteins and 1,217 phosphopeptides (corresponding to 645 proteins), respectively, altered in abundance. Functional characterization of the phosphopeptides from the wild-type strain exposed to caspofungin showed enrichment for transcription factors, protein kinases, and cytoskeleton proteins. Proteomic analysis of the Δ and Δ mutants indicated that control of proteins involved in metabolism, such as in production of secondary metabolites, was highly represented in both mutants. Results of functional categorization of phosphopeptides from both mutants were very similar and showed a high number of proteins with decreased phosphorylation of proteins involved in transcriptional control, DNA/RNA binding, cell cycle control, and DNA processing. This report reveals novel transcription factors involved in caspofungin tolerance. is an opportunistic human-pathogenic fungus causing allergic reactions or systemic infections, such as invasive pulmonary aspergillosis in immunocompromised patients. Caspofungin is an echinocandin that impacts the construction of the fungal cell wall by inhibiting the biosynthesis of the β-1,3-glucan polysaccharide. Caspofungin is a fungistatic drug and is recommended as a second-line therapy for treatment of aspergillosis. Treatment at high concentrations induces an increase of fungal growth, a phenomenon called the aspofungin aradoxical ffect (CPE). Collaboration between the mitogen-activated protein kinases (MAPK) of the cell wall integrity (MapkA) and high-osmolarity glycerol (SakA) pathways is essential for CPE. Here, we investigate the global proteome and phosphoproteome of wild-type, Δ, and Δ strains upon CPE. This study showed intense cross talk between the two MAPKs for the CPE and identified novel protein kinases and transcription factors possibly important for CPE. Increased understanding of how the modulation of protein phosphorylation may affect the fungal growth in the presence of caspofungin represents an important step in the development of new strategies and methods to combat the fungus inside the host.
Topics: Antifungal Agents; Aspergillus fumigatus; Caspofungin; Fungal Proteins; Mass Spectrometry; Phosphopeptides; Phosphorylation; Proteome; Proteomics; Signal Transduction; Transcription Factors
PubMed: 32461274
DOI: 10.1128/mSphere.00365-20 -
Emerging Microbes & Infections Dec 2017This study investigated the triazole phenotype and genotypic of clinical Aspergillus fumigatus isolates from China. We determined the triazole susceptibility profiles of...
This study investigated the triazole phenotype and genotypic of clinical Aspergillus fumigatus isolates from China. We determined the triazole susceptibility profiles of 159 A. fumigatus isolates collected between 2011 and 2015 from four different areas in China tested against 10 antifungal drugs using the Clinical Laboratory Standard Institute M38-A2 method. For the seven itraconazole-resistant A. fumigatus isolates identified in the study, the cyp51A gene, including its promoter region, was sequenced and the mutation patterns were characterized. The resistant isolates were genotyped by microsatellite typing to determine the genetic relatedness to isolates from China and other countries. The frequency of itraconazole resistance in A. fumigatus isolates in our study was 4.4% (7/159). Six of the seven triazole-resistant isolates were recovered from the east and southeast of China, and one from was recovered from the west of China. No resistant isolates were found in the north. Three triazole-resistant isolates exhibited the TR/L98H mutation, two carried the TR/L98H/S297T/F495I mutation and one harbored a G54V mutation in the cyp51A gene. Analysis of the microsatellite markers from seven non-wild-type isolates indicated the presence of five unique genotypes, which clustered into two major genetic groups. The cyp51A gene mutations TR/L98H and TR/L98H/S297T were the most frequently found mutations, and the G54V mutation was reported for the first time in China. The geographic origin of the triazole-resistant isolates appeared to concentrate in eastern and south-eastern areas, which suggests that routine antifungal susceptibility testing in these areas should be performed for all clinically relevant A. fumigatus isolates to guide antifungal therapy and for epidemiological purposes.
Topics: Antifungal Agents; Aspergillosis; Aspergillus fumigatus; China; Fungal Proteins; Genotype; Humans; Microbial Sensitivity Tests; Phenotype; Triazoles
PubMed: 29209054
DOI: 10.1038/emi.2017.97 -
BMC Systems Biology Apr 2015Aspergillus fumigatus is a ubiquitous airborne fungal pathogen that presents a life-threatening health risk to individuals with weakened immune systems. A. fumigatus...
BACKGROUND
Aspergillus fumigatus is a ubiquitous airborne fungal pathogen that presents a life-threatening health risk to individuals with weakened immune systems. A. fumigatus pathogenicity depends on its ability to acquire iron from the host and to resist host-generated oxidative stress. Gaining a deeper understanding of the molecular mechanisms governing A. fumigatus iron acquisition and oxidative stress response may ultimately help to improve the diagnosis and treatment of invasive aspergillus infections.
RESULTS
This study follows a systems biology approach to investigate how adaptive behaviors emerge from molecular interactions underlying A. fumigatus iron regulation and oxidative stress response. We construct a Boolean network model from known interactions and simulate how changes in environmental iron and superoxide levels affect network dynamics. We propose rules for linking long term model behavior to qualitative estimates of cell growth and cell death. These rules are used to predict phenotypes of gene deletion strains. The model is validated on the basis of its ability to reproduce literature data not used in model generation.
CONCLUSIONS
The model reproduces gene expression patterns in experimental time course data when A. fumigatus is switched from a low iron to a high iron environment. In addition, the model is able to accurately represent the phenotypes of many knockout strains under varying iron and superoxide conditions. Model simulations support the hypothesis that intracellular iron regulates A. fumigatus transcription factors, SreA and HapX, by a post-translational, rather than transcriptional, mechanism. Finally, the model predicts that blocking siderophore-mediated iron uptake reduces resistance to oxidative stress. This indicates that combined targeting of siderophore-mediated iron uptake and the oxidative stress response network may act synergistically to increase fungal cell killing.
Topics: Aspergillus fumigatus; Biological Transport; Cell Death; Cell Proliferation; Environment; Fungal Proteins; Gene Knockout Techniques; Homeostasis; Iron; Models, Biological; Oxidative Stress; Oxygen; Phenotype; Siderophores; Stochastic Processes; Superoxides; Systems Biology
PubMed: 25908096
DOI: 10.1186/s12918-015-0163-1 -
Virulence 2018Gliotoxin contributes to the virulence of the fungus Aspergillus fumigatus in non-neutropenic mice that are immunosuppressed with corticosteroids. To investigate how the...
UNLABELLED
Gliotoxin contributes to the virulence of the fungus Aspergillus fumigatus in non-neutropenic mice that are immunosuppressed with corticosteroids. To investigate how the absence of gliotoxin affects both the fungus and the host, we used a nanoString nCounter to analyze their transcriptional responses during pulmonary infection of a non-neutropenic host with a gliotoxin-deficient ΔgliP mutant. We found that the ΔgliP mutation led to increased expression of aspf1, which specifies a secreted ribotoxin. Prior studies have shown that aspf1, like gliP, is not required for virulence in a neutropenic infection model, but its role in a non-neutropenic infection model has not been fully investigated. To investigate the functional significance of this up-regulation of aspf1, a Δaspf1 single mutant and a Δaspf1 ΔgliP double mutant were constructed. Both Δaspf1 and ΔgliP single mutants had reduced lethality in non-neutropenic mice, and a Δaspf1 ΔgliP double mutant had a greater reduction in lethality than either single mutant. Analysis of mice infected with these mutants indicated that the presence of gliP is associated with massive apoptosis of leukocytes at the foci of infection and inhibition of chemokine production. Also, the combination of gliP and aspf1 is associated with suppression of CXCL1 chemokine expression. Thus, aspf1 contributes to A. fumigatus pathogenicity in non-neutropenic mice and its up-regulation in the ΔgliP mutant may partially compensate for the absence of gliotoxin.
ABBREVIATIONS
PAS: periodic acid-Schiff; PBS: phosphate buffered saline; ROS: reactive oxygen species; TUNEL: terminal deoxynucleotidyl transferase dUTP nick-end labeling.
Topics: Allergens; Animals; Aspergillosis; Aspergillus fumigatus; Fungal Proteins; Gene Deletion; Gliotoxin; Humans; Male; Mice; Mice, Inbred BALB C; Virulence
PubMed: 30052103
DOI: 10.1080/21505594.2018.1482182 -
Virulence Nov 2016Invasive aspergillosis is a life-threatening infection caused by the opportunistic filamentous fungus Aspergillus fumigatus. Patients undergoing haematopoietic stem cell... (Review)
Review
Invasive aspergillosis is a life-threatening infection caused by the opportunistic filamentous fungus Aspergillus fumigatus. Patients undergoing haematopoietic stem cell transplant (HSCT) for the treatment of hematological malignancy are at particularly high risk of developing this fatal infection. The susceptibility of HSCT patients to infection with A. fumigatus is a consequence of a complex interplay of both fungal and host factors. Here we review our understanding of the host-pathogen interactions underlying the susceptibility of the immunocompromised host to infection with A. fumigatus with a focus on the experimental validation of fungal and host factors relevant to HSCT patients. These include fungal factors such as secondary metabolites, cell wall constituents, and metabolic adaptations that facilitate immune evasion and survival within the host microenvironment, as well as the innate and adaptive immune responses involved in host defense against A. fumigatus.
Topics: Aspergillus fumigatus; Hematologic Neoplasms; Host-Pathogen Interactions; Humans; Immune Evasion; Immunity, Innate; Immunocompromised Host; Invasive Pulmonary Aspergillosis; Stem Cell Transplantation; Virulence
PubMed: 27687755
DOI: 10.1080/21505594.2016.1231278