Screening and Evaluation of Streptomyces Species as a Potential Biocontrol Agent against a Wood Decay Fungus, Gloeophyllum trabeum

Two-hundred and fifty-five strains of actinomycetes isolated from soil samples were screened for their antagonistic activities against four well-known wood decay fungi (WDF), including a brown rot fungus, Gloeophyllum trabeum and three white rot fungi Donkioporia expansa, Trametes versicolor, and Schizophyllum commune. A dual culture assay using culture media supplemented with heated or unheated culture filtrates of selected bacterial strains was used for the detection of their antimicrobial activity against four WDF. It was shown that Streptomyces atratus, S. tsukiyonensis, and Streptomyces sp. greatly inhibited the mycelial growth of the WDF tested compared with the control. To evaluate the biocontrol efficacy of S. atratus, S. tsukiyonensis, and Streptomyces sp., wood blocks of Pinus densiflora inoculated with three selected Streptomyces isolates were tested for weight loss, compression strength (perpendicular or parallel to the grain), bending strength, and chemical component changes. Of these three isolates used, Streptomyces sp. exhibited higher inhibitory activity against WDF, especially G. trabeum, as observed in mechanical and chemical change analyses. Scanning electron microscopy showed that cell walls of the wood block treated with Streptomyces strains were thicker and collapsed to a lesser extent than those of the non-treated control. Taken together, our findings indicate that Streptomyces sp. exhibits the potential to be used as a biocontrol agent for wood decay brown rot fungus that causes severe damage to coniferous woods.

Phenolic acids isolated from the fungus Schizophyllum commune exert analgesic activity by inhibiting voltage-gated sodium channels / 中国天然药物

The present study was designed to search for compounds with analgesic activity from the Schizophyllum commune (SC), which is widely consumed as edible and medicinal mushroom world. Thin layer chromatography (TLC), tosilica gel column chromatography, sephadex LH 20, and reverse-phase high performance liquid chromatography (RP-HPLC) were used to isolate and purify compounds from SC. Structural analysis of the isolated compounds was based on nuclear magnetic resonance (NMR). The effects of these compounds on voltage-gated sodium (NaV) channels were evaluated using patch clamp. The analgesic activity of these compounds was tested in two types of mouse pain models induced by noxious chemicals. Five phenolic acids identified from SC extracts in the present study included vanillic acid, m-hydroxybenzoic acid, o-hydroxybenzeneacetic acid, 3-hydroxy-5-methybenzoic acid, and p-hydroxybenzoic acid. They inhibited the activity of both tetrodotoxin-resistant (TTX-r) and tetrodotoxin-sensitive (TTX-s) NaV channels. All the compounds showed low selectivity on NaV channel subtypes. After intraperitoneal injection, three compounds of these compounds exerted analgesic activity in mice. In conclusion, phenolic acids identified in SC demonstrated analgesic activity, facilitating the mechanistic studies of SC in the treatment of neurasthenia.

Characterization and biological activities of extracellular melanin produced by Schizophyllum commune (Fries).

Melanins are enigmatic pigments produced by a wide variety of microorganisms including bacteria and fungi. Here, we have isolated and characterized extracellular melanin from mushroom fungus, Schizophyllum commune. The extracellular dark pigment produced by the broth culture of S. commune, after 21 days of incubation was recovered by hot acid-alkali treatment. The melanin nature of the pigment was characterized by biochemical tests and further, confirmed by UV, IR, EPR, NMR and MALDI-TOF Mass Spectra. Extracellular melanin, at 100 µg/ml, showed significant antibacterial activity against Escherichia coli, Bacillus subtilis, Klebsiella pneumoniae and Pseudomonas fluorescens and antifungal activity against Trichophyton simii and T. rubrum. At a concentration of 50 µg/ml, melanin showed high free radical scavenging activity of DPPH (2,2-diphenyl-1-picrylhydrazyl) indicating its antioxidant potential. It showed concentration dependent inhibition of cell proliferation of Human Epidermoid Larynx Carcinoma Cell Line (HEP-2). This study has demonstrated characterization of melanin from basidiomycetes mushroom fungus, Schizophyllum commune and its applications.

The Mycobiota of Air Inside and Outside the Meju Fermentation Room and the Origin of Meju Fungi

The fungi on Meju are known to play an important role as degrader of macromolecule of soybeans. In order to elucidate the origin of fungi on traditional Meju, mycobiota of the air both inside and outside traditional Meju fermentation rooms was examined. From 11 samples of air collected from inside and outside of 7 Meju fermentation rooms, 37 genera and 90 species of fungi were identified. In outside air of the fermentation room, Cladosporium sp. and Cladosporium cladosporioides were the dominant species, followed by Cladosporium tenuissimum, Eurotium sp., Phoma sp., Sistotrema brinkmannii, Alternaria sp., Aspergillus fumigatus, Schizophyllum commune, and Penicillium glabrum. In inside air of the fermentation room, Cladosporium sp., Aspergillus oryzae, Penicillium chrysogenum, Asp. nidulans, Aspergillus sp., Cla. cladosporioides, Eurotium sp., Penicillium sp., Cla. tenuissimum, Asp. niger, Eur. herbariorum, Asp. sydowii, and Eur. repens were collected with high frequency. The concentrations of the genera Aspergillus, Eurotium, and Penicillium were significantly higher in inside air than outside air. From this result and those of previous reports, the origin of fungi present on Meju was inferred. Of the dominant fungal species present on Meju, Lichtheimia ramosa, Mucor circinelloides, Mucor racemosus, and Scopulariopsis brevicaulis are thought to be originated from outside air, because these species are not or are rarely isolated from rice straw and soybean; however, they were detected outside air of fermentation room and are species commonly found in indoor environments. However, Asp. oryzae, Pen. polonicum, Eur. repens, Pen. solitum, and Eur. chevalieri, which are frequently found on Meju, are common in rice straw and could be transferred from rice straw to Meju. The fungi grow and produce abundant spores during Meju fermentation, and after the spores accumulate in the air of fermentation room, they could influence mycobiota of Meju fermentation in the following year. This could explain why concentrations of the genera Aspergillus, Eurotium, and Penicillium are much higher inside than outside of the fermentation rooms.

Optimization of manganese peroxidase production from Schizophyllum sp. F17 in solid-state fermentation of agro-industrial residues / 生物工程学报

Manganese peroxidase (MnP), a crucial enzyme in lignin degradation, has wide potential applications in environmental protection. However, large-scale industrial application of this enzyme is limited due to several factors primarily related to cost and availability. Special attention has been paid to the production of MnP from inexpensive sources, such as lignocellulosic residues, using solid-state fermentation (SSF) systems. In the present study, a suitable SSF medium for the production of MnP by Schizophyllum sp. F17 from agro-industrial residues has been optimized. The mixed solid medium, comprising pine sawdust, rice straw, and soybean powder at a ratio of 0.52:0.15:0.33, conferred a maximum enzyme activity of 11.18 U/g on the sixth day of SSF. The results show that the use of wastes such as pine sawdust and rice straw makes the enzyme production more economical as well as helps solve environmental problems.

Molecular Identification of Schizophyllum commune as a Cause of Allergic Fungal Sinusitis

Schizophyllum commune, a basidiomycetous fungus, rarely causes disease in humans. We report a rare case of allergic fungal sinusitis caused by S. commune in a 14-yr-old girl. The patient presented with nasal obstruction and a purulent nasal discharge. Materials obtained during endoscopic surgery of the frontal recess revealed allergic mucin and a few fungal hyphae. A potato dextrose agar (PDA) culture from the allergic mucin yielded a rapidly growing white woolly mold. Although no distinctive features including hyphae bearing spicules or a clamp connection were present, the case isolate disclosed compatible mycological features including growth at 37degrees C, susceptibility to cycloheximide, and production of a tart and disagreeable smell. S. commune was confirmed by sequence analysis of the internal transcribed spacer region and D1/D2 regions of the 26S ribosomal DNA. We believe this is the first report of allergic fungal sinusitis caused by S. commune in Korea. Moreover, this report highlights the value of gene sequencing as an identification tool for non-sporulating isolates of S. commune.

Schizophyllum commune sinusitis in an immunocompetent host.

Schizophyllum commune is widely distributed in the nature, but it rarely causes human infection. We have isolated this mould in a 46-year-old immunocompetent, non-diabetic patient with chronic sinusitis, previously treated with multiple antibiotics and topical steroid nasal drops with no response. Materials obtained from the nasal sinus during the endoscopic surgery, on KOH mount and histopathological study revealed broad septed hyaline hyphae. Growth on the Sabouraud's dextrose agar and potato dextrose agar produced white moulds with microscopic and macroscopic characters of S. commune. Till date there are few reports of S. commune sinusitis in immunocompetent individuals Worldwide. This is the first reported case in India to the best of our knowledge.

Growth and Cultural Characteristics of Ophiocordyceps longissima Collected in Korea

We investigated the effect of nutritional and environmental factors on Ophiocordyceps longissima mycelial growth. The longest colony diameter was observed on Schizophyllum (mushroom) genetics complete medium plus yeast extract, Schizophyllum (mushroom) genetics minimal medium, and Sabouraud dextrose agar (SDA); however, malt-extract yeast-extract agar, SDA plus yeast extract, yeast-extract malt-extract peptone dextrose agar, SDA, oatmeal agar, and potato dextrose agar showed higher mycelia density. A temperature of 25degrees C was optimum and 7.0 was the optimum pH for mycelial growth. Colony diameter was similar under light and dark conditions. Maltose and yeast extract showed the highest mycelial growth among carbon and nitrogen sources respectively. The effect of mineral salts was less obvious; however, K3PO4 showed slightly better growth than that of the other mineral salts tested. Among all nutrition sources tested, complex organic nitrogen sources such as yeast extract, peptone, and tryptone were best for mycelial growth of O. longissima. Ophiocordyceps longissima composite medium, formulated by adding maltose (2% w/v), yeast extract (1% w/v), and K3PO4 (0.05% w/v) resulted in slightly longer colony diameter. In vitro mycelial O. longissima growth was sustainable and the production of fruiting bodies could be used for commercial purposes in the future.

Producción de inóculo de cepas nativas del hongo comestible Asam (Schizophyllum commune Fr) en diferentes sustratos y temperaturas / Inoculum production of edible mushroom native strains (Schizophyllum commune Fr) Asam on different substrates and temperatures

En este estudio se evaluó la producción de inóculo de cinco cepas nativas de S. commune, utilizando granos de sorgo, trigo, cebada y arroz, los cuales fueron inoculados con el micelio de cada una de las cepas, incubándose tanto a 26 oC como a 18 oC. Se determinó que el sustrato donde se obtuvieron los menores tiempos de producción de inóculo fue el trigo, seguido de sorgo, cebada y arroz, tanto a 18 oC como 26 oC, observándose diferentes diferencia significativa (p <0.001 solamente entre el primero respecto a los demás. Las cepas con menores tiempos de producción fueron la...

Growth and Cultural Characteristics of Cordyceps cardinalis Collected from Korea

Cordyceps cardinalis was reported in Japan and the USA in 2004, and its fruiting bodies have recently been cultured in Korea. Herbarium specimens preserved at the Cordyceps Research Institute, Mushtech, Korea were revised and identified as C. cardinalis, based on morphological characters and conidial structures. Most of the C. cardinalis specimens were collected from Mt. Halla in Jeju-do. The effects of various nutritional sources and environmental conditions such as temperature and pH on mycelial growth of C. cardinalis were studied. Oatmeal agar, Martin's peptone dextrose agar, and Schizophyllum (mushroom) genetics complete medium plus yeast extract resulted in the best mycelial growth. Among carbon sources, cereals, and nitrogen sources, maltose, oatmeal, and peptone resulted in the best mycelial growth respectively. Mineral salts helped to increase growth rate but only resulted in thin mycelial density, similar to water agar. A temperature of 25degrees C and a pH of 7 resulted in the highest mycelial growth. Based on these results, a Cordyceps cardinalis composite medium (CCM) was formulated with 1% maltose, 2% oatmeal, 1% peptone, and 2% agar. Use of the CCM resulted in slightly better mycelial growth than that of other commonly used agar media. Only organic nitrogen sources imparted a reddish pigmentation to the agar media, but this character diminished after several subcultures. A 7 day culture duration resulted in the best mycelial growth.

Cultural Conditions for Mycelial Growth and Molecular Phylogenetic Relationship in Different Wild Strains of Schizophyllum commune

The common split-gilled mushroom, Schizophyllum commune is found throughout the world on woody plants. This study was initiated to evaluate conditions for favorable vegetative growth and to determine molecular phylogenetic relationship in twelve different strains of S. commune. A suitable temperature for mycelial growth was obtained at 30degrees C. This mushroom grew well in acidic conditions and pH 5 was the most favorable. Hamada, glucose peptone, Hennerberg, potato dextrose agar and yeast malt extract were favorable media for growing mycelia, while Lilly and glucose tryptone were unfavorable. Dextrin was the best and lactose was the less effective carbon source. The most suitable nitrogen sources were calcium nitrate, glycine, and potassium nitrate, whereas ammonium phosphate and histidine were the least effective for the mycelial growth of S. commune. The genetic diversity of each strain was investigated in order to identify them. The internal transcribed spacer (ITS) regions of rDNA were amplified using PCR. The size of the ITS1 and ITS2 regions of rDNA from the different strains varied from 129 to 143 bp and 241 to 243 bp, respectively. The sequence of ITS1 was more variable than that of ITS2, while the 5.8S sequences were identical. A phylogenetic tree of the ITS region sequences indicated that the selected strains were classified into three clusters. The reciprocal homologies of the ITS region sequences ranged from 99 to 100%. The strains were also analyzed by random amplification of polymorphic DNA (RAPD) with 20 arbitrary primers. Twelve primers efficiently amplified the genomic DNA. The number of amplified bands varied depending on the primers used or the strains tested. The average number of polymorphic bands observed per primer was 4.5. The size of polymorphic fragments was obtained in the range of 0.2 to 2.3 kb. These results indicate that the RAPD technique is well suited for detecting the genetic diversity in the S. commune strains tested.

Characteristics of Metacordyceps yongmunensis, a New Species from Korea

Metacordyceps yongmunensis is a newly reported species from Korea, which is very similar to Cordyceps species in morphological characters. It grows on large lepidopteran pupa, and numerous white stromata grow on a single host. Mycelial growth characteristics of M. yongmunensis isolates were studied in different media and at different temperatures. Also, different carbon sources, nitrogen sources, and mineral salts were tested for mycelial growth of M. yongmunensis. Schizophyllum (mushroom) genetics complete medium plus yeast extract, Schizophyllum (mushroom) genetics minimal medium, and Martin's peptone dextrose agar produced longer colony diameters and more compact mycelial density than other media. The optimum temperature for mycelial growth was 25degrees C. Carbon sources such as sucrose, soluble starch, dextrose, glucose, dextrin, maltose, and fructose showed better mycelial growth, whereas peptone, yeast extract and tryptone resulted in the best mycelial growth of all of the nitrogen sources tested. All of the mineral salts tested showed similar growth as the control, except K2HPO4 which showed longer colony diameter and more compact mycelial density. The compact colonies were white and cottony with a greenish margin. The results showed that M. yongmunensis is an easy fungus to growas it grew from 30 to more than 50 mm in 2 wk.

Cultural Characteristics of Shimizuomyces paradoxus Collected from Korea

This study investigated the cultural characteristics of Shimizuomyces paradoxus in different nutritional and environmental conditions. The highest mycelial growth was observed in Schizophyllum (mushroom) genetics complete medium plus yeast extract agar medium, and the optimal temperature and pH were 25degrees C and pH 8.0, respectively. The optimal carbon and nitrogen sources were 1% dextrose and 1% peptone in agar. However, in liquid culture the highest dry mycelium weight was found for the potato dextrose agar and potato sucrose agar broths. The optimum inoculum size was five mycelial discs (5 mm) per 100 mL of broth, and the optimum liquid culture period was 25 days. This is the first ever report of S. paradoxus cultural characteristics.

Optimization of process variables for the manganese peroxidase of the white-rot fungus Schizophyllum sp. F17 by full factorial central composite design / 生物工程学报

White-rot fungus manganese peroxidase (MnP) that has great potential in degrading azo dyes is one of the extracellular glycolsylated heme proteins. MnP from Schizophyllum sp. F17 was isolated and purified by Sephadex G-75 gel filtration chromatography followed by DEAE-cellulose anion exchange chromatography. The molecular weight of the puried enzyme was 49.2 kDa, while the half-life of the MnP in the presence of 0.1 mmol/L H2O2 was 5-6 min. The efficiency of MnP-catalyzed reactions were determined by three key factors: the concentrations of Mn2+, H2O2, and the amount of MnP. Using single factor analysis, an optimized concentration of Mn2+, H2O2 and enzyme were optimized to be 1.2 mmol/L, 0.1 mmol/L, and 0.4 mL, respectively. A response surface methodology (RSM) employing two-level-three-factor full factorial central composite design was used to optimize the catalytic conditions. The result showed that the concentration of H2O2 and the interaction between H2O2 and MnP mostly affect the MnP catalytic efficiency. Finally, we show that the azo dyes could be efficiently decolorized by the purified MnP under optimized conditions.

Physicochemical Requirement for the Vegetative Growth of Schizophyllum commune Collected from Different Ecological Origins

Schizophyllum commune is an edible and medicinal mushroom widely distributed in the world. The optimal growth conditions for the mycelia of 10 strains of the fungus were investigated. The temperature suitable for the mycelial growth and density was obtained at 30~35degrees C. Among the tested conditions, the minimum mycelial growth was found at 15degrees C. In case of pH, the most favorable growth was found at pH 5. The results indicated that this mushroom well adapted to high temperature and low pH for its mycelial growth. Considering growth phenotype of mycelia, Hamada, Hennerberg, PDA and YM were the most suitable and Lilly, Glucose triptone, Glucose peptone and Hoppkins were the most unfavorable among tested media for the mycelial growth of S. commune. Out of tested carbon sources, dextrin and fructose were the most suitable and lactose, mannose and sorbitol were the unsuitable for the fungus. Compact mycelial density was obtained from most of the carbon sources. Among used nitrogen sources, calcium nitrate, potassium nitrate and alanine were the most appropriate and the most incompatible were ammonium phosphate, histidine, urea and arginine for mycelial growth of S. commune on the culture media. Calcium nitrate, histidine and potassium nitrate showed moderately thin or thin, and rest of nitrogen sources showed compact or moderately compact mycelial density.

Studies on the properties and co-immobilization of manganese peroxidase / 生物工程学报

White-rot fungus manganese peroxidase (MnP) oxidizes a wide range of substrates, rendering it an interesting enzyme for potential applications. The stability of MnP can be improved by immobilization. With sodium alginate, gelatin, or chitosan as a carrier, and glutaraldehyde as the crosslinking agent, MnP was co-immobilized using the embed-crosslinked method and the adsorb-crosslinked method. The immobilization conditions and the partial properties of the three immobilized enzymes were investigated. When compared with the free enzyme, the optimum pH values and the temperatures of the three immobilized MnPs carried by alginate, gelatin, and chitosan were respectively shifted from 7.0 to 5.0, 5.0, 3.0 and from 35 degrees C to 75 degrees C , 55 degrees , 75 degrees C . The thermostabilities of the three immobilized MnPs were considerably better than that of the native enzyme. The chitosan-decreased by less than 5% even after repeated use for 6 - 9 times. The ability of decolorizing azo dyes in static and shaky situation by gelatin-immobilized MnP approached to the free enzyme, and there was no loss of enzyme activity during 2 repeated batch reactions.

T-DNA transfer from Agrobacterium tumefaciens to the ectomycorrhizal fungus Pisolithus microcarpus / Transferencia de T-DNA de Agrobacterium tumefaciens al hongo ectomicorrícico Pisolithus microcarpus

The model ectomycorrhizal fungus Pisolithus microcarpus isolate 441 was transformed by using Agrobacterium tumefaciens LBA1100 and AGL-1. The selection marker was the Shble gene of Streptoallotecius hidustanus, conferring resistance to phleomycin, under the control of the gpd gene promoter and terminator of Schizophyllum commune. Transformation resulted in phleomycin resistant clones which were confirmed by PCR to contain the resistance cassette. A. tumefaciens-mediated gene transfer would allow the development of RNA interference technology in P. microcarpus.

El hongo ectomicorrícico modelo Pisolithus microcarpus aislamiento 441 fue transformado utilizando Agrobacterium tumefaciens LBA 1100 y AGL-1. El marcador de selección fue el gen Shble de Streptoallotecius hidustanus, el cual confiere resistencia a fleomicina, bajo el control del promotor y terminador del gen gpd de Schizophyllum commune. La transformación resultó en clones resistentes a fleomicina comprobándose por PCR la presencia del transgen. La transferencia génica mediada por Agrobacterium podría permitir el desarrollo de la tecnología de interferencia por ARN en P. microcarpus.

Transformation of the Edible Basidiomycete, Pleurotus ostreatus to Phleomycin Resistance

For transformation of Pleurotus ostreatus, two novel vectors, pPhKM1 and pPhKM2, were constructed, using the regulatory sequences of the P. sajor-caju beta-tubulin gene (TUB1) and the ble gene encoding phleomycin binding protein. pPhKM1 contains ble fused to the TUB1 promoter and the Schizophyllum commune GPD terminator. pPhKM2 contains ble fused to the promoter and terminator regions of P. sajor-caju TUB1. To confirm phleomycin-resistance activity, each vector was cotransformed with pTRura3-2 into the P. ostreatus homokaryotic ura - strain. The transforming DNA was stably integrated into the genomic DNA. Subsequently, phleomycin resistance was conferred on wild-type dikaryotic P. ostreatus by transformation with pPhKM1 or pPhKM2. This transformation system generated stable phleomycin-resistant transformants.

Fill up to the antineoplastic fungi group in Vietnam: Schizophyllum commune

Schizophyllum commune (Fr:Fr.) rick were collected from various regions in Vietnam, thoroughly described and isolated for fruitful cultivations and mycelial fermentations to supply precious material for antitumor preparations