The regulatory network of the White Collar complex during early mushroom development in Schizophyllum commune.

Blue light is an important signal for fungal development. In the mushroom-forming basidiomycete Schizophyllum commune, blue light is detected by the White Collar complex, which consists of WC-1 and WC-2. Most of our knowledge on this complex is derived from the ascomycete Neurospora crassa, where both WC-1 and WC-2 contain GATA zinc-finger transcription factor domains. In basidiomycetes, WC-1 is truncated and does not contain a transcription factor domain, but both WC-1 and WC-2 are still important for development. We show that dimerization of WC-1 and WC-2 happens independent of light in S. commune, but that induction by light is required for promoter binding by the White Collar complex. Furthermore, the White Collar complex is a promoter of transcription, but binding of the complex alone is not always sufficient to initiate transcription. For its function, the White Collar complex associates directly with the promoters of structural genes involved in mushroom development, like hydrophobins, but also promotes the expression of other transcription factors that play a role in mushroom development.

Anaerobic production and biosynthesis mechanism of exopolysaccharides in Schizophyllum commune 20R-7-F01.

Exopolysaccharides (EPS) produced by microorganisms play a vital role in physiological and ecological processes. However, the mechanisms of EPS synthesis and release in anaerobic environments remain poorly understood. Here, we provide the first evidence of anaerobic EPS synthesis by the fungus Schizophyllum commune 20R-7-F01, isolated from coal-bearing sediments ~2.0 km below the seafloor. Under anaerobic conditions, the fungus exhibited significantly higher specific EPS production (1.57 times) than under aerobic conditions. Transcriptomic analysis revealed 2057 differentially expressed genes (DEGs) in the strain cultured anaerobically for 7 days compared to aerobically. Among these genes, 642 were significantly upregulated, while 1415 were significantly downregulated, mainly associated with carbon metabolism pathways. Genes involved in glycolysis and EPS synthesis, including hexokinase (HK), phosphoglucomutase (PGM), and (1 â†’ 3)-ß-glucan synthase (GLS), were significantly upregulated, while those related to the TCA cycle, respiratory chain, and pentose phosphate pathway were downregulated under anaerobic conditions. These findings highlight the oxygen-dependent regulation of EPS synthesis and suggest that EPS may serve as a key mechanism for fungal adaptation to anaerobic environments.

Successful management of nasopharyngitis caused by Schizophyllum commune in a captive cheetah (Acinonyx jubatus).

Herein, we describe the management of nasopharyngitis caused by Schizophyllum commune infection in a captive cheetah. Computed tomography revealed a nodule in the nasal cavity and pharynx, and an endoscopic biopsy was performed. As a result, the nodule was surgically resected because of a suspected carcinoma. However, the surgical specimen was histologically re-evaluated and a fungal granuloma was diagnosed. Sequence analysis of DNA from formalin-fixed, paraffin-embedded samples revealed S. commune infection. The cheetah was administered fluconazole orally for 73 days. However, the drug was ineffective and itraconazole was administered for 14 days. Symptoms such as nasal discharge and sneezing have completely resolved for 4 years.

Production, purification, and characterization of p-diphenol oxidase (PDO) enzyme from lignolytic fungal isolate Schizophyllum commune MF-O5.

Fungi are producers of lignolytic extracellular enzymes which are used in industries like textile, detergents, biorefineries, and paper pulping. This study assessed for the production, purification, and characterization of novel p-diphenol oxidase (PDO; laccase) enzyme from lignolytic white-rot fungal isolate. Fungi samples collected from different areas of Pakistan were initially screened using guaiacol plate method. The maximum PDO producing fungal isolate was identified on the basis of ITS (internal transcribed spacer sequence of DNA of ribosomal RNA) sequencing. To get optimum enzyme yield, various growth and fermentation conditions were optimized. Later PDO was purified using ammonium sulfate precipitation, size exclusion, and anion exchange chromatography and characterized. It was observed that the maximum PDO producing fungal isolate was Schizophyllum commune (MF-O5). Characterization results showed that the purified PDO was a monomeric protein with a molecular mass of 68 kDa and showed stability at lower temperature (30 °C) for 1 h. The Km and Vmax values of the purified PDO recorded were 2.48 mM and 6.20 U/min. Thermal stability results showed that at 30 °C PDO had 119.17 kJ/K/mol Ea value and 33.64 min half-life. The PDO activity was stimulated by Cu2+ ion at 1.0 mM showing enhanced activity up to 111.04%. Strong inhibition effect was noted for Fe2+ ions at 1 mM showing 12.04% activity. The enzyme showed stability against 10 mM concentration oxidizing reducing agents like DMSO, EDTA, H2O2, NaOCl, and urea and retained more than 75% of relative activity. The characterization of purified PDO enzyme confirmed its tolerance against salt, metal ions, organic solvents, and surfactants indicating its ability to be used in the versatile commercial applications.

An endophytic fungus Schizophyllum commune isolated from Panax ginseng enhances hairy roots growth and ginsenoside biosynthesis.

Using endophytic fungal elicitors to increase the accumulation of valuable secondary metabolites in plant tissue culture is an effective biotechnology strategy. In this study, a collection of 56 strains of endophytic fungi were isolated from different organs of cultivated Panax ginseng, of which seven strains can be symbiotically co-cultured with the hairy roots of P. ginseng. Further experiments observed that strain 3R-2, identified as endophytic fungus Schizophyllum commune, can not only infect hairy roots but also promote the accumulation of specific ginsenosides. This was further verified because S. commune colonization significantly affected the overall metabolic profile of ginseng hairy roots. By comparing the effects of S. commune mycelia and its mycelia extract (EM) on ginsenoside production in P. ginseng hairy roots, the EM was confirmed to be a relatively better stimulus elicitor. Additionally, the introduction of EM elicitor can significantly enhance the expressions of key enzyme genes of pgHMGR, pgSS, pgSE, and pgSD involved in the biosynthetic pathway of ginsenosides, which was deemed the most relevant factor for promoting ginsenosides production during the elicitation period. In conclusion, this study is the first to show that the EM of endophytic fungus S. commune can be considered as an effective endophytic fungal elicitor for increasing the biosynthesis of ginsenosides in hairy root cultures of P. ginseng.

The trends in the spread of simple sequence repeats in the genomes of Schizophyllum commune.

Due to their high number and variety, simple sequence repeats in DNA are a valuable source of genetic markers widely used in population genetics, genetic diversity, and fingerprinting. Schizophyllum commune is a cosmopolitan basidiomycete and a valuable object for biotechnology and mycology. It is often used as a model organism in genetic and population studies. Numerous genome sequences of S. commune are available in the National Center for Biotechnology Information (NCBI) database, allowing for a meticulous study of the distribution of simple tandem nucleotide repeats in the genomes of different strains, based on their origin. Analysis of the average density of simple tandem repeats in the genomes of S. commune fungi reveals the division of cultures into groups based on the USA, Ru (Russian Federation), and EA (South Korea, Japan, China) populations. Intensive point mutations lead to the accumulation and dominance of mononucleotide (C)n and dinucleotide (CG)n motifs in fungal genomes. Results show that the GCT motif contributes more to the divergence of the Ru population, whereas CTC, GAG, and GGA motifs contribute to the divergence of the USA population. Strains MG53 (China) and IUM1114-SS01 (South Korea: Incheon) from the EA population occupy a separate position, mainly because of the CGA motif. The divergence in quantitative traits of motifs between populations provides evidence of the evolutionary transformation of fungal genomes under environmental factors.

Pulmonary infection of Schizophyllum commune diagnosed by metagenomic next- generation sequencing: A case report.

RATIONALE: Fungal infection is common and difficult to be diagnosed timely in clinical, for its various kinds and similar manifestations. The rare pulmonary fungal infection such as Schizophyllum commune was one of the harder ones and misdiagnosed in usual. PATIENT CONCERNS: We report a 32-year-old female which was diagnosed with Metagenomic Next-Generation Sequencing (mNGS). She was hospitalized with the complaint of 4 months and more of repeated cough and expectorating. The chest computer tomography revealed left lower lobe pathological changes, but antibiotics were ineffective. No positive results were found in laboratory tests, including sputum culture and the pathology of lung puncture biopsy. DIAGNOSES: mNGS of lung biopsy was performed and detected the sequence number of Schizophyllum for 11. INTERVENTIONS: The patient was treated with voriconazole and itraconazole successively. OUTCOMES: She recovered to health. There was no recurrence during follow-up. LESSONS: mNGS as a diagnostic method could quickly detect pathogens through the processing of fragment, synthesis, comparison, and analysis of sample genes. It is suitable for detecting especially rare and polymicrobial infections. To our best knowledge, infection of Schizophyllum commune have not been reported in English literature with diagnostic method of mNGS.

Post-transplant Schizophyllum commune abscess in a pediatric patient with chronic granulomatous disease.

Schizophyllum commune is a widely distributed basidiomycete fungus that occasionally causes sinusitis or allergic bronchopulmonary mycosis. The invasive infection mostly occurs in immunocompromised adults. The number of reports on S. commune infection have increased in this decade due to the expansion of diagnostic techniques and awareness in clinical practice. However, S.commune infection in patients with primary immunodeficiencies has not been reported yet. Here, we described S. commune-abscesses developed in the brain and lung of a boy with chronic granulomatous disease (CGD) after allogenic hematopoietic cell transplantation (HCT). A 12-year-old CGD patient developed febrile neutropenia from day 4 after HCT, followed by chest pain on day 23. He had no obvious infection before HCT. Diagnostic imaging revealed disseminated lung and brain abscesses. He received administration of voriconazole, and his symptoms improved after engraftment. Chronic administration of voriconazole had also a favorable therapeutic response to brain lesion. A part of the fungus ball exhaled by the patient was cultured to develop a filamentous fungus. S. commune was identified by the analysis of the 28S rRNA gene. The catalase test was positive for S. commune, indicating that S. commune had virulence in this patient with CGD. The assessment of specific-IgG to S. commune suggested peri-transplant infection, although colonization was not excluded. This rare pediatric case of S. commune infection highlights that CGD patients are vulnerable to invasive infection, especially when undergoing HCT.

Identification of novel SSR markers for predicting the geographic origin of fungus Schizophyllum commune Fr.

The fungus Schizophyllum commune is a cosmopolitan basidiomycete, which is popular as an edible, medical mushroom. It causes wood rot and often used as a model object in research. In this study, we analyzed thirty-two genomes of S. commune strains from the NCBI database and designed forty-seven unique SSR DNA markers. The detailed analysis revealed the enrichment of the S. commune genome for CG, GC, CTC, GAG, and TCG motifs. Principal components analysis confirmed the effectiveness of novel SSR DNA markers that preserve the initial heterogeneity of populations. The construction of a network between strains showed single one at a maximum similarity of 38%, and increasing the similarity to 55% breaks the linkage between large groups while separating two new groups containing strains of the population Ru and test cultures S. commune. The amplicons' presence was identified as a sufficient sign of relation of the culture to a specific population. Testing the novel SSR markers allowed to establish a clear delimitation of all groups by geographic location and to differentiate the H4-8 (GCA_000143185.1) strain from the USA population. This research is the basis for the further analysis of S. commune populations and improvement of approaches to determine its genetic diversity.

The Transcription Factor Roc1 Is a Key Regulator of Cellulose Degradation in the Wood-Decaying Mushroom Schizophyllum commune.

Wood-decaying fungi of the class Agaricomycetes (phylum Basidiomycota) are saprotrophs that break down lignocellulose and play an important role in nutrient recycling. They secrete a wide range of extracellular plant cell wall degrading enzymes that break down cellulose, hemicellulose, and lignin, the main building blocks of plant biomass. Although the production of these enzymes is regulated mainly at the transcriptional level, no activating regulators have been identified in any wood-decaying fungus in the class Agaricomycetes. We studied the regulation of cellulase expression in the wood-decaying fungus Schizophyllum commune. Comparative genomics and transcriptomics on two wild isolates revealed a Zn2Cys6-type transcription factor gene (roc1) that was highly upregulated during growth on cellulose, compared to glucose. It is only conserved in the class Agaricomycetes. A roc1 knockout strain showed an inability to grow on medium with cellulose as sole carbon source, and growth on cellobiose and xylan (other components of wood) was inhibited. Growth on non-wood-related carbon sources was not inhibited. Cellulase gene expression and enzyme activity were reduced in the Δroc1 strain. ChIP-Seq identified 1474 binding sites of the Roc1 transcription factor. Promoters of genes involved in lignocellulose degradation were enriched with these binding sites, especially those of LPMO (lytic polysaccharide monooxygenase) CAZymes, indicating that Roc1 directly regulates these genes. A conserved motif was identified as the binding site of Roc1, which was confirmed by a functional promoter analysis. Together, Roc1 is a key regulator of cellulose degradation and the first identified in wood-decaying fungi in the phylum Basidiomycota. IMPORTANCE Wood-degrading fungi in the phylum Basidiomycota play a crucial role in nutrient recycling by breaking down all components of wood. Fungi have evolved transcriptional networks that regulate expression of wood-degrading enzymes, allowing them to prioritize one nutrient source over another. However, to date all these transcription factors have been identified in the phylum Ascomycota, which is only distantly related to the phylum Basidiomycota. Here, we identified the transcription factor Roc1 as a key regulator of cellulose degradation in the mushroom-forming and wood-degrading fungus Schizophyllum commune. Roc1 is highly conserved in the phylum Basidiomycota. Using comparative genomics, transcriptomics, ChIP-Seq and promoter analysis we have identified direct targets of Roc1, as well as other aspects of the transcriptional response to cellulose.

Metal adaptation and transport in hyphae of the wood-rot fungus Schizophyllum commune.

Fungi living in heavy metals and radionuclides contaminated environments, namely the Chernobyl Exclusion Zone need to be able to cope with these pollutants. In this study, the wood-rot fungus Schizophyllum commune was investigated for its metal tolerance mechanisms, and for its ability to transport such metals through its hyphae. Effects of temperature and pH on tolerance of Cs, Sr, Cd, and Zn were tested. At concentrations allowing for half-maximal growth, adapted strains were raised. The strontium-adapted strain, S. commune 12-43 Sr, showed transport of specifically Sr over distances on a cm-scale using split plates. The adaptation did not yield changes in cell or colony morphology. Intracellular metal localization was not changed, and gene expression profiles under metal stress growing on soil versus artificial medium showed a higher impact of a structured surface for growth on soil than with different metal concentrations. In the transcriptome, transporter genes were mostly down-regulated, while up-regulation was seen for genes involved in the secretory pathway under metal stress. A comparison of wildtype and adapted strains could confirm lower cellular stress levels leading to lack of glutathione S-transferase up-regulation in the adapted strain. Thus, we could show metal transport as well as specific mechanisms in metal stress avoidance.

Comparison of cell wall polysaccharides in Schizophyllum commune after changing phenotype by mutation.

The Agaricomycetes fungi produce various compounds with pharmaceutical, medicinal, cosmetic, environmental and biotechnological properties. In addition, some polysaccharides extracted from the fungal cell wall have antitumor and immunomodulatory actions. The aim of this study was to use genetic modification to transform Schizophyllum commune and identify if the phenotype observed (different from the wild type) resulted in changes of the cell wall polysaccharides. The plasmid pUCHYG-GPDGLS, which contains the Pleurotus ostreatus glucan synthase gene, was used in S. commune transformations. Polysaccharides from cell wall of wild (ScW) and mutants were compared in this study. Polysaccharides from the biomass and culture broth were extracted with hot water. One of the mutants (ScT4) was selected for further studies and, after hydrolysis/acetylation, the GLC analysis showed galactose as the major component in polysaccharide fraction from the mutant and glucose as the major monomer in the wild type. Differences were also found in the elution profiles from HPSEC and NMR analyses. From the monosaccharide composition it was proposed that mannogalactans are components of S. commune cell wall for both, wild and mutant, but in different proportions. To our knowledge, this is the first time that mannogalactans are isolated from S. commune liquid culture.

A Schizophyllum commune fungus ball in a lung cancer cavity: a case report.

BACKGROUND: Schizophyllum commune is a basidiomycete that lives in the environment and can cause infections, mainly those of the respiratory system. Although S. commune is increasingly reported as a cause of allergic bronchopulmonary mycosis and sinusitis, cases of fungal ball formation are extremely uncommon. Identification of S. commune is difficult using routine mycological diagnostic methods, and in clinically suspicious cases, internal transcribed spacer sequencing should be used for diagnosis. Here, we report a first case of lung cancer with a fungal ball formation of S. commune, confirmed by analyzing the internal transcribed spacer. CASE PRESENTATION: A 76-year-old man with diabetes and hypertension was admitted to the hospital with a chief complaint of hemosputum, which he had for about 19 months. A computed tomography image of the patient's chest showed a cavity and internal nodule in the left upper lobe of his lung. A left upper lobectomy was performed, and histopathological examination revealed squamous cell carcinoma of the lung and a fungal ball. The isolate from the surgical specimen was identified as S. commune by analyzing the internal transcribed spacer. The patient had no recurrence of the infection during 5 months of follow-up. CONCLUSIONS: Only three cases of lung fungal balls caused by S. commune have been previously reported, and this is the first case of lung cancer cavity with a fungal ball formation. In cases of fungal ball formation in the lung, S. commune should be considered a possible causative microorganism.

Submerged cultivation, characterization and in vitro antitumor activity of polysaccharides from Schizophyllum radiatum.

Production of polysaccharides by white-rot-fungi in submerged cultivation has several advantages due to process control. This work deals with the submerged cultivation, extraction and antitumor activity of polysaccharides from a wild strain of Schizophyllum radiatum isolated from a tropical forest of Colombia. The mushroom was cultivated in laboratory conditions, and classified by classical and molecular taxonomy. Submerged cultivation was performed in a bioreactor of 5 L using a ligninolytic residue as substrate. The fermentation conditions were 30 ± 1 °C, pH 4.5, 300 rpm and 1.5 vvm of air for 4 days. The yields were 16.8 g/L (w/v) of biomass, and after extraction, 0.6 g/L of water-soluble exopolysaccharide (SEPS) and 2.01 % (w/w) of water-soluble intrapolysaccharide (SIPS) were obtained. In each extract total carbohydrate, glucans and protein contents were determined. Also, nuclear magnetic resonance (NMR), scanning electron microscopy (SEM), Fourier transform infrared (FTIR) spectroscopy, X-ray diffractometry (XRD), high performance liquid chromatography with refraction index detection (HPLC-RI), high performance gel permeation chromatography (HPGPC) and Nuclear Magnetic Resonance (NMR) analysis were performed. Results indicated that SEPS and SIPS are heteropolysaccharides with amorphous structure and high molecular weights. Antitumor and immunostimulant activity was evaluated in different cancer cell lines. The results suggest these polysaccharides have direct and indirect antitumor activity activating immune cells such as macrophages. These findings enhance our knowledge about new sources of fungal metabolites that serve as adjuvant, cheaper and less harmful alternatives to cancer treatment.

H3K4me2 ChIP-Seq reveals the epigenetic landscape during mushroom formation and novel developmental regulators of Schizophyllum commune.

Mushroom formation represents the most complex multicellular development in fungi. In the model mushroom Schizophyllum commune, comparative genomics and transcriptomics have previously resulted in a regulatory model of mushroom development. However, little is known about the role of epigenetic regulation. We used chromatin immunoprecipitation sequencing (ChIP-Seq) to determine the distribution of dimethylation of lysine 4 on histone H3 (H3K4me2), a mark for transcriptionally active genes, during monokaryotic and dikaryotic development. We identified a total of 6032 and 5889 sites during monokaryotic and dikaryotic development, respectively. The sites were strongly enriched near translation initiation sites of genes. Although the overall epigenetic landscape was similar between both conditions, we identified 837 sites of differential enrichment during monokaryotic or dikaryotic development, associated with 965 genes. Six transcription factor genes were enriched in H3K4me2 during dikaryotic development, indicating that these are epigenetically regulated during development. Deletion of two of these genes (fst1 and zfc7) resulted in arrested development of fruiting bodies, resulting in immature mushrooms. Together these results indicate that H3K4me2 ChIP-Seq is a powerful new tool to map the restructuring of the epigenetic landscape during mushroom development. Moreover, it can be used to identify novel developmental regulators.

A case of allergic fungal rhinosinusitis caused by Schizophyllum commune identified in both patient's nasal sputum and veranda's soil samples.

This is a case report of allergic fungal rhinosinusitis caused by Schizophyllum commune (S. commune) identified in a patient's nasal mucus and environmental soil sample using (r)DNA sequencing. Although filamentous basidiomycetes, including S. commune, are known as environmental pathogens causing allergic respiratory diseases worldwide, many patients with infections caused by S. commune have not been correctly diagnosed. Repeated exposures to environmental floating fungi supposedly make an easy sensitization and colonization of fungi in the nasal passages, resulting in the onset of allergic fungal rhinosinusitis due to S. commune in our living environments. This report indicates the importance of reconsidering allergic respiratory diseases associated with our living environments.

The anaerobic survival mechanism of Schizophyllum commune 20R-7-F01, isolated from deep sediment 2 km below the seafloor.

Fungi dominated the eukaryotic group in the anaerobic sedimentary environment below the ocean floor where they play an essential ecological role. However, the adaptive mechanism of fungi to these anaerobic environments is still unclear. Here, we reported the anaerobic adaptive mechanism of Schizophyllum commune 20R-7-F01, isolated from deep coal-bearing sediment down to ~2 km below the seafloor, through biochemical, metabolomic and transcriptome analyses. The fungus grows well, but the morphology changes obviously and the fruit body develops incompletely under complete hypoxia. Compared with aerobic conditions, the fungus has enhanced branched-chain amino acid biosynthesis and ethanol fermentation under anaerobic conditions, and genes related to these metabolisms have been significantly up-regulated. Additionally, the fungus shows novel strategies for synthesizing ethanol by utilizing both glycolysis and ethanol fermentation pathways. These findings suggest that the subseafloor fungi may adopt multiple mechanisms to cope with lack of oxygen.

Genetic Diversity and Dye-Decolorizing Spectrum of Schizophyllum commune Population.

Synthetic dyes are widely used in various industries and their wastage causes severe environmental problems while being hazardous to human health, leading to the need for eco-friendly degradation techniques. The split-gill fungus Schizophyllum commune, which is found worldwide, has the potential to degrade all components of the lignocellulosic biomass and is a candidate for the treatment of synthetic dyes. A systematic molecular analysis of 75 Korean and 6 foreign S. commune strains has revealed the high genetic diversity of this population and its important contribution to the total diversity of S. commune. We examined the dye decolorization ability of this population and revealed 5 excellent strains that strongly decolorized 3 dyes: Crystal Violet, Congo Red and Methylene Blue. Finally, comparison of dye decolorization ability and the phylogenetic identification of these strains generalized their genetic and physiological diversity. This study provides an initial resource for physiological and genetic research projects as well as the bioremediation of textile dyes.

Rapid Accumulation of Mutations in Growing Mycelia of a Hypervariable Fungus Schizophyllum commune.

The basidiomycete Schizophyllum commune has the highest level of genetic polymorphism known among living organisms. In a previous study, it was also found to have a moderately high per-generation mutation rate of 2×10-8, likely contributing to its high polymorphism. However, this rate has been measured only in an experiment on Petri dishes, and it is unclear how it translates to natural populations. Here, we used an experimental design that measures the rate of accumulation of de novo mutations in a linearly growing mycelium. We show that S. commune accumulates mutations at a rate of 1.24×10-7 substitutions per nucleotide per meter of growth, or ∼2.04×10-11 per nucleotide per cell division. In contrast to what has been observed in a number of species with extensive vegetative growth, this rate does not decline in the course of propagation of a mycelium. As a result, even a moderate per-cell-division mutation rate in S. commune can translate into a very high per-generation mutation rate when the number of cell divisions between consecutive meiosis is large.