Effect of freezing and thawing on cell membranes of Lentinus edodes,the shiitake mushroom

When cell membranes of Lentinus edodes mycelium were rapidly frozen at either 50 or 160°C/min, viability was lost and this correlated with rupture of the plasmalemma and residual membrane material and with alterations in the organelles. Although with slow cooling (1°C/min) 80% of the samples recovered viability, some cells still showed similar changes to those cooled rapidly, indicating that individual cells of the mycelium do not respond in the same way.     

Studies on purification of a lectin from fruiting bodies of the edible shiitake mushroom Lentinus edodes

A lectin, with a specificity for N-acetylgalactosamine and N-acetylglucosamine and a molecular weight of 43 kDa, was isolated from fruiting bodies of the edible shiitake mushroom Lentinus edodes. The purification procedure entailed extraction with aqueous buffer, ammonium sulfate precipitation, gel filtration on Sephadex G-100 and affinity chromatography on N-acetylgalactosamine-agarose. The lectin was unique in that it was tenaciously bound on anion exchangers including DEAE-cellulose, DEAE-Sepharose, DEAE-Sephadex, Q-Sepharose, Dowex and PEI-cellulose and also on hydroxyapatite and phenyl Sepharose. It was largely unadsorbed on cation exchangers including CM-cellulose, CM-Sepharose, SP-Sepharose and Amberlite, and also on protein G-Sepharose, Red Sepharose and Affi-gel Blue gel, wheat germ lectin-Sepharose and p-aminophenyl-d-glucopyranoside agarose.     

Polyisoprenoid alcohols from the mushroom Lentinus edodes

Lipids extracted from the shiitake mushroom Lentinus edodes contain dolichols composed of 15 up to 19 isoprene units with Dol-17 as the dominating prenologue. Identification of dolichols was achieved by the application of 2D-TLC, HPLC and electrospray ionization mass spectrometry. Additionally a family of polyprenols (-unsaturated counterparts) with the same chain-length was also detected. Dolichols comprised approximately 0.002% of the fresh weight of the mushroom. Dolichols accompanied by traces of polyprenols are for the first time found in the mushroom tissue.     

Lentinan from shiitake mushroom (Lentinus edodes) suppresses expression of cytochrome P450 1A subfamily in the mouse liver

Induction of xenobiotics metabolizing enzymes is related to the formation and chemoprevention of cancer. Since cytochrome P450s (CYPs) including CYP1A subfamily metabolize certain pro-carcinogens to their ultimate forms, down-regulation of CYP1As by food factors leads to the prevention of cancer. Mushroom polysaccharides, especially beta-glucans such as lentinan from Lentinus edodes, possess the anti-tumor and immunomodulating activities through the cytokine production from immunocytes. Recent our studies have demonstrated that lentinan suppresses hepatic CYP1As expression in the both constitutive and inducible levels through the production of tumor necrosis factor-alpha and an increase in the DNA-binding activity of nuclear factor-kappaB. This paper discusses on the effective lentinan dosage and route of administration for suppression of CYP1As.     

Detection and distribution of six linear mitochondrial plasmids in the shiitake mushroom,Lentinula edodes

The presence of plasmids was surveyed in 90 wild isolates ofLentinula edodes collected from geographically different world regions. DNA plasmids of different sizes were found in about 80% of the isolates. The plasmids detected were of six kinds, designated as pLE1 (9.0 kb), pLE2 (11.1 kb,=pLLE1 described by other authors), pLE3A (9.8 kb), pLE3B (10.8 kb), pLE3C (12.1 kb), and pLE3D (12.3 kb). Hybridization analysis suggested that pLE1 and pLE2 were distinct plasmid types of different homology groups to each other, and the four other plasmids were variant types belonging to a third homology group. These plasmids had no homology with their host's and non-host's nuclear and mitochondrial genome DNAs. Restriction analysis and electron microscopy indicated that the plasmids are linear in form. Since all six plasmids were transmitted uniparentally in sexual crosses and were consistently associated with the DNA preparations from mitochondria fractionated from mycelia of representative isolates, they were suggested to be located in mitochondria, similar to many other known fungal DNA plasmids. Geographically, pLE1 and pLE2 were widely distributed in natural populations ofL. edodes, while the remaining four plasmids were uniquely present in delimited natural populations. Contribution No. 322 from the Tottori Mycological Institute.     

Carboxyl Proteinase Essential for Maturation of Basidiospores of Lentinus edodes (shiitake)

The effects of eicosapentaenoic acid (EPA, 20: 5n-3) on essential fatty acid (EFA)-deficient rats were studied. After low growth and scaly dermatitis in the hind legs due to dietary EFA deficiency were induced by feeding rats an EFA-free 25 % casein diet (25C) containing 30 % hydrogenated coconut oil with 1 % cholesterol (HCO ? CHOL) for 8 weeks, they received the 25C diet with 0.19 or 0.57 % EPA ethyl ester concentrate added, or 0.02 % or 0.38 % linoleic acid (LA, 18: 2n-6) concentrate (Exp. I), and the HCO ? CHOL meal including any one of 0.25, 0.50, or 1.00 % EPA concentrate, and 0.12 and 0.48 % LA concentrate (Exp. II) for an additional 6 weeks. When EFA-deficient rats were fed the EPA in both experiments, body weight was gained to almost reach those of the 0.38 or 0.48 % LA-fed group (control), and the dermal symptoms of the hind legs were relieved, though the degree of healing was less than those of the controls. The ratios of eicosatrienoic acid (20: 3n-9) to arachidonic acid (20: 4n-6) characteristically increased due to EFA deficiency were reduced to the level of the control in the liver and heart by addition of the EPA concentrate.     

Biodegradation of 2,4-dichlorophenol by shiitake mushroom (Lentinula edodes) using vanillin as an activator

The white-rot shiitake mushroom, Lentinula edodes, was used to degrade an environmentally hazardous compound, 2,4-dichlorophenol (DCP), using vanillin as an activator. Vanillin increased the mycelial growth from 74 to 118 mg/150 ml culture and accelerated laccase and Mn-peroxidase production from the maximum on days 24–28 without vanillin to days 10–14. It eliminated 92 % of 100 mM DCP with 50 mg vanillin/l compared with only 15 % without vanillin. GC–MS revealed that a diaryl ether dimer of DCP was formed in the culture without vanillin, whereas dimer formation was diminished with vanillin addition. This indicates that vanillin enhances the degradation of DCP and disrupts the formation of the toxic dimer. Therefore, lignin-derived phenol such as vanillin can be used as natural and eco-friendly activators to control white-rot mushrooms, thereby facilitating the effective degradation of environmentally hazardous compounds.     

Construction of a genetic linkage map of shiitake mushroom Lentinula edodes strain L-54

From fruiting bodies of L. edodes strain L-54, single-spore isolates (SSIs) were collected. Two parental types of L-54 were regenerated via monokaryotization. By means of random-amplified polymorphic DNA (RAPD), DNA samples from L-54, its two parental types, and 32 SSIs were amplified with arbitrary primers. Dedikaryotization was demonstrated, and 91 RAPD-based molecular markers were generated. RAPD markers that were segregated at a 1:1 ratio were used to construct a linkage map of L. edodes. This RAPD-linkage map greatly enhanced the mapping of other inheritable and stable markers [such as those that are linked to a phenotype (the mating type), a known gene (priA) and a sequenced DNA fragment (MAT)] with the aid of mating tests, bulked-segregant analysis, and PCR-single-strand conformational polymorphism. These markers comprised a genetic map of L. edodes with 14 linkage groups and a total length of 622.4 cM.     

Hazelnut husk as a substrate for the cultivation of shiitake mushroom (Lentinula edodes)

The possibility of using hazelnut husk (HH) as a new basal ingredient for substrate preparation in Lentinula edodes cultivation was investigated. Some chemical properties of the substrates prepared by HH alone and its mixtures with wheat straw (WS), beech wood-chip (BWC) and wheat bran (WB) in different ratios were compared, and their effects on spawn run time, days to first harvest (earliness), yield and biological efficiency (BE) were determined. The N content of the substrate prepared from HH alone was very high (0.82%), and thus the C:N ratio of substrates decreased with an increase in the rate of HH in the mixtures. Yield and BE in the HH alone substrate was considerably low compared with the controls (80BWC:10WS:10M and 60BWC:20WS:20WB), and decreased with an increase in the rate of HH in the mixtures. However, when the HH content in the mixtures was kept below 50%, the yield was relatively high (50HH:50WS and 50HH:50BWC). Even when the HH content increased to 75% in the mixture, the comparable yield and BE to the controls could be obtained by adding 10% of WB as nutrients (75HH:15WS:10WB and 75HH:15BWC:10WB). The results revealed that HH could be used as a new basal ingredient for substrate preparation in L. edodes cultivation.     

Immunoprophylactic effects of shiitake mushroom (Lentinula edodes) against Bordetella bronchiseptica in mice

Antimicrobials are used as feed additives to improve growth performance and to prevent subclinical disease challenge in industrial animals. However, these drugs can lead to the development of resistant strains of bacteria. Shiitake mushrooms (SM) (Lentinula edodes) have long been popular as a health food in East Asia. Moreover, SM-derived polysaccharides are well-known as immunostimulants that possess antimicrobial properties. The aim of the present study was to evaluate the immunoprophylactic effects of SM against Bordetella bronchiseptica infection in mice as an initial step towards the development of eco-friendly feed additives to reduce the use of antimicrobials. Although SM had no effect on body weight gain under the un-infected conditions, SM alleviated progressive weight loss and helped in the recovery of body weight in B. bronchiseptica infected mice. Dietary supplementation with SM reinforced bacterial clearance in the infected mice. Of note, SM markedly increased the percentage of various T lymphocytes and the relative mRNA expression levels of tumor necrosis factor-α and interferon-γ in the bronchial lymph node early in the infection. Taken together, these findings suggest that SM could help in the improvement of body weight gain during B. bronchiseptica infection and may enhance the protective immune activity against a subclinical disease challenge, such as B. bronchiseptica infection in mice, probably by a strong stimulation of non-specific immune responses. Hence, SM may provide an alternative to reduce use of antimicrobials. Confirmation of the beneficial effects of SM as a feed additive is now required in industrial animals.     

Lentinula edodes (shiitake mushroom): An assessment of in vitro anti-atherosclerotic bio-functionality

Mushrooms have been highly regarded as possessing enormous nutritive and medicinal values. In the present study, we evaluated the anti-oxidative and anti-atherosclerotic potential of shiitake mushroom (Lentinula edodes) using its solvent–solvent partitioned fractions that consisted of methanol:dichloromethane (M:DCM), hexane (HEX), dichloromethane (DCM), ethyl acetate (EA) and aqueous residue (AQ). The hexane fraction (1 mg/mL) mostly scavenged (67.38%, IC₅₀ 0.55 mg/mL) the 2,2-diphenyl-1-picryl hydrazyl (DPPH) free radical, contained the highest reducing capacity (60.16 mg gallic acid equivalents/g fraction), and most potently inhibited lipid peroxidation (67.07%), low density lipo-protein oxidation and the activity of 3-hydroxy 3-methyl glutaryl co-enzyme A reductase (HMGR). GC–MS analyses of the hexane fraction identified α-tocopherol (vitamin E), oleic acid, linoleic acid, ergosterol and butyric acid as the bio-functional components present in L. edodes. Our findings suggest that L. edodes possesses anti-atherosclerotic bio-functionality that can be applied as functional food-based therapeutics against cardiovascular diseases.     

Characterization of xylanase from Lentinus edodes M290 cultured on waste mushroom logs

Extracellular enzymes from Lentinus edodes M290 on normal woods (Quercus mongolica) and waste logs from oak mushroom production were comparatively investigated. Endoglucanase, cellobiohydrolase, beta-glucosidase, and xylanase activities were higher on waste mushroom logs than on normal woods after L. edodes M290 inoculation. Xylanase activity was especially different, with a three times higher activity on waste mushroom logs. When the waste mushroom logs were used as a carbon source, a new 35 kDa protein appeared. After the purification, the optimal pH and temperature for xylanase activity were determined to be 4.0 and 50 degrees C, respectively. More than 50% of the optimal xylanase activity was retained when the temperature was increased from 20 to 60 degrees C, after a 240 min reaction. At 40 degrees C, the xylanase maintained 93% of the optimal activity, after a 240 min reaction. The purified xylanase showed a very high homology to the xylanase family 10 from Aspergillus terreus by LC/MS-MS analysis. The highest Xcorr (1.737) was obtained from the peptide KWI SQGIPIDGIG SQTHLGSGGS WTVK originated from Aspergillus terreus, indicating that the 35 kDa protein was xylanase. This protein showed low homology to a previously reported L. edodes xylanase sequence.     

Correlation of the properties of several lignocellulosic substrates to the crop performance of the shiitake mushroom Lentinula edodes

Two selected Lentinula edodes strains (S4080 and SIEF0231) were cultivated on oak-wood sawdust (OS), wheat straw (WS) and corn-cobs (CC) substrates in order to examine the influence of those residues on mycelium growth and on basidiomata production. For both strains, mycelial growth measurements conducted in race tubes demonstrated faster colonization of OS and WS media. Lag-phase and complete colonization periods were correlated to mycelium extension rates in the three substrates tested. Similar patterns of pH and electrical conductivity (Ec) changes were detected in all media and for all strains tested; the pH decreased steadily throughout the colonization process to reach values of 4.49–5.06; Ec increased by the end of mycelium colonization, and it presented the highest and lowest values in the WS and OS media respectively. In addition, a negative correlation was established between final salt content of the substrates and mycelium extension rates. Subjecting fully colonized substrates to a cold-shock treatment resulted in fruiting 58–65 days after inoculation in tubes; WS and CC promoted earlier sporophore initiation than OS. Monitoring CO₂ emissions by strain SIEF0231 in pilot-scale cultivation on synthetic blocks, revealed higher respiration rates from OS and CC than from WS, which were further correlated with substrate colonization rates. Among residues colonized by the same strain, WS appeared to promote earliness and crop productivity (BE 54.17%) by presenting shorter cropping periods and equal yield distribution among flushes, while on OS and CC maximum yields were obtained within the first two flushes. Moreover, heavier basidiomata were produced by WS and OS substrates.     

香菇蛋白质氨基酸的分析

分析结果表明,香菇的菌柄与菌盖中蛋白质氨基酸均为18种,缺少谷氨酰胺(Gln)和天冬酰胺(Asn),其中,含量最高的是谷氨酸(菌柄为11.03 mg/gDW,菌盖为12.57 mg/gDW)。菌盖与菌柄均含必需氨基酸、半必需氨基酸10种,其中精氨酸含量(菌柄为10.73 mg/gDW,菌盖为11.84 mg/gDW)。必需氨基酸的含量十分接近于非必需氨基酸的含量(比率为1.00∶1.04),无论总氨基酸含量还是必需氨基酸含量,菌盖中的皆略高于菌柄中的。     

Characterization of vitamin B12 compounds in the fruiting bodies of shiitake mushroom (Lentinula edodes) and bed logs after fruiting of the mushroom

This study determined the vitamin B₁₂ content in commercially available dried fruiting bodies of shiitake mushroom, Lentinula edodes. The vitamin B₁₂ contents in dried donko-type fruiting bodies with closed caps (5.61 ± 3.90 μg/100 g dry weight), did not significantly differ from those of dried koushin-type fruiting bodies with open caps (4.23 ± 2.42 μg/100 g dry weight). The bed logs after fruiting of the mushroom also contained the vitamin B₁₂ levels similar to that in the dried shiitake fruiting bodies. To determine whether the dried shiitake fruiting bodies and their bed logs contained vitamin B₁₂ or other corrinoid compounds that are inactive in humans, we purified corrinoid compounds using an immunoaffinity column and identified vitamin B₁₂ using vitamin B₁₂-dependent Escherichia coli 215 bioautograms and liquid chromatography-electrospray ionization tandem mass spectrometry (LC/ESI-MS/MS) chromatograms. Dried shiitake fruiting bodies rarely contained an unnatural corrinoid vitamin B₁₂[c-lactone] that is inactive in humans. Given that shiitake mushroom lacks the ability to synthesize vitamin B₁₂ de novo, the vitamin B₁₂ found in dried shiitake fruiting bodies must have been derived from the bed logs.     

Mannitol Metabolism in Lentinus edodes, the Shiitake Mushroom

Mannitol metabolism was evaluated in fruiting bodies of Lentinus edodes. Cell extracts were prepared from fruiting bodies, and key enzymes involved in mannitol metabolism were assayed, including hexokinase, mannitol dehydrogenase, mannitol-1-phosphate dehydrogenase, mannitol-1-phosphatase, and fructose-6-phosphatase. Mannitol dehydrogenase, fructose-6-phosphatase, mannitol-1-phosphatase, and hexokinase activities were found in extracts of fruiting bodies. However, mannitol-1-phosphate dehydrogenase activity was not detected. Mycelial cultures were grown in an enriched liquid medium, and enzymes of the mannitol cycle were assayed in cell extracts of rapidly growing cells. Mannitol-1-phosphate dehydrogenase activity was also not found in mycelial extracts. Hence, evidence for a complete mannitol cycle both in vegetative mycelia and during mushroom development was lacking. The pathway of mannitol synthesis in L. edodes appears to utilize fructose as an intermediate.     

PEG介导下香菇的转化

表达载体p301-bG1含有香菇（Lentinus edodes (Berk.)Sing)三磷酸甘油醛脱氢酶启动子驱动下的gus基因和除草剂抗性基因。利用PEG法实现了p301－bG1对香菇原生质体的转化。香菇原生质体与经PEG纯化的质粒DNA混合,用PEG处理后培养于含40ug/mL除草剂的CYM再生平板上,得到了抗除草剂和有GUS活性的转化菌株。虽然这种方法转化效率较低,但不需要昂贵的仪器和限制性内切酶,为蘑菇的分子育种研究提供了一种简便经济的转化方法。     

Lectin activity of Lentinus edodes

The hemagglutinating activity of submerged mycelium and culture liquid for four strains of Lentinus edodes (Berk.) Sing [L. edodes (Berk.) Pegler] was studied in the search for lectins. The hemagglutinating activity of culture liquid was substantially higher, compared with mycelium. The carbohydrate-binding capacity of the agglutinins was established, and the lectin activity of extracts from mycelia grown on several agar media was elucidated in relation to fruiting. The lectin activity of L. edodes was examined at different morphogenetic steps: mycelium, brown mycelial film, primordium, and fruiting body. Hemagglutination titers at the brown film step were higher than in the mycelium, whereas activity at the primordial and fruiting bodies steps decreased. Lectins seem to be involved in the formation of hyphal aggregates of brown mycelial film.