De novo transcriptome assembly and comprehensive assessment provide insight into fruiting body formation of Sparassis latifolia.

The genes associated with fruiting body formation of Sparasis latifolia are valuable for improving mushroom breeding. To investigate this process, 4.8 × 108 RNA-Seq reads were acquired from three stages: hyphal knot (SM), primordium (SP), and primordium differentiation (SPD). The de novo assembly generated a total of 48,549 unigenes, of which 71.53% (34,728) unigenes could be annotated by at least one of the KEGG (Kyoto Encyclopedia of Genes and Genomes), GO (Gene Ontology), and KOG (Eukaryotic Orthologous Group) databases. KEGG and KOG analyses respectively mapped 32,765 unigenes to 202 pathways and 19,408 unigenes to 25 categories. KEGG pathway enrichment analysis of DEGs (differentially expressed genes) indicated primordium initiation was significantly related to 66 pathways, such as "Ribosome", "metabolism of xenobiotics by cytochrome P450", and "glutathione metabolism" (among others). The MAPK and mTOR signal transduction pathways underwent significant adjustments during the SM to SP transition. Further, our research revealed the PI3K-Akt signaling pathway related to cell proliferation could play crucial functions during the development of SP and SPD. These findings provide crucial candidate genes and pathways related to primordium differentiation and development in S. latifolia, and advances our knowledge about mushroom morphogenesis.

Nutritional analysis and molecular characterization of hybrid mushrooms developed through intergeneric protoplast fusion between Pleurotus sajor-caju and Calocybe indica with the purpose to achieve improved strains.

Two edible mushrooms Calocybe indica and Pleurotus sajor-caju were chosen as parent strains in this study to approach the concept of hybridization through the protoplast fusion technique. Protoplast fusion in presence of polyethylene glycol (PEG) was conducted between the parent strains and by further double selection screening method, six somatic hybrid lines were developed. Those fruit bodies of the hybrid lines showed phenotypic resemblance with Pleurotus sajor-caju when grown on paddy straw under favorable conditions. The hybridity of the newly developed somatic hybrid strains was established by barrage reaction, morphological traits, fruitbody parameter and, inter single sequence repeat (ISSR) profiling. One-way analysis of variance (ANOVA) was used for the analysis of phenotypic data of hybrid lines and parents. Five ISSR primers were used to generate 51 amplified DNA fragments ranged between 250 and 3000 bp in size in six hybrids and two parents with 90.19% polymorphism. Some of the hybrids contain some non-parental bands which indicate that recombination might happen in the hybrid genome hence confirming the hybridity of newly developed strains. The dendrogram was created using the Average Linkage (Between Groups) method based on ISSR profiling and genetic distance between parent-hybrids and hybrid-hybrid was analyzed by Jaccard's proximity matrix. A definite improvement in nutritional properties and biological activity was observed in the study. Due to ease in their cultivation, it can play a significant role in the rural economic development.

A phylogenetic assessment of Pholiota and the new genus Pyrrhulomyces.

Multigene data sets were assembled to evaluate the phylogeny of species attributed to the genus Pholiota sensu A.H. Sm. & Hesler. This effort included generation of just more than 200 new sequences from 19 type collections of Pholiota and recent samples from East Asia. Phylogenetic analyses reinforced the autonomous phylogenetic positions of pholiotoid taxa in the genera Flammula (Hymenogastraceae) and Kuehneromyces (Strophariaceae). Samples of Pholiota astragalina from diverse geographic regions split into two species-level lineages but occupied an isolated phylogenetic position apart from Pholiota sensu stricto. The new genus Pyrrhulomyces is described to accommodate P. astragalina and a new cryptic species from the Southern Appalachians, Pyrrhulomyces amariceps. Pyrrhulomyces is distinguished from other genera of Strophariaceae by the blackening basidiomata with a bitter taste, smooth basidiospores without a germ pore under light microscopy, presence of pleurochrysocystidia, an ixocutis, rugulose spore ornamentation under scanning electron microscope (SEM), and association with late stages of conifer wood decay. Pholiota subochracea was found to be sister to a clade containing samples of Hypholoma and Bogbodia, but this portion of the Strophariaceae will require further taxon and gene sampling to resolve relationships between these three taxa. Pholiota sensu stricto comprised at least two major groups, but several residual poorly placed lineages were also noted depending on the data set analyzed. New combinations are made in the genera Flammula, Kuehneromyces, and Stropharia for three species of Pholiota-P. abieticola, P. obscura, and P. scabella, respectively, based on molecular annotation of type collections. Overall, 20 new synonymies are proposed, mostly in Pholiota. Illustrations of Pyrrhulomyces are provided along with a key to genera of Strophariaceae and Hymenogastraceae.

Biochemical characterization and inhibition of the alternative oxidase enzyme from the fungal phytopathogen Moniliophthora perniciosa.

Moniliophthora perniciosa is a fungal pathogen and causal agent of the witches' broom disease of cocoa, a threat to the chocolate industry and to the economic and social security in cocoa-planting countries. The membrane-bound enzyme alternative oxidase (MpAOX) is crucial for pathogen survival; however a lack of information on the biochemical properties of MpAOX hinders the development of novel fungicides. In this study, we purified and characterised recombinant MpAOX in dose-response assays with activators and inhibitors, followed by a kinetic characterization both in an aqueous environment and in physiologically-relevant proteoliposomes. We present structure-activity relationships of AOX inhibitors such as colletochlorin B and analogues which, aided by an MpAOX structural model, indicates key residues for protein-inhibitor interaction. We also discuss the importance of the correct hydrophobic environment for MpAOX enzymatic activity. We envisage that such results will guide the future development of AOX-targeting antifungal agents against M. perniciosa, an important outcome for the chocolate industry.

UPLC-QqQ/MS combined with similarity assessment of 17 nucleic acid constituents in 147 edible fungi from Sichuan Basin, China.

Nucleic acid constituents are the main functional ingredients in edible fungi, therefore understanding the nucleic acid constituents of edible fungi often eaten on the table by the Chinese people is of significant value. In this study, Sichuan Provincial Center for Disease Control and Prevention collected 147 samples of edible fungi (including certain species that the Chinese often eat) from different parts of the Sichuan Basin. A new UPLC-QqQ/MS method has been developed to determine the 17 nucleic acid constituents in these 147 samples, including guanosine, adenosine, uridine, cytidine, inosine, thymidine, xanthosine dehydrate, 2'-deoxyguanosine, 2'-deoxyadenosine, 2'-deoxyuridine, 2'-deoxycytidine, 2'-deoxyinosine, guanosine 5'-monophosphate, adenosine 5'-monophosphate, uridine 5'-monophosphate, cytidine 5'-monophosphate, and inosine 5'-monophosphate. Finally, similarity assessment of the main edible fungus was performed using vectorial angle cosine method, and hierarchical cluster analysis was used to classify all the 147 samples. The results showed that some edible fungi have high similarities, especially in Lentinula edodes (Berk.) Pegler, the monogenic nucleotides content of which (55.84 ±â€¯8.4 mg/100 g) is far greater than any other edible fungus, which is directly related to its taste. For quality control, this paper proposed to use the reference values of total nucleic acid compounds in edible fungi computed by percentile threshold method. This is the first time a comprehensive evaluation of nucleic acid constituents of different edible fungi of daily consumption was conducted for a large region, and the results is conducive to the quality evaluation and quality control of edible fungus.

SQUAT: a Sequencing Quality Assessment Tool for data quality assessments of genome assemblies.

BACKGROUND: With the rapid increase in genome sequencing projects for non-model organisms, numerous genome assemblies are currently in progress or available as drafts, but not made available as satisfactory, usable genomes. Data quality assessment of genome assemblies is gaining importance not only for people who perform the assembly/re-assembly processes, but also for those who attempt to use assemblies as maps in downstream analyses. Recent studies of the quality control, quality evaluation/ assessment of genome assemblies have focused on either quality control of reads before assemblies or evaluation of the assemblies with respect to their contiguity and correctness. However, correctness assessment depends on a reference and is not applicable for de novo assembly projects. Hence, development of methods providing both post-assembly and pre-assembly quality assessment reports for examining the quality/correctness of de novo assemblies and the input reads is worth studying. RESULTS: We present SQUAT, an efficient tool for both pre-assembly and post-assembly quality assessment of de novo genome assemblies. The pre-assembly module of SQUAT computes quality statistics of reads and presents the analysis in a well-designed interface to visualize the distribution of high- and poor-quality reads in a portable HTML report. The post-assembly module of SQUAT provides read mapping analytics in an HTML format. We categorized reads into several groups including uniquely mapped reads, multiply mapped, unmapped reads; for uniquely mapped reads, we further categorized them into perfectly matched, with substitutions, containing clips, and the others. We carefully defined the poorly mapped (PM) reads into several groups to prevent the underestimation of unmapped reads; indeed, a high PM% would be a sign of a poor assembly that requires researchers' attention for further examination or improvements before using the assembly. Finally, we evaluate SQUAT with six datasets, including the genome assemblies for eel, worm, mushroom, and three bacteria. The results show that SQUAT reports provide useful information with details for assessing the quality of assemblies and reads. AVAILABILITY: The SQUAT software with links to both its docker image and the on-line manual is freely available at https://github.com/luke831215/SQUAT .

Phytogeographical and sociolinguistical patterns of the diversity, distribution, and uses of wild mushrooms in Côte d'Ivoire, West Africa.

BACKGROUND: Many fungal species in tropical Africa are useful, with high added value, and play essential roles in the structure and dynamic of ecosystems. However, the diversity, distribution, and uses by local populations of these non-timber forest products (NTFPs) and their respective habitats are still very poorly understood in sub-Saharan Africa in general and more specifically in Côte d'Ivoire. This study aims at (i) inventorying the wild useful mushrooms of Côte d'Ivoire within its major protected areas and their respective surrounding sociolinguistical groups, according to climatic and phytogeographical gradients, and (ii) recording ethnomycological knowledge and considerations of these local people. METHODS: Field and ethnomycological surveys were conducted in the main and highest protected areas of Côte d'Ivoire (Comoé, Marahoué, and Taï national parks) and a set of their respective surrounding villages, along climatic and phytogeographical gradients. Standardized methods (permanent plots and opportunistic searches) were used for field surveys. In addition, a total 748 respondents belonging to 13 ethnic groups were interviewed at a rate of 300 interviewees during the preliminary investigations and 448 persons during the proper ethnomycological surveys. RESULTS: Sixty-eight useful wild fungal species, belonging to 17 families and 23 genera, were listed and collected. Four categories of usage were reported by the rural people (food, medicinal, belief and recreational), with a dominance of food and medicinal uses. Fifty-six species were reported to be used as food and 16 species as medicinal fungi. These uses varied not only from one sociolinguistical group to another but also from a visited village to another. The high number (41) of the reported useful species was found in the Sudano-Guinean savanna zone while 28 species were collected in the forest zone and 22 species in the forest-savanna mosaic zone. These mushrooms were either saprotrophic or symbiotic (ectomycorrhizal or termitophilic). Auricularia sp3, Psathyrella tuberculata, and Termitomyces spp. were found as the most commonly used mushrooms. CONCLUSIONS: These national scale field and ethnomycological surveys give one of the more complete but non-exhaustive list of useful mushrooms of Côte d'Ivoire. Mushrooms are relatively well known and used by the Ivorian people within the main phytogeographical zone of the country. These people also have an interest in all the functional groups with an important phytogeographical zone-fungal-specific used species. However, protected areas of the visited zones seem to represent the last sanctuaries of these organisms due to high rate of loss of natural habitats.

Phylogenetic assessment of Chromocyphellaceae (Agaricineae, Basidiomycota) and a new lamellate species of Chromocyphella.

Cyphelloid fungi represent a polyphyletic assemblage of reduced agarics, including the brown-spored family Chromocyphellaceae. In order to elucidate the phylogenetic position of Chromocyphellaceae, newly generated sequences of Chromocyphella were included in a multigene alignment of the Agaricineae and phylogenetically analyzed. The current analyses show that the Chromocyphella muscicola specimen used to phylogenetically place Chromocyphellaceae in its original description was misidentified and that the Chromocyphellaceae nests in the Hymenogastraceae, Chromocyphella being sister to Flammula. Chromocyphella is emended, including now a new species with lamellate and stipitate basidiomata, C. lamellata. The name Cymbella crouanii, type species of Chromocyphella, is lecto- and epitypified. Our analyses support a new origin of cyphelloid fungi. The shift to a cyphelloid basidioma from an agaric ancestor is suggested to have occurred in two evolutionary steps in Chromocyphella, an initial reduction in basidioma size and a subsequent loss of lamellae and stipe.

Safety assessment of mushrooms in dietary supplements by combining analytical data with in silico toxicology evaluation.

Despite growing popularity in dietary supplements, many medicinal mushrooms have not been evaluated for their safe human consumption using modern techniques. The multifaceted approach described here relies on five key principles to evaluate the safety of non-culinary fungi for human use: (1) identification by sequencing the nuclear ribosomal internal transcribed spacer (ITS) region (commonly referred to as ITS barcoding), (2) screening an extract of each fungal raw material against a database of known fungal metabolites, (3) comparison of these extracts to those prepared from grocery store-bought culinary mushrooms using UHPLCPDA-ELS-HRMS, (4) review of the toxicological and chemical literature for each fungus, and (5) evaluation of data establishing presence in-market. This weight-of-evidence approach was used to evaluate seven fungal raw materials and determine safe human use for each. Such an approach may provide an effective alternative to conventional toxicological animal studies (or more efficiently identifies when studies are necessary) for the safety assessment of fungal dietary ingredients.

Induction of Cbl-dependent epidermal growth factor receptor degradation in Ling Zhi-8 suppressed lung cancer.

Activating mutation of epidermal growth factor receptor (EGFR) is correlated with malignant lung tumor. In our study, we demonstrated that recombinant LZ-8 (rLZ-8), a medicinal mushroom Ganoderma lucidum protein, induced cell cycle arrest and apoptosis by downregulating the expression of wild-type and mutated EGFR and inhibiting EGFR downstream effectors, AKT and ERK1/2 in lung cancer cells. We showed that rLZ-8 effectively inhibited lung cancer progression and suppressed EGFR expression of lung tumor lesions in mouse model. Functional studies revealed that rLZ-8 reduced the amount of EGFR in cell membranes by altering EGFR localization to enhance the EGF-induced degradation of EGFR. Mechanistically, we demonstrated that rLZ-8 bound to EGFR to induce EGFR autophosphorylation at tyrosine1045 and trigger ubiquitination by inducing the formation of EGFR/Cbl complexes, resulting in the degradation of EGFR; however, Cbl-shRNA abolished rLZ-8-induced EGFR degradation. We provide the first evidence showing that rLZ-8 inhibits growth and induces apoptosis of lung cancer cells by promoting EGFR degradation. The current findings therefore suggest a novel anti-cancer function of rLZ-8 that targeting EGFR overexpression or mutation as well as EGFR-dependent processes in cancer cells.

The evolution of defense mechanisms correlate with the explosive diversification of autodigesting Coprinellus mushrooms (Agaricales, Fungi).

Bursts of diversification are known to have contributed significantly to the extant morphological and species diversity, but evidence for many of the theoretical predictions about adaptive radiations have remained contentious. Despite their tremendous diversity, patterns of evolutionary diversification and the contribution of explosive episodes in fungi are largely unknown. Here, using the genus Coprinellus (Psathyrellaceae, Agaricales) as a model, we report the first explosive fungal radiation and infer that the onset of the radiation correlates with a change from a multilayered to a much simpler defense structure on the fruiting bodies. We hypothesize that this change constitutes a key innovation, probably relaxing constraints on diversification imposed by nutritional investment into the development of protective tissues of fruiting bodies. Fossil calibration suggests that Coprinellus mushrooms radiated during the Miocene coinciding with global radiation of large grazing mammals following expansion of dry open grasslands. In addition to diversification rate-based methods, we test the hard polytomy hypothesis, by analyzing the resolvability of internal nodes of the backbone of the putative radiation using Reversible-Jump MCMC. We discuss potential applications and pitfalls of this approach as well as how biologically meaningful polytomies can be distinguished from alignment shortcomings. Our data provide insights into the nature of adaptive radiations in general by revealing a deceleration of morphological diversification through time. The dynamics of morphological diversification was approximated by obtaining the temporal distribution of state changes in discrete traits along the trees and comparing it with the tempo of lineage accumulation. We found that the number of state changes correlate with the number of lineages, even in parts of the tree with short internal branches, and peaks around the onset of the explosive radiation followed by a slowdown, most likely because of the decrease in available niches.

The systematic relevance of conidiogenesis modes in the gilled Agaricales.

Dikaryotic and haploid mycelia of more than 150 gilled species of euagarics were studied morphologically and by molecular phylogenetic methods. The morphological investigations revealed anamorphs in more than 90 species that were often specific at the genus or family level. Thallic conidiogenesis dominated and varied from fragmentation of normally branched hyphae to the formation of differentiated sympodially branched conidiophores. Secession modes, coiling of the conidiogenous hyphae or the swelling of the conidia were additional distinguishing features. Phylogenetic analysis of the D1-D3 domains of the nuclear gene for the ribosomal large subunit using a Bayesian Markov chain Monte Carlo approach resulted in several well-supported groups that are consistent with anamorph morphology. These results indicate that the anamorphs provide valuable characters for a natural classification of the Agaricales.

Modern aspects of mushroom culture technology.

The production and culture of new species of mushrooms is increasing. The breeding of new strains has significantly improved, allowing the use of strains with high yield and resistance to diseases, increasing productivity and diminishing the use of chemicals for pest control. The improvement and development of modern technologies, such as computerized control, automated mushroom harvesting, preparation of compost, production of mushrooms in a non-composted substrate, and new methods of substrate sterilization and spawn preparation, will increase the productivity of mushroom culture. All these aspects are crucial for the production of mushrooms with better flavor, appearance, texture, nutritional qualities, and medicinal properties at low cost. Mushroom culture is a biotechnological process that recycles ligninocellulosic wastes, since mushrooms are food for human consumption and the spent substrate can be used in different ways.

Assessment the genetic diversity of Auricularia strains by two PCR-based typing methods.

Two PCR-based methods, enterobacterial repetitive intergenic consensus-PCR (ERIC-PCR) and randomly amplified polymorphic DNA (RAPD), were adopted for differentiating Auricularia strains. Taken the similarity coefficient as 75%, 29 strains of three Auricularia species were grouped into 6 and 9 clusters by RAPD and ERIC, respectively. The dendrogram from ERIC exhibited two distinct parts, one representing A. auricula and the other A. polytricha, but the dendrogram from RAPD failed to clearly distinguish between these two species. However, both methods similarly revealed high homology between A. fuscosuccinea and A. auricula. The homology relationships among the three species obtained from ERIC were validated by Southern hybridization. The analyses showed that RAPD is able to differentiate mainly at the species level, while ERIC is effective at the strain level and therefore more consistent with cultivation characteristics. The results indicate that the method of ERIC-PCR is more rapid and reliable than RAPD, and may substitute for RAPD in research related to the genetic identification and genetic diversity in Auricularia.