Unroughing the cat's tongue mushrooms: Four new species of Pseudohydnum from Brazil based on morphological and molecular phylogenetic evidence.

Pseudohydnum, commonly known as cat's tongue mushrooms, is a monophyletic assemblage within Auriculariales, which encompasses species with gelatinous basidiomata, spathulate, flabellate, or shell-shaped pileus, hydnoid hymenophore, globose to ellipsoidal basidiospores, and longitudinally cruciate-septate basidia. According to the available literature, 16 species have been described in Pseudohydnum, mostly represented in temperate-boreal forests of the Northern Hemisphere. However, the limited morphological, molecular, and ecological information, especially from the Southern Hemisphere ecosystems, does not presently allow a reliable assessment of its taxonomic boundaries nor provide a complete picture of the species diversity in the genus. In an ongoing effort to examine specimens collected in dense and mixed ombrophilous forest fragments (Atlantic Rainforest domain) from Southeastern and Southern Brazil, additional taxa assigned to Pseudohydnum were identified. Four new species are recognized based mostly on characters of the pileus surface, stipe, hymenium, and basidiospores. Molecular phylogenetic analyses based on nuc rDNA internal transcribed spacer region ITS1-5.8S-ITS2 (ITS barcode), partial nuc rDNA 28S, and partial RNA polymerase II largest subunit (RPB1) sequences supported the description of these new taxa. Here, we propose Pseudohydnum brasiliense, P. brunneovelutinum, P. cupulisnymphae, and P. viridimontanum as new species. Morphological descriptions, line drawings, habitat photos, and comparisons with closely related taxa are provided. A dichotomous key for identification of currently known Southern Hemisphere Pseudohydnum species is presented.

New Cortinariaceae species associated with Dicymbe, Aldina, and Pakaraimaea in Guyana.

Species of the ectomycorrhizal (ECM) family Cortinariaceae (Agaricales, Agaricomycetes, Basidiomycota) have long been considered impoverished or absent from lowland tropical rainforests. Several decades of collecting in forests dominated by ECM trees in South America's Guiana Shield is countering this view, with discovery of numerous Cortinariaceae species. To date, ~12 morphospecies of this family have been found in the central Pakaraima Mountains of Guyana. Here, we describe three of these as new species of Cortinarius and two as new species of Phlegmacium from forests dominated by the ECM tree genera Dicymbe (Fabaceae subfam. Detarioideae), Aldina (Fabaceae subfam. Papilionoideae), and Pakaraimaea (Cistaceae). Macromorphological, micromorphological, habitat, and DNA sequence data are provided for each new species.

Optimized methodology for obtention of high-yield and -quality RNA from the mycelium of the bioluminescent fungus Neonothopanus gardneri.

Neonothopanus gardneri, also known as coconut flower mushroom (flor-de-coco), is a Brazilian bioluminescent basidiomycete found in Palm Forest, a transitional biome between the Amazonian Forest and Caatinga (Savanna-like vegetation) in Northeast Brazil, especially in Piauí State. Recent advances toward the elucidation of fungal bioluminescence have contributed to the discovery of four genes (hisps, h3h, luz and cph) involved with the bioluminescence process, the so-called Caffeic Acid Cycle (CAC) and to develop biotechnological applications such autoluminescent tobacco plants and luciferase-based reporter genes. High-yield and -quality RNA-extraction methods are required for most of these purposes. Herein, four methods for RNA isolation from the mycelium of N. gardneri were evaluated: RNeasy® kit (QIAGEN), TRI+, TRI18G+, and TRI26G+. Highest RNA yield was observed for TRI18G+ and TRI26G+ methods, an increase of ~130% in comparison to the RNeasy® method and of ~40% to the TRI+ protocol. All the RNA samples showed good purity and integrity, except by gDNA contamination in RNA samples produced with the RNeasy® method. High quality of RNA samples was confirmed by successful cDNA synthesis and PCR amplification of the coding sequence of h3h gene, responsible for the hydroxylation of the precursor of fungal luciferin (3-hydroxyhispidin). Similarly, RT-qPCR amplification of ef-tu gene, related to the protein biosynthesis in the cell, was demonstrated from RNA samples. This is the first report of a reproducible, time-saving and low-cost optimized method for isolation of high-quality and -yield, DNA-free RNA from a bioluminescent fungus, but that can also be useful for other basidiomycetes.

Growth of Leucoagaricus gongylophorus Möller (Singer) and production of key enzymes in submerged and solid-state cultures with lignocellulosic substrates.

The aim of this study was to characterize the growth of the fungus Leucoagaricus gongylophorus LEU18496, isolated from the fungus garden of the nest of leaf cutter ants Atta mexicana. The fungus garden was cultivated in an artificial laboratory nest and the fungus further grown in submerged (SmC) and solid state (SSC) cultures with sugarcane bagasse, grass or model substrates containing CM-cellulose, xylan or lignin. The CO2 production rate with grass in SmC (Vmax 34.76 mg CO2 Lgas-1 day- 1) was almost four times than SSC (Vmax 9.49 mg CO2 Lgas-1 day- 1), while the production rate obtained in sugarcane bagasse in SmC (Vmax 16.02 mg CO2 Lgas-1 day- 1) was almost three times than that for SSC (Vmax 5.42 mg CO2 Lgas-1 day- 1). In addition, the fungus grew with defined carbon substrates mixtures in SmC, but at different rates, first xylan, followed by CM-cellulose and lignin. Endoglucanase and xylanase activities (U mgprotein-1) were detected in all cultures, the specific activity was higher in the fungus-garden, 5.2 and 1.8; followed by SSC-grass, 1.5 and 0.8, and SSC-bagasse, 0.9 and 0.8, respectively. Laccase activity in the fungus-garden was 44.8 U L- 1 and 10.9 U L- 1 in the SSC-grass. The gongylidia structures observed by environmental scanning electron microscopy were ca. 40 µm and the hyphae width ca. 5 µm. The results show that L. gongylophorus from A. mexicana have promising applications for the treatment of plant residues to release fermentable sugars and the production of high value lignocellulolytic enzymes such as endoglucanase, xylanase or laccases.

Biodegradation of atrazine and ligninolytic enzyme production by basidiomycete strains.

BACKGROUND: Atrazine is one of the most widespread chlorinated herbicides, leaving large bulks in soils and groundwater. The biodegradation of atrazine by bacteria is well described, but many aspects of the fungal metabolism of this compound remain unclear. Thus, we investigated the toxicity and degradation of atrazine by 13 rainforest basidiomycete strains. RESULTS: In liquid medium, Pluteus cubensis SXS320, Gloelophyllum striatum MCA7, and Agaricales MCA17 removed 30, 37, and 38%, respectively, of initial 25 mg L- 1 of the herbicide within 20 days. Deficiency of nitrogen drove atrazine degradation by Pluteus cubensis SXS320; this strain removed 30% of atrazine within 20 days in a culture medium with 2.5 mM of N, raising three metabolites; in a medium with 25 mM of N, only 21% of initial atrazine were removed after 40 days, and two metabolites appeared in culture extracts. This is the first report of such different outcomes linked to nitrogen availability during the biodegradation of atrazine by basidiomycetes. The herbicide also induced synthesis and secretion of extracellular laccases by Datronia caperata MCA5, Pycnoporus sanguineus MCA16, and Polyporus tenuiculus MCA11. Laccase levels produced by of P. tenuiculus MCA11 were 13.3-fold superior in the contaminated medium than in control; the possible role of this enzyme on atrazine biodegradation was evaluated, considering the strong induction and the removal of 13.9% of the herbicide in vivo. Although 88% of initial laccase activity remained after 6 h, no evidence of in vitro degradation was observed, even though ABTS was present as mediator. CONCLUSIONS: This study revealed a high potential for atrazine biodegradation among tropical basidiomycete strains. Further investigations, focusing on less explored ligninolytic enzymes and cell-bound mechanisms, could enlighten key aspects of the atrazine fungal metabolism and the role of the nitrogen in the process.

Cortinarius section Thaumasti in South American Nothofagaceae forests.

We studied the South American species of Cortinarius section Thaumasti based on morphological and molecular data. Members of this group can easily be identified in the field because the basidiomata are small and Phlegmacium-like with a bulbous stipe and the universal veil in most species forms a distinct volva at the base of the stipe. The phylogenetic delimitation of the clade was mostly in concordance with the earlier, morphology-based grouping of the South American taxa except that C. chrysophaeus was resolved outside of the clade. Altogether nine species were recognized in the section. Four species, C. chlorophanus, C. coleopus, C. cosmoxanthus, and C. vaginatus, were previously described by other authors, whereas three species, C. chlorosplendidus, C. olivaceovaginatus, and C. subcosmoxanthus, are described here as new. We were able to identify two remaining taxa, but we do not have sufficient morphological data to allow for a formal description. All of the species in C. section Thaumasti form ectomycorrhizal associations with Nothofagaceae. They have been documented from South America and New Zealand. The Patagonian species are considered endemic to the region. A key to the described species is provided.

New species and records of bioluminescent Mycena from Mexico.

Seven species of bioluminescent fungi are recorded from the cloud forests in Mexico. Six represent new species belonging to the genus Mycena, whereas Mycena globulispora is a new distribution record for the country. Descriptions, illustrations, photographs, and an identification key to bioluminescent fungi species from Mexico are provided. Sequences of nuc rDNA internal transcribed spacer (ITS1-5.8S-ITS2 = ITS) region were generated for barcoding purposes and comparisons with similar species.

Taxonomy and phylogeny of Macrolepiota: two new species from Brazil.

Macrolepiota is a poorly known genus in the Neotropics. In order to increase knowledge about this group, we collected specimens from the Atlantic Forest in southern and northeastern Brazil. Macrolepiota cyanolamellata and M. sabulosa from subtropical and tropical regions, respectively, are proposed as new species. We performed molecular phylogenetic analyses of the nuc rDNA internal transcribed spacer region ITS1-5.8S-ITS2 (ITS) and the combined data set ITS + nuclear large subunit rDNA (28S) + RNA polymerase II second largest (RPB2), as well as morphological analyses. Two lineages with unique morphotypes were found. The species proposed were strongly supported as the sister lineage closely related to M. clelandii and M. subcitrophylla. Detailed descriptions and illustrations of their macro- and microscopic characters are provided.

Genetic characterization, evaluation of growth and production of biomass of strains from wild edible mushrooms of Lyophyllum of Central Mexico

The present study conducted a genetic characterization and determined growth rate and biomass production in solid and liquid media, using strains obtained from wild edible sporomes of Lyophyllum that grow in high mountains. Vegetative isolation was used to obtain a total of four strains, which were divided into two clades within the section Difformia: Lyophyllum sp. and Lyophyllum aff. shimeji. Growth rate and biomass production were influenced by both the culture media and the strains. In a potato dextrose agar medium, the strains presented a higher growth rate, while in a malt extract-peptone and yeast agar medium, the growth rate was lower, but with a higher biomass production that was equal to that in the malt extract-peptone and yeast liquid medium.(AU)

Genome sequence and effectorome of Moniliophthora perniciosa and Moniliophthora roreri subpopulations.

BACKGROUND: The hemibiotrophic pathogens Moniliophthora perniciosa (witches' broom disease) and Moniliophthora roreri (frosty pod rot disease) are among the most important pathogens of cacao. Moniliophthora perniciosa has a broad host range and infects a variety of meristematic tissues in cacao plants, whereas M. roreri infects only pods of Theobroma and Herrania genera. Comparative pathogenomics of these fungi is essential to understand Moniliophthora infection strategies, therefore the detection and in silico functional characterization of effector candidates are important steps to gain insight on their pathogenicity. RESULTS: Candidate secreted effector proteins repertoire were predicted using the genomes of five representative isolates of M. perniciosa subpopulations (three from cacao and two from solanaceous hosts), and one representative isolate of M. roreri from Peru. Many putative effectors candidates were identified in M. perniciosa: 157 and 134 in cacao isolates from Bahia, Brazil; 109 in cacao isolate from Ecuador, 92 and 80 in wild solanaceous isolates from Minas Gerais (Lobeira) and Bahia (Caiçara), Brazil; respectively. Moniliophthora roreri showed the highest number of effector candidates, a total of 243. A set of eight core effectors were shared among all Moniliophthora isolates, while others were shared either between the wild solanaceous isolates or among cacao isolates. Mostly, candidate effectors of M. perniciosa were shared among the isolates, whereas in M. roreri nearly 50% were exclusive to the specie. In addition, a large number of cell wall-degrading enzymes characteristic of hemibiotrophic fungi were found. From these, we highlighted the proteins involved in cell wall modification, an enzymatic arsenal that allows the plant pathogens to inhabit environments with oxidative stress, which promotes degradation of plant compounds and facilitates infection. CONCLUSIONS: The present work reports six genomes and provides a database of the putative effectorome of Moniliophthora, a first step towards the understanding of the functional basis of fungal pathogenicity.

Analysis of the cocobiota and metabolites of Moniliophthora perniciosa-resistant Theobroma cacao beans during spontaneous fermentation in southern Brazil.

BACKGROUND: Cocoa bean fermentation is a spontaneous process involving a succession of microbial activities, yeasts, lactic acid, and acetic acid bacteria. The spontaneous fermentation of cocoa beans by Theobroma cacao TSH565 clonal variety, a highly productive hybrid resistant to Moniliophthora perniciosa and Phytophthora spp., was investigated. The natural cocobiota involved in the spontaneous fermentation of this hybrid in southern Brazil, was investigated by using both a culture-dependent microbiological analysis and a molecular analysis. The changes in the physicochemical characteristics and the kinetics of substrate utilization and metabolite production during fermentation were also evaluated. RESULTS: Yeasts (178) and bacteria (244) isolated during fermentation were identified by partial sequencing of the ITS and 16S rDNAs, respectively. After 144 h of fermentation, the indigenous yeast community was composed of Hanseniaspora spp., Saccharomyces spp., and Pichia spp. The bacterial population comprised Lactococcus spp., Staphylococcus spp., Acetobacter spp. and Lactobacilli strains. The kinetics of substrate transformation reflected the dynamic composition of the cocobiota. Substrates such as glucose, fructose, sucrose, and citric acid, present at the beginning of fermentation, were metabolized to produce ethanol, acetic acid, and lactic acid. CONCLUSION: The results described here provide new insights into microbial diversity in cocoa bean-pulp mass fermentation and the kinetics of metabolites synthesis, and pave the way for the selection of starter cultures to increase efficiency and consistency to obtain homogeneous and best quality cocoa products. © 2018 Society of Chemical Industry.

Growth and enzymatic activity of Leucoagaricus gongylophorus, a mutualistic fungus isolated from the leaf-cutting ant Atta mexicana, on cellulose and lignocellulosic biomass.

A mutualistic fungus of the leaf-cutting ant Atta mexicana was isolated and identified as Leucoagaricus gongylophorus. This isolate had a close phylogenetic relationship with L. gongylophorus fungi cultivated by other leaf-cutting ants as determined by ITS sequencing. A subcolony started with ~500 A. mexicana workers could process 2 g day-1 of plant material and generate a 135 cm3 fungus garden in 160 days. The presence of gongylidia structures of ~35 µm was observed on the tip of the hyphae. The fungus could grow without ants on semi-solid cultures with α-cellulose and microcrystalline cellulose and in solid-state cultures with grass and sugarcane bagasse, as sole sources of carbon. The maximum CO2 production rate on grass (Vmax  = 17·5 mg CO2  Lg-1  day-1 ) was three times higher than on sugarcane bagasse (Vmax  = 6·6 mg CO2  Lg-1 day-1 ). Recoveries of 32·9 mgglucose  gbiomass-1 and 12·3 mgglucose  gbiomass-1 were obtained from the fungal biomass and the fungus garden, respectively. Endoglucanase activity was detected on carboxymethylcellulose agar plates. This is the first study reporting the growth of L. gongylophorus from A. mexicana on cellulose and plant material. SIGNIFICANCE AND IMPACT OF THE STUDY: According to the best of our knowledge, this is the first report about the growth of Leucoagaricus gongylophorus, isolated from the colony of the ant Atta mexicana, on semisolid medium with cellulose and solid-state cultures with lignocellulosic materials. The maximum CO2 production rate on grass was three times higher than on sugarcane bagasse. Endoglucanase activity was detected and it was possible to recover glucose from the fungal gongylidia. The cellulolytic activity could be used to process lignocellulosic residues and obtain sugar or valuable products, but more work is needed in this direction.

Lichenomphalia altoandina, a new species of Hygrophoraceae from the Chilean Altiplano.

Lichenomphalia is a lichenized agaric genus characterized by its omphalinoid basidiomes. Lichenomphalia species are associated with unicellular green algae in the genus Coccomyxa and are mainly distributed in polar and alpine habitats. The aim of this work is to describe L. altoandina, a new species from northern Chile that grows among cushion plants over 3000 m above sea level in the Andes Mountains. The species is remarkable for living in highly saline environments, in some cases virtually on salt crusts. Lichenomphalia altoandina differs from other known species and particularly from L. aurantiaca, the most morphologically similar species, in its smooth and broader stipe and its slightly larger spores. Lichenomphalia altoandina is also morphologicaly and ecologically more similar to the core Lichenomphalia clade. Our phylogenetic study based on nuclear rDNA ITS and partial 28S sequences shows that L. altoandina belongs to the Protolichenomphalia clade and is sister to an unknown lineage, L. aff. umbellifera, from New Zealand.

A new stipitate species of Crepidotus from India and Thailand, with notes on other tropical species.

A new Asian species of Crepidotus (Basidiomycota, Agaricales), C. asiaticus, is presented based on morphological and nuc rDNA internal transcribed spacer (ITS1-5.8S-ITS2 = ITS) and large subunit (28S) sequence data. This new species, found in India and Thailand, is characterized by the centrally stipitate medium-sized basidiomata, orange to reddish brown pileus, white to brownish orange lamellae, and white stipe. Based on morphology, C. asiaticus is similar to the neotropical C. thermophilus. However, the microscopic characters, especially the size and shape of the basidiospores, can be used to distinguish these two taxa, as well as their geographic distributions. Further, the phylogenetic position of C. asiaticus is unique based on ITS and 28S nuc rDNA sequences. Melanomphalia argipoda, described by Singer from Ecuador, is also a stipitate Crepidotus based on an ITS sequence of the type specimen, so the new combination is proposed here. Phylogenetically, the three species form a monophyletic group with the Asiatic C. asiaticus forming the sister lineage to the neotropical C. argipodus and C. thermophilus.

Age-dependent mycorrhizal specificity in an invasive orchid, Oeceoclades maculata.

PREMISE OF THE STUDY: Oeceoclades maculata is a naturalized, invasive, terrestrial orchid in Puerto Rico and elsewhere in the neotropics. We asked whether its success might be partly explained by its mycorrhizal associations, hypothesizing a relationship with many fungal partners or with one widely distributed partner. METHODS: Oeceoclades maculata roots were collected throughout Puerto Rico, and the degree of mycorrhizal colonization was measured. For identification of fungi, the ITS region was sequenced from pure cultures and directly from roots. Representative fungi were used for symbiotic seed germination experiments. KEY RESULTS: Colonization of O. maculata roots was very variable. The most common fungus identified by BLAST searches was Psathyrella cf. candolleana, but typical orchid mycorrhizal fungi (Ceratobasidium and Tulasnella) were also found, as were a range of saprotrophs. Seeds germinated in vitro only in the presence of Psathyrella. CONCLUSIONS: These results are surprising in two respects. First, O. maculata appears to be highly specific for fungi during seed germination, but unusually promiscuous as adult plants. Second, mycorrhizal associations with Psathyrella and with other saprotrophic fungi have been previously reported, but only from mycoheterotrophic (i.e., nonphotosynthetic) orchids, not from green orchids like Oeceoclades. This combination may partly explain the success of Oeceoclades.

Guyanagarika, a new ectomycorrhizal genus of Agaricales from the Neotropics.

A new genus and three new species of Agaricales are described from the Pakaraima Mountains of Guyana in the central Guiana Shield. All three of these new species fruit on the ground in association with species of the ectomycorrhizal (ECM) tree genus Dicymbe (Fabaceae subfam. Caesalpinioideae) and one species has been shown to form ectomycorrhizas. Multi-locus molecular phylogenetic analyses place Guyanagarika gen. nov. within the Catathelasma clade, a lineage in the suborder Tricholomatineae of the Agaricales. We formally recognize this 'Catathelasma clade' as an expanded family Catathelasmataceae that includes the genera Callistosporium, Catathelasma, Guyanagarika, Macrocybe, Pleurocollybia, and Pseudolaccaria. Within the Catathelasmataceae, Catathelasma and Guyanagarika represent independent origins of the ectomycorrhizal habit. Guyanagarika is the first documented case of an ECM Agaricales genus known only from the Neotropics.

New luminescent mycenoid fungi (Basidiomycota, Agaricales) from São Paulo State, Brazil.

Four species of mycenoid fungi are reported as luminescent (or putatively luminescent) on the basis of specimens collected from São Paulo State, Brazil. Two of them represent new species (Mycena oculisnymphae, Resinomycena petarensis), and two represent new reports of luminescence in previously described species (M. deformis, M. globulispora). Comprehensive descriptions, illustrations, photographs, and comparisons with phenetically similar species are provided. Sequences of nuc rDNA internal transcribed spacer regions were generated for barcoding purposes and for comparisons with similar species.

Medicinal Mushrooms in Guatemala.

Guatemala, located in Central America, has a long and rich history in the traditional use of edible, medicinal, and hallucinogenic mushrooms. This article describes the use of these mushrooms and presents studies on the scientific validation of native and foreign species.

Fungal communities in pressmud composting harbour beneficial and detrimental fungi for human welfare.

Pressmud is a substrate derived from sugarcane juice filtrate, and around 26-40 kg of this residue are produced per ton of sugarcane. It is mainly used as fertilizer in crops, and its application in the field is often made without any prior treatment, but, in this research, it was studied for the risk this practice poses for human health. This research was stimulated by previous results indicating the presence of opportunistic pathogens in residues used in various composting systems and the extensive use of fresh pressmud in agriculture. Here, It was assessed the fungal diversity present in both fresh and composting pressmud using 454 pyrosequencing. In addition, heat-tolerant fungi were isolated and surveyed for their enzymatic repertoire of biomass-degrading enzymes (cellulase, xylanase, laccase and polygalacturonase). A wide range of opportunistic pathogens was found among the most abundant taxa in the fresh pressmud, such as Lomentospora prolificans (43.13 %), Trichosporon sp. (10.07 %), Candida tropicalis (7.91 %), and Hormographiella aspergillata (8.19 %). This indicates that fresh pressmud might be a putative source of human pathogenic fungi, presenting a potential threat to human health if applied as fertilizer without any treatment. With regard to the heat-tolerant fungi found in this substrate, all the 110 isolates screened were able to produce at least one of the tested enzymes. The pressmud composting process not only effectively reduces the load of pathogenic fungi, but also creates an interesting environment for fungi able to produce thermostable hydrolytic and oxidative enzymes with biotechnological applications.

Gymnopanella nothofagi, a new genus and species of gymnopoid fungi (Omphalotaceae) from Chilean Nothofagus forest.

A novel, lignicolous agaric from Nothofagus forests of southern Chile is described as a new genus and species, Gymnopanella nothofagi This taxon falls within the family Omphalotaceae as a sister group to Gymnopus in phylogenetic analyses based on sequences spanning the internal transcribed spacer region and D1/D2 region of nuclear 28S rDNA. Morphologically it is characterized by convex to flabellate basidiomata with distinctly gelatinized trama, pileipellis in the form of a cutis with erect fascicles of cylindrical, spirally incrusted hyphae and nonamyloid, broadly ellipsoid basidiospores. This combination of features, in particular the lack of a rameales structure, serve to distinguish Gymnopanella from Gymnopus, Marasmiellus and other similar genera of the Omphalotaceae or Marasmiaceae. The new taxon is known only from Chilean Nothofagus forests at approximately 45-46° south latitude, but concerted searching in similar habitats in surrounding areas or in New Zealand may extend the known range considerably.