The impact of continuous cultivation of Ganoderma lucidum on soil nutrients, enzyme activity, and fruiting body metabolites.

To explore the impacts of continuous Ganoderma lucidum cultivation on soil physicochemical factors, soil enzyme activity, and the metabolome of Ganoderma lucidum fruiting bodies, this study conducted two consecutive years of cultivation on the same plot of land. Soil physicochemical factors and enzyme activity were assessed, alongside non-targeted metabolomic analysis of the Ganoderma lucidum fruiting bodies under continuous cultivation. The findings unveiled that in the surface soil layer (0-15 cm), there was a declining trend in organic matter, ammonium nitrogen, available phosphorus, available potassium, pH, polyphenol oxidase, peroxidase, alkaline phosphatase, and sucrase, whereas nitrate nitrogen, electrical conductivity (EC), and salt content exhibited an upward trend. Conversely, in the deeper soil layer (15-30 cm), organic matter, ammonium nitrogen, available potassium, alkaline phosphatase, and sucrase demonstrated a decreasing trend, while nitrate nitrogen, available phosphorus, pH, EC, salt content, polyphenol oxidase, and soil peroxidase showed an increasing trend. Metabolomic analysis of Ganoderma lucidum fruiting bodies distinguished 64 significantly different metabolites between the GCK and GT groups, with 39 components having markedly higher relative contents in GCK and 25 components having significantly lower relative contents in GCK compared to GT. Moreover, among these metabolites, there were more types with higher contents in the fruiting bodies harvested in the first year (GCK) compared to those harvested in the second year (GT), with pronounced differences. KEGG pathway analysis revealed that GCK exhibited more complex metabolic pathways compared to GT. The metabolites of Ganoderma lucidum fruiting bodies were predominantly influenced by soil physicochemical factors and soil enzyme activity. In the surface soil layer (0-15 cm), the metabolome was significantly affected by soil pH, soil organic matter, available phosphorus, and soil alkaline phosphatase, while in the deeper soil layer (15-30 cm), differences in the Ganoderma lucidum metabolome were more influenced by soil alkaline phosphatase, soil catalase, pH, nitrate nitrogen, and soil sucrase.

Changes of the Physicochemical Properties and Structural Characteristics of Alkali-Extracted Polysaccharides from Agrocybe cylindracea Across the Growth Process.

Polysaccharides derived from Agrocybe cylindracea have been demonstrated to exhibit various bioactivities. However, studies on their structural characteristics during the growth process are limited. This study aimed to compare the physicochemical properties and structural characteristics of alkali-extracted polysaccharides from A. cylindracea fruiting bodies (JACP) across four growth stages. Results showed that the extraction yields and protein levels of JACP declined along with the growth of A. cylindracea, while the contents of neutral sugar and glucose increased significantly. However, JACP exhibited structural characteristics similar to those across the four stages. Four polysaccharide subfractions were isolated from each growth stage, including JACP-Et30, JACP-Et50, JACP-Et60, and JACP-Et70. JACP-Et30 from the four stages and JACP-Et50 from the initial three stages were identified as heteroglucans with ß-1,3-d-Glcp and ß-1,6-d-Glcp residues as main chains, respectively. However, other subfractions were considered as ß-1,6-d-glucans containing minor glucuronic acid. These subfractions were predominantly replaced by Glcp residues at the O-3 and O-6 positions. Overall, while JACP exhibited variable physicochemical properties, its structural characteristics remained stable during the growth process, offering new insights into its potential applications in the food and medicinal industries.

Comprehensive untargeted lipidomics study of black morel (Morchella sextelata) at different growth stages.

The black morel (Morchella sextelata) is a valuable edible and medicinal mushroom appreciated worldwide. Here, lipidomic profiles and lipid dynamic changes during the growth of M. sexletata were analyzed using ultra-performance liquid chromatography coupled with mass spectrometry. 203 lipid molecules, including four categories and fourteen subclasses, were identified in mature fruiting bodies, with triacylglycerol being the most abundant (37.00 %). Fatty acid composition analysis revealed that linoleic acid was the major fatty acid among the free fatty acids, glycerolipids and glycerophospholipids. The relative concentration of lipids in M. sextelata changed significantly during its growth, from which 12 and 29 differential lipid molecules were screened out, respectively. Pathway analysis based on these differential lipids showed that glycerophospholipid metabolism was the major pathway involved in the growth of M. sextelata. Our study provides a comprehensive understanding of the lipids in M. sextelata and will facilitate the development and utilization of M. sextelata.

Identification of two genes essential for basidiospore formation during the postmeiotic stages in Pleurotus ostreatus.

A sporeless strain is an important breeding target in the mushroom industry. However, basidiospore production in the oyster mushroom Pleurotus ostreatus has been shown to be impaired by single-gene mutations in only two meiosis-related genes, mer3 and msh4. This study proposed a strategy for identifying the genes essential for basidiospore formation after meiotic division to determine new targets for molecular breeding. RNA-seq analysis was performed to identify P. ostreatus genes that are specifically expressed in the gill tissue of fruiting bodies, where basidiospore formation occurs. Transcriptome data during fruiting development of Coprinopsis cinerea, in which the meiotic steps progress synchronously, were then used to identify genes that are active in the postmeiotic stages. Based on these comparative analyses, five P. ostreatus genes were identified. Plasmids containing expression cassettes for hygromycin B-resistance screening, Cas9, and single-guide RNA targeting each gene were introduced into the protoplasts of dikaryotic strain, PC9×#64, to generate dikaryotic gene disruptants. Among the obtained transformants, three dikaryotic pcl1 disruptants and two cro6c disruptants did not produce basidiospores. Microscopic analyses indicated that spore formation was arrested at particular stages in these gene disruptants. These results indicate that these two genes are essential for mature spore formation in this fungus.

High Throughput Identification of the Potential Antioxidant Peptides in Ophiocordyceps sinensis.

Ophiocordyceps sinensis, an ascomycete caterpillar fungus, has been used as a Traditional Chinese Medicine owing to its bioactive properties. However, until now the bio-active peptides have not been identified in this fungus. Here, the raw RNA sequences of three crucial growth stages of the artificially cultivated O. sinensis and the wild-grown mature fruit-body were aligned to the genome of O. sinensis. Both homology-based prediction and de novo-based prediction methods were used to identify 8541 putative antioxidant peptides (pAOPs). The expression profiles of the cultivated mature fruiting body were similar to those found in the wild specimens. The differential expression of 1008 pAOPs matched genes had the highest difference between ST and MF, suggesting that the pAOPs were primarily induced and play important roles in the process of the fruit-body maturation. Gene ontology analysis showed that most of pAOPs matched genes were enriched in terms of 'cell redox homeostasis', 'response to oxidative stresses', 'catalase activity', and ' integral component of cell membrane'. A total of 1655 pAOPs was identified in our protein-seqs, and some crucial pAOPs were selected, including catalase, peroxiredoxin, and SOD [Cu-Zn]. Our findings offer the first identification of the active peptide ingredients in O. sinensis, facilitating the discovery of anti-infectious bio-activity and the understanding of the roles of AOPs in fungal pathogenicity and the high-altitude adaptation in this medicinal fungus.

Functional characterization of the developmental genes asm2, asm3, and spt3 required for fruiting body formation in the filamentous ascomycete Sordaria macrospora.

The formation of fruiting bodies is one of the most complex developmental processes in filamentous ascomycetes. It requires the development of sexual structures that give rise to meiosporangia (asci) and meiotic spores (ascospores) as well as surrounding structures for protection and dispersal of the spores. Previous studies have shown that these developmental processes are accompanied by significant changes of the transcriptome, and comparative transcriptomics of different fungi as well as the analysis of transcriptome changes in developmental mutants have aided in the identification of differentially regulated genes that are themselves involved in regulating fruiting body development. In previous analyses, we used transcriptomics to identify the genes asm2 and spt3, which result in developmental phenotypes when deleted in Sordaria macrospora. In this study, we identified another gene, asm3, required for fruiting body formation, and performed transcriptomics analyses of Δasm2, Δasm3, and Δspt3. Deletion of spt3, which encodes a subunit of the SAGA complex, results in a block at an early stage of development and drastic changes in the transcriptome. Deletion mutants of asm2 and asm3 are able to form fruiting bodies, but have defects in ascospore maturation. Transcriptomics analysis of fruiting bodies revealed a large overlap in differentially regulated genes in Δasm2 and Δasm3 compared to the wild type. Analysis of nuclear distribution during ascus development showed that both mutants undergo meiosis and postmeiotic divisions, suggesting that the transcriptomic and morphological changes might be related to defects in the morphogenesis of structural features of the developing asci and ascospores.

Courtship Ritual of Male and Female Nuclei during Fertilization in Neurospora crassa.

Sexual reproduction is a key process influencing the evolution and adaptation of animals, plants, and many eukaryotic microorganisms, such as fungi. However, the sequential cell biology of fertilization and the associated nuclear dynamics after plasmogamy are poorly understood in filamentous fungi. Using histone-fluorescent parental isolates, we tracked male and female nuclei during fertilization in the model ascomycete Neurospora crassa using live-cell imaging. This study unravels the behavior of trichogyne resident female nuclei and the extraordinary manner in which male nuclei migrate up the trichogyne to the protoperithecium. Our observations raise new fundamental questions about the modus operandi of nucleus movements during sexual reproduction, male and female nuclear identity, guidance of nuclei within the trichogyne and, unexpectedly, the avoidance of "polyspermy" in fungi. The spatiotemporal dynamics of male nuclei within the trichogyne following plasmogamy are also described, where the speed and the deformation of male nuclei are of the most dramatic observed to date in a living organism. IMPORTANCE Using live-cell fluorescence imaging, for the first time we have observed live male and female nuclei during sexual reproduction in the model fungus Neurospora crassa. This study reveals the specific behavior of resident female nuclei within the trichogyne (the female organ) after fertilization and the extraordinary manner in which male nuclei migrate across the trichogyne toward their final destination, the protoperithecium, where karyogamy takes place. Importantly, the speed and deformation of male nuclei were found to be among the most dramatic ever observed in a living organism. Furthermore, we observed that entry of male nuclei into protoperithecia may block the entry of other male nuclei, suggesting that a process analogous to polyspermy avoidance could exist in fungi. Our live-cell imaging approach opens new opportunities for novel research on cell-signaling during sexual reproduction in fungi and, on a broader scale, nuclear dynamics in eukaryotes.

Effects of exogenous ascorbic acid on the mycelia growth and primordia formation of Pleurotus ostreatus.

Primordia formation is the first and most critical step in the development of fruiting bodies of edible fungi. In this study, the effects of exogenous ascorbic acid (ASA) on the Pleurotus ostreatus mycelia growth and primordia formation were researched and the results showed that the growth rate of P. ostreatus mycelia was accelerated and the time of primordia formation was advanced. The protein content and ascorbate oxidase (AAO) activity analysis showed that with the increase of ASA concentration, the protein content of mycelia first decreased and then increased, and in a certain concentration range, exogenous ASA could significantly promote the activity of AAO. Further expression analysis of the development regulating genes (Pofst3 and Pofst4) as well as blue light receptor coding genes (PoWC-1 and PoWC-2) showed the expression levels of those four genes all changed after the exogenous ASA addition, which indicated that the expression changes of PoWC-1 and PoWC-2, two key genes in the light morphogenesis, might affect the expression levels of development regulating genes Pofst3 and Pofst4, so as to lead to the formation of primordia in advance.

Evaluación del crecimiento radial de Lactarius volemus en medio semisólido / Evaluation of radial growth of Lactarius volemus in semisolid medium

Objetivo: Evaluar el crecimiento radial de Lactarius volemus en cinco medios de cultivo semisólidos in vitro.Materiales y métodos: Cuerpos fructíferos de L. volemus provenientes de la Sierra Norte del estado de Oaxaca, México, se cultivaron en laboratorio en medios Agar Papa Dextrosa, Agar Czapek-Dox, Agar Extracto de Malta, Agar Papa Sacarosa y Agar Dextrosa Saboraud; mediante dos técnicas de sembrado. Se evaluaron las características morfológicas de colonias obtenidas de distintas muestras del cuerpo fructífero, así como el crecimiento radial de cada una.Resultados: El crecimiento colonial evaluado permitió seleccionar un medio que reúne las condiciones óptimas para el cultivo de Lactarius volemus in vitro. No todas las muestras utilizadas desarrollan un crecimiento abundante: la muestra proveniente del látex presenta un crecimiento escaso.Conclusiones: Con la evaluación del crecimiento radial de Lactarius volemus se obtiene una referencia directa del ciclo de crecimiento de esta especie; es posible identificar las fases exponencial y estacionaria pero las condiciones del medio no permiten evaluar la fase de muerte debido a la deshidratación y reducción del agar.(AU)

Objective: To evaluate the radial growth of Lactarius volemus in five semi-solid culture media in vitro.Materials and methods: Fruitful bodies of L. volemus from the Sierra Norte of the state of Oaxaca, Mexico, were cultured in the laboratory in Potato Dextrose Agar Papa, Czapek-Dox Agar, Malt Extract Agar, Potato Sucrose Agar and Dextrose Saboraud Agar; using two seeding techniques. The morphological characteristics of colonies obtained from different samples of the fruiting body were evaluated, as well as the radial growth of each one.Results: The evaluated colonial growth allowed to select a culture medium that meets the optimal conditions for the cultivation of Lactarius volemus in vitro. Not all samples used develop abundant growth: the sample from latex shows little growth.Conclusions: With the evaluation of the radial growth of Lactarius volemus a direct reference to the growth cycle of this species is obtained; it is possible to identify the exponential and stationary phases but the conditions of the medium do not allow evaluating the phase of death due to dehydration and reduction of the agar.(AU)

Endogenous bacteria inhabiting the Ophiocordyceps highlandensis during fruiting body development.

BACKGROUND: The genus Ophiocordyceps, which includes Ophiocordyceps sinensis, has been demonstrated to be one of the most valuable medicinal taxa. The low rate of larval infection and slow development that characterize the cultivation of this genus should be urgently addressed. To identify potential bioinoculants that stimulate the growth of Ophiocordyceps, O. highlandensis was selected as a model system, and a total of 72 samples were collected to systematically compare the microbial communities present during fruiting body development. By applying high-throughput 16S and ITS2 amplicon sequencing technology, the bacterial and fungal communities were identified in O. highlandensis and its surrounding soil, and the functional dynamics of the bacteria were explored. RESULTS: The results indicate that the most abundant bacteria across all the samples from O. highlandensis were Proteobacteria, Firmicutes and Bacteroidetes, while members of Ascomycota were detected among the fungi. The pathways enriched in the developmental stages were associated with carbohydrate degradation, nucleotides and pyridoxal biosynthesis, and the TCA cycle. Compared with that in the fungal community, an unexpectedly high taxonomic and functional fluctuation was discovered in the bacterial community during the maturation of O. highlandensis. Furthermore, bipartite network analysis identified four potential supercore OTUs associated with O. highlandensis growth. CONCLUSIONS: All the findings of this study suggest unexpectedly high taxonomic and functional fluctuations in the bacterial community of O. highlandensis during its maturation. O. highlandensis may recruit different endogenous bacteria across its life cycle to enhance growth and support rapid infection. These results may facilitate Ophiocordyceps cultivation and improve the development of strategies for the identification of potential bioinoculant resources.

Transcriptome data reveal conserved patterns of fruiting body development and response to heat stress in the mushroom-forming fungus Flammulina filiformis.

Mushroom-forming fungi are complex multicellular organisms that form the basis of a large industry, yet, our understanding of the mechanisms of mushroom development and its responses to various stresses remains limited. The winter mushroom (Flammulina filiformis) is cultivated at a large commercial scale in East Asia and is a species with a preference for low temperatures. This study investigated fruiting body development in F. filiformis by comparing transcriptomes of 4 developmental stages, and compared the developmental genes to a 200-genome dataset to identify conserved genes involved in fruiting body development, and examined the response of heat sensitive and -resistant strains to heat stress. Our data revealed widely conserved genes involved in primordium development of F. filiformis, many of which originated before the emergence of the Agaricomycetes, indicating co-option for complex multicellularity during evolution. We also revealed several notable fruiting-specific genes, including the genes with conserved stipe-specific expression patterns and the others which related to sexual development, water absorption, basidium formation and sporulation, among others. Comparative analysis revealed that heat stress induced more genes in the heat resistant strain (M1) than in the heat sensitive one (XR). Of particular importance are the hsp70, hsp90 and fes1 genes, which may facilitate the adjustment to heat stress in the early stages of fruiting body development. These data highlighted novel genes involved in complex multicellular development in fungi and aid further studies on gene function and efforts to improve the productivity and heat tolerance in mushroom-forming fungi.

Study on the community structure and function of symbiotic bacteria from different growth and developmental stages of Hypsizygus marmoreus.

BACKGROUND: The symbiotic bacteria associated with edible fungi are valuable microbial resources worthy of in-depth exploration. It is important to analyze the community structure and succession of symbiotic bacteria in mushrooms. This can assist in the isolation of growth-promoting strains that have an essential relationship with the cultivation cycle as well as the agronomic traits and yields of fruiting bodies. RESULTS: In all of the samples from cultivation bags of Hypsizygus marmoreus, 34 bacterial phyla were detected. Firmicutes was the most abundant bacterial phylum (78.85%). The genus Serratia showed an exponential increase in abundance in samples collected from the cultivation bags in the mature period, reaching a peak abundance of 55.74% and the dominant symbiotic flora. The most predominant strain was Serratia odorifera HZSO-1, and its abundance increased with the amount of hyphae of H. marmoreus. Serratia odorifera HZSO-1 could reside in the hyphae of H. marmoreus, promote growth and development, shorten the fruiting cycle by 3-4 days, and further increase the fruiting body yield by 12%. CONCLUSIONS: This study is a pioneering demonstration of the community structure of the symbiotic microbiota and bacteria-mushroom interaction in the growth and development of edible fungi. This work lays a theoretical foundation to improve the industrial production of mushrooms with symbiotic bacteria as assisting agents.

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.

Characterization of morphological changes within stromata during sexual reproduction in Aspergillus flavus.

Aspergillus flavus contaminates agricultural products worldwide with carcinogenic aflatoxins that pose a serious health risk to humans and animals. The fungus survives adverse environmental conditions through production of sclerotia. When fertilized by a compatible conidium of an opposite mating type, a sclerotium transforms into a stroma within which ascocarps, asci, and ascospores are formed. However, the transition from a sclerotium to a stroma during sexual reproduction in A. flavus is not well understood. Early events during the interaction between sexually compatible strains of A. flavus were visualized using conidia of a green fluorescent protein (GFP)-labeled MAT1-1 strain and sclerotia of an mCherry-labeled MAT1-2 strain. Both conidia and sclerotia of transformed strains germinated to produce hyphae within 24 h of incubation. Hyphal growth of these two strains produced what appeared to be a network of interlocking hyphal strands that were observed at the base of the mCherry-labeled sclerotia (i.e., region in contact with agar surface) after 72 h of incubation. At 5 wk following incubation, intracellular green-fluorescent hyphal strands were observed within the stromatal matrix of the mCherry-labeled strain. Scanning electron microscopy of stromata from a high- and low-fertility cross and unmated sclerotia was used to visualize the formation and development of sexual structures within the stromatal and sclerotial matrices, starting at the time of crossing and thereafter every 2 wk until 8 wk of incubation. Morphological differences between sclerotia and stromata became apparent at 4 wk of incubation. Internal hyphae and croziers were detected inside multiple ascocarps that developed within the stromatal matrix of the high-fertility cross but were not detected in the matrix of the low-fertility cross or the unmated sclerotia. At 6 to 8 wk of incubation, hyphal tips produced numerous asci, each containing one to eight ascospores that emerged out of an ascus following the breakdown of the ascus wall. These observations broaden our knowledge of early events during sexual reproduction and suggest that hyphae from the conidium-producing strain may be involved in the early stages of sexual reproduction in A. flavus. When combined with omics data, these findings could be useful in further exploration of the molecular and biochemical mechanisms underlying sexual reproduction in A. flavus.

Transcriptomic analysis of the orchestrated molecular mechanisms underlying fruiting body initiation in Chinese cordyceps.

Chinese cordyceps, the fruiting body of the Chinese caterpillar fungus (Ophiocordyceps sinensis, syn. Cordyceps sinensis), is among the most valuable traditional Chinese medicine fungi. Transcriptomic analysis of O. sinensis has revealed several aspects of its life cycle and ecological importance. However, the molecular mechanisms involved in fruiting body initiation remain unclear. The developmental transcriptomes were analyzed from three tissues at the fruiting body initiation stage, namely, the mycelium, sclerotium and primordium. Principal component analysis showed that in the three tissues, the gene expression patterns differed from each other. The functional analysis of differentially expressed genes showed that DNA synthesis and cell division were active in the primordium. In addition, the function of the mycelium was to absorb certain substances from the environment and the sclerotium was the metabolism center of O. sinensis. Genes participating in the mitogen-activated protein kinase (MAPK) signal pathway were involved in fruiting body initiation. Two environmental sensing genes, including a pheromone receptor gene (OSIN6252) and an amino acid sensing gene (OSIN6398), were highly expressed in the primordium, suggesting their important roles in initiation. These results provided insights into the orchestrated functions and gene profiles of different O. sinensis tissues at the key stage. These findings will aid in revealing the underlying mechanisms of fruiting body initiation, which will further benefit artificial cultivation.

Nutritional and Biochemical Characterization of Panus lecomtei Mushroom (Agaricomycetes) from India and Its Cultivation.

Panus lecomtei is emerging as an edible mushroom found worldwide and particularly in the Northern Hemisphere. The mushroom contains a substantial amount of useful nutritional and medicinal compounds. In the present study, we have examined a specimen of P. lecomtei submitted to the ICAR-Directorate of Mushroom Research gene bank. The specimen was examined for taxonomical characters using classical and molecular tools. Attempts were made for cultivation of this mushroom under controlled conditions using sawdust-based substrate. The specimen was characterized by its purplish fruiting body having coarse, rigid, dense hairs on the cap, pubescent stipe, and abundant metuloids. Molecular identification through conserved ITS region was done and the sequence was deposited in NCBI GenBank under accession number MN332200. Nutritional profiling and biochemical analysis showed that the mushroom contained high carbohydrate but low fat contents. The mushroom showed the presence of phenolics, ß-carotene, and lycopene. The analysis also showed substantial antioxidant properties in the mushroom. The findings presented herein point out that P. lecomtei can be used as a potential edible mushroom for diversification of mushroom production in India.

Characterization and expression pattern analysis of pheromone receptor-like genes in Winter Mushroom Flammulina filiformis.

Pheromone receptor-like genes (PRLGs) belong to the G protein-coupled receptors (GPCRs) family that interacts with biotic and abiotic stimulants and transmits signals to intracellular downstream pathways in eukaryotic cells. In this study, we investigated the structure and expressions patterns of PRLGs in Winter Mushroom Flammulina filiformis. Based on the alignment analysis, the structure of PRLGs was found conserved in F. filiformis strains expect few single-nucleotide polymorphism (SNP) sites. Six PRLGs were found at five different unlinked loci, scattered in the genomes of F. filiformis strains. These genes contain 2-5 introns; however, the introns were not found in the same relative positions regarding the encoded protein sequences in tested strains of F. filiformis. Three conserved motifs were identified in peptides structures of PRLGs, however, FfSte3.s6 contained only two types, suggests its difference in evolution and function. We have further analyzed the expression patterns of each PRLGs in different developmental stages of the fruiting body in F. filiformis by quantitative real-time polymerase chain reaction (qRT-PCR). The results exhibited expression variation of PRLGs at different developmental stages of the F. filiformis. Especially, FfSte3.s1 and FfSte3.s2 exhibited maximum expression level in mycelia stage. Other PRLGs exhibited high expression level in fruiting body stages. This study suggests that PRLGs could be vital genes involving in fruiting body development in F. filiformis. However, further studies could be performed to reveal their specific functional pathways in the fruiting body development.

Carbon metabolism and transcriptome in developmental paths differentiation of a homokaryotic Coprinopsis cinerea strain.

The balance and interplay between sexual and asexual reproduction is one of the most intriguing mysteries in the study of fungi. The choice of developmental strategy reflects the ability of fungi to adapt to the changing environment. However, the evolution of developmental paths and the metabolic regulation during differentiation and morphogenesis are poorly understood. Here, an analysis was performed of carbohydrate metabolism and gene expression regulation during the early differentiation process from the vegetative mycelium, to the differentiated structures, fruiting body, oidia and sclerotia, of a homokaryotic fruiting Coprinopsis cinerea strain A43mutB43mut pab1-1 #326. Changes during morphogenesis and the evolution of developmental strategies were followed. Conversion between glucose and glycogen and between glucose and beta-glucan were the main carbon flows in the differentiation processes. Genes related to carbohydrate transport and metabolism were significantly differentially expressed among paths. Sclerotia displayed a set of specifically up-regulated genes that were enriched in the carbon metabolism and energy production and conversion processes. Evolutionary transcriptomic analysis of four developmental paths showed that all transcriptomes were under the purifying selection, and the more stressful the environment, the younger the transcriptome age. Oidiation has the lowest value of transcriptome age index (TAI) and transcriptome divergence index (TDI), while the fruiting process has the highest of both indexes. These findings provide new insights into the regulations of carbon metabolism and gene expressions during the early stages of fungal developmental paths differentiation, and improve our understanding of the evolutionary process of life history and reproductive strategy in fungi.

Hymenophore configuration of the oak mazegill (Daedalea quercina).

The complex hymenophore configuration of the oak mazegill (Daedalea quercina, Polyporales) is rarely quantified, although quantifications are important analytical tools to assess form and growth. We quantified the hymenophore configuration of the oak mazegill by manual counting of tubes and tubular branches and ends. Complementary measurements were made with the software AngioTool. We found that the number of tubular branches and ends varied substantially between specimens, with a positive correlation with hymenophore area (5-51 cm2). We then measured complexity as tubular branches and ends per area, and complexity was not correlated with the size of the basidiocarps. Basidiocarps from two locations were compared (Hald ege, N = 11; Hvidding krat, N = 7), and the prevalence of branches and that of ends were greater in the Hvidding krat hymenophores (P < 0.001 and P = 0.029, respectively). Additionally, lacunarity, a measure of complexity ("gappiness"), gave a higher score for the Hald ege hymenophores (P = 0.002). Lacunarity analysis of multiple species of Polyporales showed that the oak mazegill hymenophore is comparatively complex. Concerning factors that affect hymenophore complexity of the oak mazegill, we observed that greater hymenophore complexity was associated with abrupt boundaries between growth zones on the pileus surface. Several years of monitoring documented that basidiocarps can remodel to gravitational changes and heal from damage. In conclusion, intra- and interspecies differences of hymenophore configuration can be quantified. In oak mazegill, hymenophore complexity is not dependent on size per se, although abrupt borders between growth zones are associated with increased complexity. Some of the variation between basidiocarps may reflect aspects of the ecology of the individual fungus.

Comparative Studies on Bioactive Compounds, Ganoderic Acid Biosynthesis, and Antioxidant Activity of Pileus and Stipes of Lingzhi or Reishi Medicinal Mushroom, Ganoderma lucidum (Agaricomycetes) Fruiting Body at Different Growth Stages.

Total phenolics, flavonoids, and polysaccharides, and individual ganoderic acid (GA) contents, antioxidant capacity, and transcription levels of key enzyme genes involved in GA biosynthesis in pileus and stipes of Ganoderma lucidum fruiting body at different growth stages were investigated in this study. Results showed that the highest total phenolics and total flavonoids contents were determined in stipes at spore maturity stage, resulting in high antioxidant activity, while the highest total polysaccharide content was found in pileus at the same stage. The pileus contained more GA than the stipes, and higher contents of ganoderic acid A and D were found at fruiting body mature stage while that of ganoderic acid B, C2, and G were found at bud elongation stage. Results from quantitative real-time PCR indicated that higher gene transcription levels of hydroxyl methylglutaryl-CoA reductase (hmgr), farnesyl pyrophosphate synthase (fps), squalene synthase (sqs), and oxidosqualene cyclase (osc) were found in pileus at bud elongation stage. Our findings will be helpful for understanding the biosynthesis of bioactive components and determining the harvest time for the desired G. lucidum fruiting bodies.