Mature Fruiting Bodies Research Articles

Geographical-based variations in white truffle Tuber magnatum aroma is explained by quantitative differences in key volatile compounds.

The factors that vary the aroma of Tuber magnatum fruiting bodies are poorly understood. The study determined the headspace aroma composition, sensory aroma profiles, maturity and bacterial communities from T.magnatum originating from Italy, Croatia, Hungary, and Serbia, and tested if truffle aroma is dependent on provenance and if fruiting body volatiles are explained by maturity and/or bacterial communities. Headspace volatile profiles were determined using gas chromatography-mass spectrometry-olfactometry (GC-MS-O) and aroma of fruiting body extracts were sensorially assessed. Fruiting body maturity was estimated through spore melanisation. Bacterial community was determined using 16S rRNA amplicon sequencing. Main odour active compounds were present in all truffles but varied in concentration. Aroma of truffle extracts were sensorially discriminated by sites. However, volatile profiles of individual fruiting bodies varied more within sites than across geographic area, while maturity level did not play a role. Bacterial communities varied highly and were partially explained by provenance. A few rare bacterial operational taxonomical units associated with a select few nonodour active volatile compounds. Specificities of the aroma of T.magnatum truffles are more likely to be linked to individual properties than provenance. Some constituents of bacteria may provide biomarkers of provenance and be linked to nonodour active volatiles.

ITRAQ-based comparative proteome analyses of different growth stages revealing the regulatory role of reactive oxygen species in the fruiting body development of Ophiocordyceps sinensis.

In this study, using an isobaric tags for relative and absolute quantitation (iTRAQ ) approach coupled with LC-MS / MS and bioinformatics, the proteomes were analyzed for the crucial three stages covering the fruiting body development of Ophiocordyceps sinensis, including sclerotium (ST), primordium (PR) and mature fruiting body (MF), with a focus on fruiting body development-related proteins and the potential mechanisms of the development. A total of 1,875 proteins were identified. Principal Component Analysis (PCA) demonstrated that the protein patterns between PR and MF were more similar than ST. Differentially accumulated proteins (DAPs) analysis showed that there were 510, 173 and 514 DAPs in the comparisons of ST vs. PR, PR vs. MF and ST vs. MF, respectively. A total of 62 shared DAPs were identified and primarily enriched in proteins related to ‘carbon transport and mechanism’, ‘the response to oxidative stress’, ‘antioxidative activity’ and ‘translation’. KEGG and GO databases showed that the DAPs were enriched in terms of ‘primary metabolisms (amino acid/fatty acid/energy metabolism)’, ‘the response to oxidative stress’ and ‘peroxidase’. Furthermore, 34 DAPs involved in reactive oxygen species (ROS) metabolism were identified and clustered across the three stages using hierarchical clustering implemented in hCluster R package . It was suggested that their roles and the underlying mechanisms may be stage-specific. ROS may play a role in fungal pathogenicity in ST, the fruit-body initiation in PR, sexual reproduction and highland adaptation in MF. Crucial ROS-related proteins were identified, such as superoxide dismutase (SOD, T5A6F1), Nor-1 (T5AFX3), electron transport protein (T5AHD1), histidine phosphotransferase (HPt, T5A9Z5) and Glutathione peroxidase (T5A9V1). Besides, the accumulation of ROS at the three stages were assayed using 2,7-dichlorofuorescin diacetate (DCFH-DA) stanning. A much stronger ROS accumulation was detected at the stage MF, compared to the stages of PR and ST. Sections of ST and fruit-body part of MF were stained by DCFH-DA and observed under the fluorescencemicroscope, showing ROS was distributed within the conidiospore and ascus. Besides, SOD activity increased across the three stages, while CAT activity has a strong increasement in MF compared to the stages of ST and PR. It was suggested that ROS may act in gradient-dependent manner to regulate the fruiting body development. The coding region sequences of six DAPs were analyzed at mRNA level by quantitative real-time PCR (qRT-PCR). The results support the result of DAPs analysis and the proteome sequencing data. Our findings offer the perspective of proteome to understand the biology of fruiting body development and highland adaptation in O. sinensis, which would inform the big industry of this valuable fungus.

Aroma profile of two commercial truffle species from Yunnan and Sichuan, China: inter- and intraspecific variability and shared key compounds

Aroma is central to the worldwide success of truffles as gourmet food and the high prices paid for these edible mushrooms. In this study, volatile organic compounds (VOCs) from fruiting bodies of two Chinese truffles of commercial relevance, Tuber indicum and Tuber pseudohimalayense, were analyzed using headspace solid-phase microextraction (HS-SPME) coupled with gas chromatography-mass spectrometry (GC-MS). We aimed to characterize the aroma profile and determine whether it would be influenced by provenance and stage of maturation. We thus collected and analyzed young, middle mature and mature fruiting bodies of each species from different locations in Yunnan and Sichuan provinces, located in southwestern China. Overall, 76 VOCs were identified, belonging to different chemical classes, i.e. alcohols and phenols, aldehydes and ketones, benzenes and methoxy compounds, hydrocarbons and amines. A large number of volatiles identified in T. indicum and T. pseudohimalayense are reported here for the first time for these truffles. While more than 50% of identified VOCs were produced by both truffle species, considerable differences were present in the aroma profiles of fruiting bodies collected at various maturation stages, revealing a dynamic pattern in the biosynthesis of VOCs. Furthermore, truffles of different provenance had distinct proportions of volatile constituents, suggesting that, besides genetic factors, edaphic and microclimatic conditions influence the synthesis of VOCs in a complex manner.

Bioactivities of Lentinus polychrous and Ganoderma lucidum (Agaricomycetes) Polysaccharide Extracts.

This study aimed to compare the bioactivities of Lentinus polychrous polysaccharide extracts with Ganoderma lucidum. Three hot water-extracted polysaccharide fractions of L. polychrous (LPE), including immature fruiting bodies (im-LPE), mature fruiting bodies (ma-LPE), and mycelium (my-LPE), were analyzed for their phytochemical contents and bioactivities (e.g., anticancer, antiviral, anti-inflammatory, and immunomodulatory effects) compared with G. lucidum extract (GLE). Although GLE had the highest total phenolic and protein contents and lower carbohydrate content than LPE, im-LPE showed strong inhibition on HepG2 cell proliferation as did GLE. GLE and LPE showed antiviral effects on herpes simplex virus-1 and the relative antiproliferative activity was from im-LPE > my-LPE > ma-LPE. However, im-LPE showed the best protective activity on the viral attachment step with some virucidal effects. Moreover, all LPE showed selective and stronger inhibitory effects on LPS-induced RAW264.7 macrophages than those of GLE on genetic expression. Considering the 50% inhibitory concentration values, my-LPE possessed the strongest inhibitory activity on the expression of cyclooxygenase-2 and inducible nitric oxide synthase. However, ma-LPE had the strongest inhibitory effects on interleukin-1β and tumor necrosis factor-α gene expression. The extracts increased splenocyte proliferation under mycohaemagglutinin induction with a relative order of my-LPE > GLE > ma-LPE = im-LPE. In conclusion, LPE showed positive effects and stronger activity than G. lucidum. L. polychrous may have potential for use as an ingredient in functional foods.

Application of digital colorimeter for preliminary characterization of gold nanoparticle swarms produced by Termitomyces heimii using a novel Bioinspired Microfluidics assay

In our laboratory work extending over several years we have successfully studied the biogeochemical role of termite mounds and their occupants the termites and the exosymbiont fungus-Termitomyces. Fungi appear to be promising for large scale production of nanoparticles (NPs) as these are simpler to grow both in laboratory and at industrial scale. This paper reports a novel microfluidics based assay system to detect Gold bioreduction capacity of different tissues in tissue based and cell free environment. Using sterile microtest well, different tissues such as umbo, pileus, lamellae, stipe context, stipe epicutis, pseudorrhiza context, pseudorrhiza epicutis of Termitomyces heimii mature fruitbodies were tested with 200μl chloroauric acid (one mM) and after an interval of 5,10, 15, 30, 45, 60, 120 min and 12, 24 and 48 hours. The results in terms production of distinct nanoparticles were directly visualized microscopically and using mobile based digital colorimeter. Membrane filtered sterile water soluble extracts (SWSE) from the same tissues were similarly screened. The results manifested by mono and polydisperse GNPs and microparticles of mixed size groups demonstrated that cell free system can be potentially useful for bioinspired fabrication of GNPs. Further work in this direction is in progress using several termitomyces cultures.

COMPOSICIÓN MICOQUÍMICA Y ACTIVIDAD ANTIOXIDANTE DE LA SETA Pleurotus ostreatus EN DIFERENTES ESTADOS DE CRECIMIENTO

Pleurotussp. es un género de basidiomicetos ampliamente distribuido a nivel mundial, con especies que revisten importancia, no solo desde el punto de vista comercial por su valor nutricional, sino también por sus propiedades medicinales. Sin embargo, aún son insuficientes los estudios micoquímicos y se desconoce su relación con las propiedades antioxidantes de bioproductos obtenidos de estas setas durante diferentes estados de crecimiento. En el presente trabajo, se determinó el contenido de proteínas, carbohidratos, azúcares reductores, fenoles totales y flavonoides, así como la actividad antioxidante in vitro(ensayos de captación de radicales DPPH y estimación del poder reductor) de extractos acuosos de Pleurotus ostreatusen dos estados de crecimiento (primordios y cuerpos fructíferos maduros) durante la fermentación sólida. El contenido de proteínas, azúcares reductores y flavonoides fue superior en el extracto acuoso de cuerpos fructíferos maduros, mientras que los carbohidratos y fenoles totales fueron mayores en el extracto de primordios. Los valores de EC50en los ensayos de DPPH y los de poder reductor fueron de 1,72 mg/mL y 3,27 mg/mL para el extracto de primordios y de 0,44 mg/mL y 0,48 mg/mL para el de cuerpos fructíferos maduros, respectivamente. Los resultados sugieren que las propiedades antioxidantes de extractos acuosos de Pleurotus ostreatus(primordios y cuerpos fructíferos maduros) reflejan las variaciones en la concentración de moléculas bioactivas, teniendo en cuenta las particularidades fisiológicas de las fases de crecimiento. Ello podría resultar de utilidad para el diseño de protocolos de obtención de bioproductos de Pleurotus ostreatuscon aplicaciones potenciales como antioxidantes en las industrias alimentaria y farmacéutica.

Arcyria versicolor of western mountains, U.S.A. (Myxomycetes: Trichiales: Trichiaceae): a morphological and taxonomic study with observations of nivicolous species

Arcyria versicolor (Trichiales: Trichiaceae) is a distinct myxomycete species described by William Phillips in 1877. The genus Arcyria dates back to Linnaeus in 1753 through the species A. denudata. Arcyria sporangia are brightly colored red, yellow, grey or white, mostly stalked, often in large groups easily seen with the naked eye. Approximately 54 species are known, many are common, and distributed worldwide. Collectors often encounter these colorful species on decaying logs as clusters of many sporangia often covering extensive areas. Arcyria versicolor, collected in the Valles Caldera National Preserve located in the Jemez Mountains of north central New Mexico, is a new record for the state. The nomenclatural history of this species is reviewed and the justification for selection of the species name versicolor is discussed. Arcyria versicolor is accepted as the valid species name and A. vitellina a synonym after examination of type specimens. Environmental parameters for coloration are discussed in general for fruiting bodies of Arcyria and more specifically for nivicolous snowbank species. Transitional stages of plasmodial color to mature fruiting body formation are described for Arcyria versicolor. More than 140 specimens of Arcyria versicolor fruiting bodies were examined with light microscopy and in part illustrated with multifocal computer stacked imaging. Higher magnifications were highlighted using scanning electron microscopy. A more complete and accurate species description is provided for Arcyria versicolor. Differences of fruiting body morphology including habit, color, dehiscence, peridial inner and outer surface features, capillitial ornamentation and size, spore color, size, and ornamentation, and stalk spore-like bodies are described and illustrated. Observation of type specimens from the type locality is illustrated, discussed, and nomenclatural evaluation given for the name selected. Mountain myxomycetes are reviewed based on the observations of T.H. Macbride and his early 1914 paper published in Mycologia. Collection data is presented that compares the dark-spored and light spored nivicolous myxomycetes in the French Alps. The history of renown collectors of nivicolous myxomycetes in western mountains of U.S.A. documents the discovery and study of this special ecological group of myxomycetes. This current paper is the first in a series from an ongoing research project entitled Myxomycetes of New Mexico.

The STRIPAK signaling complex regulates dephosphorylation of GUL1, an RNA-binding protein that shuttles on endosomes.

The striatin-interacting phosphatase and kinase (STRIPAK) multi-subunit signaling complex is highly conserved within eukaryotes. In fungi, STRIPAK controls multicellular development, morphogenesis, pathogenicity, and cell-cell recognition, while in humans, certain diseases are related to this signaling complex. To date, phosphorylation and dephosphorylation targets of STRIPAK are still widely unknown in microbial as well as animal systems. Here, we provide an extended global proteome and phosphoproteome study using the wild type as well as STRIPAK single and double deletion mutants (Δpro11, Δpro11Δpro22, Δpp2Ac1Δpro22) from the filamentous fungus Sordaria macrospora. Notably, in the deletion mutants, we identified the differential phosphorylation of 129 proteins, of which 70 phosphorylation sites were previously unknown. Included in the list of STRIPAK targets are eight proteins with RNA recognition motifs (RRMs) including GUL1. Knockout mutants and complemented transformants clearly show that GUL1 affects hyphal growth and sexual development. To assess the role of GUL1 phosphorylation on fungal development, we constructed phospho-mimetic and -deficient mutants of GUL1 residues. While S180 was dephosphorylated in a STRIPAK-dependent manner, S216, and S1343 served as non-regulated phosphorylation sites. While the S1343 mutants were indistinguishable from wild type, phospho-deficiency of S180 and S216 resulted in a drastic reduction in hyphal growth, and phospho-deficiency of S216 also affects sexual fertility. These results thus suggest that differential phosphorylation of GUL1 regulates developmental processes such as fruiting body maturation and hyphal morphogenesis. Moreover, genetic interaction studies provide strong evidence that GUL1 is not an integral subunit of STRIPAK. Finally, fluorescence microscopy revealed that GUL1 co-localizes with endosomal marker proteins and shuttles on endosomes. Here, we provide a new mechanistic model that explains how STRIPAK-dependent and -independent phosphorylation of GUL1 regulates sexual development and asexual growth.

Quantitative proteomic and metabolomic analysis of Dictyophora indusiata fruiting bodies during post-harvest morphological development

The differences in Dictyophora indusiata fruiting bodies between peach-shaped and mature stage during the postharvest were systematically investigated through quantitative proteomic and metabolomic analyses. A total of 951 differentially expressed proteins were identified, 571 upregulated and 380 downregulated in the mature fruiting body; additionally, 173 upregulated and 165 downregulated differential abundance metabolites were screened. Integrated proteome and metabolome analyses showed that, during the maturation of D. indusiata fruiting bodies, glycerophospholipids were hydrolyzed and drastically decreased, the degradation of glucan was upregulated, the degradation and synthesis of chitin were simultaneously enhanced, and proteins were dominated via catabolism. Along with vigorous material metabolism, energy production was enhanced through the upregulated TCA-cycles and oxidative phosphorylation. In addition, the synthesis of antioxidant substances and the decomposition of peroxides were enhanced in mature fruiting bodies. These omics analyses of D. indusiata provide high-throughput data and reveal the changes in the post-harvest morphological development.

Mycoparasite Hypomyces odoratus infests Agaricus xanthodermus fruiting bodies in nature

Mycopathogens are serious threats to the crops in commercial mushroom cultivations. In contrast, little is yet known on their occurrence and behaviour in nature. Cobweb infections by a conidiogenous Cladobotryum-type fungus identified by morphology and ITS sequences as Hypomyces odoratus were observed in the year 2015 on primordia and young and mature fruiting bodies of Agaricus xanthodermus in the wild. Progress in development and morphologies of fruiting bodies were affected by the infections. Infested structures aged and decayed prematurely. The mycoparasites tended by mycelial growth from the surroundings to infect healthy fungal structures. They entered from the base of the stipes to grow upwards and eventually also onto lamellae and caps. Isolated H. odoratus strains from a diseased standing mushroom, from a decaying overturned mushroom stipe and from rotting plant material infected mushrooms of different species of the genus Agaricus while Pleurotus ostreatus fruiting bodies were largely resistant. Growing and grown A. xanthodermus and P. ostreatus mycelium showed degrees of resistance against the mycopathogen, in contrast to mycelium of Coprinopsis cinerea. Mycelial morphological characteristics (colonies, conidiophores and conidia, chlamydospores, microsclerotia, pulvinate stroma) and variations of five different H. odoratus isolates are presented. In pH-dependent manner, H. odoratus strains stained growth media by pigment production yellow (acidic pH range) or pinkish-red (neutral to slightly alkaline pH range).

Comparative study on growth parameters of Pleurotus florida cultivated on different substrates at Tawang, Arunachal Pradesh, India

The aim of the study was to evaluate effect of different substrates namely finger millet (Eleusine coracana) straw (FMS), paddy (Oryza sativa) straw (PS) and their combination (50% FMS + 50% PS) on growth parameters such as spawn run, pin head appearance, maturation of fruiting bodies, flushing intervals, cropping period, total yield (g) and biological efficiency (B.E %) of Pleurotus florida cultivated at Tawang, Arunachal Pradesh, India. A total of three treatments replicated five times were taken under complete randomize design. The minimum average interval between flushes was recorded on FMS (7.8 days) followed by 50% FMS + 50% PS (12.1 days) for P. florida. The total maximum yield and biological efficiency were noted on FMS (1155.40g and 92.43% respectively) which were significantly (P≤0.5) higher than other tested substrates. This was the first comparative study conducted to evaluate the efficiency of locally available finger millet straw with paddy straw (conventionally used) for the cultivation of oyster mushroom (P. florida) under the high altitude condition of Tawang, Arunachal Pradesh, India.

MAT1-1-3, a Mating Type Gene in the Villosiclava virens, Is Required for Fruiting Bodies and Sclerotia Formation, Asexual Development and Pathogenicity.

Villosiclava virens is the prevalent causative pathogen of rice false smut, a destructive rice disease. Mating-type genes play a vital role in the evolution of mating systems in fungi. Some fungi have lost MAT1-1-3, one of the mating-type genes, during evolution, whereas others still retain MAT1-1-3. However, how MAT1-1-3 regulates the sexual development of heterothallic V. virens remains unknown. Here, we generated the MAT1-1-3 mutants, which exhibited defects in vegetative growth, stress response, pathogenicity, sclerotia formation and fruiting body maturation. An artificial outcrossing inoculation assay showed that the Δmat1-1-3 mutant was unable to produce sclerotia. Unexpectedly, the Δmat1-1-3 mutant could form immature fruiting bodies without mating on potato sucrose agar medium (PSA) compared with the wild-type strain, most likely by activating the truncated MAT1-2-1 transcription to regulate the sexual development. Moreover, RNA-seq data showed that knockout of MAT1-1-3 results in misregulation of a subset of genes involved in sexual development, MAPK signaling, cell wall integrity, autophagy, epigenetic modification, and transcriptional regulation. Collectively, this study reveals that MAT1-1-3 is required for asexual and sexual development, and pathogenicity of V. virens, thereby provides new insights into the function of mating-type genes in the fungi life cycle and infection process.

Genome-wide mRNA and miRNA analysis in the early stages of germ tube outgrowth in Coprinopsis cinerea

Coprinopsis cinerea is a model mushroom-forming basidiomycete which produces basidiospores during sexual reproduction. This fungus is widely used to study fruiting body formation and development. Molecular mechanisms controlling its growth from vegetative mycelium to multicellular mature fruiting body have been studied extensively. However, little is known about the underlying biological processes during germ tube outgrowth or the transition from basidiospores to multinucleate hyphae. To better understand sexual spore germination in fungi, here we examined the time-dependent cellular events at resting, germinating and fully germinated basidiospores of C. cinerea by genome-wide transcriptional and post-transcriptional analyses and by carbohydrate composition analysis. Our results revealed a high demand of protein degradation, and biosynthesis of various compounds at the early stage of basidiospore gemination and dynamic changes of carbohydrate metabolism throughout the germination process. Seven microRNA-like RNAs (milRNAs) were identified in the resting basidiospores of C. cinerea, six of which were basidiospore-specific. Glycogen and trehalose were shown to be the carbon sources supporting the initiation of germ tube outgrowth. One basidiospore-specific milRNA, cci-milR-37, was found to be a potential regulator of glycogen metabolic pathways related to vegetative hyphal growth. Our results demonstrated the mRNA and miRNA-mediated regulation on energy production, protein and carbohydrate metabolisms at the early developmental stages of germ tube to form totipotent hyphae. To our knowledge, this is the first study to show the roles of miRNAs in mushroom basidiospore germination and out-growth.

Optimization of processing parameters for vacuum fried oyster mushroom (Pleurotus ostreatus (Jacquin) P. kummer)

This study is aimed to optimize the processing parameters for the production of a vacuum fried product from oyster mushroom (Pleurotus ostreatus (Jacquin) P. kummer). A 33 factorial experiment was employed following the Central Composite Design (CCD) with fifteen experimental treatments. The treatments were subjected to proximate analysis, total phenolics content, DPPH assay and sensory evaluation. Contour plots of the results of various analyses were superimposed to identify the optimum process and formulation. Variables considered for optimization were maturity, salt concentration and frying time. Response surface regression analysis revealed that maturity significantly affected the crude protein, crude fiber, total phenolics content (TPC) and radical scavenging activity (RSA) while salt concentration significantly affected the ash, crude fat, crude fiber, TPC, aroma and crispiness of the vacuum fried product. On the other hand, frying time significantly affected the ash content. Interaction between variables also significantly affected ash, crude fat, crude fiber, TPC, RSA, crispiness and taste. Parameters considered in the identification of the optimum region were ash content, crude protein, crude fiber, radical scavenging activity and sensory attributes. Minimum values were set for each parameter as follows: ≥ 3% for ash, ≥ 11% for crude protein, ≥ 10% for crude fiber, and ≥ 6.75 for sensory attributes. Superimposed contour plots showed that the optimum region based on set of criteria is at treatment combination of 3 days fruiting body maturity, 3% salt concentration and 35 mins vacuum frying time. Verification test reveals that the optimum vacuum fried product was highly acceptable than the product found outside the optimum region.

Selection and validation of reliable reference genes for Tolypocladium guangdongense gene expression analysis under differentially developmental stages and temperature stresses

Tolypocladium guangdongense, formerly known as Cordyceps guangdongensis, is a widely cultivated fungus of the Cordyceps s.l. species that has been investigated over the last 12 years. It has the potential to be used in a number of applications in the health and pharmaceutical industries for it has shown its high nutritional and medicinal values according to previous animal studies. qRT-PCR (quantitative reverse transcription polymerase chain reaction) is extensively used to analyze the expression pattern and molecular mechanisms of functional genes under differentially experimental conditions. The expression stability of reference genes used for normalization determines the reliability of qRT-PCR results, indicating the importance of selection and validation of reference genes before gene expression analysis. In the present study, three statistical algorithms, geNorm, NormFinder and BestKeeper, were used for analyzing the expression stability of nineteen candidate reference genes (CRGs) in T. guangdongense. Investigation were carried out under differentially experimental conditions, which included differentially developmential stages (mycelia, primordia, young and mature fruiting bodies), different carbon sources, cold and heat stresses. The results showed that histone H4 and tubulin beta chain 2 (β-tub2) were the most and least stable genes, respectively, across all the experimental samples. Moreover, analysis of individual data sets exhibited different stability and expression profiles of reference genes. The vacuolar protein sorting gene VPS was the most stable gene expressed under the differentially developmental stages and temperature stresses, whereas H4 was the most stably expressed gene under different carbon sources. Therefore, it can be proposed that VPS and H4 are the preferred reference genes for normalization of gene expression under different experimental conditions. The results of our present study will enable more accurate evaluation of gene expression in T. guangdongense using the optimal reference gene for qRT-PCR analysis.

Comparative transcriptome analysis revealed genes involved in the fruiting body development of Ophiocordyceps sinensis.

Ophiocordyceps sinensis is a highly valued fungus that has been used as traditional Asian medicine. This fungus is one of the most important sources of income for the nomadic populations of the Tibetan Plateau. With global warming and excessive collection, the wild O. sinensis resources declined dramatically. The cultivation of O. sinensis hasn’t been fully operational due to the unclear genetic basis of the fruiting body development. Here, our study conducted pairwise comparisons between transcriptomes acquired from different growth stages of O. sinensis including asexual mycelium (CM), developing fruiting body (DF) and mature fruiting body (FB). All RNA-Seq reads were aligned to the genome of O. sinensis CO18 prior to comparative analyses. Cluster analysis showed that the expression profiles of FB and DF were highly similar compared to CM. Alternative splicing analysis (AS) revealed that the stage-specific splicing genes may have important functions in the development of fruiting body. Functional enrichment analyses showed that differentially expressed genes (DEGs) were enriched in protein synthesis and baseline metabolism during fruiting body development, indicating that more protein and energy might be required for fruiting body development. In addition, some fruiting body development-associated genes impacted by ecological factors were up-regulated in FB samples, such as the nucleoside diphosphate kinase gene (ndk), β subunit of the fatty acid synthase gene (cel-2) and the superoxide dismutase gene (sod). Moreover, the expression levels of several cytoskeletons genes were significantly altered during all these growth stages, suggesting that these genes play crucial roles in both vegetative growth and the fruiting body development. Quantitative PCR (qPCR) was used to validate the gene expression profile and the results supported the accuracy of the RNA-Seq and DEGs analysis. Our study offers a novel perspective to understand the underlying growth stage-specific molecular differences and the biology of O. sinensis fruiting body development.

Molecular cloning and bioinformatics analyses of a GH3 beta-glucosidase from Auricularia heimuer

The present study reports the isolation and analysis of a beta-glucosidase gene (accession number MN816859) from the fungus Auricularia heimuer. The Aa-bgl gene comprises 2583 bp and encodes a putative polypeptide of 860 amino acids. BlastP analysis revealed conserved glycoside hydrolase family 3 domains. Recombinant plasmid pET-32a-bgl was constructed to overexpress the target protein in Escherichia coli BL21, and sodium dodecyl sulphate polyacrylamide gel electrophoresis analysis revealed target bands between 100 kDa and 135 kDa, consistent with the expected molecular weight. Additionally, the transcription levels of Aa-bgl at different developmental stages were investigated, and the results showed that the expression levels increased with the maturation of the fruiting bodies. In mature fruiting bodies, the levels were 6.69-fold higher than in controls (mycelium at 6 days). We therefore speculate that Aa-bgl may be correlated with development in A. heimuer. The findings provide a theoretical basis for studying the function of this beta-glucosidase in future work.

Ligno(hemi)cellulolytic Enzyme Profiles during the Developmental Cycle of the Royal Oyster Medicinal Mushroom Pleurotus eryngii (Agaricomycetes) Grown on Supplemented Agri-Wastes.

We have determined the production profiles of major ligno(hemi)cellulolytic enzymes at different stages of the mushroom development cycle during industrial scale cultivation of Pleurotus eryngii on supplemented agri-wastes. Endo-1,4-β-glucanase, cellobiohydrolase and endoxylanase levels remained relatively low during substrate colonization, increased sharply when small fruit bodies appeared, and peaked at maturation. β-Glucosidase and β-xylosidase levels decreased when substrate colonization was complete, increased with the appearance of small fruit bodies and primordia, respectively, and reached maxima at maturation. Laccase peaked along with substrate colonization but, after falling sharply in the upper substrate layers, remained relatively low until postinduction. Levels increased slightly when primordia appeared, fell to minimal values during the small and mature fruit body stages, and increased again postharvest. Manganese peroxidase (Mn-P) exhibited a similar pattern initially but high enzyme levels also coincided with primordia formation. Laccase and Mn-P activity patterns were compatible with a lignin-degradation function associated with substrate colonization and, in the former case, a putative role in fruit body morphogenesis. Based on the relatively low levels of polysaccharidases recorded during the initial stages of substrate colonization, we conclude that reducing sugar levels in noncolonized substrate were adequate for sustainable vegetative growth at that stage. We further conclude that the increase in enzyme production later in the developmental cycle was consistent with the replenishment of depleted reducing sugar from cellulose in the growth substrate to levels required for fruit body formation. These data provide new information describing combined temporal and spatial enzyme production profiles throughout the mushroom development cycle under a set of conditions used in industrial scale production.

Combining transcriptomics and metabolomics to reveal the underlying molecular mechanism of ergosterol biosynthesis during the fruiting process of Flammulina velutipes

BackgroundFlammulina velutipes has been recognized as a useful basidiomycete with nutritional and medicinal values. Ergosterol, one of the main sterols of F. velutipes is an important precursor of novel anticancer and anti-HIV drugs. Therefore, many studies have focused on the biosynthesis of ergosterol and have attempted to upregulate its content in multiple organisms. Great progress has been made in understanding the regulation of ergosterol biosynthesis in Saccharomyces cerevisiae. However, this molecular mechanism in F. velutipes remains largely uncharacterized.ResultsIn this study, nine cDNA libraries, prepared from mycelia, young fruiting bodies and mature fruiting bodies of F. velutipes (three replicate sets for each stage), were sequenced using the Illumina HiSeq™ 4000 platform, resulting in at least 6.63 Gb of clean reads from each library. We studied the changes in genes and metabolites in the ergosterol biosynthesis pathway of F. velutipes during the development of fruiting bodies. A total of 13 genes (6 upregulated and 7 downregulated) were differentially expressed during the development from mycelia to young fruiting bodies (T1), while only 1 gene (1 downregulated) was differentially expressed during the development from young fruiting bodies to mature fruiting bodies (T2). A total of 7 metabolites (3 increased and 4 reduced) were found to have changed in content during T1, and 4 metabolites (4 increased) were found to be different during T2. A conjoint analysis of the genome-wide connection network revealed that the metabolites that were more likely to be regulated were primarily in the post-squalene pathway.ConclusionsThis study provides useful information for understanding the regulation of ergosterol biosynthesis and the regulatory relationship between metabolites and genes in the ergosterol biosynthesis pathway during the development of fruiting bodies in F. velutipes.

De novo transcriptome and proteome analysis of Dictyophora indusiata fruiting bodies provides insights into the changes during morphological development

De novo transcriptome assembly and shotgun proteome analysis of Dictyophora indusiata fruiting bodies were performed. A total of 19,704 unigenes were sequenced, and 4380 proteins were identified. Annotation and functional analysis of the identified proteins were significantly enriched in small molecule synthetic and metabolic processes, protein modification regulation (phosphorylation and ubiquitination), and vesicle transport. Furthermore, quantitative developmental transcriptome analysis was performed between the peach-shaped and mature fruiting bodies, and the results revealed that the metabolism and transport activities were upregulated in the mature stage, while protein translation was downregulated; this regulation is likely the main reason for the significant changes in the nutrients of fruiting bodies. Furthermore, the cell wall stress-dependent MAPK sub-pathway was activated in the mature stage, and fungal cell wall degradation-related genes were upregulated, which could promote reconstruction of the cell wall and might play a key role in the morphological development of D. indusiata fruiting bodies.