Fungal Biology Research Articles

Use their names: there are no basal, lower, or early diverging fungi

ABSTRACT Fungal biologists have embraced phylogenies for understanding the biology of this diverse group in an evolutionary framework. In an attempt to highlight lineages of fungi that are distinct from the most speciose subphylum Dikarya (Ascomycota + Basidiomycota), the terms “early diverging fungi [lineages]” and “basal fungi” have been introduced, typically to refer to any phylum of fungi outside Dikarya. However, these terms are problematic, because they implicitly assume that the traits and taxa outside of Dikarya are ancestral by invoking a “ladder of progress.” A simple rearrangement of the tree to deemphasize the species-rich Dikarya shows that the logic that these taxa are “early branching” or “basal” is a fallacy, because it ignores two facts: (i) that all extant lineages of fungi have evolved an equivalent amount of time since a last common fungal ancestor, and (ii) that the “early diverging lineages” are no more related to each other than they are to Dikarya. To support the many mycologists who want to celebrate the understudied lineages outside of Dikarya while ensuring that these lineages are not mistakenly perceived as “less evolved,” “more ancient,” or of “lower complexity,” we propose that the community abandon these terms and simply use formal taxonomic names, e.g. Mucoromycota. Doing so will promote knowledge of these often overlooked branches of the tree of fungal life.

Genomic insights and comparative analysis of Colletotrichum species associated with anthracnose fruit rot and crown rot of strawberry in North Carolina.

Colletotrichum is a large genus of fungal phytopathogens responsible for significant economic losses in numerous crops globally. These pathogens exhibit varying host specificities; some have a broad host range, while others are more limited. To explore the genetic composition and underlying factors of fungal virulence and pathogenicity, we sequenced the genomes of seven isolates of Colletotrichum spp.: three from the C. acutatum and four from the C. gloeosporioides. These isolates were sourced from anthracnose fruit rot and crown rot of strawberry in North Carolina. Phylogenetic and phylogenomic analyses classified the isolates within the C. acutatum as C. nymphaeae, while those in the C. gloeosporioides were identified as C. siamense. The genome sizes of the C. nymphaeae isolates ranged from 50.3 Mb to 50.7 Mb, with 14,235 to 14,260 predicted protein-coding gene models. In contrast, the genome sizes of the C. siamense isolates ranged from 55.7 Mb to 58.6 Mb, with predicted protein-coding gene models ranging from 17,420 to 17,729. The GC content across all genomes spanned from 51.9 to 53.7%. The predicted gene models included effectors (339 to 480), secondary metabolic gene clusters (67 to 90), and carbohydrate-active enzymes (800 to 1,060), with C. siamense isolates exhibiting the highest numbers in these categories. The genomic resources from this study will aid in resolving taxonomic challenges associated with Colletotrichum spp., elucidate their evolutionary history, and enhance the understanding of fungal biology and ecology, which is crucial for developing effective disease management strategies.

A Plant Health View on Endohyphal Bacteria and Their Fungal Hosts

Microbe-microbe interactions within the phytobiome, including those between endosymbiotic bacteria and their fungal hosts, influence plant health outcomes. Endohyphal bacteria (EHB) can modulate the virulence and survival of phytopathogenic fungi, making them both potential targets for fungal control and potential tools for exploring fungal biology. Fungal hosts may increase the fitness of bacterial symbionts, some of which act as plant pathogens themselves. These relationships can also result in an increase of microbially produced phytohormones, influencing the plant host and other symbioses. However, the functional outcome of EHB metabolites and mechanisms of change remain largely uninvestigated in most of the EHB-fungal relationships identified. Additionally, morphological change experienced by some bacterial cells within fungi represents just one of the knowledge gaps in this field, with factors that determine bacterial and fungal partner specificity being another. Here, we compile studies of EHB from across diverse plant-associated microbes to provide a primer for EHB studies, considered through a phytobiomes lens, and comment on the directions for this emerging field.

Optimizing a modified cetyltrimethylammonium bromide protocol for fungal DNA extraction: Insights from multilocus gene amplification.

Genomic DNA (gDNA) extraction is an important step in many molecular studies of fungal biology, and it is necessary to evaluate the efficiency, cost-effectiveness, and efficacy of different extraction methods to ensure successful amplification of the target gene and minimize deoxyribonucleic acid (DNA) degradation. The modified cetyltrimethylammonium bromide (CTAB) method was found to be effective in releasing high molecular weight gDNA with minimal protein contamination. Based on anticipated gDNA yield and quality, extraction time, cost effectiveness, successful amplification, and waste management, our findings serve as a guide for selecting techniques of gDNA extraction from fungal species. This study presents a modified CTAB method for extracting DNA from a variety of fungal species including Aspergillus, Penicillium, Alternaria, Dothiorella, and Fusarium. Comparison of three cell crushing methods reveals similar gDNA yields, demonstrating the method's effectiveness. Furthermore, the modified CTAB method is cost-effective and safe, eliminating the need for grinding with liquid nitrogen or bead beating. The method has a potential use for nucleic-based fungal disease diagnosis such as fish fungal diseases, plant pathogens, fruit rot associated pathogens, and human fungal diseases as we were successful in PCR amplifying several gene loci from varied fungal pathogens.

Novel Strategies for Preventing Fungal Infections-Outline.

Fungal infections are a significant global health challenge, causing approximately 3.8 million deaths annually, with immunocompromised populations particularly at risk. Traditional antifungal therapies, including azoles, echinocandins, and polyenes, face limitations due to rising antifungal resistance, toxicity, and inadequate treatment options. This review explores innovative strategies for preventing and managing fungal infections, such as vaccines, antifungal peptides, nanotechnology, probiotics, and immunotherapy. Vaccines offer promising avenues for long-term protection, despite difficulties in their development due to fungal complexity and immune evasion mechanisms. Antifungal peptides provide a novel class of agents with broad-spectrum activity and reduced resistance risk, whilst nanotechnology enables targeted, effective drug delivery systems. Probiotics show potential in preventing fungal infections, particularly vulvovaginal candidiasis, by maintaining microbial balance. Immunotherapy leverages immune system modulation to enhance antifungal defenses, and omics technologies deliver comprehensive insights into fungal biology, paving the way for novel therapeutic and vaccine targets. While these approaches hold immense promise, challenges such as cost, accessibility, and translational barriers remain. A coordinated effort among researchers, clinicians, and policymakers is critical to advancing these strategies and addressing the global burden of fungal infections effectively.

Fungal Metabolomics: A Comprehensive Approach to Understanding Pathogenesis in Humans and Identifying Potential Therapeutics.

Metabolomics has emerged as a transformative tool in the study of microbes, including pathogenic fungi, facilitating the identification of unique metabolic profiles that elucidate their pathogenic mechanisms, host interactions, and treatment resistance. This review highlights key applications of metabolomics in understanding fungal metabolites essential for human virulence, such as mycotoxins produced by various fungal species, including Aspergillus fumigatus (gliotoxin, fumagillins) and Candida species (phenylethyl alcohol, TCA cycle metabolites), and secondary metabolites that contribute to pathogenicity. It also explores the metabolic adaptations of fungi in relation to drug resistance and biofilm formation, revealing alterations in key metabolic pathways during infection, as seen in C. albicans and C. auris. Furthermore, metabolomics aids in deciphering host-pathogen interactions, showcasing how fungi like Cryptococcus neoformans and Candida modify host metabolism to promote survival and evade immune responses. The study of antifungal resistance mechanisms has also benefited from metabolomic approaches, identifying specific metabolite patterns that signify resistance, such as in Candida albicans and Candidozyma (Candida) auris, and informing new therapeutic strategies. The integration of metabolomics with other omics technologies is paving the way for a comprehensive understanding of fungal biology and pathogenesis. Such multi-omics approaches are crucial for discovering new therapeutic targets and developing innovative antifungal treatments. Thus, the purpose of this review is to provide an overview of how metabolomics is revolutionizing our understanding of fungal pathogenesis, drug resistance, and host interactions, and to highlight its potential for identifying new therapeutic targets and improving antifungal strategies.

Research progress on the function and regulatory pathways of amino acid permeases in fungi.

Nitrogen sources are pivotal for the formation of fungal mycelia and the biosynthesis of metabolites, playing a crucial role in the growth and development of fungi. Amino acids are integral to protein construction, constitute an essential nitrogen source for fungi. Fungi actively uptake amino acids from their surroundings, a process that necessitates the involvement of amino acid permeases (AAPs) located on the plasma membrane. By sensing the intracellular demand for amino acids and their extracellular availability, fungi activate or suppress relevant pathways to precisely regulate the genes encoding these transporters. This review aims to illustrate the function of fungal AAPs on uptake of amino acids and the effect of AAPs on fungal growth, development and virulence. Additionally, the complex mechanisms to regulate expression of aaps are elucidated in mainly Saccharomyces cerevisiae, including the Ssy1-Ptr3-Ssy5 (SPS) pathway, the Nitrogen Catabolite Repression (NCR) pathway, and the General Amino Acid Control (GAAC) pathway. However, the physiological roles of AAPs and their regulatory mechanisms in other species, particularly pathogenic fungi, merit further exploration. Gaining insights into these aspects could reveal how AAPs facilitate fungal adaptation and survival under diverse stress conditions, shedding light on their potential impact on fungal biology and pathogenicity.

Unveiling mycoviral diversity in Ophiocordyceps sinensis through transcriptome analyses.

Ophiocordyceps sinensis, an entomopathogenic fungus, infects larvae from the Lepidoptera: Hepialidae family, forming the valuable Chinese cordyceps. Mycoviruses are widespread across major lineages of filamentous fungi, oomycetes, and yeasts and have the potential to influence fungal biology and ecology. This study aimed to detect mycovirus within O. sinensis by isolating double-stranded RNA from six stains for transcriptomic sequencing and analyzing publicly available transcriptome data from 13 O. sinensis representative samples. Our analysis revealed 13 mycoviruses, with nine reported for the first time in O. sinensis. These mycoviruses are distributed across five families-Partitiviridae, Mitoviridae, Narnaviridae, Botourmiaviridae, Deltaflexiviridae-and two unclassified lineages, Ormycovirus and Vivivirus. This study also revealed frequent coinfections within individual O. sinensis strains and dynamic shifts in viral composition during fungal development. These findings enhance our knowledge of mycovirus diversity within O. sinensis and provide new insights into their taxonomy.

Dietary Mycotoxins: An Overview on Toxicokinetics, Toxicodynamics, Toxicity, Epidemiology, Detection, and Their Mitigation with Special Emphasis on Aflatoxicosis in Humans and Animals.

Mycotoxins are secondary metabolites of filamentous fungi and ubiquitous dietary contaminants. Aflatoxins, a group of mycotoxins with high prevalence and toxicity, have raised a high level of public health concern, the most prevalent and toxic being aflatoxin B1 (AFB1). Many aspects appertaining to AFB1 poisoning are not well understood. Yet this information is necessary to devise appropriate surveillance and mitigation strategies against human and animal aflatoxicosis. This review provides an in-depth update of work carried out on mycotoxin poisoning, particularly aflatoxicosis in humans and animals, to identify gaps in knowledge. Hypotheses explaining the functional significance of mycotoxins in fungal biology and their dietary epidemiological data are presented and briefly discussed. The toxicology of aflatoxins and the challenges of their mitigation are discussed in depth. It was concluded that the identification of potential mycotoxin-hazard-prone food items and quantification of the associated risk of cancer ailments in humans is a prime priority. There is a dearth of reliable sampling methodologies for estimating AFB1 in animal feed. Data update on AFB1 in animal feed and its implication in animal production, mitigation strategies, and elucidation of risk factors to this hazard is required. To reduce the burden of aflatoxins, surveillance employing predictive technology, and biocontrol strategies seem promising approaches.

A new leaf inhabiting ascomycete from the Jurassic (ca 170 Mya) of Yorkshire, UK, and insights into the appearance and diversification of filamentous Ascomycota

Leaf-associated fungi, the fungi that depend on leaves to sporulate, have a rich Cenozoic record, however their earlier diversity is poorly characterized. Here we describe Harristroma eboracense gen. et sp. nov., a Middle Jurassic leaf-associated fungus colonizing the leaf cuticle of Nilssonia tenuicaulis (cycadophyte). To place our newly described species into a picture of the diversification of Mesozoic fungi, we reassess fossils with leaf-associated stromata in the context of fungal molecular phylogeny. Being melanized, with radiate stromata, and on leaves, H. eboracense and other fossils from the Jurassic and earlier periods are probably related to filamentous Ascomycota in the superclass Leotiomyceta. Characters needed for further resolution of leaf-associated fungal biology and classification, such as the presence of an ostiole for spore discharge and appressoria for entry into leaf tissue first appear in the Mesozoic fossil record. Among Early Cretaceous fossils, Spataporthe taylorii represents the oldest unambiguous evidence of perithecial Sordariomycetes while Protographum luttrellii and Bleximothyrium ostiolatum are the oldest Dothideomycetes thyriothecia. Environmental observations show that broad leaved gymnosperms (especially cycadophytes) growing in warm temperate wet forests might have been the first environment for the radiation of Leotiomyceta.

Diversity of Pleosporalean Fungi Isolated from Rice (Oryza sativa L.) in Northern Thailand and Descriptions of Five New Species.

Pleosporales represents the largest order within the class Dothideomycetes (Fungi), comprising phytopathogenic, saprobic, and endophytic taxa with a widespread presence in terrestrial and aquatic environments. Rice (Oryza sativa) is a primary economic crop in numerous tropical countries, particularly in Thailand. Studying fungal species associated with rice holds the potential to enhance our understanding of fungal diversity, lifestyles, and biology of rice, offering valuable insights for future research aimed at disease management and yield improvement. Thirty-nine pleosporalean isolates were obtained from various parts of rice plants collected across diverse regions in Chiang Rai Province, Thailand. Species identification involved a combination of morphology and molecular phylogeny, utilizing multi-locus sequence analyses of the ITS, LSU, SSU, gapdh, rpb2, tef1, and tub2 genes. The isolates were identified in 18 taxa distributed across five families and ten genera, including five new species (Bipolaris chiangraiensis, Ophiosphaerella oryzae, Paraphaeosphaeria oryzae, Pyrenochaetopsis oryzicola, and Setophoma oryzicola). Additionally, six new host records and two new geographical records are documented. Photoplates, detailed morphological descriptions, and phylogenetic trees are provided to elucidate the placement of both known and novel taxa.

Quantitative Characterization of Gene Regulatory Circuits Associated With Fungal Secondary Metabolism to Discover Novel Natural Products.

Microbial genetic circuits are vital for regulating gene expression and synthesizing bioactive compounds. However, assessing their strength and timing, especially in multicellular fungi, remains challenging. Here, an advanced microfluidic platform is combined with a mathematical model enabling precise characterization of fungal gene regulatory circuits (GRCs) at the single-cell level. Utilizing this platform, the expression intensity and timing of 30 transcription factor-promoter combinations derived from two representative fungal GRCs, using the model fungus Aspergillus nidulans are determined. As a proof of concept, the selected GRC combination is utilized to successfully refactor the biosynthetic pathways of bioactive molecules, precisely control their production, and activate the expression of the silenced biosynthetic gene clusters (BGCs). This study provides insights into microbial gene regulation and highlights the potential of platform in fungal synthetic biology applications and the discovery of novel natural products.

Genome-Wide Identification and Expression Analysis of the Cys2His2 Zinc Finger Protein Gene Family in Flammulina filiformis.

Zinc finger proteins (ZFPs) are essential transcription factors in eukaryotes, particularly the extensively studied C2H2 family, which is known for its involvement in various biological processes. This research provides a thorough examination and analysis of the C2H2-ZFP gene family in Flammulina filiformis. Using bioinformatics tools, 58 FfC2H2-ZFP genes spread across 11 chromosomes were identified and scrutinized in detail for their gene structures, protein characteristics, and phylogenetic relationships. The study of phylogenetics and synteny sheds light on the evolutionary relationships among C2H2-ZFPs in F. filiformis and other fungi, revealing a complex evolutionary past. The identification of conserved cis-regulatory elements in the gene promoter regions suggests intricate functionalities, particularly in the developmental and stress response pathways. By utilizing RNA-seq and qRT-PCR techniques, the expression patterns of these genes were explored across different developmental stages and tissues of F. filiformis, unveiling distinct expression profiles. Notably, significant expression variations were observed in the stipe elongation region and pilei of various sizes, indicating potential roles in fruiting body morphogenesis. This study enhances our knowledge of the C2H2-ZFP gene family in F. filiformis and lays the groundwork for future investigations into their regulatory mechanisms and applications in fungal biology and biotechnology.

Emerging Roles of Exocyst Complex in Fungi: A Review.

The exocyst complex, an evolutionarily conserved octameric protein assembly, plays a central role in the targeted binding and fusion of vesicles at the plasma membrane. In fungal cells, this transport system is essential for polarized growth, morphogenesis, cell wall maintenance and virulence. Recent advances have greatly improved our understanding of the role and regulation of the exocyst complex in fungi. This review synthesizes these developments and focuses on the intricate interplay between the exocyst complex, specific fungal cargos and regulatory proteins. Insights into thestructure of the exocyst and its functional dynamics have revealed new dimensions of its architecture and its interactions with the cellular environment. Furthermore, the regulation of exocyst activity involves complex signaling pathways and interactions with cytoskeletal elements that are crucial for its role in vesicle trafficking. By exploring these emerging themes, this review provides a comprehensive overview of the multifaceted functions of the exocyst complex in fungal biology. Understanding these mechanisms offers potential avenues for novel therapeutic strategies against fungal pathogens and insights into the general principles of vesicle trafficking in eukaryotic cells. The review therefore highlights the importance of the exocyst complex in maintaining cellular functions and its broader implications in fungal pathogenicity and cell biology.

A deep learning-guided automated workflow in LipidOz for detailed characterization of fungal fatty acid unsaturation by ozonolysis.

Understanding fungal lipid biology and metabolism is critical for antifungal target discovery as lipids play central roles in cellular processes. Nuances in lipid structural differences can significantly impact their functions, making it necessary to characterize lipids in detail to understand their roles in these complex systems. In particular, lipid double bond (DB) locations are an important component of lipid structure that can only be determined using a few specialized analytical techniques. Ozone-induced dissociation mass spectrometry (OzID-MS) is one such technique that uses ozone to break lipid DBs, producing pairs of characteristic fragments that allow the determination of DB positions. In this work, we apply OzID-MS and LipidOz software to analyze the complex lipids of Saccharomyces cerevisiae yeast strains transformed with different fatty acid desaturases from Histoplasma capsulatum to determine the specific unsaturated lipids produced. The automated data analysis in LipidOz made the determination of DB positions from this large dataset more practical, but manual verification for all targets was still time-consuming. The DL model reduces manual involvement in data analysis, but since it was trained using mammalian lipid extracts, the prediction accuracy on yeast-derived data was reduced. We addressed both shortcomings by retraining the DL model to act as a pre-filter to prioritize targets for automated analysis, providing confident manually verified results but requiring less computational time and manual effort. Our workflow resulted in the determination of detailed DB positions and enzymatic specificity.

Broad Protection against Invasive Fungal Disease from a Nanobody Targeting the Active Site of Fungal β‐1,3‐Glucanosyltransferases

AbstractInvasive fungal disease accounts for about 3.8 million deaths annually, an unacceptable rate that urgently prompts the discovery of new knowledge‐driven treatments. We report the use of camelid single‐domain nanobodies (Nbs) against fungal β‐1,3‐glucanosyltransferases (Gel) involved in β‐1,3‐glucan transglycosylation. Crystal structures of two Nbs with Gel4 from Aspergillus fumigatus revealed binding to a dissimilar CBM43 domain and a highly conserved catalytic domain across fungal species, respectively. Anti‐Gel4 active site Nb3 showed significant antifungal efficacy in vitro and in vivo prophylactically and therapeutically against different A. fumigatus and Cryptococcus neoformans isolates, reducing the fungal burden and disease severity, thus significantly improving immunocompromised animal survival. Notably, C. deneoformans (serotype D) strains were more susceptible to Nb3 and genetic Gel deletion than C. neoformans (serotype A) strains, indicating a key role for β‐1,3‐glucan remodelling in C. deneoformans survival. These findings add new insight about the role of β‐1,3‐glucan in fungal biology and demonstrate the potential of nanobodies in targeting fungal enzymes to combat invasive fungal diseases.

Broad Protection against Invasive Fungal Disease from a Nanobody Targeting the Active Site of Fungal β-1,3-Glucanosyltransferases.

Invasive fungal disease accounts for about 3.8 million deaths annually, an unacceptable rate that urgently prompts the discovery of new knowledge-driven treatments. We report the use of camelid single-domain nanobodies (Nbs) against fungal β-1,3-glucanosyltransferases (Gel) involved in β-1,3-glucan transglycosylation. Crystal structures of two Nbs with Gel4 from Aspergillus fumigatus revealed binding to a dissimilar CBM43 domain and a highly conserved catalytic domain across fungal species, respectively. Anti-Gel4 active site Nb3 showed significant antifungal efficacy in vitro and in vivo prophylactically and therapeutically against different A. fumigatus and Cryptococcus neoformans isolates, reducing the fungal burden and disease severity, thus significantly improving immunocompromised animal survival. Notably, C. deneoformans (serotype D) strains were more susceptible to Nb3 and genetic Gel deletion than C. neoformans (serotype A) strains, indicating a key role for β-1,3-glucan remodelling in C. deneoformans survival. These findings add new insight about the role of β-1,3-glucan in fungal biology and demonstrate the potential of nanobodies in targeting fungal enzymes to combat invasive fungal diseases.

Harnessing Fungi Signaling in Living Composites.

Signaling pathways in fungi offer a profound avenue for harnessing cellular communication and have garnered considerable interest in biomaterial engineering. Fungi respond to environmental stimuli through intricate signaling networks involving biochemical and electrical pathways, yet deciphering these mechanisms remains a challenge. In this review, an overview of fungal biology and their signaling pathways is provided, which can be activated in response to external stimuli and direct fungal growth and orientation. By examining the hyphal structure and the pathways involved in fungal signaling, the current state of recording fungal electrophysiological signals as well as the landscape of fungal biomaterials is explored. Innovative applications are highlighted, from sustainable materials to biomonitoring systems, and an outlook on the future of harnessing fungi signaling in living composites is provided.

Diagnosis and Molecular Characterization of Dermatophytosis: An Observational Study

Dermatophytes are the keratinophilic fungi which infect humans and is the most recurring type of disease. The high level of transmissibility creates an epidemiological risk and emphasises the significance of these illnesses. However, a growing number of reports describing dermatophytes can cause deep infections in diabetic and immunocompromised patients, by invading deep layers like the dermis and hypodermis. Despite the prevalence and significance of dermatophytes in clinical mycology, it is not always possible to accurately diagnose this specific infection due to its overlapping structures among species of dermatophytes. Since it is difficult to identify species that exhibit weak characteristics in the morphological highlights, identification of the dermatophyte is often relied on its morphological analysis, which is a laborious process and demands skill. The massive shift in genetic variation, the source of infection, and epidemiological research can be discovered using molecular approaches. Therefore, the development of an accurate laboratory test for dermatophyte species identification is essential for the prevention and efficient management of dermatophytoses. One such methodology allows use of PCR technology which has many methods for molecular level characterization which is rapid, efficient, and capable of producing DNA polymorphisms specific to various dermatophyte species based on distinctive band patterns seen by agarose gel electrophoresis. The RAPD-PCR approach will be used in this study protocol to molecularly characterize the dermatophytes for precise speciation of the sample. In addition to improving knowledge of fungal biology and pathology with a focus on adaptive mechanisms to combat difficult conditions from host counteractions, there is a need to improve awareness of the importance of these diseases through accurate epidemiological data. The advantages of molecular approaches for characterizing objects over traditional methods are their sensitivity and specificity.

Review of the Clinical Types of Dermatophytes

Fungi known as dermatophytes require keratin in order to develop. "Ringworm" is the common term for superficial fungus infections that affect the skin's keratinized layers and its limbs in both humans and animals. These fungi have the potential to superficially infect nails, hair, and skin. Ringworm fungi have the ability to penetrate all layers of skin. Dermatophytes can be transmitted directly by people (anthropophilic organisms), indirectly by fomites, and by animals (zoophilic organisms), soil (geophilic organisms), and other people. Dermatophytes are the cause of the medical condition known as dermatophytosis. Approximately 25% of the global population suffers from dermatophytosis, and no human population is immune to these mycotic disorders. There are three genera in total. the more than twenty species of dermatophytes: Trichophyton, Microsporum, and Epidermophyton. By providing reliable epidemiological data, it is imperative that we expand broaden public knowledge of the importance of these illnesses and deepen our understanding of fungal biology and pathology. Ecological classifications, clinical potential for pathogenicity manifestations, and "chemical and natural" dermatophytosis therapy alternatives are some of the review themes.