J. Fungi, Volume 9, Issue 8 (August 2023) – 83 articles

The endophytic fungal community of the Amazonian medicinal plant Arrabidaea chica (Bignoniaceae) was evaluated based on the hypothesis that microbial communities associated with plant species in the Amazon region may produce metabolites with interesting bioactive properties. Therefore, the antimicrobial and antioxidant activities of the fungal extracts were investigated. A total of 107 endophytic fungi were grown in liquid medium and the metabolites were extracted with ethyl acetate. In the screening of fungal extracts for antimicrobial activity, the fungus identified as Botryosphaeria mamane CF2-13 was the most promising, with activity against E. coli, S. epidermidis, P. mirabilis, B. subtilis, S. marcescens, K. pneumoniae, S. enterica, A. brasiliensis, C. albicans, C. tropicalis and, especially, against S. aureus and C. parapsilosis (MIC = 0.312 mg/mL). Screening for antioxidant potential using the DPPH elimination assay showed that the Colletotrichum sp. CG1-7 endophyte extract exhibited potential activity with an EC₅₀ of 11 µg/mL, which is equivalent to quercetin (8 µg/mL). The FRAP method confirmed the antioxidant potential of the fungal extracts. The presence of phenolic compounds and flavonoids in the active extracts was confirmed using TLC. These results indicate that two of the fungi isolated from A. chica exhibit significant antimicrobial and antioxidant potential. Full article

The litchi downy blight disease of litchi caused by Peronophythora litchii accounts for severe losses in the field and during storage. While ample quantitative studies have shown that 6-pentyl-2H-pyran-2-one (6PP) possesses antifungal activities against multiple plant pathogenic fungi, the regulatory mechanisms of 6PP-mediated inhibition of fungal pathogenesis and growth are still unknown. Here, we investigated the potential molecular targets of 6PP in the phytopathogenic oomycetes P. litchii through integrated deployment of RNA-sequencing, functional genetics, and biochemical techniques to investigate the regulatory effects of 6PP against P. litchii. Previously we demonstrated that 6PP exerted significant oomyticidal activities. Also, comparative transcriptomic evaluation of P. litchii strains treated with 6PP Revealed significant up-regulations in the expression profile of TOR pathway-related genes, including PlCytochrome C and the transcription factors PlYY1. We also noticed that 6PP treatment down-regulated putative negative regulatory genes of the TOR pathway, including PlSpm1 and PlrhoH12 in P. litchii. Protein-ligand binding analyses revealed stable affinities between PlYY1, PlCytochrome C, PlSpm1, PlrhoH12 proteins, and the 6PP ligand. Phenotypic characterization of PlYY1 targeted gene deletion strains generated in this study using CRISPR/Cas9 and homologous recombination strategies significantly reduced the vegetative growth, sporangium, encystment, zoospore release, and pathogenicity of P. litchii. These findings suggest that 6PP-mediated activation of PlYY1 expression positively regulates TOR-related responses and significantly influences vegetative growth and the virulence of P. litchii. The current investigations revealed novel targets for 6PP and underscored the potential of deploying 6PP in developing management strategies for controlling the litchi downy blight pathogen. Full article

Species of Amanita sect. Vaginatae (Fr.) Quél. are challenging to delimitate due to the morphological similarity or morphostasis among different taxa. In this study, a multi-locus (nuc rDNA region encompassing the internal transcribed spacers 1 and 2 with the 5.8S rDNA, the D1–D3 domains of nuc 28S rDNA, partial sequences of translation elongation factor 1-a, and the second largest subunit of RNA polymerase II) phylogeny was employed to investigate the species diversity of the section in eastern China. Sixteen species were recognized, including four new species; namely, A. circulata, A. multicingulata, A. orientalis, and A. sinofulva. They were documented with illustrated descriptions, ecological evidence, and comparisons with similar species. A key to the species of the section from eastern China is provided. Full article

Clonostachys rosea is an excellent biocontrol fungus against numerous fungal plant pathogens. The cAMP signaling pathway is a crucial signal transduction pathway in fungi. To date, the role of the cAMP signaling pathway in C. rosea mycoparasitism remains unknown. An adenylate cyclase-encoding gene, crac (an important component of the cAMP signaling pathway), was previously screened from C. rosea 67-1, and its expression level was dramatically upregulated during the C. rosea mycoparasitization of the sclerotia of Sclerotinia sclerotiorum. In this study, the function of crac in C. rosea mycoparasitism was explored through gene knockout and complementation. The obtained results show that the deletion of crac influenced the growth rate and colony morphology of C. rosea, as well as the tolerance to NaCl and H₂O₂ stress. The mycoparasitic effects on the sclerotia of S. sclerotiorum and the biocontrol capacity on soybean Sclerotinia stem rot in ∆crac-6 and ∆crac-13 were both attenuated compared with that of the wild-type strain and complementation transformants. To understand the regulatory mechanism of crac during C. rosea mycoparasitism, transcriptomic analysis was conducted between the wild-type strain and knockout mutant. A number of biocontrol-related genes, including genes encoding cell wall-degrading enzymes and transporters, were significantly differentially expressed during C. rosea mycoparasitism, suggesting that crac may be involved in C. rosea mycoparasitism by regulating the expression of these DEGs. These findings provide insight for further exploring the molecular mechanism of C. rosea mycoparasitism. Full article

Plant biomass is one of the most abundant renewable carbon sources, which holds great potential for replacing current fossil-based production of fuels and chemicals. In nature, fungi can efficiently degrade plant polysaccharides by secreting a broad range of carbohydrate-active enzymes (CAZymes), such as cellulases, hemicellulases, and pectinases. Due to the crucial role of plant biomass-degrading (PBD) CAZymes in fungal growth and related biotechnology applications, investigation of their genomic diversity and transcriptional dynamics has attracted increasing attention. In this project, we systematically compared the genome content of PBD CAZymes in six taxonomically distant species, Aspergillus niger, Aspergillus nidulans, Penicillium subrubescens, Trichoderma reesei, Phanerochaete chrysosporium, and Dichomitus squalens, as well as their transcriptome profiles during growth on nine monosaccharides. Considerable genomic variation and remarkable transcriptomic diversity of CAZymes were identified, implying the preferred carbon source of these fungi and their different methods of transcription regulation. In addition, the specific carbon utilization ability inferred from genomics and transcriptomics was compared with fungal growth profiles on corresponding sugars, to improve our understanding of the conversion process. This study enhances our understanding of genomic and transcriptomic diversity of fungal plant polysaccharide-degrading enzymes and provides new insights into designing enzyme mixtures and metabolic engineering of fungi for related industrial applications. Full article

Echinocandins, used for the prevention and treatment of invasive fungal infections, have led to a rise in breakthrough infections caused by resistant Candida species. Among these species, those belonging to the Candida haemulonii complex are rare multidrug-resistant (MDR) yeasts that are frequently misidentified but have emerged as significant healthcare-associated pathogens causing invasive infections. The objectives of this study were to investigate the evolutionary pathways of echinocandin resistance in C. haemulonii by identifying mutations in the FKS1 gene and evaluating the impact of resistance on fitness. After subjecting a MDR clinical isolate of C. haemulonii (named Ch4) to direct selection using increasing caspofungin concentrations, we successfully obtained an isolate (designated Ch4′r) that exhibited a high level of resistance, with MIC values exceeding 16 mg/L for all tested echinocandin drugs (caspofungin, micafungin, and anidulafungin). Sequence analysis revealed a specific mutation in the resistant Ch4′r strain, leading to an arginine-histidine amino acid substitution (R1354H), occurring at the G4061A position of the HS2 region of the FKS1 gene. Compared to the wild-type strain, Ch4′r exhibited significantly reduced growth proliferation, biofilm formation capability, and phagocytosis ratio, indicating a decrease in fitness. Transmission electron microscopy analysis revealed alterations in cell wall components, with a notable increase in cell wall thickness. The resistant strain also exhibited higher amounts (2.5-fold) of chitin, a cell wall-located molecule, compared to the wild-type strain. Furthermore, the resistant strain demonstrated attenuated virulence in the Galleria mellonella larval model. The evolved strain Ch4′r maintained its resistance profile in vivo since the treatment with either caspofungin or micafungin did not improve larval survival or reduce the fungal load. Taken together, our findings suggest that the acquisition of pan-echinocandin resistance occurred rapidly after drug exposure and was associated with a significant fitness cost in C. haemulonii. This is particularly concerning as echinocandins are often the first-line treatment option for MDR Candida species. Full article

The exploration of the western forests of Algeria led to the remarkable discovery of the first occurrence of Lepista sordida, an edible wild mushroom of significant culinary importance for the local community, traditionally consumed in its natural state. This discovery was made possible through the use of various methods, including macroscopic observations (revealing a violet color) as well as microscopic observations conducted using scanning electron microscopy (SEM), revealing a cylindrical shape with distinct contours. Additionally, molecular analyses were conducted. Genomic DNA was extracted from the mycelium, followed by DNA amplification using specific primers targeting the internal transcribed spacer region (ITS1 and ITS2). After PCR reactions and sequencing of the obtained amplicons, the nucleotide sequences of the mycelium were submitted to the GenBank database of NCBI with the assigned accession number: MZ928450.1. These sequences were subsequently used to construct the phylogenetic tree. Furthermore, an in-depth study of physicochemical parameters was undertaken to determine the optimal conditions for cultivating the mycelium of this edible wild mushroom, including pH, temperature, relative humidity, and light. Different temperatures were examined: 20, 25, 30, 35, 40, and 45 °C. The effect of pH on mycelium growth was studied using a PDA agar medium with buffered values of 4, 5, 5.6, 6, 7, and 8. Similarly, six levels of relative humidity were tested: 14, 50, 74, 80, 95, and 100%. A study on the impact of light on mycelium growth was conducted by exposing Petri dishes inoculated with PDA to a light intensity of 500 lux for 5, 10, 15, 20, and 24 h. The results clearly demonstrated that variations in these different physicochemical parameters significantly influenced mycelium growth. For the Lepista sordida strain, growth was favored at pH levels of 4, 5, 6, and 6, with no growth observed at pH 7 and 8. The optimal temperature range for mycelium growth of Lepista sordida was 20–25 °C, while no growth was observed at 30, 35, 40, and 45 °C. Relative humidity levels of 74, 80, and 95% showed no significant differences. Optimization of mycelium growth and primordia production in Lepista sordida were successfully achieved. Optimal conditions for the primordia phase were identified as 25 °C, with humidity ranging from 90 to 95%. A nutritional analysis of fresh sporophores was conducted using established analytical methods. Notably, the nutritional composition of Lepista sordida sporophores exhibited high significance for the following parameters: moisture content (67.23 ± 1.90%), ash content (9.35 ± 0.66%), fat content (3.25 ± 0.24%), protein content (17.22 ± 0.38%), and carbohydrate content (63.83 ± 1.23%). Full article

The global environmental issue of arsenic (As) contamination in drinking water is a significant problem that requires attention. Therefore, the aim of this research was to address the application of a sustainable methodology for arsenic removal through mycoremediation aerated with micro-nanobubbles (MNBs), leading to bioscorodite (FeAsO₄·2H₂O) generation. To achieve this, the fungus Trichoderma atroviride was cultivated in a medium amended with 1 g/L of As(III) and 8.5 g/L of Fe(II) salts at 28 °C for 5 days in a tubular reactor equipped with an air MNBs diffuser (TR-MNBs). A control was performed using shaking flasks (SF) at 120 rpm. A reaction was conducted at 92 °C for 32 h for bioscorodite synthesis, followed by further characterization of crystals through Fourier–Transform Infrared Spectroscopy (FTIR), Scanning Electron Microscopy (SEM), and X-ray diffraction (XRD) analyses. At the end of the fungal growth in the TR-MNBs, the pH decreased to 2.7–3.0, and the oxidation-reduction potential (ORP) reached a value of 306 mV at 5 days. Arsenic decreased by 70%, attributed to possible adsorption through rapid complexation of oxidized As(V) with the exchangeable ferrihydrite ((Fe(III))_4-5(OH,O)₁₂), sites, and the fungal biomass. This mineral might be produced under oxidizing and acidic conditions, with a high iron concentration (As:Fe molar ratio = 0.14). The crystals produced in the reaction using the TR-MNBs culture broth and characterized by SEM, XRD, and FTIR revealed the morphology, pattern, and As-O-Fe vibration bands typical of bioscorodite and römerite (Fe(II)(Fe(III))₂(SO₄)₄·14H₂O). Arsenic reduction in SF was 30%, with slight characteristics of bioscorodite. Consequently, further research should include integrating the TR-MNBs system into a pilot plant for arsenic removal from contaminated water. Full article

The community structure of macrofungi is influenced by multiple complex factors, including climate, soil, vegetation, and human activities, making it challenging to discern their individual contributions. To investigate the dynamic changes in macrofungal diversity in an Ancient Tree Park located in Northeast China and explore the factors influencing this change, we collected 1007 macrofungi specimens from different habitats within the park and identified 210 distinct fungal species using morphological characteristics and ITS sequencing. The species were classified into 2 phyla, 6 classes, 18 orders, 55 families, and 94 genera. We found macrofungal compositions among different forest types, with the mixed forest displaying the highest richness and diversity. Climatic factors, particularly rainfall and temperature, positively influenced macrofungal species richness and abundance. Additionally, by analyzing the soil fungal community structure and comparing aboveground macrofungi with soil fungi in this small-scale survey, we found that the soil fungal bank is not the main factor leading to changes in the macrofungal community structure, as compared to the influence of climate factors and forest types. Our findings provide valuable insights into the dynamic nature of macrofungal diversity in the Ancient Tree Park, highlighting the influence of climate and forest type. Full article

Morels are one of the most highly prized edible and medicinal mushrooms worldwide. Therefore, historically, there has been a large international interest in their cultivation. Numerous ecological, physiological, genetic, taxonomic, and mycochemical studies have been previously developed. At the beginning of this century, China finally achieved artificial cultivation and started a high-scale commercial development in 2012. Due to its international interest, its cultivation scale and area expanded rapidly in this country. However, along with the massive industrial scale, a number of challenges, including the maintenance of steady economic profits, arise. In order to contribute to the solution of these challenges, formal research studying selection, species recognition, strain aging, mating type structure, life cycle, nutrient metabolism, growth and development, and multi-omics has recently been boosted. This paper focuses on discussing current morel cultivation technologies, the industrial status of cultivation in China, and the relevance of basic biological research, including, e.g., the study of strain characteristics, species breeding, mating type structure, and microbial interactions. The main challenges related to the morel cultivation industry on a large scale are also analyzed. It is expected that this review will promote a steady global development of the morel industry based on permanent and robust basic scientific knowledge. Full article

Background: The treatment of paracoccidioidomycosis (PCM) is a challenge, and the discovery of new antifungal compounds is crucial. The phenacylideneoxindoles exhibited promising antifungal activity against Paracoccidioides spp., but their mode of action remains unknown. Methods: Through proteomic analysis, we investigated the effects of (E)-3-(2-oxo-2-phenylethylidene)indolin-2-one on P. brasiliensis. In addition, we investigated the metabolic alterations of P. brasiliensis in response to the compound. Furthermore, the effects of the compound on the membrane, ethanol production, and reactive oxygen species (ROS) production were verified. Results: We identified differentially regulated proteins that revealed significant metabolic reorganization, including an increase in ethanol production, suggesting the activation of alcoholic fermentation and alterations in the rigidity of fungal cell membrane with an increase of the ergosterol content and formation of ROS. Conclusions: These findings enhance our understanding of the mode of action and response of P. brasiliensis to the investigated promising antifungal compound, emphasizing its potential as a candidate for the treatment of PCM. Full article

The family Phaeosphaeriaceae is a diverse group of ascomycetous fungi that are commonly found in terrestrial habitats worldwide. In recent years, there has been increasing interest in the biodiversity of Phaeosphaeriaceae in China, particularly in Sichuan Province, which has not been fully explored for its high fungal diversity. In our study, we conducted extensive surveys in Sichuan Province to identify and describe new species of Ascomycota with diverse habitats. Here, we present a taxonomic revision of Phaeosphaeriaceae with taxonomic novelties from terrestrial habitats in Sichuan Province based on a multi-gene phylogenetic approach. Our study focuses on the description of four new species of Phaeosphaeriaceae, representing novel additions to the diversity of this fungal family. Using a combination of morphological and molecular data, we provide detailed descriptions of the new taxa and their placement within the family. Additionally, we discuss the phylogenetic relationships of these new taxa with other members of Phaeosphaeriaceae, providing insights into the correct taxonomic classification of the family. Our taxonomic revision contributes to understanding fungal diversity in China and provides a foundation for future studies investigating the taxonomy and ecological roles of Phaeosphaeriaceae fungi. Furthermore, our multi-gene phylogenetic approach provides increased resolution and accuracy in the delimitation of species boundaries within the family. Our study highlights the importance of continued exploration and taxonomic revision in order to fully understand the diversity and distribution of fungal species in China and beyond. New species: Paraloratospora sichuanensis, Phaeosphaeria chengduensis, P. sichuanensis, and Septoriella shoemakeri. New combinations: Paraloratospora breonadiae, P. fructigena, Septoriella ammophilicola, S. asparagicola, S. festucae, S. luzulae, and S. verrucispora. New names: Septoriella paradactylidis, and S. neomuriformis. Full article

In nature, arbuscular mycorrhizal fungi (AMF) play a crucial role in the root systems of plants. They can help enhance the resistance of host plants by improving the compartmentalization of toxic metal contaminants in the cell walls (CWs). However, the functions and responses of various CW subfractions to mycorrhizal colonization under Cd exposure remain unknown. Here we conducted a study to investigate how Cd is stored in the cell walls of maize roots colonized by Funneliformis mosseae. Our findings indicate that inoculating the roots with AMF significantly lowers the amount of Cd in the maize shoots (63.6 ± 6.54 mg kg⁻¹ vs. 45.3 ± 2.19 mg kg⁻¹, p < 0.05) by retaining more Cd in the mycorrhized roots (224.0 ± 17.13 mg kg⁻¹ vs. 289.5 ± 8.75 mg kg⁻¹, p < 0.01). This reduces the adverse effects of excessive Cd on the maize plant. Additional research on the subcellular distribution of Cd showed that AMF colonization significantly improves the compartmentalization of 88.2% of Cd in the cell walls of maize roots, compared to the 80.8% of Cd associated with cell walls in the non-mycorrhizal controls. We observed that the presence of AMF did not increase the amount of Cd in pectin, a primary binding site for cell walls; however, it significantly enhanced the content of lignin and the proportion of Cd in the total root cell walls. This finding is consistent with the increased activity of lignin-related enzymes, such as PAL, 4CL, and laccase, which were also positively impacted by mycorrhizal colonization. Fourier transform infrared spectroscopy (FTIR) results revealed that AMF increased the number and types of functional groups, including −OH/−NH and carboxylate, which chelate Cd in the lignin. Our research shows that AMF can improve the ability of maize plants to tolerate Cd by reducing the amount of Cd transferred from the roots to the shoots. This is achieved by increasing the amount of lignin in the cell walls, which binds with Cd and prevents it from moving through the plant. This is accomplished by activating enzymes related to lignin synthesis and increasing the exposure of Cd-binding functional groups of lignin. However, more direct evidence on the immobilization of Cd in the mycorrhiza-altered cell wall subfractions is needed. Full article

In this study, a halotolerant yeast that is capable of efficiently decolorizing and detoxifying azo dyes was isolated, identified and characterized for coping with the treatment of azo-dye-containing wastewaters. A characterization of the yeast, including the optimization of its metabolism and growth conditions, its detoxification effectiveness and the degradation pathway of the target azo dye, as well as a determination of the key activities of the enzyme, was performed. Finally, the possible halotolerance mechanisms of the yeast were proposed through a comparative transcriptome analysis. The results show that a halotolerant yeast, A4, which could decolorize various azo dyes, was isolated from a marine environment and was identified as Meyerozyma guilliermondii. Its optimal conditions for dye decolorization were ≥1.0 g/L of sucrose, ≥0.2 g/L of (NH₄)₂SO₄, 0.06 g/L of yeast extract, pH 6.0, a temperature of 35 °C and a rotation speed of ≥160 rpm. The yeast, A4, degraded and detoxified ARB through a series of steps, relying on the key enzymes that might be involved in the degradation of azo dye and aromatic compounds. The halotolerance of the yeast, A4, was mainly related to the regulation of the cell wall components and the excessive uptake of Na⁺/K⁺ and/or compatible organic solutes into the cells under different salinity conditions. The up-regulation of genes encoding Ca²⁺-ATPase and casein kinase II as well as the enrichment of KEGG pathways associated with proteasome and ribosome might also be responsible for its halotolerance. Full article

Fusarium species are among the filamentous fungi with the most pronounced impact on agricultural production and human health. The mycotoxins produced by pathogenic Fusarium not only attack various plants including crops, causing various plant diseases that lead to reduced yields and even death, but also penetrate into the food chain of humans and animals to cause food poisoning and consequent health hazards. Although sporadic studies have revealed some of the biosynthetic pathways of Fusarium toxins, they are insufficient to satisfy the need for a comprehensive understanding of Fusarium toxin production. In this study, we focused on 35 serious pathogenic Fusarium species with available genomes and systematically analyzed the ubiquity of the distribution of identified Fusarium- and non-Fusarium-derived fungal toxin biosynthesis gene clusters (BGCs) in these species through the mining of core genes and the comparative analysis of corresponding BGCs. Additionally, novel sesterterpene synthases and PKS_NRPS clusters were discovered and analyzed. This work is the first to systematically analyze the distribution of related mycotoxin biosynthesis in pathogenic Fusarium species. These findings enhance the knowledge of mycotoxin production and provide a theoretical grounding for the prevention of fungal toxin production using biotechnological approaches. Full article

Candida auris is a globally emerging fungal pathogen that is associated with healthcare-related infections. The accurate and rapid detection of C. auris is crucial for effective infection prevention, control, and patient management. This study aimed to validate the analytical and diagnostic performance of the DiaSorin Molecular C. auris Detection Kit. The analytical specificity, sensitivity, and reproducibility of the assay were evaluated. The limit of detection (LOD) was determined to be 266 CFU/µL using the ZeptoMetrix Candida auris Z485 strain and standard calibration curves. The assay demonstrated high analytical specificity and showed no amplification against a diverse panel of bacteria and fungi. Clinical validation was conducted using deidentified residual axillary/groin surveillance culture specimens from C. auris culture-positive and culture-negative patients. The DiaSorin Molecular Detection Kit exhibited 100% agreement in sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) when compared to cultures coupled with MALDI-TOF identification. Intra- and inter-reproducibility testing demonstrated consistent and reliable diagnostic performance. This validated assay offers rapid and accurate detection of C. auris, facilitating timely implementation of infection control measures and appropriate patient care. The DiaSorin Molecular C. auris Detection Kit has the potential to aid in controlling the outbreaks caused by this emerging fungal pathogen. Providing a reliable diagnostic tool can contribute to the effective management and containment of C. auris infections in healthcare settings and ultimately improve patient outcomes. Full article

Austropuccinia psidii is a biotrophic fungus that causes myrtle rust. First described in Brazil, it has since spread to become a globally important pathogen that infects more than 480 myrtaceous species. One of the most important commercial crops affected by A. psidii is eucalypt, a widely grown forestry tree. The A. psidii–Eucalyptus spp. interaction is poorly understood, but pathogenesis is likely driven by pathogen-secreted effector molecules. Here, we identified and characterized a total of 255 virulence effector candidates using a genome assembly of A. psidii strain MF-1, which was recovered from Eucalyptus grandis in Brazil. We show that the expression of seven effector candidate genes is modulated by cell wax from leaves sourced from resistant and susceptible hosts. Two effector candidates with different subcellular localization predictions, and with specific gene expression profiles, were transiently expressed with GFP-fusions in Nicotiana benthamiana leaves. Interestingly, we observed the accumulation of an effector candidate, Ap28303, which was upregulated under cell wax from rust susceptible E. grandis and described as a peptidase inhibitor I9 domain-containing protein in the nucleus. This was in accordance with in silico analyses. Few studies have characterized nuclear effectors. Our findings open new perspectives on the study of A. psidii–Eucalyptus interactions by providing a potential entry point to understand how the pathogen manipulates its hosts in modulating physiology, structure, or function with effector proteins. Full article

Cucumber plants commonly suffer from Fusarium wilt disease, which is caused by Fusarium oxysporum f. sp. cucumerinum (Foc). Although resistant cultivars assist with Fusarium wilt disease control, enhancement of the virulence of Foc has been identified after monoculture of wilt-resistant cultivars. To investigate the biological characteristics that contribute to the virulence evolution of Foc, a wildtype strain foc-3b (WT) and its virulence-enhanced variant Ra-4 (InVir) were compared in terms of their growth, reproduction, stress tolerance, and colonization in cucumber plants. The InVir strain showed similar culture characteristics on PDA media to the WT strain but produced significantly more conidia (>two fold), with a distinctly higher germination rate (>four fold) than the WT strain. The colony diameter of the InVir strain increased faster than the WT strain on PDA plates; however, the mycelia dry weight of the InVir was significantly lower (<70%) than that of the WT harvested from PDB. The InVir strain exhibited a significant increase in tolerance to osmolality (1 M NaCl, 1 M KCl, etc.). The GFP-labeled InVir strain propagated in the cucumber vascular faster than the WT strain. These results suggest that increased conidia production and germination in vitro may correlate with virulence enhancement in Fusarium oxysporum f. sp. cucumerinum. This study will provide an insight into its virulence evolution and help us understand the mechanisms underlying the evolutionary biology of F. oxysporum. Full article

Sharp eyespot is a crucial disease affecting cereal plants, such as bread wheat (Triticum aestivum) and barley (Hordeum vulgare), and is primarily caused by the pathogenic fungus Rhizoctonia cerealis. As disease severity has increased, it has become imperative to find an effective and reasonable control strategy. One such strategy is the use of the trehalose analog, validamycin, which has been shown to have a potent inhibitory effect on several trehalases found in both insects and fungi, and is widely used as a fungicide in agriculture. In this study, we demonstrated that 0.5 μg/mL validamycin on PDA plates had an inhibitory effect on R. cerealis strain R0301, but had no significant impact on Fusarium graminearum strain PH-1. Except for its inhibiting the trehalase activity of pathogenic fungi, little is known about its mechanism of action. Six trehalase genes were identified in the genome of R. cerealis, including one neutral trehalase and five acidic trehalase genes. Enzyme activity assays indicated that treatment with 5 μg/mL validamycin significantly reduces trehalase activity, providing evidence that validamycin treatment does indeed affect trehalase, even though the expression levels of most trehalase genes, except Rc17406, were not obviously affected. Transcriptome analysis revealed that treatment with validamycin downregulated genes involved in metabolic processes, ribosome biogenesis, and pathogenicity in the R. cerealis. KEGG pathway analysis further showed that validamycin affected genes related to the MAPK signaling pathway, with a significant decrease in ribosome synthesis and assembly. In conclusion, our results indicated that validamycin not only inhibits trehalose activity, but also affects the ribosome synthesis and MAPK pathways of R. cerealis, leading to the suppression of fungal growth and pesticidal effects. This study provides novel insights into the mechanism of action of validamycin. Full article

Monitoring the dynamics of the spore bank of arbuscular mycorrhizal fungi (AMF) is essential for the sustainable management and protection of agroecosystems. The most common method for extracting AMF spores from soil is the wet-sieving technique (WST). However, this method has many disadvantages. In this study, we modified the WST using new approaches: the ultrasound wet-sieving technique (UWST) and the ultrasound centrifuge technique (UCT). We enumerated and compared the numbers and quality of spores obtained from WST, UWST, and UCT to validate the new modified techniques. We extracted AMF spores from the rhizospheres of different plants, including wheat (Triticum aestivum L.), bean (Phaseolus vulgaris L.), tomato (Solanum lycopersicum L.), pepper (Piper nigrum L.), parsley (Petroselinum crispum Mill.), and turfgrass (Lolium perenne L.) collected from the Van Lake basin, Turkey. The highest and lowest AMF spore numbers were observed in wheat and turfgrass rhizospheres. The UCT allowed for the extraction of the highest number of spores from all rhizospheres, followed by the UWST and WST. The UWST and WST allowed for the extraction of similar spore numbers from wheat, pepper, parsley, and turfgrass rhizospheres. Beyond the high extracted spore number, UCT was shown to be a fast and low-material-consuming approach. These findings demonstrate that the UCT can be used to efficiently extract AMF spores in future research. Full article

The present study aimed to describe the clinical, epidemiological and laboratory characteristics of invasive candidiasis by C. parapsilosis complex (CPC) in a Brazilian tertiary pediatric hospital during the COVID-19 pandemic. Clinical samples were processed in the BACT/ALERT^® 3D system or on agar plates. Definitive identification was achieved by MALDI-TOF MS. Antifungal susceptibility was initially analyzed by the VITEK 2 system (AST-YS08 card) and confirmed by the CLSI protocol. Patient data were collected from the medical records using a structured questionnaire. CPC was recovered from 124 patients over an 18-month period, as follows: C. parapsilosis (83.87%), C. orthopsilosis (13.71%) and C. metapsilosis (2.42%). Antifungal resistance was not detected. The age of the patients with invasive CPC infections ranged from <1 to 18 years, and most of them came from oncology-related sectors, as these patients were more affected by C. parapsilosis. C. orthopsilosis infections were significantly more prevalent in patients from critical care units. Invasive infections caused by different pathogens occurred in 75 patients up to 30 days after the recovery of CPC isolates. Overall, 23 (18.55%) patients died within 30 days of CPC diagnosis. Catheter removal and antifungal therapy were important measures to prevent mortality. COVID-19 coinfection was only detected in one patient. Full article

Fusarium dry rot is one of the major potato diseases during storage after harvest, which not only results in quality degradation but also causes great economic losses. The disease can be elicited by some species of Fusarium, and the pathogenic fungi of Fusarium causing potato dry rot are considerably diverse in various countries and regions. The disease caused by Fusarium spp. is associated with mycotoxins accumulation, which has phytotoxic and mycotoxic effects on humans and animals. Chemical synthetic fungicide is considered the main control measure for the Fusarium dry rot of potato; nevertheless, it is unfortunate that persistent application inevitably results in the emergency of a resistant strain and environmental contamination. A comprehensive disease control strategy includes potato cultivar selection, appropriate cultural practices (crop rotation, cultivate pattern, fertilization, and irrigation), harvesting processes and postharvest treatments (harvesting, classification, packaging, wound healing), and storage conditions (environmental disinfection, temperature, humidity and gas composition) along with the application of fungicide pre-harvest or post-harvest. Recently, emerging studies have indicated that eco-friendly strategies include physical control, chemical methods (such as the application of generally-recognised-as-safe (GRAS) compounds or chemical (elicitors) and biological control have been introduced to combat the Fusarium dry rot of potato. Full article

Aspergillus mold is a ubiquitously found, airborne pathogen that can cause a variety of diseases from mild to life-threatening in severity. Limitations in diagnostic methods combined with anti-fungal resistance render Aspergillus a global emerging pathogen. In industry, Aspergilli produce toxins, such as aflatoxins, which can cause food spoilage and pose public health risk issues. Here, we report a multiplex qPCR method for the detection and identification of the five most common pathogenic Aspergillus species, Aspergillus fumigatus, Aspergillus flavus, Aspergillus niger, Aspergillus terreus, and Aspergillus nidulans. Our approach exploits species-specific nucleotide polymorphisms within their ITS genomic regions. This novel assay combines multiplex single-color real time qPCR and melting curve analysis and provides a straight-forward, rapid, and cost-effective detection method that can identify five Aspergillus species simultaneously in a single reaction using only six unlabeled primers. Due to their unique fragment lengths, the resulting amplicons are directly linked to certain Aspergillus species like fingerprints, following either electrophoresis or melting curve analysis. Our method is characterized by high analytical sensitivity and specificity, so it may serve as a useful and inexpensive tool for Aspergillus diagnostic applications both in health care and the food industry. Full article

The present study, initially to resolve the cryptic species within Corollospora maritima, is to determine how to attain taxonomic discrimination at species and generic levels. Multiple sequence alignments (MSAs) of the ITS, 28S, and 18S regions of the nuclear ribosomal cistron were separately subjected to pairwise distance assessments, Bayesian, and Maximum likelihood phylogenetic analyses. Morphological descriptions of 15 type strains of Corollospora species, along with MSAs involving representatives of the whole genus Corollospora (268 isolates, many from C. maritima sensu lato) totaling 355 published sequences, allowed phylogenetic assessments conducted to the following p-distance thresholds in the ITS/28S regions: ≥3%/1% for species segregation and ≥8%/2% for generic segregation. This resulted in the introduction of 10 new genera encompassing 13 new combinations of current Corollospora species: Ajigaurospora pseudopulchella, Corollosporella anglusa, Corollosporella ramulosa, Corollosporopsis portsaidica, Garethelia parvula, Honshuriella fusca, Keraliethelia pulcehlla, Nakagariella filiformis, Paracorollospora angusta, Paracorollospora luteola, Paracorollospora marina, Shirahamella gracilis, and Tokuratelia colossa. Furthermore, seven undefined genera considered putative new genera (pNGenus A to G), and 16 undefined putative new species (seven spp. come from the resolution of the C. maritima complex), await re-assessment of their morphology and additional molecular data, which may result in the recognition of new taxa. Full article

Native wild yeasts from forest ecosystems represent an interesting potential source of biocontrol organisms in synergy with disease-tolerant forest materials. Yeasts have a combination of competitive mechanisms and low requirements for their biotechnological application as biocontrol agents. The current study aimed to increase the number of biocontrol candidates against Fusarium circinatum and Diplodia sapinea. The enzymatic and antagonistic activities of the biocontrol candidates were evaluated using different screening methods, in which the direct impact on the growth of the pathogen was measured as well as some properties such as cellulose and lignin degradation, tolerance to biocides, volatile compound production, or iron effect, which may be of interest in biotechnological processes related to the management of forest diseases. A total of 58 yeast strains belonging to 21 different species were obtained from oak forest and vineyard ecosystems and evaluated. The application of yeast treatment behaved differently depending on the pathogen and the plant clone. The 2g isolate (Torulaspora delbrueckii) showed the highest inhibitory activity for D. sapinea and 25q and 90q (Saccharomyces paradoxus) for F. circinatum. Clones IN416 and IN216 were the most susceptible and the most tolerant to D. sapinea, respectively, while the opposite was observed for F. circinatum. Full article

Background: Alternaria alternata is a melanic fungus capable of causing a wide variety of infections, some of which have lethal potential. It is a ubiquitous fungus and a well-known plant pathogen. Cutaneous infections with Alternaria alternata most often occur in the extremities of patients who perform conventional agriculture, thus being exposed to occupational hazards leading to the disruption of the skin barrier. Methods: This paper presents the first case report from Romania of an itraconazole nonresponsive cutaneous alternariosis in a patient without any type of immunosuppression. Results: After an initial misdiagnosis regarding the etiology of the patient’s skin infection, two successive punch biopsies, followed by mycologic examination, lead to the final diagnosis of cutaneous alternariosis. Treatment guided by antifungal susceptibility testing has been instituted, leading to the gradual healing of the patient’s skin ulcerations. Conclusions: The ability of Alternaria alternata to infect immunocompetent human hosts and to develop resistance to antifungal drugs highlight the importance of correctly diagnosing the etiology of skin ulcerations and instituting appropriate treatment guided by antifungal susceptibility testing whenever the suspicion of a fungal skin infection is plausible. Full article

Background: Very few cases of Pneumocystis jirovecii pneumonia (PJP) have been reported in COVID-19 so far, and mostly in patients with concomitant HIV infection or in solid-organ transplant recipients. Despite COVID-19 being associated with lymphopenia and the use of steroids, there are no studies specifically aimed at investigating the risk factors for PJP in COVID-19. Methods: A retrospective case-control study was performed. We matched PJP cases with controls with a 1:2 ratio, based on age ± 10 years, solid-organ transplantation (SOT), hematological malignancies, and in the setting of PJP development (ICU vs. non-ICU). A direct immunofluorescence assay on bronchoalveolar lavage fluid was used to diagnose PJP. Results: We enrolled 54 patients. Among 18 cases of PJP, 16 were diagnosed as “proven”. Seven of the eighteen cases were immunocompromised, while the other patients had no previous immunological impairment. Patients with PJP had significantly lower median lymphocyte values (p = 0.033), longer COVID-19 duration (p = 0.014), a higher dose of steroid received (p = 0.026), higher CRP values (p = 0.005), and a lower SARS-CoV-2 vaccination rate than the controls (p = 0.029). Cumulative steroid dose is the independent risk factor for PJP development (OR = 1.004, 95%CI = 1–1.008, p = 0.042). Conclusions: PJP develops in COVID-19 patients regardless of immunosuppressive conditions and the severity of disease, and it is correlated to the corticosteroid dose received. Full article

Candida auris is an opportunistic human pathogen that has rapidly spread to multiple countries and continents and has been associated with a high number of nosocomial outbreaks. Herein, we report the first case of C. auris in Portugal, which was associated with a patient transferred from Angola to an ICU in Portugal for liver transplantation after a SARS-CoV-2 infection. C. auris was isolated during the course of bronchoalveolar lavage, and it was subjected to antifungal susceptibility testing and whole-genome sequence analysis. This isolate presents low susceptibility to azoles and belongs to the genetic clade III with a phylogenetic placement close to African isolates. Although clade III has already been reported in Europe, taking into account the patient’s clinical history, we cannot discard the possibility that the patient’s colonization/infection occurred in Angola, prior to admission in the Portuguese hospital. Considering that C. auris is a fungal pathogen referenced by WHO as a critical priority, this case reinforces the need for continuous surveillance in a hospital setting. Full article

Cryptococcosis is the leading cause of death among people with HIV in Sub-Saharan Africa. The lack of optimum diagnoses and medications significantly impair the management of the disease. We investigated the burden of cryptococcosis and related mortality among people with HIV and suspected sepsis in Ethiopia. We conducted a prospective study at (1) Adama Hospital Medical College and (2) Asella Referral and Teaching Hospital from September 2019 to November 2020. We enrolled adult, HIV-infected patients presenting with suspected sepsis and assessed their 28-day survival rates. We performed blood cultures and cryptococcal antigen (CrAg) testing. In total, 82 participants were enrolled with a median age of 35 years, and 61% were female. Overall, eleven (13%) had positive CrAg tests, of which five grew Cryptococcus in blood cultures. Despite high-dose fluconazole (1200 mg/d) monotherapy being given to those with positive CrAg tests, the 28-day mortality was 64% (7/11), with mortality being significantly higher than among the CrAg-negative patients (9% (6/71); p < 0.001). Cryptococcosis was the leading cause of mortality among HIV-infected sepsis patients in this Ethiopian cohort. The CrAg screening of HIV-infected patients attending an emergency department can minimize the number of missed cryptococcosis cases irrespective of the CD4 T cell count and viral load. These findings warrant the need for a bundle approach for the diagnosis of HIV-infected persons presenting with sepsis in low- and middle-income countries. Full article

Tribolium castaneum, Trogoderma granarium, Oryzaephilus surinamensis, Sitophilus oryzae, Rhyzopertha dominica, and Cryptolestes ferrugineus are all major pests of stored grains. In this study, the efficiency of single and joint applications of the entomopathogenic nematode (EPN) Steinernema carpocapsae at two different doses (50 and 100 IJs cm⁻²) and the entomopathogenic fungus (EPF) Beauveria bassiana for the management of the aforementioned pests was estimated. At single treatments, both doses of S. carpocapsae caused higher mortality rates to all six pest species compared to B. bassiana. The combined treatment of EPF and EPN resulted in higher mortality compared to single treatments. Mortality was strongly influenced by the exposure interval and the application dose of the EPN at both single and combined treatments. Maximum mortality was observed for the application of the combined treatment at the high dose of S. carpocapsae and B. bassiana. Among the different insect species tested, the maximum mortality rate was observed for R. dominica (96.62%), followed by S. oryzae (90.48%), T. castaneum (87.23%), C. ferrugineus (76.05%), O. surinamensis (70.74%), and T. granarium (57.71%). The outcomes of this study demonstrate the potential of utilizing specific combinations of EPF and EPN as effective natural enemies against stored-grain pests. Full article