Population structure of Guyanagaster necrorhizus supports termite dispersal for this enigmatic fungus.

Understanding how species accomplish dispersal of their propagules can shed light on how they are adapted for their ecosystem. Guyanagaster necrorhizus is a sequestrate fungus, meaning its dispersal propagules, or spores, are entirely enclosed within a fruiting body, termed a sporocarp. This fungus is most closely related to Armillaria and its allies. While Armillaria species form mushrooms and have forcibly discharged spores, G. necrorhizus spores have lost this ability, and by necessity, must be passively dispersed. However, G. necrorhizus does not possess characteristics of other sequestrate fungi with known dispersal mechanisms. Repeated observations of termites feeding on G. necrorhizus sporocarps, and spores subsequently adhering to their exoskeletons, led to the hypothesis that termites disperse G. necrorhizus spores. To test this hypothesis, we used microsatellite markers and population genetics analyses to understand patterns of clonality and population structure of G. necrorhizus. While Armillaria individuals can spread vegetatively over large areas, high genotypic diversity in G. necrorhizus populations suggests spores are the primary mode of dispersal. Spatial genetic structure analyses show that G. necrorhizus sporocarps within 238 m of each other are more closely related than would be expected by chance and conservative estimates from population assignment tests suggest gene flow no longer occurs between sporocarps separated by 2 km. These distances are consistent with previous studies analysing foraging distances of the termites found associated with G. necrorhizus sporocarps. Termites have rarely been recorded to specifically target fungal sporocarps, making this a potentially novel fungal-insect interaction.

Resolved phylogeny and biogeography of the root pathogen Armillaria and its gasteroid relative, Guyanagaster.

BACKGROUND: Armillaria is a globally distributed mushroom-forming genus composed primarily of plant pathogens. Species in this genus are prolific producers of rhizomorphs, or vegetative structures, which, when found, are often associated with infection. Because of their importance as plant pathogens, understanding the evolutionary origins of this genus and how it gained a worldwide distribution is of interest. The first gasteroid fungus with close affinities to Armillaria-Guyanagaster necrorhizus-was described from the Neotropical rainforests of Guyana. In this study, we conducted phylogenetic analyses to fully resolve the relationship of G. necrorhizus with Armillaria. Data sets containing Guyanagaster from two collecting localities, along with a global sampling of 21 Armillaria species-including newly collected specimens from Guyana and Africa-at six loci (28S, EF1α, RPB2, TUB, actin-1 and gpd) were used. Three loci-28S, EF1α and RPB2-were analyzed in a partitioned nucleotide data set to infer divergence dates and ancestral range estimations for well-supported, monophyletic lineages. RESULTS: The six-locus phylogenetic analysis resolves Guyanagaster as the earliest diverging lineage in the armillarioid clade. The next lineage to diverge is that composed of species in Armillaria subgenus Desarmillaria. This subgenus is elevated to genus level to accommodate the exannulate mushroom-forming armillarioid species. The final lineage to diverge is that composed of annulate mushroom-forming armillarioid species, in what is now Armillaria sensu stricto. The molecular clock analysis and ancestral range estimation suggest the most recent common ancestor to the armillarioid lineage arose 51 million years ago in Eurasia. A new species, Guyanagaster lucianii sp. nov. from Guyana, is described. CONCLUSIONS: The armillarioid lineage evolved in Eurasia during the height of tropical rainforest expansion about 51 million years ago, a time marked by a warm and wet global climate. Species of Guyanagaster and Desarmillaria represent extant taxa of these early diverging lineages. Desarmillaria represents an armillarioid lineage that was likely much more widespread in the past. Guyanagaster likely evolved from a gilled mushroom ancestor and could represent a highly specialized endemic in the Guiana Shield. Armillaria species represent those that evolved after the shift in climate from warm and tropical to cool and arid during the late Eocene. No species in either Desarmillaria or Guyanagaster are known to produce melanized rhizomorphs in nature, whereas almost all Armillaria species are known to produce them. The production of rhizomorphs is an adaptation to harsh environments, and could be a driver of diversification in Armillaria by conferring a competitive advantage to the species that produce them.

Three new species of Physalacria from China, with a key to the Asian taxa.

Three new and one previously described species of Physalacria (Physalacriaceae, Agaricales) are reported from China. Specimens of two additional species described from Malaysia and North America were studied for comparison. Placements of these species were corroborated based on morphological observations and molecular evidence from partial sequences of the nuc rDNA internal transcribed spacer regions (ITS) and the 28S D1-D3 region, and genes for translation elongation factor 1-α (tef1α) and the second largest subunit of RNA polymerase II (rpb2). These new species of Physalacria distributed in subtropical China were found on rotten wood of broadleaf trees or bamboo and possess stipitate-capitate basidiomata with four-spored basidia, clamp connections and smooth, inamyloid basidiospores. To facilitate studies of the genus in Asia, a key is provided for all Physalacria species reported from this region.

A new species and new records of Hymenopellis and Xerula (Agaricales, Physalacriaceae) from China.

Hymenopellis is the genus that exhibits the highest number of species within the Xerula/Oudemansiella complex. Numerous species of Hymenopellis demonstrate edibility, and some of these species have been domesticated and cultivated. During an extensive survey carried out in Henan and Jilin Provinces, China, a substantial quantity of Hymenopellis specimens was gathered as a component of the macrofungal resource inventory. Based on the findings of morphological and molecular phylogenetic studies, a new species, Hymenopellis biyangensis, has been identified. A new record species, Hymenopellis altissima, has been discovered in China. Additionally, two new record species, Hymenopellis raphanipes and Xerula strigosa, have been found in Henan Province. Internal transcribed spacer (ITS) and large subunit ribosomal (nrLSU) were used to establish a phylogeny for species identification. Detailed descriptions, field habitat maps and line drawings of these species are presented. The discussion focuses on the relationships between newly discovered species and other related taxa. Additionally, this study provides and a key to the documented species of Hymenopellis and Xerula found in China.

Desarmillaria caespitosa, a North American vicariant of D. tabescens.

Desarmillaria caespitosa, a North American vicariant species of European D. tabescens, is redescribed in detail based on recent collections from the USA and Mexico. This species is characterized by morphological features and multilocus phylogenetic analyses using portions of nuc rDNA 28S (28S), translation elongation factor 1-alpha (tef1), the second largest subunit of RNA polymerase II (rpb2), actin (act), and glyceraldehyde-3-phosphate dehydrogenase (gpd). A neotype of D. caespitosa is designated here. Morphological and genetic differences between D. caespitosa and D. tabescens were identified. Morphologically, D. caespitosa differs from D. tabescens by having wider basidiospores, narrower cheilocystidia, which are often irregular or mixed (regular, irregular, or coralloid), and narrower caulocystidia. Phylogenetic analyses of five independent gene regions show that D. caespitosa and D. tabescens are separated by nodes with strong support. The new combination, D. caespitosa, is proposed.

Symbiotic nitrogen fixation in the reproductive structures of a basidiomycete fungus.

Nitrogen (N) fixation is a driving force for the formation of symbiotic associations between N2-fixing bacteria and eukaryotes.1 Limited examples of these associations are known in fungi, and none with sexual structures of non-lichenized species.2-6 The basidiomycete Guyanagaster necrorhizus is a sequestrate fungus endemic to the Guiana Shield.7 Like the root rot-causing species in its sister genera Armillaria and Desarmillaria, G. necrorhizus sporocarps fruit from roots of decaying trees (Figures 1A-1C),8 and genome sequencing is consistent with observations that G. necrorhizus is a white-rotting decomposer. This species also represents the first documentation of an arthropod-dispersed sequestrate fungus. Numerous species of distantly related wood-feeding termites, which scavenge for N-rich food, feed on the mature spore-bearing tissue, or gleba, of G. necrorhizus. During feeding, mature spores adhere to termites for subsequent dispersal.9 Using chemical assays, isotope analysis, and high-throughput sequencing, we show that the sporocarps harbor actively N2-fixing Enterobacteriaceae species and that the N content within fungal tissue increases with maturation. Untargeted proteomic profiling suggests that ATP generation in the gleba is accomplished via fermentation. The use of fermentation-an anaerobic process-indicates that the sporocarp environment is anoxic, likely an adaptation to protect the oxygen-sensitive nitrogenase enzyme. Sporocarps also have a thick outer covering, possibly to limit oxygen diffusion. The enriched N content within mature sporocarps may offer a dietary inducement for termites in exchange for spore dispersal. These results show that the flexible metabolic capacity of fungi may facilitate N2-fixing associations, as well as higher-level organismal associations.

Ergostane-Type Steroids from Korean Wild Mushroom Xerula furfuracea that Control Adipocyte and Osteoblast Differentiation.

As part of our current work to discover structurally and/or biologically novel compounds from Korean wild mushrooms, we isolated five ergostane-type steroids (1-5) from the fruiting bodies of Xerula furfuracea via repeated column chromatographic separations and HPLC purification. The chemical structures of the isolated steroids were shown to be (22E,24R)-24-methylcholesta-4,22- diene-3,6-dione (1), ergosta-7,22-diene-3ß,5α,6ß-triol (2), ergosta-7,22-diene-3ß,5α,6ß,9α-tetraol (3), (22E,24R)-5α,8α-epidioxyergosta-6,22-diene-3ß-ol-3-O-ß-D-glucopyranoside (4), and (22E,24R)-5α,8α-epidioxyergosta-6,9,22-triene-3ß-ol-3-O-ß-D-glucopyranoside (5) based on comparison of the data regarding their spectroscopic and physical properties with those of previous studies. Notably, this is the first report on the presence of the identified steroids (1-5) in this mushroom. We tested compounds 1-5 to determine their effects on adipogenesis and osteogenesis in the mouse mesenchymal stem cell line C3H10T1/2 and found that compounds 4 and 5 suppressed the differentiation of stem cells into adipocytes. Notably, in addition to its suppressive effect on adipogenesis, compound 5 was also shown to promote the osteogenic differentiation of stem cells. These findings demonstrate that the bioactive compounds isolated might be effective for the treatment of menopause-associated syndromes, such as osteoporosis and obesity, as the isolated compounds were shown to suppress adipogenesis and/or promote osteogenesis of stem cells.

Species diversity, distribution patterns, and substrate specificity of Strobilurus.

The fungal genus Strobilurus belongs to Physalacriaceae and contains approximately 11 species worldwide. Species of this genus grow and reproduce on cones of various conifers, seed pods or fruits of Magnolia and Liquidambar, and branches and wood of conifers. Previous studies focused mainly on samples from Europe and North America. And no genus-specific phylogenetic analysis has been carried out to date. The monophyly, degree of species diversity and substrate specificity, and overall distribution patterns are addressed here using morphological and molecular evidence. The authors collected samples of Strobilurus from much of its known distribution ranges and carried out morphological observations and multilocus phylogenetic analyses using five molecular markers. The results show that Strobilurus is a monophyletic group but may exclude one species, S. ohshimae. A total of 13 species was identified, with two, S. orientalis and S. pachycystidiatus, described as new from China. Several species were shown to be specific to certain substrates, whereas a few less so. Biogeographic analyses indicated that historical exchanges of species between East Asia, Europe, and North America, later vicariance events, and substrate specificity have contributed jointly to diversification of Strobilurus.

Three New Recorded Species of the Physalacriaceae on Ulleung Island, Korea.

Most known species in the Physalacriaceae are saprotrophs that grow on decaying leaves and wood, and approximately 21 genera in the Physalacriaceae have been reported worldwide. During an ongoing survey of indigenous fungi in Korea, four specimens belonging to the Physalacriaceae were collected on Ulleung Island. These specimens were identified as three species based on morphological characteristics and molecular analysis of rDNA-internal transcribed spacer sequences. Three species in three genera were shown to be new records in Korea: Hymenopellis orientalis, Paraxerula hongoi, and Ponticulomyces orientalis. The latter two are the first records of these genera in Korea. In this study, we provide detailed morphological descriptions of these species and describe their phylogenetic position within the Physalacriaceae.

Cytotoxic and antifungal activities of melleolide antibiotics follow dissimilar structure-activity relationships.

The fungal genus Armillaria is unique in that it is the only natural source of melleolide antibiotics, i.e., protoilludene alcohols esterified with orsellinic acid or its derivatives. This class of natural products is known to exert antimicrobial and cytotoxic effects. Here, we present a refined relationship between the structure and the antimicrobial activity of the melleolides. Using both agar diffusion and broth dilution assays, we identified the Δ(2,4)-double bond of the protoilludene moiety as a key structural feature for antifungal activity against Aspergillus nidulans, Aspergillus flavus, and Penicillium notatum. These findings contrast former reports on cytotoxic activities and may indicate a different mode of action towards susceptible fungi. We also report the isolation and structure elucidation of five melleolides (6'-dechloroarnamial, 6'-chloromelleolide F, 10-hydroxy-5'-methoxy-6'-chloroarmillane, and 13-deoxyarmellides A and B), along with the finding that treatment with an antifungal melleolide impacts transcription of A. nidulans natural product genes.

Three New Recorded Species of the Physalacriaceae on Ulleung Island, Korea

Most known species in the Physalacriaceae are saprotrophs that grow on decaying leaves and wood, and approximately 21 genera in the Physalacriaceae have been reported worldwide. During an ongoing survey of indigenous fungi in Korea, four specimens belonging to the Physalacriaceae were collected on Ulleung Island. These specimens were identified as three species based on morphological characteristics and molecular analysis of rDNA-internal transcribed spacer sequences. Three species in three genera were shown to be new records in Korea: Hymenopellis orientalis, Paraxerula hongoi, and Ponticulomyces orientalis. The latter two are the first records of these genera in Korea. In this study, we provide detailed morphological descriptions of these species and describe their phylogenetic position within the Physalacriaceae.