Fungal Metabolism Research Articles

Dynamics of gliotoxin and bis(methylthio)gliotoxin production during the course of Aspergillus fumigatus infection.

Aspergillus fumigatus secondary metabolites may contribute to fungal survival. A new chromatographic method was applied to simultaneously characterize two relevant metabolites. Presence of toxic gliotoxin may indicate an early phase of development, whereas the detection of the inactive derivate may represent an advanced infection stage.

The phosphate language of fungi.

Phosphate is an essential macronutrient for fungal proliferation as well as a key mediator of antagonistic, beneficial, and pathogenic interactions between fungi and other organisms. In this review, we summarize recent insights into the integration of phosphate metabolism with mechanisms of fungal adaptation that support growth and survival. In particular, we highlight aspects of phosphate sensing important for responses to stress and regulation of cell-surface changes with an impact on fungal pathogenesis, host immune responses, and disease outcomes. Additionally, new studies provide insights into the influence of phosphate availability on cooperative or antagonistic interactions between fungi and other microbes, the associations of mycorrhizal and endophytic fungi with plants, and connections with plant immunity. Overall, phosphate homeostasis is emerging as an integral part of fungal metabolism and communication to support diverse lifestyles.

Relevance of Nutrient-Sensing in the Pathogenesis of Trichophyton rubrum and Trichophyton interdigitale.

The growth and development of organisms depend on nutrient availability. Dermatophytes must sense nutrient levels and adapt to the host environment to colonize human and animal keratinized tissues. Owing to the clinical importance of the Trichophyton genus, this study compared the expression profile of genes involved in metabolism, cell cycle control, and proteases in two Trichophyton species, Trichophyton rubrum, and Trichophyton interdigitale, in response to nutrients and environmental pH. In addition, we evaluated the activity of enzymes in the tricarboxylic acid, glyoxylate, and methylcitrate cycles. Moreover, the effects of interruption of the transcription factor pacC on T. interdigitale in the same conditions as for the wild-type strain were determined. Our analyses revealed specific responses in each species to the nutritional and pH variation. An improved adaptation of T. interdigitale to keratin was observed, compared with that of T. rubrum. T. rubrum growth in buffered keratin media indicated pH 8.0 as an optimal pH condition for metabolic activity, which differed from that for T. interdigitale. Tricarboxylic acid components in T. rubrum showed increased enzymatic activity and transcript accumulation. In T. interdigitale, a higher activity of enzymes in glyoxylate and methylcitrate cycles was observed, with no direct correlation to the transcriptional profile. T. interdigitale fungal metabolism suggests the requirement of anaplerotic pathways in the late cultivation period. The identified differences between T. rubrum and T. interdigitale may represent determinants for adaptation to the host and the incidence of infection with each species.

Sulfur Metabolism as a Promising Source of New Antifungal Targets.

Fungal infections are a growing threat to human health. Despite their clinical relevance, there is a surprisingly limited availability of clinically approved antifungal agents, which is seriously aggravated by the recent appearance and fast spread of drug resistance. It is therefore clear that there is an urgent need for novel and efficient antifungals. In this context, metabolism is recognized as a promising source for new antifungal targets and, indeed, there are new drugs in development that target metabolic pathways. Fungal sulfur metabolism is particularly interesting, as many of its processes are essential for viability and/or pathogenicity and it shows substantial differences with human metabolism. This short-review will summarize our current knowledge of sulfur-related genes and routes that are important for Aspergillus fumigatus virulence, which consequently could be pursued for drug development.

Emerging Antifungal Targets and Strategies.

Despite abundant research in the field of antifungal drug discovery, fungal infections remain a significant healthcare burden. There is an emerging need for the development of novel antifungals since those currently available are limited and do not completely provide safe and secure protection. Since the current knowledge regarding the physiology of fungal cells and the infection mechanisms is greater than ever, we have the opportunity to use this for the development of novel generations of antifungals. In this review, we selected and summarized recent studies describing agents employing different antifungal mechanisms. These mechanisms include interference with fungal resistance, including impact on the efflux pumps and heat shock protein 90. Additionally, interference with virulence factors, such as biofilms and hyphae; the impact on fungal enzymes, metabolism, mitochondria, and cell wall; and antifungal vaccines are explored. The agents investigated belong to different classes of natural or synthetic molecules with significant attention given also to plant extracts. The efficacy of these antifungals has been studied mainly in vitro with some in vivo, and clinical studies are needed. Nevertheless, there is a large quantity of products employing novel antifungal mechanisms that can be further explored for the development of new generation of antifungals.

Metrics of Antifungal Effects of Ciprofloxacin on Aspergillus fumigatus Planktonic Growth and Biofilm Metabolism; Effects of Iron and Siderophores.

Pseudomonas aeruginosa and Aspergillus fumigatus frequently coexist in the airways of immunocompromised patients or individuals with cystic fibrosis. Ciprofloxacin (CIP) is a synthetic quinolone antibiotic commonly used to treat bacterial infections, such as those produced by Pseudomonas aeruginosa. CIP binds iron, and it is unclear what effect this complex would have on the mycobiome. The effects of CIP on Aspergillus were dependent on the iron levels present, and on the presence of Aspergillus siderophores. We found that CIP alone stimulated wildtype planktonic growth, but not biofilm metabolism. At high concentrations, CIP antagonized a profungal effect of iron on wildtype Aspergillus metabolism, presumably owing to iron chelation. CIP interfered with the metabolism and growth of an Aspergillus siderophore mutant, with the effect on metabolism being antagonized by iron. CIP acted synergistically with iron on the growth of the mutant, and, to a lesser extent, the wildtype. In summary, CIP can increase fungal growth or affect fungal metabolism, depending on the local iron concentration and available siderophores. Therefore, high local CIP concentrations during treatment of Pseudomonas–Aspergillus co-infections may increase the fungal burden.

Sequential Visible Light-Induced Reactions Using Different Photocatalysts: Transformation of Furans into 2-Pyridones via γ-Lactams Using a New Ring Expansion Reaction.

A new photocatalytic ring expansion reaction that transforms γ-lactams into 2-pyridones is described. The reaction is radical-triggered and was inspired by a late-stage oxidation commonly observed in fungal metabolism. The reaction is particularly powerful because it can be included in a one pot process which converts readily accessible furans directly into 2-pyridones through sequential photo-induced reactions (energy transfer and electron transfer). The study also unveiled unprecedented reactivity for the mild and metal-free photocatalyst eosin, wherein it is used to cleave a C-I bond activated only by an electron donor on the neighbouring carbon.

Roles of AaVeA on Mycotoxin Production via Light in Alternaria alternata.

Alternaria alternata is a principal plant pathogen responsible for the biosynthesis of mycotoxins, including tenuazonic acid (TeA), alternariol (AOH), and alternariol monomethyl ether (AME). The velvet gene VeA is involved in fungal growth, development, and secondary metabolism, including mycotoxin biosynthesis via light regulation. In this study, the detailed regulatory roles of AaVeA in A. alternata with various light sources were investigated from the comparative analyses between the wild type and the gene knockout strains. In fungal growth and conidiation, mycelial extension was independent of light regulation in A. alternata. Red light favored conidiation, but blue light repressed it. The absence of AaVeA caused the marked reduction of hyphae extension and conidiophore formation even though red light could not induce more spores in ΔAaVeA mutant. The differentially expressed genes (DEGs) enriched in hyphal growth and conidiation were drastically transcribed from the comparatively transcriptomic profile between the wild type and ΔAaVeA mutant strains with or without light. In mycotoxin production, TeA biosynthesis seems no obvious effect by light regulation, but AOH and AME formation was significantly stimulated by blue light. Nevertheless, the disruption of AaVeA resulted in a marked decrease in mycotoxin production and the action of the stimulation was lost via blue light for the abundant accumulation of AOH and AME in the ΔAaVeA strain. From DEG expression and further verification by RT-qPCR, the loss of AaVeA caused the discontinuous supply of the substrates for mycotoxin biosynthesis and the drastic decline of biosynthetic gene expression. In addition, pathogenicity depends on AaVeA regulation in tomato infected by A. alternata in vivo. These findings provide a distinct understanding of the roles of AaVeA in fungal growth, development, mycotoxin biosynthesis, and pathogenicity in response to various light sources.

Lauric Acid Induces Apoptosis of Rice Sheath Blight Disease Caused by Rhizoctonia solani by Affecting Fungal Fatty Acid Metabolism and Destroying the Dynamic Equilibrium of Reactive Oxygen Species

Rice sheath blight, caused by Rhizoctonia solani, is one of the major rice diseases. In order to better understand the inhibitory mechanism of lauric acid on the disease, RNA sequencing (RNA-Seq) was used to analyze the transcriptome changes in Rhizoctonia solani treated with lauric acid for 3 h, 6 h, 18 h, and 24 h, including 2306 genes; 1994 genes; 2778 genes; and 2872 genes. Based on gene ontology (GO) enrichment and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses, we found that protein processing in endoplasmic reticulum (KO04141), carbon metabolism (KO01200), and starch and sucrose metabolism were significantly enriched. Most oxidoreductase, dehydrogenase, reductase, and transferase genes are downregulated in this process. Lauric acid can affect ergosterol content, mitochondrial membrane potential collapse, hydrogen peroxide content, electrolyte leakage, reactive oxygen species balance, and can induce endoplasmic reticulum (ER) stress. Lauric acid also increased the expression levels of ER chaperone glucose regulatory protein Grp78 (BIP), protein disulfide isomerase (PDI), and Calpain (CNX), and decreased the expression levels of HSP40, HSP70, and HSP90 genes. Lauric acid affected the ergosterol content in the cell membrane of R. solani, which induces ER stress and increases the BiP level to induce the apoptosis of Rhizoctonia solani. These results indicated that lauric acid could be used to control rice sheath blight.

Identification of fungal trehalose for the diagnosis of invasive candidiasis by mass spectrometry

The rapidity of the diagnosis of invasive candidiasis (IC) is crucial to allow the early introduction of antifungal therapy that dramatically increases the survival rate of patients. Early diagnosis is unfortunately often delayed because Candida blood culture, the gold standard diagnostic test, is positive in only 50% of cases of IC and takes several days to obtain this result. Complementary non-culture-based methods relying on the detection of Candida cell wall polysaccharides in the serum, β-glucans and mannans, by enzymatic and immunological reagents have been successfully developed to allow a more efficient patients care. We have previously demonstrated that detection of circulating glycans by mass spectrometry could provide a reliable and cost-effective early diagnosis method called MS-DS for Mass Spectrometry of Di-Saccharide. Here, by comparing patient's sera and Candida albicans strains deficient in carbohydrates synthesis, we demonstrate that trehalose derived from fungal metabolism can be specifically targeted by MS-DS to allow early diagnosis. In particular, the use of C. albicans strains deficient in the synthesis of trehalose synthesizing enzymes Tps1 and Tps2 show that MS-DS results were correlated to the metabolism of trehalose. Finally, we demonstrate that the performance of the IC diagnosis can be significantly improved by using high resolution mass spectrometry, which opens new perspectives in the management of the disease.

Bacterial volatile organic compounds induce adverse ultrastructural changes and DNA damage to the sugarcane pathogenic fungus Thielaviopsis ethacetica.

Due to an increasing demand for sustainable agricultural practices, the adoption of microbial volatile organic compounds (VOCs) as antagonists against phytopathogens has emerged as an eco-friendly alternative to the use of agrochemicals. Here, we identified three Pseudomonas strains that were able to inhibit, in vitro, up to 80% of mycelial growth of the phytopathogenic fungus Thielaviopsis ethacetica, the causal agent of pineapple sett rot disease in sugarcane. Using GC/MS, we found that these bacteria produced 62 different VOCs, and further functional validation revealed compounds with high antagonistic activity to T. ethacetica. Transcriptomic analysis of the fungal response to VOCs indicated that these metabolites downregulated genes related to fungal central metabolism, such as those involved in carbohydrate metabolism. Interestingly, genes related to the DNA damage response were upregulated, and micro-FTIR analysis corroborated our hypothesis that VOCs triggered DNA damage. Electron microscopy analysis showed critical morphological changes in mycelia treated with VOCs. Altogether, these results indicated that VOCs hampered fungal growth and could lead to cell death. This study represents the first demonstration of the molecular mechanisms involved in the antagonism of sugarcane phytopathogens by VOCs and reinforces that VOCs can be a sustainable alternative for use in phytopathogen biocontrol.

Advances in Production of Medicinal Mushrooms Biomass in Solid State and Submerged Bioreactors.

Production of mushroom fruit bodies using farming technology could hardly meet the increasing demand of the world market. During the last several decades, there have been various basic and applied studies on fungal physiology, metabolism, process engineering, and (pre)clinical studies. The fundamental aspects of solid-state cultivation of various kinds of medicinal mushroom mycelia in various types of bioreactors were established. Solid-state cultivation of medicinal mushrooms for their biomass and bioactive metabolites production appear very suitable for veterinary use. Development of comprehensive submerged technologies using stirred tank and airlift bioreactors is the most promising technology for fast and large-scale production of medicinal fungi biomass and their pharmaceutically active products for human need. The potentials initiate the development of new drugs and some of the most attractive over-the-counter human and veterinary remedies. This article is to overview the engineering achievements in solid state and submerged cultivations of medicinal mushrooms in bioreactors.

Fungal Secondary Metabolism

Fungal secondary metabolites (SMs) comprise a vast collection of compounds expendable for these organisms under laboratory conditions. They exhibit enormous chemical diversity, and usually belong to four major families: terpenoids, polyketides, non-ribosomal peptides, or a combination of the last two. Their functions are very diverse and are normally associated with a greater fitness of the producing fungi in their environment, which often compete with other microorganisms or interact with host plants. Many SMs have beneficial applications, e.g., as antibiotics or medical drugs, but others, known as mycotoxins, are harmful to health.

Metabolomic profiles of the liquid state fermentation in co-culture of A. oryzae and Z. rouxii

Aspergillus oryzae and Zygosaccharomyces rouxii are the perquisites microorganisms in food fermentation due to the broad application prospects of their secondary metabolites. The co-culture strategy simulates the naturally occurring ecology by developing artificial microbial communities. This strategy has also been widely adopted to investigate the novel secondary metabolites. In the present study, the effects of co-culture on extracellular and intracellular secondary metabolites of fungi in liquid culture were investigated through UPLC-QTOF-MS. Notably, A. oryzae could significantly inhibit the growth of Z. rouxii when A. oryzae and Z. rouxii were co-cultured. The results further indicated that the co-culture of fungi could affect the secondary metabolites and produce various metabolic pathways. Overall, this study will provide insights into fungal interactions and cell metabolism mechanisms, contributing to supplementing and enriching the fermentation potential of fungi.

Infection metallomics for critical care in the post-COVID era.

Infection metallomics is a mass spectrometry (MS) platform we established based on the central concept that microbial metallophores are specific, sensitive, noninvasive, and promising biomarkers of invasive infectious diseases. Here we review the in vitro, in vivo, and clinical applications of metallophores from historical and functional perspectives, and identify under-studied and emerging application areas with high diagnostic potential for the post-COVID era. MS with isotope data filtering is fundamental to infection metallomics; it has been used to study the interplay between "frenemies" in hosts and to monitor the dynamic response of the microbiome to antibiotic and antimycotic therapies. During infection in critically ill patients, the hostile environment of the host's body activates secondary bacterial, mycobacterial, and fungal metabolism, leading to the production of metallophores that increase the pathogen's chance of survival in the host. MS can reveal the structures, stability, and threshold concentrations of these metal-containing microbial biomarkers of infection in humans and model organisms, and can discriminate invasive disease from benign colonization based on well-defined thresholds distinguishing proliferation from the colonization steady state.

Carbon-rich substrates altered microbial communities with indication of carbon metabolism functional shifting in a degraded salt marsh of the Yellow River Delta, China

Coastal salt marshes have been significantly suffering degradation worldwide, turning these wetlands from carbon sink to carbon source. The additions of carbon-rich substrates in degraded coastal wetlands have been documented to alleviate this situation; however, the microbial responses as well as their carbon metabolisms to the input are not well explored. Here, we investigated the effects of different addition levels (0.1%, 1%, and 3%) of reed straw and biochar addition on microbial communities (bacteria and fungi) and predicted functions of carbon metabolism in degraded coastal salt marshes. The effects of the carbon-rich substrates on microbial communities are more pronounced under 3% of reed straw and biochar than other levels, straw increased opportunistic bacteria (e.g., Bacillus and Cellvibrionaceae) and biochar increased low carbon turnover bacteria (e.g., Acidobacteria and Actinobacteria). The addition of higher doses of reed straw and biochar promoted the carbon metabolism of soil bacteria as predicted by the PICRUSt 2. Interestingly, the addition of reed straw increased the abundance involved in fungal carbon metabolism, but this was not the case for the biochar input. The addition of reed straw and biochar promoted the growth of some bacteria (e.g., Rhizobiales and Bacillus) that are beneficial to plant growth, and decreased some fungal pathogen (e.g., Magnaporthaceae and Phaeosphaeria). Our findings revealed that the carbon-rich substrates addition can reshape the microbial structure and shift their carbon metabolisms, specially, biochar has a higher potential to improve the ecosystem services, such as blue carbon sequestration, of these degraded coastal salt marshes.

Energy Potential of Oil Palm Empty Fruit Bunch (EFB) Fiber from Subsequent Cultivation of Volvariella volvacea (Bull.) Singer

EFB and EFB-based mushroom compost (SMC) from Volvariella volvacea cultivation is a promising energy feedstock because it has adequate nutrient quality. The biochemical methane potential (BMP) and calorific value (CV) of this biomass are investigated. Other analyses such as proximate, compositional, and final analysis; thermogravimetric analysis (TGA); and Fourier transform infrared spectroscopy (FTIR) are also performed. The biomass samples consist of two types of EFB, namely fibers (F) and pellets (P) and SMC from the subsequent cultivation of Volvariella volvacea, with samples FS and PS from the first cultivation and FS2 and PS2 from the second cultivation. P produces the highest biological efficiency (BE) of 28% compared to 9.83% for F. Subsequent cultivation with FS and PS then produces only 2.9 and 6.83% of BE. A higher amount of methane is measured in samples P and PS2, while better biodegradability is observed in PS2 and FS2, suggesting that subsequent cultivation is a good pretreatment of the substrate for anaerobic digestion (AD). CV is highest in F (20.57 MJ/kg), followed by P (19.06 MJ/kg), which is comparable to commercial wood pellet. Samples F, FS, and FS2 have higher ash content, which is due to higher mineral content. The cellulose composition is reduced to almost 50% during cultivation due to fungal metabolism, which is also evidenced by FTIR analysis. TGA analysis revealed that EFB-based SMC exhibits higher weight loss during combustion compared to EFB, which reduces its thermal properties. SMC of EFB is a high potential biomethane feedstock, but not recommended as a fuel pellet.

Inhibitory Effects of Sulfonamide Derivatives on the β-Carbonic Anhydrase (MpaCA) from Malassezia pachydermatis, a Commensal, Pathogenic Fungus Present in Domestic Animals.

Fungi are exposed to various environmental variables during their life cycle, including changes in CO2 concentration. CO2 has the potential to act as an activator of several cell signaling pathways. In fungi, the sensing of CO2 triggers cell differentiation and the biosynthesis of proteins involved in the metabolism and pathogenicity of these microorganisms. The molecular machineries involved in CO2 sensing constitute a promising target for the development of antifungals. Carbonic anhydrases (CAs, EC 4.2.1.1) are crucial enzymes in the CO2 sensing systems of fungi, because they catalyze the reversible hydration of CO2 to proton and HCO3-. Bicarbonate in turn boots a cascade of reactions triggering fungal pathogenicity and metabolism. Accordingly, CAs affect microorganism proliferation and may represent a potential therapeutic target against fungal infection. Here, the inhibition of the unique β-CA (MpaCA) encoded in the genome of Malassezia pachydermatis, a fungus with substantial relevance in veterinary and medical sciences, was investigated using a series of conventional CA inhibitors (CAIs), namely aromatic and heterocyclic sulfonamides. This study aimed to describe novel candidates that can kill this harmful fungus by inhibiting their CA, and thus lead to effective anti-dandruff and anti-seborrheic dermatitis agents. In this context, current antifungal compounds, such as the azoles and their derivatives, have been demonstrated to induce the selection of resistant fungal strains and lose therapeutic efficacy, which might be restored by the concomitant use of alternative compounds, such as the fungal CA inhibitors.

MaNmrA, a Negative Transcription Regulator in Nitrogen Catabolite Repression Pathway, Contributes to Nutrient Utilization, Stress Resistance, and Virulence in Entomopathogenic Fungus Metarhizium acridum.

Simple SummaryNutrient metabolism is closely related to the growth, development, and pathogenicity of pathogenic fungi. The nitrogen catabolite repression (NCR) pathway is a well-known fungal nitrogen source regulation path, in which NmrA plays an important regulatory role. Here, we reported a negative regulatory protein MaNmrA, the NmrA homologous protein, in the entomopathogenic fungus Metarhizium acridum, and found that it played important roles in carbon and nitrogen metabolism, growth, stress tolerance, and virulence of M. acridum. Our work will provide a theoretical basis for further exploring the functions of NCR pathway related genes in entomopathogenic fungi.The NCR pathway plays an important regulatory role in the nitrogen metabolism of filamentous fungi. NmrA, a central negative regulatory protein in the NCR pathway and a key factor in sensing to the carbon metabolism, plays important roles in pathogenic fungal nutrition metabolism. In this study, we characterized the functions of MaNmrA in the insect pathogenic fungus M. acridum. Multiple sequence alignments found that the conserved domain (NAD/NADP binding domain) of MaNmrA was highly conservative with its homologues proteins. Deletion of MaNmrA improved the utilization of multiple carbon sources (such as glucose, mannose, sucrose, and trehalose) and non-preferred nitrogen sources (such as NaNO3 and urea), significantly delayed the conidial germination rate and reduced the conidial yield. The MaNmrA-disruption strain (ΔMaNmrA) significantly decreased tolerances to UV-B and heat-shock, and it also increased the sensitivity to the hypertonic substance sorbitol, oxygen stress substance H2O2, and cell wall destroyer calcofluor white, indicating that loss of MaNmrA affected cell wall integrity, tolerances to hypertonic and oxidative stress. Bioassays demonstrated that disruption of MaNmrA decreased the virulence in both topical inoculation and intrahemocoel injection tests. Further studies revealed that the appressorium formation, turgor pressure, and colonization in hemolymph were significantly reduced in the absence of MaNmrA. Our work will deepen the functional cognition of MaNmrA and make a contribution to the study of its homologous proteins.

Nitric Oxide in Fungal Metabolism (Review)

Nitric Oxide in Fungal Metabolism (Review)