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Fungi are diverse organisms with various characteristics and functions. Some play a role in recycling essential elements, such as nitrogen and carbon, while others are utilized in the food and drink production industry. Some others are known to cause diseases in various organisms, including humans. Fungal pathogens cause superficial, subcutaneous, and systemic infections. Consequently, many scientists have focused on studying the factors contributing to the development of human diseases. Therefore, multiple approaches have been assessed to examine the biology of these intriguing organisms. The genome-scale metabolic models (GEMs) have demonstrated many advantages to microbial metabolism studies and the ability to propose novel therapeutic alternatives. Despite significant advancements, much remains to be elucidated regarding the use of this tool for investigating fungal metabolism. This review aims to compile the data provided by the published GEMs of human fungal pathogens. It gives specific examples of the most significant contributions made by these models, examines the advantages and difficulties associated with using such models, and explores the novel approaches suggested to enhance and refine their development.

Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The author(s) declared that they were an editorial board member of Frontiers, at the time of submission. This had no impact on the peer review process and the final decision.

Fusarium and Neocosmospora are two fungal genera recently recognized in the list of fungal priority pathogens. They cause a wide range of diseases that affect humans, animals, and plants. In clinical laboratories, there is increasing concern about diagnosis due to limitations in sample collection and morphological identification. Despite the advances in molecular diagnosis, due to the cost, some countries cannot implement these methodologies. However, recent changes in taxonomy and intrinsic resistance to antifungals reveal the necessity of accurate species-level identification. In this review, we discuss the current phenotypic and molecular tools available for diagnosis in clinical laboratory settings and their advantages and disadvantages.

Introduction. Malassezia globosa is a yeast species that belongs to the mycobiota of humans and animals, associated with dermatological disorders, such as dandruff. This is a chronic scalp skin disorder characterized by flaking and itching. Treatments include commercial shampoo with different formulations that contain antifungal activities like zinc pyrithione (ZPT) or piroctone olamine (PO). The effectiveness of these formulations has been evaluated for decades for dandruff symptom relief of volunteers. To date, non-mammalian, in vivo methods exist to test formulations of these actives. Aim. To evaluate in vivo in Galleria mellonella larva, two commercial antifungal shampoos (shampoo with 1 % ZPT and 1.6 % zinc Carbonate and shampoo with 0.5 % PO) against this species. Methodology. G. mellonella larvae were inoculated with M. globosa on abraded cuticular surface. Then, integument cell viability, histological changes, and fungal burden were evaluated. Results. Larvae inoculated with M. globosa showed higher lesion melanization and tissue damage. In addition, M. globosa population showed to increase over time. Concerning the shampoo's effectiveness, both formulations significantly reduced M. globosa burden and tissue damage. Conclusion. G. mellonella larvae were allowed to evaluate M. globosa superficial infection and antifungal effectiveness. Shampoos with ZPT and PO showed a positive effect on inoculated larvae., Competing Interests: The CeMoP research group has received funding from Procter and Gamble to finance this study. M.G.D. and J.R.S. are employees of Procter and Gamble., (Copyright © 2024 The Authors.)

The Terminal Fusarium Clade (TFC) is a group in the Nectriaceae family with agricultural and clinical relevance. In recent years, various phylogenies have been presented in the literature, showing disagreement in the topologies, but only a few studies have conducted analyses on the divergence time scale of the group. Therefore, the evolutionary history of this group is still being determined. This study aimed to understand the evolutionary history of the TFC from a phylogenomic perspective. To achieve this objective, we performed a phylogenomic analysis using the available genomes in GenBank and ran eight different pipelines. We presented a new robust topology of the TFC that differs at some nodes from previous studies. These new relationships allowed us to formulate new hypotheses about the evolutionary history of the TFC. We also inferred new divergence time estimates, which differ from those of previous studies due to topology discordances and taxon sampling. The results suggested an important diversification process in the Neogene period, likely associated with the diversification and predominance of terrestrial ecosystems by angiosperms. In conclusion, we presented a robust time-scale phylogeny that allowed us to formulate new hypotheses regarding the evolutionary history of the TFC., (© 2024. The Author(s).)

Introduction: Malassezia is a lipophilic and lipid-dependent yeast genus belonging to the skin microbiota of humans and other animals. However, due to dysbiosis processes or other factors in the host, this yeast can cause different pathologies, ranging from skin diseases, such as seborrheic dermatitis, to fungemia. Isolation of Malassezia furfur has been reported in HIV-positive patients with or without skin lesions. Due to its opportunistic nature and its variable resistance to antifungal compounds, it is relevant to know the Malassezia sensitivity profiles., Objective: To determine the sensitivity to different antifungal agents, of clinical isolates of M. furfur obtained from HIV-positive or negative patients, with or without seborrheic dermatitis., Materials and Methods: Assessment of isolates sensitivity to itraconazole, voriconazole, fluconazole, and amphotericin B was performed by two techniques: (1) Broth microdilution using Clinical and Laboratory Standards Institute (CLSI) protocol M27-A3 with modifications; and (2) agar tests using Etest®., Results: Isolates obtained from HIV patients showed an increase in the minimum inhibitory concentration of fluconazole, voriconazole, and amphotericin B, compared with those of non-HIV patients. Itraconazole was the antifungal with the lowest minimum inhibitory concentration (MIC) in most isolates., Conclusion: We observed differences in the sensitivity profiles of M. furfur isolates according to the context of the patient. High MIC of antifungals like fluconazole, commonly used for treating pathologies caused by Malassezia, were identified.

Malassezia synthesizes and releases volatile organic compounds (VOCs), small molecules that allow them to carry out interaction processes. These lipid-dependent yeasts belong to the human skin mycobiota and are related to dermatological diseases. However, knowledge about VOC production and its function is lacking. This study aimed to determine the volatile profiles of Malassezia globosa , Malassezia restricta , and Malassezia sympodialis in the exponential and stationary growth phases. The compounds were separated and characterized in each growth phase through headspace solid-phase microextraction (HS-SPME) and gas chromatography-mass spectrometry (GC-MS). We found a total of 54 compounds, 40 annotated. Most of the compounds identified belong to alcohols and polyols, fatty alcohols, alkanes, and unsaturated aliphatic hydrocarbons. Unsupervised and supervised statistical multivariate analyses demonstrated that the volatile profiles of Malassezia differed between species and growth phases, with M. globosa being the species with the highest quantity of VOCs. Some Malassezia volatiles, such as butan-1-ol, 2-methylbutan-1-ol, 3-methylbutan-1-ol, and 2-methylpropan-1-ol, associated with biological interactions were also detected. All three species show at least one unique compound, suggesting a unique metabolism. The ecological functions of the compounds detected in each species and growth phase remain to be studied. They could interact with other microorganisms or be an important clue in understanding the pathogenic role of these yeasts.

Malassezia, a lipophilic and lipid-dependent yeast, is a microorganism of current interest to mycobiologists because of its role as a commensal or pathogen in health conditions such as dermatological diseases, fungemia, and, as discovered recently, cancer and certain neurological disorders. Various novel approaches in the study of Malassezia have led to increased knowledge of the cellular and molecular mechanisms of this yeast. However, additional efforts are needed for more comprehensive understanding of the behavior of Malassezia in interactions with the host. This article reviews advances useful in the experimental field for Malassezia., Competing Interests: The authors have declared that no competing interests exist.

Malassezia spp. are lipid-dependent yeasts that have been related to skin mycobiota and dermatological and systemic diseases. Study of lipid droplets (LDs) is relevant to elucidate the unknown role of these organelles in Malassezia and to gain a broader overview of lipid metabolism in Malassezia. Here, we standardized two protocols for the analysis of LDs in M. pachydermatis and M. globosa. The first describes co-staining for confocal laser-scanning fluorescence microscopy, and the second details extraction and purification of LDs. The double stain is achieved with three different neutral lipid fluorophores, namely Nile Red, BODIPY™ 493/503, and HCS LipidTOX™ Deep Red Neutral, in combination with Calcofluor White. For LD extraction, cell wall rupture is conducted using Trichoderma harzianum enzymes and cycles of vortexing with zirconium beads. LD purification is performed in a three-step ultracentrifugation process. These standardizations will contribute to the study of the dynamics, morphology, and composition of LDs in Malassezia. © 2021 Wiley Periodicals LLC. Basic Protocol 1: Lipid droplet fluorescence staining Basic Protocol 2: Lipid droplet extraction and purification Support Protocol: Malassezia spp. culture conditions., (© 2021 Wiley Periodicals LLC.)

Background: Microorganisms synthesize and release a large diversity of small molecules like volatile compounds, which allow them to relate and interact with their environment. Volatile organic compounds (VOCs) are carbon-based compounds with low molecular weight and generally, high vapor pressure; because of their nature, they spread easily in the environment. Little is known about the role of VOCs in the interaction processes, and less is known about VOCs produced by Malassezia, a genus of yeasts that belongs to the human skin mycobiota. These yeasts have been associated with several dermatological diseases and currently, they are considered as emerging opportunistic yeasts. Research about secondary metabolites of these yeasts is limited. The pathogenic role and the molecular mechanisms involved in the infection processes of this genus are yet to be clarified. VOCs produced by Malassezia yeasts could play an important function in their metabolism; in addition, they might be involved in either beneficial or pathogenic host-interaction processes. Since these yeasts present differences in their nutritional requirements, like lipids to grow, it is possible that these variations of growth requirements also define differences in the volatile organic compounds produced in Malassezia species., Aim of Review: We present a mini review about VOCs produced by microorganisms and Malassezia species, and hypothesize about their role in its metabolism, which would reveal clues about host-pathogen interaction., Key Scientific Concepts of Review: Since living organisms inhabit a similar environment, the interaction processes occur naturally; as a result, a signal and a response from participants of these processes become important in understanding several biological behaviors. The efforts to elucidate how living organisms interact has been studied from several perspectives. An important issue is that VOCs released by the microbiota plays a key role in the setup of relationships between living micro and macro organisms. The challenge is to determine what is the role of these VOCs produced by human microbiota in commensal/pathogenic scenarios, and how these allow understanding the species metabolism. Malassezia is part of the human mycobiota, and it is implicated in commensal and pathogenic processes. It is possible that their VOCs are involved in these behavioral changes, but the knowledge about this remains overlocked. For this reason, VOCs produced by microorganisms and Malassezia spp. and their role in several biological processes are the main topic in this review.

Some species of fusaria are well-known pathogens of humans, animals and plants. Fusarium oxysporum and Neocosmospora solani (formerly Fusarium solani ) cause human infections that range from onychomycosis or keratitis to severe disseminated infections. In general, these infections are difficult to treat due to poor therapeutic responses in immunocompromised patients. Despite that, little is known about the molecular mechanisms and transcriptional changes responsible for the antifungal resistance in fusaria. To shed light on the transcriptional response to antifungals, we carried out the first reported high-throughput RNA-seq analysis for F. oxysporum and N. solani that had been exposed to amphotericin B (AMB) and posaconazole (PSC). We detected significant differences between the transcriptional profiles of the two species and we found that some oxidation-reduction, metabolic, cellular and transport processes were regulated differentially by both fungi. The same was found with several genes from the ergosterol synthesis, efflux pumps, oxidative stress response and membrane biosynthesis pathways. A significant up-regulation of the C-22 sterol desaturase ( ERG5 ), the sterol 24-C-methyltransferase ( ERG6 ) gene, the glutathione S-transferase ( GST ) gene and of several members of the major facilitator superfamily ( MSF ) was demonstrated in this study after treating F. oxysporum with AMB. These results offer a good overview of transcriptional changes after exposure to commonly used antifungals, highlights the genes that are related to resistance mechanisms of these fungi, which will be a valuable tool for identifying causes of failure of treatments.

Malassezia is a lipid-dependent genus of yeasts known for being an important part of the skin mycobiota. These yeasts have been associated with the development of skin disorders and cataloged as a causal agent of systemic infections under specific conditions, making them opportunistic pathogens. Little is known about the host-microbe interactions of Malassezia spp., and unraveling this implies the implementation of infection models. In this mini review, we present different models that have been implemented in fungal infections studies with greater attention to Malassezia spp. infections. These models range from in vitro (cell cultures and ex vivo tissue), to in vivo (murine models, rabbits, guinea pigs, insects, nematodes, and amoebas). We additionally highlight the alternative models that reduce the use of mammals as model organisms, which have been gaining importance in the study of fungal host-microbe interactions. This is due to the fact that these systems have been shown to have reliable results, which correlate with those obtained from mammalian models. Examples of alternative models are Caenorhabditis elegans , Drosophila melanogaster , Tenebrio molitor , and Galleria mellonella . These are invertebrates that have been implemented in the study of Malassezia spp. infections in order to identify differences in virulence between Malassezia species.

Malassezia yeasts are lipid dependent and part of the human and animal skin microbiome. However, they are also associated with a variety of dermatological conditions and even cause systemic infections. How these yeasts can live as commensals on the skin and switch to a pathogenic stage has long been a matter of debate. Lipids are important cellular molecules, and understanding the lipid metabolism and composition of Malassezia species is crucial to comprehending their biology and host-microbe interaction. Here, we investigated the lipid composition of Malassezia strains grown to the stationary phase in a complex Dixon medium broth. In this study, we perform a lipidomic analysis of a subset of species; in addition, we conducted a gene prediction analysis for the detection of lipid metabolic proteins. We identified 18 lipid classes and 428 lipidic compounds. The most commonly found lipids were triglycerides (TAG), sterol (CH), diglycerides (DG), fatty acids (FAs), phosphatidylcholine (PC), phosphatidylethanolamine (PE), ceramides, cholesteryl ester (CE), sphingomyelin (SM), acylcarnitine, and lysophospholipids. Particularly, we found a low content of CEs in Malassezia furfur , atypical M. furfur , and Malassezia pachydermatis and undetectable traces of these components in Malassezia globosa, Malassezia restricta , and Malassezia sympodialis . Remarkably, uncommon lipids in yeast, like diacylglyceryltrimethylhomoserine and FA esters of hydroxyl FAs, were found in a variable concentration in these Malassezia species. The latter are bioactive lipids recently reported to have antidiabetic and anti-inflammatory properties. The results obtained can be used to discriminate different Malassezia species and offer a new overview of the lipid composition of these yeasts. We could confirm the presence and the absence of certain lipid-biosynthesis genes in specific species. Further analyses are necessary to continue disclosing the complex lipidome of Malassezia species and the impact of the lipid metabolism in connection with the host interaction., (Copyright © 2020 Celis Ramírez, Amézquita, Cardona Jaramillo, Matiz-Cerón, Andrade-Martínez, Triana, Mantilla, Restrepo, Barrios and Cock.)

Malassezia furfur and Malassezia pachydermatis are lipophilic and lipid dependent yeasts, associated with the skin microbiota in humans and domestic animals, respectively. Although they are commensals, under specific conditions they become pathogens, causing skin conditions, such as pityriasis versicolor, dandruff/seborrheic dermatitis, folliculitis in humans, and dermatitis and otitis in dogs. Additionally, these species are associated with fungemia in immunocompromised patients and low-weight neonates in intensive care units with intravenous catheters or with parenteral nutrition and that are under-treatment of broad-spectrum antibiotics. The host-pathogen interaction mechanism in these yeasts is still unclear; for this reason, it is necessary to implement suitable new host systems, such as Galleria mellonella . This infection model has been widely used to assess virulence, host-pathogen interaction, and antimicrobial activity in bacteria and fungi. Some advantages of the G. mellonella model are: (1) the immune response has phagocytic cells and antimicrobial peptides that are similar to those in the innate immune response of human beings; (2) no ethical implications; (3) low cost; and (4) easy to handle and inoculate. This study aims to establish G. mellonella as an in vivo infection model for M. furfur and M. pachydermatis . To achieve this objective, first, G. mellonella larvae were first inoculated with different inoculum concentrations of these two Malassezia species, 1.5 × 10 6 CFU/mL, 1.5 × 10 7 CFU/mL, 1.5 × 10 8 CFU/mL, and 11.5 × 10 9 CFU/mL, and incubated at 33 and 37°C. Then, for 15 days, the mortality and melanization were evaluated daily. Finally, the characterization of hemocytes and fungal burden assessment were as carried out. It was found that at 33 and 37°C both M. furfur and M. pachydermatis successfully established a systemic infection in G. mellonella . M. pachydermatis proved to be slightly more virulent than M. furfur at a temperature of 37°C. The results suggest that larvae mortality and melanization is dependent on the specie of Malassezia , the inoculum concentration and the temperature. According to the findings, G. mellonella can be used as an in vivo model of infection to conduct easy and reliable approaches to boost our knowledge of the Malassezia genus., (Copyright © 2020 Torres, Pinzón, Rey, Martinez, Parra Giraldo and Celis Ramírez.)

The opportunistic pathogen Malassezia pachydermatis causes bloodstream infections in preterm infants or individuals with immunodeficiency disorders and has been associated with a broad spectrum of diseases in animals such as seborrheic dermatitis, external otitis and fungemia. The current approaches to treat these infections are failing as a consequence of their adverse effects, changes in susceptibility and antifungal resistance. Thus, the identification of novel therapeutic targets against M. pachydermatis infections are highly relevant. Here, Gene Essentiality Analysis and Flux Variability Analysis was applied to a previously reported M. pachydermatis metabolic network to identify enzymes that, when absent, negatively affect biomass production. Three novel therapeutic targets (i.e., homoserine dehydrogenase (MpHSD), homocitrate synthase (MpHCS) and saccharopine dehydrogenase (MpSDH)) were identified that are absent in humans. Notably, L-lysine was shown to be an inhibitor of the enzymatic activity of MpHCS and MpSDH at concentrations of 1 mM and 75 mM, respectively, while L-threonine (1 mM) inhibited MpHSD. Interestingly, L- lysine was also shown to inhibit M. pachydermatis growth during in vitro assays with reference strains and canine isolates, while it had a negligible cytotoxic activity on HEKa cells. Together, our findings form the bases for the development of novel treatments against M. pachydermatis infections.