Earticle

This experiment was carried out to clarify the effect of light qualities on the growth characteristics and yield of fruiting body in the cultivation of Lyophyllum ulmarium. The intensity of illumination by light qualities was in the order of white light(2,270Lux), yellow light(1,750Lux), blue light(460Lux) and red light(400Lux). An investigation of fruiting body showed these results that the pileus size and stipe diameter of fruiting body on CBM(Chungbuk mushroom)-1757 were much larger than Hypsizigus marmoreus, and an effect of yellow light seemed to be better than those of another light. In comparison with Hypsizigus marmoreus, the growth duration of CBM-1757 was shortened by 8 days which included 2 days for mycelial culture, 1 day for first pinning requirement and 1 day for growth. The growth duration in yellow light illumination was about 70 days showing the tendency of 2~4 days reduction. There were no differences in results such as number of effective stem and fresh weight. The yield of fruiting body per bottle in CBM-1757(95.6g) was little higher than Hypsizigus marmoreus(94.8g). By a white light’s standard, the yields of blue and red light illumination were decreased by 2~9%, but that of yellow light illumination was increased by 8%. The chromaticity results showed that brightness, red and yellow coloration of CBM-1757 were higher than those of Hypsizigus marmoreus, and yellow light treatment was more effective than another light.

This study was conducted to investigate the physiological activities of the ethanol extracts from 10 different strains of Pholiota nameko. In addition, the β-glucan and polyphenol contents from these strains were also measured. Total polyphenol contents from all the strains were more than 40 mg% with the highest content of 61.50±0.59 mg% and β-glucan contents were 30% with the highest content of 37.20±1.12%. The highest inhibitory activities on α-amyloglucosidase activity and nitric oxide production were 13.78±0.56% and 56.59±7.11%, respectively. However, ethanol extracts from all the strains have little effects on DPPH radical and nitrite scavenging rates and the activity of angiotensin I converting enzyme. The cytotoxic activity of ethanol extracts from all the strains on A549 cell was shown upto 30% with the highest effect of 47.96±8.46% treated at 1 mg/mL concentration.

This study was carried out to investigate the physiological activities of the ethanol extracts from 11 strains of Flammulina velutipes. In addition, the β-glucan and polyphenol contents were also measured. The contents of polyphenol and β-glucan of Flammulina velutipes were found to be more than 100 mg% and 20%, respectively, one of which showed 244.74 mg% and 27.37%, respectively. Further, The highest inhibitory activities on DPPH radical, α-amyloglucosidase activity and nitric oxide production were 91.74%, 63.57% and 61.44%, respectively. However, ethanol extracts from all the strains have little effects on Angiotensin I converting enzyme(ACE)-inhibitory effects. Finally, the highest cytotoxic activities of ethanol extracts from all the strains on A549 and HT-29 cell were 76.07% and 67.05%, respectively.

Agaricus blazei Murill is a important medicinal mushroom for a powerful immune system builder and tonic. Currently, it is known about a new disease phenomenon that appears to be occurring on a number of mushroom farms. We described a straightforward approach in which molecular methods was used to survey the presence of potentially endo- and epiphytic bacteria infected with the Agaricus blazei. The 16S rDNA was amplified with universal eubacterial primers directly from pure cultures of Agaricus blazei mycelium and fruit body. The 16S rDNA sequences were almost identical (96 to 97% similarity), and phylogenetic analysis placed them in a single unique rRNA branch belong to the uncultural bacterium phylogroup. PCR detection of uncultural bacterium in the malformed tissues of Agaricus blazei were carried out by using 16S rRNA sequenced specific probe. It was strongly amplified at the malformed pileus region of fruit body and also spore print was impossible.

For the development of a sporeless strain of P. ostreatus we used sporeless strain ASI 2069. We have recovered both nuclear types of strain ASI 2069 as monokaryons of nh9, nh15, nh26 and nh36 (here after referred to as neohaplonts) by protoplasting the mycelium. Crosses between neohaplonts and SSI’s(single spore isolates) obtained from a sporulating commercial strain ASI 2180. Five excellent strains are selected from 30 bred strains by quality of fruitbodies and spore number. To development of molecular markers linked to sporeless strain of P. ostreatus, we are screened helicase, recombinase(DMC1) and topoisomerase(Spo11) genes related meiosis by PCR and sequencing. Among three genes, Spo11 gene was identified into molecular marker of sporeless from neohaplonts and bred strains of P. ostreatus.

Related research on artificial cultivation of the edible fungi in modern China can be traced back to 100 years ago. The earliest article about the cultivation techniques of the edible fungi was published in 1897 on Agricultural Study Newspaper sponsored by Shanghai Agricultural Society. From the late 1800s to 1940s, the elder generation of scholars including Zou Bingwen, Hu Changzhi, Pan Zhinong, Li Shiyi, Sun Yunwei, and Yu Xiaotie, etc. not only introduced many foreign techniques in edible fungi cultivation a nd disseminated scientific knowledge and cultivation techniques, but also held various edible fungi talents training courses, set up experimental bases, and conduct edible fungi cultivation experiments. Those have laid a preliminary foundation for the modernization of China’s edible fungi cultivation techniques. After the founding of the PRC, the edible fungi cultivation industry has gained more space for development, and has achieved many milestone achievements, mainly including the tremella artificial cultivation technique, the hedgehog hydnum artificial cultivation technique, the Xianggu artificial cultivation with crushed-wood material technique, the white mushroom fine breed selection and breeding and cultivation technique improvement, the black fungus cultivation with crushed-wood material and fine breed selection and breeding, and the golden needle mushroom find breed selection and year-round cultivation technique innovation, as well as the acclimatization of wild edible fungi and the development of new varieties of edible fungi. These inventions and innovations have provided solid technical support to the development of the edible fungi industry in China. The reform and opening-up starting 1978 has provided a favorable policy environment for the development of the edible fungi industry in China. In the period of more than 30 years thereafter, the edible fungi industry in China has been developing rapidly, with the annual yield rocketing from 60,000 tons in 1978 to 20.2 million tons in 2009, and the proportion of the yield against total world yield growing from 5% in the past to more than 70% at present. During this historical period, many research institutions and scientific research staffs have made important contributions to the development of the edible fungi industry in China. Among them, the most important achievements are made by Researcher Chen Meipeng from the Institute of Edible Fungi of Shanghai Academy of Agricultural Sciences, Professor Yang Xinmei from Huazhong Agricultural University, Researcher Huang Nianlai from Fujian Sanming Mycological Institute, and Professor Zhang Shuting from Chinese University of Hong Kong. When we look back, we treasure the outstanding achievements made by the elder generation of scholars on the development of the edible fungi industry in China. When we look into the future, we are geared with enthusiasm and confidence, and we will work hard to achieve higher in the edible fungi industry in China.

The genes encoding cellulases, which belong to glycosyl hydrolase families have been cloned from the basidiomycetous mushrooms. The transcripts of cellulase genes are strongly induced when the mycelia are grown in medium containing crystalline cellulose, and they are not expressed in medium containing glucose, but how insoluble substrates such as microcrystalline cellulose are recognized by these fungal cells is not clear. The polypore mushroom Polyporus arcularius is a wood-decomposing basidiomycete that produces at least three types (I, II, and IIIa) of carboxymethyl cellulase (CMCase) when the medium contains crystalline cellulose as the sole carbon source and produced mainly cellobiose in the medium. The genomic and cDNA clones encoding the family 12 endoglucanase (CMCase IIIa) gene (cel3A) of P. arcularius have been sequenced, and Cel3A has been expressed as an active enzyme in Escherichia coli. To determine the role and function of each type of cellulase in the degradation of crystalline cellulose by basidiomycetous mushrooms, the structure of all of the cellulase genes should be investigated, but the nucleotide sequences of the other cellulase genes in P. arcularius have not yet been reported. In the current study, the genomic and cDNA clones encoding the endoglucanases (cel4), and the two cellobiohydrolases (cel1 and cel2) of P. arcularius sequenced and characterized. The predicted amino acid sequence of Cel1 Cel2, Cel3a and Cel4 are similar to glycosyl hydrolase family 7, 6 12 and 5 protein, respectively. The expressions of the all cellulase genes (cel1 cel2, cel3a and cel4) were induced by Avicel (microcrystalline cellulose) and cellopentaose but repressed by glucose, cellobiose, cellotriose, and cellotetraose. There was a low level of transcription of both genes regardless of the carbon source. These results suggest that P. arcularius cells constitutively express a very low level of cellulase that can degrade insoluble crystalline cellulose and that the transcription of celluases in the cells is induced by products produced by these endoglucanases such as cellooligosaccharides. From our findings, we propose a possible mechanism for the recognition and degradation of insoluble crystalline cellulose by fungal cells.

Basidiomycetes can degrade lignocellulosic biomass, and some basidiomycetes produce alcohol dehydrogenase, so it is feasible to produce alcohol from basidiomycetes. Agaricus blazei, Flammulina velutipes and Tricholoma matsutake have been used for mushroom fermentation to produce alcohol. To investigate whether Pholilta nameko can be used for mushroom fermentation, and find out the relationship between mannitol-1-phosphate dehydrogenase and alcohol dehydrogenase, we cloned mannitol-1-phosphate dehydrogenase gene from P. nameko, which is a zinc-containing long- chain alcohol dehydrogenase. Mpd, the gene encoding mannitol-1-phosphate dehydrogenase (MPD), has been sequenced and characterized from basiodiomycete P. nameko. The length of the coding region is 1360bp. The gene encodes a putative protein of 359 amino acids; predicted protein molecular weight is 38.6 kDa and an isoelectric point is PI = 7.34. The locations of exons and introns in the gene were deduced on the basis of interruptions in the amino acid sequence that were homologous to those in the MPD of Laccaria bicolor, the coding region was split into 6 exons and 5 introns. The protein deduced from the gene MPD showed more than 46% sequence identity to 20 fungal MPDs or alcohol dehydrogenases documented in the Gene bank protein database, based on BLASTP analysis, and was phylogenetically close to the MPDs of L. bicolor and Coprinopsis cinerea. This protein shared the same conserved domain with the alcohol dehydrogenase.

Phenotypic traits (physiological characteristics and somatic incompatibility) and genotypic traits (target region amplification polymorphism TRAP) were used to survey the diversity in Chinese Auricularia auricula systematically, which consisted of 32 main cultivated strains. The 27 important and stable physiological indexes were evaluated; somatic incompatibility test (SIT) reaction was described from three aspects: type, pigment, and intensity while intensity clustering alone revealed the phenotypic diversity among the 32 strains;16 pairs of TRAP primer combinations produced 535 unambiguous and reproducible DNA fragments, of these 524 (97.9%) were polymorphic. The phylogenetic trees were constructed by Unweighted Pair-group Method with Arithmetic Averages (UPGMA), which distributed the test strains into four to six major groups respectively. The principal coordinate analysis (PCO) of the three methods (physiological characteristics, SIT intensity and TRAP) exhibited highly similar clustering patterns, revealing that all the test strains can be divided into three groups considered significantly correlated with geographical regions. Results revealed the occurrence of relatively low diversity of A. auricula in the study. The cultivated strains in the same region are more similar physiologically and some strains are suspected to be synonymous. This study suggests that the value for estimating diversity are represented by TRAP＞SIT reaction intensity＞physiological characteristics in A. auricula, and PCO analysis can provided more effective and visible information than the UPGMA clustering dendrogram. Our finding will facilitate future germplasm resources management and breeding program of A. auricula in China.

Effect of trehalose for the mycelial growth of Tricholoma matsutake were examined. When T. matsutake Z-1 strain was cultured in the partially modified matsutake liquid (PMML) medium and the Hamada matsutake liquid (HML) medium supplemented with trehalose at 24 ℃ for 80days, the vegetative mycelial dry weights showed higher value compared with those of PMML medium and HML medium supplemented with glucose (control). The range of the effect of 1.0-8.0% carbohydrate substrate on vegetative mycelial growth was investigated. The optimal concentration for mycelial growth was 2.0% for the glucose medium but 8.0% for the trehalose medium. To evaluate the potential of the production of trehalase from T. matsutake, the extracellular trehalase activity during the vegetative mycelial growth was measured. The activity of the extracellular trehalase increased during vegetative mycelial growth of T. matsutake and was maximal 70 days after inoculation. This extracellular enzyme was investigated for the purification and the characterization. The partially purified trehalase was obtained from about 1.53l static culture filtrate, with 19.1% recovery, and about 2,940 fold purification. The molecular mass was about 62.6kDa (SDS-PAGE) and 70kDa (Gel-filtration). The enzyme was most active around 40℃ and pH 5.0 and stable over a pH of 4.5~ 6.5 for 30min at 37℃. The enzyme readily hydrolyzed trehalose having α -1,1 glucosidic bond. However, it did not hydrolyze disaccharides such as maltose, iso-maltose, cellobiose, saccharose and lactose.

Flammulina velutipes, amongst others known as Winter Mushroom or Enokitake is an important economic crop in Asia. The tetrapolarity (having four mating types) of this mushroom obligates mating and results in self-sterile progeny that carries unique genetic traits, making understanding of the genetic base desirable for breeding. Moreover, mating type genes are significant for evolutionary studies as their high polymorphism benefits phylogenetic comparisons. This polymorphism further makes mating type genes interesting candidates for genetic markers that allow identification of specific strains. Mating type loci in Agaricomycotina are classically termed A and B and control two different developmental pathways [for a review see 1]. They consist of tightly linked subloci that encode multiallelic genes. MatA loci contain two groups of divergently transcribed homeodomain proteins (HD1 and HD2) and heterodimerization of HD1 with a non-self HD2 protein forms a functional transcription factor that activates the A pathway. MatB loci hold pheromones and pheromone receptors. Pheromone genes encode small precursor proteins that are characterized by a C-terminal CAAX motif. Pheromone receptors typically contain 7 membrane spanning regions and are coupled to G-proteins. Binding of a pheromone to a receptor, triggers splicing of the (trimeric) G-protein, which activates the B pathway. New genetic data from recent genome sequences is challenging the strict concepts of old mating type models in fungi. MatB loci turn out to be rather diverse and contain considerable varying numbers of pheromone receptors and associated pheromones. To this, pheromone receptors which are not linked to matB loci have now been reported for C. cincerea, S. commune and L. bicolor [2, 3]. Also the organization of the matA locus is less strictly conserved than anticipated. Though far more tightly maintained than the matB locus, substantial differences in HD gene numbers and overall organization are reported [2, 3]. These differences stress the importance of determination of the individual mating type systems from industrially important mushrooms to assist breeding. Our analysis of F. velutipes strain 4019-20 uncovered 7 pheromone receptors together with 3 pheromones. The matB-3 locus of this strain however, is defined by only a single pheromone receptor and pheromone gene and our data strongly indicates that a 2nd pheromone receptor recently lost its function. The other receptor genes are non mating type specific. Finally, we detected three homeodomain genes distributed over two distant subloci. These subloci have been separated by two large inversions. Strikingly the distant matA subloci in S. commune seem to be separated by inversions as well. Synthenic mapping of a large regions from Coprinus cinerea, Laccaria bicolor, S. commune and F. velutipes shows that the matA loci originate from a single locus in a common ancestor of S. commune and F. velutipes that is represented by L. bicolor and C. cinerea. [1] U Kües. Micr Mol Biol Rev 64, 316 (2000) [2] H Niculita-Hirzel, J Labbé, A Kohler, F le Tacon, F Martin, IR Sanders and U Kües. New Phytol 180, 329 (2008). [3] RA Ohm, JF de Jong, LG Lugones, A aerts, E Kothe, JE Stajich, RP de Vries, E Record, A Levasseur, SE Baker, KA Bartholomew, PM Couthino, S Erdmann, TJ Fowler, AC Gathman, V Lombard, B Henrissat, N Knabe, U Kües, WW Lilly, E Lindquist, S Lucas, JK Magnuson, F Piumi, M Rausdaskoski, A Salamov, J Schmutz, FWMr Schwarze, PA van Kuyk, JS Horton, IV Grigoriev and HAB Wösten. Nat. Biotechnol ISSN: 1087-0156 (2010).

Ceriporiopsis subvermispora is a unique white rot fungus degrading plant cell wall lignin without severe loss of cellulose. Recombinant plasmids containing homologous gene expression signals fused to the coding sequence of Escherichia coli hph which encodes for hygromycin phosphotransferase were introduced in protoplasts of wild type C. subvermispora using PEG/CaCl2 protocol. A number of hygromycin B resistant strains were isolated on a screening plate containing 100㎍/ml of hygromycin B: whereas, no colonies were observed when protoplasts were treated with no DNA as a negative control. It was demonstrated that most of the isolates lost their drug resistance during successive cultivations in the presence of the same concentration of hygromycin B, but some of the isolates showed stable drug resistance after five times repeated screening. They did not lose the drug resistance even after the cultivation in the absence of hygromycin B and incorporation of the hph sequence was confirmed by specific amplification of the target sequence in PCR experiments and Southern hybridization analysis. The stable transformation system will make it possible to do molecular genetic analysis, as well as breeding of genetically modified strains, in C. subvermispora. Moreover, it was demonstrated that recombinant constructs with truncated promoter showed reduced number of the drug resistant isolates on the first screening plate, in response with the length of the remaining promoter sequence. These findings indicated that unstable drug resistance observed in these isolates should originate from transient expression of the introducing marker genes, and that a promoter assay system has been developed for the first time in basidiomycete. This system is practically not affected by the positional effect of the integrated recombinant gene in the host chromosome.

Gamma-Amino Butyric Acid (GABA), a four-carbon non protein amino acid, is widely distributed in nature and acts as the major suppressive neurotransmitter in the mammalian central nervous system. Recently, GABA has been reported to have several physiological functions such as antihypertensive, diuretic, relaxing and antidiabetic effects. Due to these unique biological functions, GABA has been used as functional ingredients for functional foods. Glutamic Acid Decarboxylase (GAD), which catalyzes decarboxylation of L-glutamic acid to GABA, has been purified from mammals, higher plants, and some microorganisms and their properties have already been reported. On the other hand, mushrooms are commonly appreciated as healthy food due to their nutritional properties, such as low calorie, rich in fiber, vitamin and mineral. L-Glutamic acid and GABA are also commonly distributed in edible mushrooms. The fact that mushrooms accumulate GABA suggests the existence of GAD. However, the enzyme property of mushroom GAD has not been reported. In the present study, we tried to purify and characterize the property of Flammulina veltipes. [Methods and Results] Mushroom (fruit-body of Flammulina veltipes) used in this study was purchased in local market. GAD activity was determined by formation of GABA from L-glutamic acid in the presence of pyridoxal-5’š- phosphate (PLP). The GAD activity was determined by the formation of GABA from L-glutamic acid. Fruit- body was crushed and then centrifuged to separate supernatant and cell debris. To investigate the localization of GAD, both supernatant and precipitate were subjected to enzyme reaction after dialysis. The cell debris showed stronger GAD activity than that of supernatant. The formation of GABA was observed between pH 4 and 6 and the maximum GAD activity was observed at pH 6. However, GAD activity was lost after dialysis for overnight against buffer of pH 6-11. These results suggest that GAD from Flammulina veltipes is stable at pH 4-5 in spite of its optimum pH for GABA production is around 6.

Laccase (Lcc; EC 1.10.3.2) belongs to a group of polyphenol oxidases, which catalyzed the oxidation of single-electron from phenolic substrates or aromatic amines. Many organisms possess several lcc encoding genes, and their biological functions diverge into many branches. There are many studies on biochemical function of Lccs, however, there are few studies about biological functions of one Lcc in detail. We researched on biological function of Lcc1, which is most abundantly secreted enzyme from vegetative mycelia into liquid culture in L. edodes. In our previous study, lcc1 gene was down regulated by RNAi method in L. edodes, and then ivrL1#32 was selected as a completely lcc1 dowaregulated transformant. We revealed that fruiting body development in ivrL1#32 was significantly suppressed compared to wild type strain. In this study, we observed the hyphal morphology of ivrL1#32. IvrL1#32 did not form thick aerial mycelium mat when grown on MYPG agar plate. From the observation using scanning electron microscope (SEM), hyphae of ivrL1#32 had many abnormal short branches and their mycelial density was lower than that of wild type strain. From transmission electron microscope observation (TEM), ivrL1#32 lacked obviously distinguishable outer and inner layer in their cell wall. In addition, the fibrous layer of ivrL1#32, which connected hyphae, obviously decreased. These morphological phenotypes would be caused by the absence of Lcc1 in L. edodes. Our results provide a clue to resolve of the biological function of Lcc1 in L. edodes.

The waste substrate from sawdust based cultivation of Heicium erinaceum was reused. This process was conducted three times. Even when the waste substrate was reused at three times, the yield of fruiting bodies was equal to that of fresh medium. However, the yield of the 1st-waste substrate was the best of all waste substrate media and the yields of waste substrate media deceased with recycling times. The yield of the 1st or the 2nd waste substrate medium increased by 1.3-1.4 times compared with that of the fresh medium. The content of low molecular α-glucan and β-glucan of the 1st or the 2nd waste substrate medium increased and C-N ratio of the 1st or the 2nd waste substrate medium decreased. These results suggest that low molecular glucan and N sources contribute to increasing fruiting bodies. It was clear that the 1st and the 2nd waste substrate were useful for the cultivation material of Heicium erinaceum.

Rhizopogon roseolus (Corda) Th. M. Fr. (=R. rubescens Tul. & Tul.), known as “shoro” in Japan, is a hypogeous basidiomycete that is an important ectomycorrhizal symbiont of Pinaceae. In order to cultivate this edible ectomycorrhizal mushroom, several researches have tried to promote mycorrhization of this mushroom on host Pinus thunbergii roots: Pine seedlings were inoculated with mycelium in vitro, or with crushed fruiting bodies in nature. However, successful cultivation of this mushroom has not been fully refined. We have developed the useful mycelial inoculum that enable to produced abundant ectomycorrhizas and then to form fruiting body under greenhouse nursery conditions. We selected the superior strain that rapidly colonized and produced a lot of ectomycorrhizas in root of P. thunbergii． The mycelial inoculum was composed of mineral solution and homogenate of mycelium that had been cultured in liquid medium. Addition of surfactant in the mycelial inoculum resulted in stimulation of mycorrhzal formations in host roots. When the mycelial inoculum containng surfactant were introduced to the mother plant system in which the colonized seedling had been planted into in the nursery, stimulatally effects were observed on not only mycorrhzation of the seedlings but also fruiting body formation. Genotype analysis using microsatellite markers for R. roseolus showed that fruiting bodies produced in the nursery were originated from the inoculated strain. These results suggest that the mycelial inoculum containg surfactant could be the model of mycelial spawn for “shoro”.

Poplar or pine sawdust is used to main substrates for oyster mushroom bottle cultivation. However, sawdusts are containing a lot of lignin that is insoluble and non digestible component. Therefore, spent substrates which are produced by oyster mushroom harvest are difficult to use as ruminant feed. In this study, we carried out to find sawdust substitute on the oyster mushroom for ruminant feed. As results of chemical analysis of cotton seed pellet, corn stem pellet, and corncob was revealed that concorb showed low crude ash(2.4%) and lignin(13.1%) contents. Mushroom yield and biological efficiency in concorb substrates is similar to and higher than that of control. Therefore we estimated that corncob could be substituted for sawdust.

World wide mushroom productions have been increased to 10-20% and more various mushrooms have been attempted to cultivate. Similar trends were also observed in Korea. More diverse mushroom varieties such as Pleurotus eryngii, Hericium erinaceus and Agrocybe aegerita have been attempted to cultivate in larger areas. In these days, 235 varieties of 33 different mushroom species have been cultivated. However only 50 varieties were protected by the law. Since 1960s, mushroom industry had been considered as one of the major export industries in Korea and mushroom production has been rapidly increased. In 2008, total mushroom production was about 198,209 ton, which were about 800 billion won (one trillion won if include mushroom factory products). Major cultivated species are Pleurotus ostreatus, P. eryngii, Flammulina velutipes, Lentinula edodes and Agaricus bisporus, which cover 90% of total production. We believed that the protection of mushroom vavarieties by the law is one of the main problems to be solved. Also, more studies to develop better mushroom production methods with low cost and improve the distribution structures are certainly required. In these days, the export of mushroom has been increased in Korea. We imported more mushrooms than exported for last 4 years, however, it has been changed that export is getting bigger in these days. Because we developed the liquid spawning and automatic cultivation systems, which lead the reduction of mushroom production cost and resulted in the increasement of export. If we develop better post-harvest system, it is certain that the mushroom export should be more important in the future. As an alternative way, some Korean companies are planning to build the plants in main export countries. Mushroom industry has promising future because mushroom is good for your health.

Comparative effect of oyster mushrooms on plasma, fecal lipid profiles, liver and kidney functions were evaluated in hyper and normocholesterolemic rats. The feeding of hypercholesterolemic rats with 5% powder of fruiting bodies of oyster mushrooms i.e., Pleurotus ostreatus, P. sajor-caju and P. florida reduced the plasma total cholesterol level by 37%, 21% and 16%, respectively and triglyceride level by 45%, 24% and 14%, respectively. LDL/HDL ratio decreased by 64%, 45% and 41% for P. sajor-caju, P. ostreatus and P. florida fed rats, respectively. Mushroom feeding also reduced body weight in hypercholesterolemic rats. However, it had no adverse effect on plasma bilirubin, creatinin and urea nitrogen level. Mushroom feeding also increased the total lipid and cholesterol excretion through the feces. The present study reveals that feeding of 5% oyster mushroom powder do not have detrimental effects on the liver and kidneys rather may provide health benefits for the cardiovascular-related complication by decreasing the atherogenic lipid profiles.

Phellinus sp. are assigned to the Basidiomycotina, Hymenomycetes, Aphyllophorales and Hymenochaetaceae, and have been shown containing various bioactive substances including triterpenoids, polysaccharides and flavones[1]. Traditional Chinese herbalists believe that Phellinus species are effective in treating many gynecopathic ailments[2] and are also reported to exhibit other pharmacological functions including tumor cell inhibition, antioxidant activity and anti-hepatic fibrosis effects[3]. Polysaccharides of Phellinus sp. has been reported to possess antitumor activities and inhibit tumor recrudescence and metastasis. There are little studies comparing the chemical composition and biological activities difference among polysaccharides from different Phellinus sp and little report about the pure polysaccharide structure analysis. In this study, eight kinds of crude polysaccharides were extracted from Phellinus fruit bodies and their chemical composition and bioactivities were researched. The polysaccharide and protein contents of eight crude polysaccharides had a certain extent differences. Monosaccharide composition and content of amino acids also existed some differences in eight crude polysaccharides. Eight different polysaccharides all showed enhancing splenocyte proliferation effect in vitro. PB-10P and JSHP had high cell proliferation rates with 50㎍/ml concentrations. The results indicated in some extent the immune activity of crude polysaccharides were correlation with the polysaccharide and protein content and composition of each sample. The crude polysaccharides of P. igniarius were further isolated and purified using DEAE Sepharose F. F. and gel-filtration chromatography (Sephacryl S-100-500 ）repeatedly. Five water-soluble homogeneous polysaccharides (P60w1、P60s1、P1SP1、P10SP1and P100SP1) were obtained. Lack of absorption at 280 nm and 260 nm by UV scanning indicated that contained no protein and nucleic acid. HPLC produced a single symmetrical peak, indicated homogeneity and their molecular weigh were 1.71×104 Da、2.07×104 Da、1.48×104 Da、2.20×104 Da and 2.56×104 Da respectively. Structural of P60w-1 were determined using sugar and methylation analysis combined with 1H and 13C NMR spectroscopy, including COSY, TOCSY, NOESY, HSQC and HMBC experiments. The effect of P60w-1 on tumor growth was examined using subcutaneously transplanted H22 and Lewis Lung Carcinoma (LLC) tumor mouse models. Cyclophosphamide or Coriolus versicolor polysaccharopeptide served as positive controls in evaluating tumor response. Results showed that P60w-1 at the most effective dose of 100 mg/kg inhibited the growth of H22 and LLC by 48% and 37%, respectively.

Phellinus gilvus(PG) is a medicinal mushroom belonging to the Hymenochaetaceae basidiomycetes, and has advantages over many Phellinus species due to its short growth period (3 mo), making it cheaper to produce. In the current investigation, we determined the major components of the ethyl acetate extract of PG responsible for its biological activities and further compared the magnitude of the antioxidant/anti-inflammatory activities of components with the various fractional extracts of PG. As the results, the average total DPPH radical scavenging activities of both Fd and Fc of PG was 10 mg/mL, > 95%. Among the fractional extracts of PG, Fd had the greatest inhibitory activity with an IC50 value of 36.70㎍/mL, whereas Fb showed the lowest activity. PCA had even greater activity of NO inhibition than Fd with an IC50 value of 19.46㎍/mL. The mRNA expression of iNOS or COX-2 was nearly undetectable in the absence of LPS. However, LPS- stimulation markedly increased the expression of both iNOS and COX-2 genes. Fd inhibited the effect of LPS in a concentration-dependent manner. Six major compounds were identified from the ethyl acetate extract of PG, and protocatechualdehyde (PCA) was supposed to be the major phenolic compound of PG responsible for its DPPH free radical scavenging activity and its inhibitory effects on LPS-induced NO production in RAW264.7 cells. Further in vitro and in vivo experiments are currently underway to confirm this observation and to investigate the detailed molecular mechanisms involved in the process as well as the biological activities of other fractions of Fd.

This study was carried out to determine the optimum dietary supplementation level of oyster mushroom in cherry salmon (Oncorhynchus masou masou). Juvenile cherry salmon averaging 5.0±0.5g (mean ± SD) were fed one of the five experimental diets containing 0, 3.5, 7.0, 10.5 and 14.0% oyster mushroom (D0, D3.5, D7.0, D10.5 and D14.0) for 12weeks. Increasing of dietary beta-glucan content were observed at a high dietary oyster mushroom powder. After the feeding trial, average weight gain (WG) of fish fed D0, D3.5 and D7.0 diets were significantly higher than those of fish fed D10.5 and D14.0 diets (P < 0.05), however there were no significant differences in WG among fish fed D0, D3.5 and D7.0 diets (P > 0.05). Average feed efficiency (FE) of fish fed D0, D3.5 and D7.0 diets were significantly higher than those of fish fed D10.5 and D14.0 diets (P < 0.05), however there were no significant differences in FE among fish fed D0, D3.5 and D7.0 diets (P > 0.05). Average hepatosomatic index (HSI) of fish fed D0, D3.5, D7.0 and D10.5 diets were significantly higher than those of fish fed D14.0 diets (P < 0.05), however there were no significant differences in HSI among fish fed D0, D3.5, D7.0 and D10.5 diets (P > 0.05). Therefore, these results indicated that the optimum dietary supplementation level could be greater than 3.5%, but less than 7.0% in juvenile cherry salmon under our experimental conditions. And additional research on the immune response will be necessary to carry out.

Sparassis crispa is an edible and a medicinal mushroom, which commonly called cauliflower mushroom and this mushroom, has recently become popular in Asian countries like Korea and Japan. S. crispa can establish to be a good source material for foods and nutraceuticals due to their rich flavor compounds and much amount of b-glucan. Basidiomycetes mushrooms hold the biologically active polysaccharides in fruit bodies, cultured mycelium, and culture broth. Many factors are amenable for the antitumor activities of polysaccharides such as water solubility, molecular size and their branch form. The antitumor activity was examined mainly in the β-glucan (1-3) branched moiety. The researchers suggest that the primary structure of purified β-glucan from the S. crispa possess the backbone structural units as β-(1,3)-D-glucan with single β- (1,6)-D-glucosyl side branching units in every three residues. Polysaccharide derived from S. crispa play an significant role in the antitumor activity. In vitro study revealed that the oral administration of S. crispa β -glucan results in suppressive effect on tumor growth and metastasis in lung cancer through the inhibition of tumor induced-angiogenesis. The researchers also suggests that this effects are not a result of direct action on the endothelial cells because cell growth, migration and capillary-like tube formation were not affected in the human umbilical vein endothelial cells by S. crispa β-glucan application.

Auricularia auricula-judae has long been used as food and traditional remedies in Asian countries such as Korea and China. In this study, we evaluated the in vitro anti-tumor activity of various fractions from the ethanol extracts of Auricularia auricula-judae using various tumor cell lines. To do this, the mesh of Auricularia auricula-judae was mixed with 70% ethanol and heated at 1000C for 6 hrs and ethanol extract (ETOH) was collected. Ethanol extract was fractionated with dichloromethane (DCM), ethyl acetate, n-butanol and a water extract at room temperature as well as concentrated in a vacuum concentrator at a controlled temperature(<500C). The P388D1 macrophage and Sarcoma 180, human NSCLC NCI H358 (bronchioalveolar) and SNU1 cells (Gastric carcinoma) were cultured in RPMI. As the results, the cytotoxicity of the fractional extracts decreased significantly (P<0.05) in a dose-dependent manner. Dichloromethane extract (1 mg/ml) was the highest (P<0.05) in all experimental cell lines. There was also a significantly different sensitivity (P<0.05) among the P388D1, Sarcoma 180, NCI H358 and SNU1 cells for the fractional extracts. According to IC50 values, the most potent cytotoxic activity of dichloromethane fraction was found in Sarcoma 180 and NCI H358 cell lines. Butanol fraction appeared more cytotoxic to SNU1 cell line and water fraction had the highest cytotoxicity in P388D1 cell line. We did not find any significant difference between MTT and SRB assays in their ability to estimate cytotoxicity in all cell lines. Our findings suggest a potent antitumor activity of various fractions from the ethanol extracts of Auricularia auricula-judae depending on the solvent fractions and tumor cell lines. Further in vitro and in vivo studies will provide more information on the active compounds responsible for these activities and their potential as an anti-cancer remedy.

Lentinula edodes (shiitake mushroom) is a very popular edible, cultivated mushroom in Japan. There are post-storage problems with shiitake mushrooms, such as gill browning and cell wall lysis of the fruiting body, which can result in loss of fresh food quality and consequent loss of value. Lentinan is a cell wall component of beta-1, 3-linked-D-glucan with beta-1, 6 branches, which was isolated as an anti-tumor active-substance from L. edodes. Lentinan content decreases following harvest as a result of increased glucanase activity. We isolated one exo-glucanase encoding genes, exg21) and two endo-glucanase encoding gene tlg12) and glu1 from L. edodes. Transcription level of the exg2, tlg1 and glu1 gene increased after harvesting. Enzymes encoded by the genes have lentinan degrading activity, therefore, these genes are involved in lentinan degradation after harvesting. We also identified several cell wall degradation- related enzyme-encoding genes3), such as mixed-linked glucanase (mlg1), chitinases (chi1, chi29), chitin deacetylase (chd1), and chitosanase (cho1). It is revealed that transcriptional levels of these genes increased after harvesting, by real-time PCR. Glucanase and chitinase activity increased following harvest as results of increased transcription of these cell wall degradation-related enzyme-encoding genes. Increase of these cell wall degradation- related enzyme activities would cause cell wall lysis and lentinan degradation during post-harvest preservation. We identified laccase and tyrosinase encoding genes (lcc4 and tyr, respectively) by PCR-subtraction. The lcc4 was a novel laccase-encoding gene in L. edodes. Transcription levels of lcc4 and tyr increased after harvesting, and these genes would be involved in browning of the fruiting body. 1) Sakamoto et al. (2005) Current Genetics, 48: 195-203 2) Sakamoto et al. (2006) Plant Physiology 141: 793-801 3) Sakamoto et al. (2009) Current Genetics 55: 409-423

Cordyceps sensu lato is known as one of the largest genera in hypocrealean fungi and largest group of entomopathogenic fungi in Ascomycota. Approximately 400 species are members of Cordyceps s. l. and most of them are obligate symbionts of 10 orders of arthropods and false truffles specifically parasitizing Elaphomyces spp. Recently, Cordyceps s. l. was reclassified into Cordyceps sensu stricto, Elaphocordyceps, MetaCordyceps, and OphioCordyceps in three families (i.e., Clavicipitaceae, Cordycipitaceae, Ophiocordycipitaceae) with the evidence of recent multigene phylogenetic analyses coupled with morphological and ecological characters. With the closely related animal, plant and fungal associated genera (e.g., Balansia, Claviceps, Hypocrella and Torrubiella) of Cordyceps s. l., Cordyceps s. l. and its related genera in Hypocreales have been considered as one of the model systems in understanding the evolution of host affiliation in Kingdom Fungi. Here the overview of molecular systematic of Cordyceps was presented with its evolutionary hypotheses of host affiliation based on the ancestral state reconstruction from 162-taxon data set. In our results, the evolution of its host affiliation is largely characterized by frequent interkingdom host-jumps and ergot and grass endophytes (e.g., Balansia, Claviceps, Epichloe and Neotyphodium) are hypothesized to be derived from an ancestor that parasitized arthropods. Around 350 taxa have been included in the molecular phylogeny of Cordyceps s. l. after the new classification was proposed. Therefore, the progress and problems in current molecular phylogeny are also presented with introduction of the future research direction in molecular systematics and genomics of Cordyceps and its related genera.

Many researchers reported biodegradation of environmental pollutants by white-rot fungi. Toward in situ bioremediation, we have investigated biodegradation of environmental pollutants by litter-decomposing fungi. In our results, lignin-degrading enzymes produced from litter-decomposing fungi are thought to participate in degradation of pentachlorophenol (PCP) and 2,4-dichlorophenoxy acetic acid (2,4-DA). Then, we examined the biodegradation of PCP, 2,4-DA and the analogs of 2,4-DA by purified laccase and the role of redox mediator on laccase reaction. Laccase purified from Calvatia craniiformis decomposed phenolic compounds, PCP and 2,4-dichlorophenol (2,4-DP), but not non-phenolic compounds, 2,4-DA and 2,4-dichlorophenoxy ethanol (2,4-DE), even in the presence of redox mediators. To clarify the reaction mechanism between the substrates and redox mediators, quantum chemical analysis was applied using MOPAC 2009 and Gaussian 03. The results of the heat of formation and the perturbation energy showed that even redox mediator radicals could not oxidize the non-phenolic compounds. Previously several reports showed that laccase-redox mediator systems decomposed non-phnolic compounds, but we propose that the system could not react on the chlorinated aromatic compounds based on the result of quantum analysis.

Addition of ammonia or any nitrogenous materials to the soil that release ammonia causing alkaline condition during decomposition stimulates the fruiting of a particular chemoecological group of fungi, called ammonia fungi (Sagara 1975). The study of ammonia fungi by artificial application of urea in forest soil has been done in diverse geographical regions such as in Japan, Taiwan, New Zealand, Western Australia, and UK. Up to date about 70 species of ammonia fungi have been recorded in those regions. However, ammonia fungi in the boreal forest of American continent have not yet been investigated. Thus, we collected the soils of A0 and the upper layer of HA horizons in plant pots from aspen forest near Edmonton, Canada. Thereafter, we applied urea (granular fertilizer; 46% nitrogen, 10 mg/g dry soil) in plant pots and incubated at 25˚C under 12 hours dark and light regime. After 40 days of incubation, several basidiomata of Coprinopsis species appeared. Among them one specimen was identified as C. rugosobispora based on macro- and microscopic features. Morphologically this species was very similar to C. phlyctidospora which was characterized by warty, ovoid basidiospores, and diverticulate veil elements. C. phlyctidospora has 4-spored basidia while C. rugosobispora had only 2-spored. In the beginning, it was thought probably it was only a 2-spored form of C. phlyctidospora. The basidiospore of C. rugosobispora (9.8-11.7×8.3-9.6㎛) was distinctly larger than that of C. phlyctidospora (8.4-10.6×6.0-7.6 ㎛). It was therefore separated from the C. phlyctidospora. Furthermore in this study we investigated its phylogenetic relationship based on the nuclear rDNA sequence in ITS regions and mating reactions among its close allies and further confirmed it as a distinct species. This is the first record of C. rugosobispora from American continent since it has been collected only from Europe (Belgium and Netherlands). Although urea effectively stimulated its occurrence but it has not yet been reported any other urea application studies so far. This indicates it is a new record in ammonia fungi as well.

[Aims] Lentinula edodes (Berk.) Pegler has many commercial strains both morphologically and physiologically similar to each other. At present, detection of polymorphism in rDNA-IGS region (Babasaki, 2006) and/or RAPD marker (Zhang and Molina, 1995) is generally used for strain typing of L. edodes. However, it is rather time-and-cost consuming. Inter-retrotransposon amplified polymorphism (IRAP)-PCR method mainly used for horticultural crops takes less time and lower in cost in strain typing (Kalendar et al, 1999). In this study, we designed IRAP primers for L. edodes and verified their strain typing efficiency. [Method] Thirty three strains were provided for this study. Either fungal cultures on PDA or fungal tissues of fruit bodies were cut into approximately 4 x 4 x 4 mm. Total DNA of each samples were extracted by DNeasy Plant Mini Kit (QIAGEN). For PCR, IRAP primer set and Pfu-X polymerase (greiner) were performed. Based on LTR (Long Terminal Repeat) sequence in L. edodes, we designed one set of primers amplifying the regions between retrotransposons. Ampricons were electrophored for 50 min at 100 V on 1.7 % agarose gel with GelRed (Biotium) and evaluated under UV irradiation. [Results] The products obtained by IRAP-PCR were determined using mini-gel electrophoresis system. The band patterns of IRAP-PCR products differ among strains except the ones having the same parental cultivar. The detected bands were bright and clear without smearing. The IRAP-PCR products of fungal cultures on PDA and correlating fungal tissues of fruit bodies showed the same band pattern, suggesting that the procedure is highly reproducible. Thus, it is considered that IRAP-PCR with short ranged (ca. 1 kb) electrophoresis is a time-efficient and practical strain typing method of L. edodes.

This study was carried to investigate the suitable conditions of Pleurotus eryngii through precooling for the more long-term freshness. The time to be same temperatures between P. eryngii and storage room through precooling at 0℃ and 4℃ were showed 2 hours and 5.5 hours, respectively. P. eryngii was cooled within 6 hours and 18 hours at 0℃ and 4℃ with two type, forced air cooling and pressure cooling. After precooling, P. eryngii was packed 400±10g with anti-fog film, and then stored at 4±1℃. In all treatment of precooling, the weight loss was peaked at 25 days and hardness of P. eryngii were decreased at 10 days during storage. There were no significant freshness differences between precooling type and conditions. It was found that the optimum stored period of P. eryngii at 4℃ after precooling was estimated to be 30~35days. Consequently, it is necessary to elucidate efficient and economic precooling conditions.