Biochemical and transcriptomic analyses of the symbiotic interaction between Cremastra appendiculata and the mycorrhizal fungus Coprinellus disseminatus.

BACKGROUND: Cremastra appendiculata is a rare terrestrial orchid with a high market value as an ornamental and medicinal plant. However, the species depends entirely on fungi for seed germination under natural conditions. In a previous study, we have successfully isolated and identified the mycorrhizal fungus Coprinellus disseminatus which was able to induce the germination of C. appendiculata seeds. We then speculated that C. disseminatus may do so by breaking the testa imposed dormancy of the seeds. In this study, biochemical and transcriptomic analyses were used to characterize the germination of C. appendiculata seeds, collected at different stages of germination, as affected by C. disseminatus. RESULTS: The lignocellulose in the seeds coat of C. appendiculata was degraded by the mycorrhizal fungus resulting in facilitated absorption of water. The rate of decline in lignin content was 67 and 73% at 6 and 12 days after sowing, respectively. The water content increased from 13 to 90% during symbiosis. A total of 15,382 genes showing significantly different levels of expression (log2 FPKM≥2.0, Qvalue≤0.05) were successfully identified among all libraries, where the highest number of DEGs was shared between 6 days versus 0 day after symbiotic germination. Gene annotation results suggested that 15 key genes related water-status, such as DHN gene family and Xero 1 were down-regulated. The genes zeaxanthin epoxidase ZEP, 9-cis-epoxycarotenoid dioxygenase NCED3 and ß-carotene hydroxylase involved in the biosynthesis of abscisic acid (ABA) were significantly down-regulated in 6 days as compared to 0 day after symbiotic germination. CONCLUSIONS: This work demonstrates that mycorrhizal fungus C. disseminatus can stimulate C. appendiculata seeds germination through a mechanism of breaking the testa imposed dormancy and inducing water absorption of the embryo.

Dillapiole in Piper holtonii as an Inhibitor of the Symbiotic Fungus Leucoagaricus gongylophorus of Leaf-Cutting Ants.

Plants of the Piperaceae family are studied for their diverse secondary metabolism with a vast array of compounds that act as chemical defense agents against herbivores. Of all the agricultural pests, the management of insects is a highly significant challenge in the Neotropics, and ants of the Attini tribe pose a major problem. Due to their symbiotic association with the fungus Leucoagaricus gongylophorus (Möller) Singer (Agaricaceae), the species of Atta and Acromyrmex have exhaustive foraging activity which has intensified as deforestation and monoculture farming have increased. The control of leaf-cutting ants is still carried out with synthetic products with negative consequences to the environment and human health. In search for natural and sustainable alternatives to synthetic pesticides, Piper holtonii C. DC. was selected among other plant species after field observations of the foraging activity of Atta cephalotes, which revealed that P. holtonii was never chosen by ants. In vitro evaluation of an ethanol extract of the leaves of P. holtonii resulted in promising inhibitory activity (IC50 102 ppm) against L. gongylophorus. Subsequently, bioassay-guided fractionation led to the isolation of the phenylpropanoid dillapiole, which was also detected in the essential oil. This compound demonstrated inhibition of the fungus with an IC50 of 38 ppm. Considering the symbiotic relationship between the Attini ants and L. gongylophorus, the negative effect on the survival of one of the organisms will affect the survival of the other, so dillapiole or standardized essential oil extracts of P. holtonii containing this active principle could be a unique and useful source as a control agent for leaf cutting-ants.

Professor Shu-Ting Chang on His 90th Birthday.

This issue is dedicated to the 90th birthday of Professor Shu-Ting Chang, a prominent scientist in the field of mushroom biology, including cultivation and nutritional values of mushrooms, medicinal mushroom science, and environmental impact of mushrooms. Professor Shu-Ting Chang is also one of the initiators and has been an editor of our journal from its inception (in 1999) until now.

Adding benefit to wetlands - Valorization of harvested common reed through mushroom production.

Wetlands have been successfully implemented as water purification systems for removal of plant nutrients and can play a significant role in nutrient recycling, depending on use of the harvested biomass. In a constructed wetland in southern Sweden examined in this study, assimilation of plant nutrients in wetland biomass corresponded to 234 kg/ha nitrogen, 22.8 kg/ha phosphorus, and 158 kg/ha potassium in the study year (2016). The harvested biomass, composed exclusively of common reed, was evaluated as a substrate for production of oyster mushrooms, one of the most widely produced edible mushrooms in the world. The biological efficiency of the substrate was 138 ±â€¯10%, corresponding to production of 1.4 kg mushrooms (fresh weight) based on 1 kg reed (dry weight). The fruiting bodies had high quality, with total protein concentration 18.3 ±â€¯2.8% and very low levels of contaminating heavy metals. Thus, nutrient assimilation in wetland biomass not only decreases the risk of eutrophication in recipient waters, but can be utilized for direct production of high-quality food. The biomass remaining after mushroom production, composed of mycelium and partly degraded wetland biomass, has potential for use in ruminant feed, i.e., as roughage.

Antiproliferative Activity and Cytotoxicity of Some Medicinal Wood-Destroying Mushrooms from Russia.

We analyzed the antiproliferative activity of 6 medicinal wood-destroying mushrooms (Fomes fomentarius, Fomitopsis pinicola, Trametes versicolor, Trichaptum biforme, Inonotus obliquus, and Coniophora puteana) that are common in deciduous and mixed coniferous forests in Central Russia. Morphological identification of strains collected from the wild was confirmed based on ribosomal DNA internal transcribed spacer phylogenetic analysis. We observed cytotoxic and cell growth-inhibitory effects of hot water extracts from mycelial biomass of 5 species-T. versicolor, C. puteana, F. fomentarius, F. pinicola, and I. obliquus-on leukemia cell lines (Jukart, K562, and THP-1); the effective extract concentrations were mostly less than 50 µg · mL-1. However, we observed no antiproliferative activity of dry biomass from methanol-chloroform (1:1) extracts of C. puteana and F. fomentarius. A chemosensitivity assay showed that the most effective polypore mushroom extract was the methanol extract of T. versicolor (strain It-1), which inhibited the growth of 6 various solid tumors (A-549 and SWi573 [lung], HBL-100 and T-47D [breast], HeLa [cervix], and WiDr [colon]) at concentrations below 45 µg · mL-1, with a concentration as low as 0.7-3.6 µg · mL-1 causing 50% reduction in the proliferation of cancer cells in lung and cervix tumors. Methanol extracts of F. pinicola and I. obliquus were less effective, with proliferation-inhibiting capacities at concentrations below 70 and 200 µg · mL-1, respectively. Thus, T. versicolor is a prospective candidate in the search for and production of new antiproliferative chemical compounds.

Proteomic analysis reveals the mechanisms of Mycena dendrobii promoting transplantation survival and growth of tissue culture seedlings of Dendrobium officinale.

AIMS: Dendrobium officinale is an important traditional Chinese medicinal herb. Its seedlings generally show low survival and growth when transferred from in vitro tissue culture to a greenhouse or field environment. In this study, the effect of Mycena dendrobii on the survival and growth of D. officinale tissue culture seedlings and the mechanisms involved was explored. METHODS AND RESULTS: Mycena dendrobii were applied underneath the roots of D. officinale tissue culture seedlings. The seedling survival and growth were analysed. The root proteins induced by M. dendrobii were identified using two-dimensional (2-D) electrophoresis and matrix-assisted laser desorption/ionization time-of-flight MS (MALDI-TOF-MS). Mycena dendrobii treatment significantly enhanced survival and growth of D. officinale seedlings. Forty-one proteins induced by M. dendrobii were identified. Among them, 10 were involved in defence and stress response, two were involved in the formation of root or mycorrhizae, and three were related to the biosynthesis of bioactive constituents. CONCLUSIONS: These results suggest that enhancing stress tolerance and promoting new root formation induced by M. dendrobii may improve the survival and growth of D. officinale tissue culture seedlings. SIGNIFICANCE AND IMPACT OF THE STUDY: This study provides a foundation for future use of M. dendrobii in the large-scale cultivation of Dendrobiums.

Simplified and lower cost methods for culinary-medicinal mushrooms cultivation.

The importance and prospect of growing mushrooms through utilization of low-cost, technologically simple methods for developing Third World countries has recently been outlined. Three different species from genus Pleurotus (P. djamor, P. pulmonarius, and P. sajor-caju) and one from genus Hypsizygus (H. ulmarius) were studied. Whole stalk wheat straw, shredded wheat straw, and ground maize cob (Zea mays) were used as the substrates. Wheat straw is the post-harvest stalk of Triticum aestivum. Biological efficiency (BE), growth dynamics, and photographs are provided for each cultivated strain, on different substrates, and substrate treatment comparisons are made. From several experiments conducted with various technologically simple methods of cultivation, it was found that all above mentioned species can be successfully cultivated at the village level in any country, be it highly developed or the poorest country on earth.

[Cloning and expression analysis of a calcium-dependent protein kinase gene in Dendrobium officinale in response to mycorrhizal fungal infection].

Calcium-dependent protein kinases (CDPKs) play an important regulatory role in the plantarbuscular mycorrhiza/rhizobium nodule symbiosis. However, the biological action of CDPKs in orchid mycorrhiza (OM) symbiosis remains unclear. In the present study, a CDPK encoding gene, designated as DoCPK1 (GenBank accession No. JX193703), was identified from D. officinale roots infected by an OM fungus-Mycena sp. using the reverse transcription polymerase chain reaction (RT-PCR) and rapid amplification of cDNA ends (RACE) methods, for the first time. The full length cDNA of DoCPK1 was 2137 bp in length and encoded a 534 aa protein with a molecular weight of 59.61 kD and an isoelectric point (pI) of 6.03. The deduced DoCPK1 protein contained the conserved serine/threonine-protein kinase catalytic domain and four Ca2+ binding EF hand motifs. Multiple sequence alignment demonstrated that DoCPK1 was highly homologous (85%) to the Panax ginseng PgCPK1 (ACY78680), followed by CDPKs genes from wheat, rice, and Arabidopsis (ABD98803, ADM14342, Q9ZSA2, respectively). Phylogenetic analysis showed that DoCPK1 was closely related to CDPKs genes from monocots, such as wheat, maize and rice. Real time quantitative PCR (qPCR) analysis revealed that DoCPK1 was constitutively expressed in the included tissues and the transcript levels were in the order of roots > stems > seeds > leaves. Furthermore, DoCPK1 transcripts were significantly accumulated in roots 30 d after fungal infection, with 5.16 fold compared to that of the mock roots, indicating involvement of DoCPK1 during the early interaction between D. officinale and Mycena sp., and a possible role in the symbiosis process. This study firstly provided important clues of a CDPK gene associated with OM symbiosis, and will be useful for further functional determination of the gene involving in D. officinale and Mycena sp. symbiosis.

Preparation of Chaga medicinal mushroom, Inonotus obliquus-fermented rice using solid-state fermentation and its taste quality and antioxidant property.

Chaga medicinal mushroom, Inonotus obliquus (Pers.:Fr.) Pilat, was inoculated into cooked embryo rice and the mycelial growth in I. obliquus-fermented embryo rice (IER) was monitored. Besides, nonvolatile taste components and antioxidant properties of fruiting bodies, mycelia, IER, and embryo rice were studied. The optimal conditions for mycelial growth were determined to be: 30°C, an inoculation rate of 1 mL/15 g, water supplementation of 60%, and no extra nitrogen source added. IER showed similar proximate composition to embryo rice but contained a substantial amount of ergothioneine (101 mg/kg dry weight). IER contained higher amounts of soluble sugars and polyols, and umami taste components, including monosodium glutamate (MSG)-like components and flavor 5'-nucleotides, than embryo rice. Besides, IER showed a second level of equivalent umami concentrations (223.73 g MSG/100 g). Fruiting bodies did not contain umami components but showed the most effective antioxidant properties. Although some of EC50 values of IER were less than those of mycelia and embryo rice, IER still showed effective antioxidant properties. Based on the results obtained, IER will be a novel functional food.

Selenium in edible mushrooms.

Selenium is vital to human health. This article is a compendium of virtually all the published data on total selenium concentrations, its distribution in fruitbody, bioconcentration factors, and chemical forms in wild-grown, cultivated, and selenium-enriched mushrooms worldwide. Of the 190 species reviewed (belonging to 21 families and 56 genera), most are considered edible, and a few selected data relate to inedible mushrooms. Most of edible mushroom species examined until now are selenium-poor (< 1 microg Se/g dry weight). The fruitbody of some species of wild-grown edible mushrooms is naturally rich in selenium; their occurrence data are reviewed, along with information on their suitability as a dietary source of selenium for humans, the impact of cooking and possible leaching out, the significance of traditional mushroom dishes, and the element's absorption rates and co-occurrence with some potentially problematic elements. The Goat's Foot (Albatrellus pes-caprae) with approximately 200 microg Se/g dw on average (maximum up to 370 microg/g dw) is the richest one in this element among the species surveyed. Several other representatives of the genus Albatrellus are also abundant in selenium. Of the most popular edible wild-grown mushrooms, the King Bolete (Boletus edulis) is considered abundant in selenium as well; on average, it contains approximately 20 microg Se/g dw (maximum up to 70 microg/g dw). Some species of the genus Boletus, such as B. pinicola, B. aereus, B. aestivalis, B. erythropus, and B. appendiculus, can also accumulate considerable amounts of selenium. Some other relatively rich sources of selenium include the European Pine Cone Lepidella (Amanita strobiliformis), which contains, on average, approximately 20 microg Se/g dw (up to 37 microg/g dw); the Macrolepiota spp., with an average range of approximately 5 to < 10 microg/g dw (an exception is M. rhacodes with < 10 microg/g dw); and the Lycoperdon spp., with an average of approximately 5 microg Se/g dw. For several wild-grown species of the genus Agaricus, the selenium content ( approximately 5 microg/g dw) is much greater than that from cultivated Champignon Mushroom; these include A. bisporus, A. bitorquis, A. campestris, A. cesarea, A. campestris, A. edulis, A. macrosporus, and A. silvaticus. A particularly rich source of selenium could be obtained from selenium-enriched mushrooms that are cultivated on a substrate fortified with selenium (as inorganic salt or selenized-yeast). The Se-enriched Champignon Mushroom could contain up to 30 or 110 microg Se/g dw, while the Varnished Polypore (Ganoderma lucidum) could contain up to 72 microg Se/g dw. An increasingly growing database on chemical forms of selenium of mushrooms indicates that the seleno-compounds identified in carpophore include selenocysteine, selenomethionine, Se-methylselenocysteine, selenite, and several unidentified seleno-compounds; their proportions vary widely. Some aspects of environmental selenium occurrence and human body pharmacokinetics and nutritional needs will also be briefly discussed in this review.

Production of polysaccharidases in different carbon sources by Leucoagaricus gongylophorus Möller (Singer), the symbiotic fungus of the leaf-cutting ant Atta sexdens Linnaeus.

Leucoagaricus gongylophorus, the fungus cultured by the leaf-cutting ant Atta sexdens, produces polysaccharidases that degrade leaf components by generating nutrients believed to be essential for ant nutrition. We evaluated pectinase, amylase, xylanase, and cellulase production by L. gongylophorus in laboratory cultures and found that polysaccharidases are produced during fungal growth on pectin, starch, cellulose, xylan, or glucose but not cellulase, whose production is inhibited during fungal growth on xylan. Pectin was the carbon source that best stimulated the production of enzymes, which showed that pectinase had the highest production activity of all of the carbon sources tested, indicating that the presence of pectin and the production of pectinase are key features for symbiotic nutrition on plant material. During growth on starch and cellulose, polysaccharidase production level was intermediate, although during growth on xylan and glucose, enzyme production was very low. We propose a possible profile of polysaccharide degradation inside the nest, where the fungus is cultured on the foliar substrate.

Flowers and fungi use scents to mimic each other.

Some flowering plants mimic the scent and appearance of mushroom fruiting bodies. Fungi may also mimic flowers. In addition, infection of plants by certain fungi can direct the plant to develop nonfunctional floral-like structures that nonetheless primarily serve the reproductive advantage of the fungus. These various mimicries may serve to attract insects that in turn spread fungal spores or plant pollen, thus facilitating sexual reproduction of the cryptic organism.

[Effects of four species of endophytic fungi on the growth and polysaccharide and alkaloid contents of Dendrobium nobile].

OBJECTIVE: To study the effects of four species of endophytic fungi on the growth and polysaccharide and alkaloid contents of cultured Dendrobium nobile. METHOD: Each fungal strain was cultured together with D. nobile, and the plant weight as well as the polysaccharide and alkaloid contents were measured. RESULT: Compared with the control group, Mycena sp. (MF23) was found to decrease the fresh weight of D. nobile by 24.9% (P < 0.01). All four species of fungi have no effect on the dry weight of this plant. On the other hand, Epulorhiza sp. (MF18) and MF23 could increase the percent of dry weight of seedlings (P < 0.05). Epulorhiza sp. (MF15) and MF24 increased the number of aerial roots by 4.25 times and 4.14 times respectively (P < 0.01). MF23 decreased the numbers of basal roots by 46.5% (P < 0.01). MF15, MF18, MF23 and MF24 were shown to increase the content of polysacchride by 153.4%, 52.1%, 18.5% and 76.7%, respectively. MF23 also increased the content of total alkaloid by 18.3%. CONCLUSION: Endophytic fungi cultured together with D. nobile could affect the growth of this plant; they also increase total alkaloid and polysaccharide contents.

Antiinflammatory activity of Tylopilus felleus (Bull. ex Fr.) P. Karst.

Tylopilus felleus lyophylised preparation (Tf) tested in the carrageenin induced oedema test in rats, exhibit a significant inhibition of inflammation at all doses above 50 mg/kg sc while oral administration produced no significant results.