Vibralactone Z4, the first chain-like vibralactone derivative from cultures of the basidiomycete Boreostereum vibrans.

The first chain-Like vibralactone derivative, vibralactone Z4 (1), together with three known ones (2-4), were isolated from cultures of the basidiomycete Boreostereum vibrans. Vibralactone Z4 (1) is likely to be a key intermediate from 3 to 4 in their biosynthetic pathway. It showed a weak cytotoxicity to human cancer cell line SW480 (IC50 29.2 ± 1.38 µM).

Hymenogaster macmurphyi and Splanchnomyces behrii are sequestrate species of Xerocomellus from the western United States.

Hymenogaster is an ectomycorrhizal genus of brown-spored sequestrate fungi that is related to the mushroom-forming genera Hebeloma and Alnicola (Agaricales). However, because of difficulties in morphological taxonomy of sequestrate fungi, Hymenogaster has become a polyphyletic repository for a variety of unrelated brown-spored sequestrate species. During studies of ectomycorrhizal ecology and sequestrate fungal evolution in the western USA, we encountered specimens of a morphologically unique species. It was originally described as Hymenogaster macmurphyi, but our morphological and molecular analyses indicate that it is not closely related to Hymenogaster. Phylogenetic analyses of multiple gene regions indicate that H. macmurphyi is actually a member of the Boletineae (Boletales, Basidiomycota) and is nested within the epigeous genus Xerocomellus, distantly related to any of the other known genera of sequestrate Boletales. While examining additional herbarium collections, we came upon isotype material of Splanchnomyces behrii, which represents a closely related species. Here we document the morphology and phylogenetic affinities of these unusual sequestrate Boletineae and transfer both species to Xerocomellus as X. macmurphyi and X. behrii. During our study, we also noted that the sequestrate taxon Rhopalogaster transversarius is nested within the epigeous genus Suillus.

Phellinopsis lonicericola and P. tibetica spp. nov. (Hymenochaetales, Basidiomycota), from Tibet, with a key to worldwide species.

Phellinopsis (Hymenochaetaceae) was recently segregated from Phellinus. Phellinopsis differs from other genera segregated from Phellinus in a combination of hymenial setae, if present, mostly originating from tramal hyphae and hyaline basidiospores becoming pale yellowish with age. In this study, two new species of Phellinopsis, P. lonicericola and P. tibetica, from Tibet in southwestern China were described and illustrated. Morphological data and sequences of the nuc rDNA ITS1-5.8S-ITS2 region (internal transcribed spacer [ITS] barcode) support the recognition of these taxa as distinct. They are morphologically compared with other species of Phellinopsis. Notably, P. lonicericola was often found on Lonicera together with Sanghuangporus alpinus, which was originally described from the same locality. In addition, Phellinopsis helwingiae, known from a single collection from Sichuan Province in midwestern China, is reported from Tibet. Eight accepted species of Phellinopsis worldwide, each forming a terminal lineage, grouped together as a strongly supported clade distinct from the clade representing Phellinus. A key to the eight species of Phellinopsis is provided.

Phylogeny and diversity of Fomitiporella (Hymenochaetales, Basidiomycota).

Fomitiporella accommodates polypores producing annual to perennial basidiocarps with an indistinct subiculum (very thin to almost lacking), mostly a dimitic hyphal structure, lacking any kind of setae, with brownish, thick-walled basidiospores, and causing a white rot. Previously, only a few samples of Fomitiporella were studied on the basis of morphological and nuc 28S rDNA (28S)-based phylogenetic analyses. In this study, we made a comprehensive study on Fomitiporella on the basis of collections from Central America, USA, Europe, and China. The phylogenetic analysis, including 28 nuc 28S rDNA and 29 nuc rDNA ITS1-5.8S-ITS2 (internal transcribed spacer [ITS]) sequences newly generated, discovered 14 new lineages. Combined with morphological evidence, 4 new lineages are described and illustrated as new species, viz., Fomitiporella americana, F. micropora, F. sinica, and F. subinermis; 10 other new lineages, each with a single collection, are still treated as unidentified taxa; three new combinations, viz., Fomitiporella tenuissima, F. chinensis, and F. resupinata, are proposed. In addition, F. inermis is redescribed. A key to the 12 known species of Fomitiporella is provided.

Global diversity and phylogeny of Onnia (Hymenochaetaceae) species on gymnosperms.

Onnia includes white rotting polypores with annual basidiocarps, a duplex context, monomitic hyphal structure, hymenial setae, and hyaline, thin-walled, smooth basidiospores. Specimens of Onnia, originating mainly from East Asia, Europe, and North America, were studied using both morphology and phylogenetic analyses. Our concatenated data set was derived from 25 collections and included (i) 25 nuc rDNA internal transcribed spacer region sequences (ITS1-5.8S-ITS2 = ITS), 17 generated in this study; and (ii) 14 nuc rDNA 28S rDNA sequences, including the D1-D2 domains, 11 of them generated in this study. The resulting maximum likelihood and Bayesian phylogenies recovered all sampled collections of Onnia as a well-supported clade. In this clade, three previously accepted species, viz., Onnia leporina, O. tomentosa, and O. triquetra, received strong support, whereas three additional lineages with strong support represent the new species described in this paper, O. subtriquetra, O. microspora, and O. tibetica. Of the six Onnia species occurring on gymnosperms, O. tomentosa and O. leporina grow mainly on Picea and have circumboreal distribution in the Northern Hemisphere. In contrast, other species that mostly grow on Pinus are geographically restricted to limited regions, viz., O. triquetra in Europe, O. subtriquetra in North America, and O. microspora and O. tibetica in Asia.

Morphological and molecular evidence for three new species of Coltricia (Hymenochaetaceae, Basidiomycota) from southern China.

Three new species of Coltricia from southern China are described on the basis of morphological characters and molecular analyses. Phylogenetic analyses based on nuc rDNA ITS1-5.8S-ITS2 and partial 28S rDNA, and partial sequences of genes for RNA polymerase II subunits 1 and 2 confirmed the generic placement of the three new species. Coltricia lateralis is characterized by small, laterally stipitate basidiocarps, obtuse pileus margin, cinnamon-buff to honey yellow pore surface, and broadly ellipsoid basidiospores that measure 7-8 × 5-6 µm. Coltricia rigida is characterized by large, woody-hard fan-shaped basidiocarps, small pores (7-8 per mm), presence of septate sclerified hyphae, and subglobose to globose basidiospores that measure 6-7 × 5-6.5 µm. Coltricia wenshanensis is characterized by centrally stipitate basidiocarps, a distinctly concentrically zonate and sulcate pileal surface, large pores (0.5-2 per mm), and broadly ellipsoid to subglobose basidiospores that measure 7.5-8 × 6-7 µm.

Three new species of Phylloporia (Hymenochaetales) with dimitic hyphal systems from tropical China.

Three new species of Phylloporia from tropical China are described in this study: P. manglietiae, P. pendula, and P. pseudopectinata. Phylloporia manglietiae is characterized by triquetrous to ungulate basidiomata, with 6-8 pores/mm, a monomitic to dimitic hyphal system, and broadly ellipsoidal basidiospores, 3-3.5 × 2-2.5 µm. Phylloporia pendula has small, imbricate, and pendent basidiomata, with 7-9 pores/mm, a dimitic hyphal system, and broadly ellipsoidal basidiospores, 3.5-4 × 2.5-3 µm. Phylloporia pseudopectinata differs from other species of Phylloporia by its applanate basidiomata, with 8-9 pores/mm, a dimitic hyphal system, and subglobose basidiospores, 3-3.5 × 2-3 µm. Phylogenetic analyses, based on sequences from the D1-D2 domains of the 28S gene and the internal transcribed spacer (ITS) region (ITS1-5.8S-ITS2 = ITS) of the nuc rDNA, support the classification of the three new species in Phylloporia.

Characterization of a Ran gene from Puccinia striiformis f. sp. tritici involved in fungal growth and anti-cell death.

Ran, an important family of small GTP-binding proteins, has been shown to regulate a variety of important cellular processes in many eukaryotes. However, little is known about Ran function in pathogenic fungi. In this study, we report the identification and functional analysis of a Ran gene (designated PsRan) from Puccinia striiformis f. sp. tritici (Pst), an important fungal pathogen affecting wheat production worldwide. The PsRan protein contains all conserved domains of Ran GTPases and shares more than 70% identity with Ran proteins from other organisms, indicating that Ran proteins are conserved in different organisms. PsRan shows a low level of intra-species polymorphism and is localized to the nucleus. qRT-PCR analysis showed that transcript level of PsRan was induced in planta during Pst infection. Silencing of PsRan did not alter Pst virulence phenotype but impeded fungal growth of Pst. In addition, heterologous overexpression of PsRan in plant failed to induce cell death but suppressed cell death triggered by a mouse BAX gene or a Pst Ras gene. Our results suggest that PsRan is involved in the regulation of fungal growth and anti-cell death, which provides significant insight into Ran function in pathogenic fungi.

Taxonomy and phylogeny of Phellinidium (Hymenochaetales, Basidiomycota): A redefinition and the segregation of Coniferiporia gen. nov. for forest pathogens.

Phellinidium, including 13 accepted polypore species mostly with resupinate basidiocarps, is one of the most aggressive forest pathogenic genera. This genus is characterized by the combination of a monomitic hyphal structure, abundant hyphoid setae in the context and trama, and hyaline and thin-walled basidiospores. To explore the relationships among the species of Phellinidium, especially those between forest pathogens and saprophytes, we examined 29 specimens representing all 13 previously known species from Asia, Europe and America from morphological and phylogenetic perspectives. A new genus, Coniferiporia, was found to segregate from Phellinidium for three aggressive forest pathogens, and three new combinations, viz. Coniferiporia qilianensis (the generic type), Coniferiporia weirii and Coniferiporia sulphurascens, were proposed. Phellinidium cryptocystidiatum was treated as a synonym of C. sulphurascens. The circumscription of Phellinidium was delimited to accommodate Phellinidium asiaticum, Phellinidium ferrugineofuscum (the generic type), Phellinidium fragrans and Phellinidium pouzarii. Accordingly, the concept of Phellinidium was emended to accommodate resupinate species bearing cylindrical to oblong-ellipsoid or allantoid basidiospores. No species of Phellinidium under the new circumscription has been reported to be a forest pathogen. Phellinidium noxium and Phellinidium rufitinctum were excluded from Phellinidium, while the taxonomical positions of Phellinidium aciferum, Phellinidium lamaënse, and Phellinidium orientale are still uncertain.

Two new species of Fomitiporia (Hymenochaetales, Basidiomycota) from Tibet, southwest China.

Two new species of Fomitiporia growing on Hippophae trees, F. norbulingka and F. subhippophaëicola, are described from southwest China based on morphological characters and phylogenetic analysis. Fomitiporia norbulingka is characterized by pileate basidiomata, mostly angular pores (6-9 per mm), slightly thick-walled generative hyphae, subglobose to globose basidiospores (6.5-7 × 5.5-7 µm), and absence of cystidioles. Fomitiporia subhippophaëicola is diagnostic by effused-reflexed to pileate basidiomata, angular pores (8-10 per mm), thick-walled generative hyphae, subglobose to obovoid basidiospores (6-8 × 5.5-7 µm), and presence of ventricose to fusoid cystidioles. Phylogenetic analysis inferred from combined sequences including the nuc rDNA ITS1-5.8-ITS2 region, 28S rDNA D1-D2 domains, partial sequences of translation elongation factor 1-α, and RNA polymerase II second largest subunits genes indicated that F. norbulingka and F. subhippophaëicola represent two new lineages which group together with F. hippophaëicola.

Global diversity and phylogeny of the Phellinus igniarius complex (Hymenochaetales, Basidiomycota) with the description of five new species.

The Phellinus igniarius complex corresponds to Phellinus s.s., a well supported clade in the polyphyletic Phellinus s.l. studied worldwide. Recently a molecular phylogeny of this complex was performed for the European and North American species. In this study we expand the taxon samplings of the P. igniarius complex to include 59, seven and 12 additional collections originating from China, Czech Republic and USA, respectively. We generated 78 nuc rDNA ITS1-5.8S-ITS2 (ITS) and 42 translation elongation factor 1-α gene (tef1α) sequences. Based on the morphological and phylogenetic (combined ITS and tef1α dataset) analyses, Phellinus monticola, P. orientoasiaticus, P. padicola, P. parmastoi and P. pomaceoides are newly described and illustrated from China and USA. Phellinus pseudoigniarius is treated as a later synonym of P. igniarius, whereas Phellinus betulinus subsp. betulinus is accepted as P. betulinus. A total of 15 species are accepted in the P. igniarius complex worldwide. Of them, 10 species are distributed in eastern Asia, eight in Europe and six in North America. The taxonomy, phylogeny, host associations and geographic distributions of these 15 species are briefly discussed.

Subcutaneous abscess due to the basidiomycete Phellinus mori in a patient with chronic granulomatous disease.

Chronic granulomatous disease (CGD), a primary immunodeficiency caused by impaired phagocyte killing of intracellular pathogens, is characterized by recurrent, life-threatening, bacterial and fungal infections. As a result of improvements in microbiologic culture and identification techniques, a number of unique filamentous fungi have been reported as significant pathogens in patients with CGD. We report a case of subcutaneous basidiomycete Phellinus mori infection in a patient with CGD. To the best of our knowledge, this is the first reported case of human infection by this fungus. The causative fungus was identified on the basis of its morphological characteristics and nucleotide sequence on the internal transcribed spacer region of the ribosomal RNA gene. This is the fifth case report of filamentous basidiomycetes infecting a patient with CGD; all of these cases have been caused by Phellinus species. We highlight the importance of recognizing filamentous basidiomycetes Phellinus species as possible agents of non-Aspergillus fungal infections in patients with CGD.

A revision of the Alpova diplophloeus complex in North America.

Alpova diplophloeus (Boletales, Paxillaceae) is the only currently recognized Alpova in North America with a brownish peridium, large gleba chambers and which forms ectomycorrhizas with Alnus. However, A. diplophloeus as currently circumscribed is a polyphyletic species, with at least three distinct genetic entities. Using a combination of molecular and morphological characters, we examined the type collections of A. diplophloeus, as well as species synonymized with it, including A. cinnamomeus and Rhizopogon parvisporus. We also examined several other collections of A. diplophloeus complex basidiomata. We describe A. diplophloeus sensu stricto; we also resurrect A. cinnamomeus, synonymized with R. parvisporus and describe a new species, A. concolor, from the complex.

Inhibition of cereal rust fungi by both class I and II defensins derived from the flowers of Nicotiana alata.

Defensins are a large family of small, cysteine-rich, basic proteins, produced by most plants and plant tissues. They have a primary function in defence against fungal disease, although other functions have been described. This study reports the isolation and characterization of a class I secreted defensin (NaD2) from the flowers of Nicotiana alata, and compares its antifungal activity with the class II defensin (NaD1) from N. alata flowers, which is stored in the vacuole. NaD2, like all other class I defensins, lacks the C-terminal pro-peptide (CTPP) characteristic of class II defensins. NaD2 is most closely related to Nt-thionin from N. tabacum (96% identical) and shares 81% identity with MtDef4 from alfalfa. The concentration required to inhibit in vitro fungal growth by 50% (IC50 ) was assessed for both NaD1 and NaD2 for the biotrophic basidiomycete fungi Puccinia coronata f. sp. avenae (Pca) and P. sorghi (Ps), the necrotrophic pathogenic ascomycetes Fusarium oxysporum f. sp. vasinfectum (Fov), F. graminearum (Fgr), Verticillium dahliae (Vd) and Thielaviopsis basicola (Tb), and the saprobe Aspergillus nidulans. NaD1 was a more potent antifungal molecule than NaD2 against both the biotrophic and necrotrophic fungal pathogens tested. NaD2 was 5-10 times less effective at killing necrotrophs, but only two-fold less effective on Puccinia species. A new procedure for testing antifungal proteins is described in this study which is applicable to pathogens with spores that are not amenable to liquid culture, such as rust pathogens. Rusts are the most damaging fungal pathogens of many agronomically important crop species (wheat, barley, oats and soybean). NaD1 and NaD2 inhibited urediniospore germination, germ tube growth and germ tube differentiation (appressoria induction) of both Puccinia species tested. NaD1 and NaD2 were fungicidal on Puccinia species and produced stunted germ tubes with a granular cytoplasm. When NaD1 and NaD2 were sprayed onto susceptible oat plants prior to the plants being inoculated with crown rust, they reduced the number of pustules per leaf area, as well as the amount of chlorosis induced by infection. Similar to observations in vitro, NaD1 was more effective as an antifungal control agent than NaD2. Further investigation revealed that both NaD1 and NaD2 permeabilized the plasma membranes of Puccinia spp. This study provides evidence that both secreted (NaD2) and nonsecreted (NaD1) defensins may be useful for broad-spectrum resistance to pathogens.

The target gene of tae-miR164, a novel NAC transcription factor from the NAM subfamily, negatively regulates resistance of wheat to stripe rust.

microRNA (miRNA) participates in various physiological and biochemical processes in plants by regulating corresponding target genes. NAC [NAM (no apical meristem), ATAF (Arabidopsis transcription activation factor) and CUC (cup-shaped cotyledon)] transcription factors, usually as the targets of miR164, play important roles in the regulation of plant development and responses to abiotic and biotic stresses. In a previous study, the target gene of tae-miR164 in wheat was sequenced through degradome sequencing. In this study, we isolated the full-length cDNA of the candidate target gene, which is a NAC transcription factor gene in the NAM subfamily, and designated it as TaNAC21/22 after bioinformatics analysis. The interaction between TaNAC21/22 and tae-miR164 was confirmed experimentally through co-transformation of both genes in tobacco leaves. Transcript accumulation of TaNAC21/22 and tae-miR164 showed contrasting divergent expression patterns in wheat response to Puccinia striiformis f. sp. tritici (Pst). TaNAC21/22 was confirmed to be located in the nucleus and could function as a transcriptional activator. Silencing of the individual gene showed that TaNAC21/22 negatively regulates resistance to stripe rust. These results indicate that the target of tae-miR164, a novel NAC transcription factor from the NAM subfamily of wheat, plays an important role in regulating the resistance of host plants to stripe rust.

Raman microspectroscopy-based identification of individual fungal spores as potential indicators of indoor contamination and moisture-related building damage.

We present an application of Raman microspectroscopy (RMS) for the rapid characterization and identification of individual spores from several species of microfungi. The RMS-based methodology requires minimal sample preparation and small sample volumes for analyses. Hence, it is suitable for preserving sample integrity while providing micrometer-scale spatial resolution required for the characterization of individual cells. We present the acquisition of unique Raman spectral signatures from intact fungal spores dispersed on commercially available aluminum foil substrate. The RMS-based method has been used to compile a reference library of Raman spectra from several species of microfungi typically associated with damp indoor environments. The acquired reference spectral library has subsequently been used to identify individual microfungal spores through direct comparison of the spore Raman spectra with the reference spectral signatures in the library. Moreover, the distinct peak structures of Raman spectra provide detailed insight into the overall chemical composition of spores. We anticipate potential application of this methodology in the fields of public health, forensic sciences, and environmental microbiology.

Molecular and morpho-anatomical description of mycorrhizas of Lactarius rimosellus on Quercus sp., with ethnomycological notes on Lactarius in Guatemala.

Guatemala is one of the richest biodiversity hotspots worldwide, bursting a wild array of ecosystems that range from pine and mixed forests in the highlands to tropical rain forests in the extensive El Petén area, bordering Belize and Mexico. Despite this biological wealth, however, current knowledge on the Guatemalan mycobiota is particularly scant, in part because of the prolonged civil war that has prevented exploration of many ecological niches. In the present paper, we report on the occurrence of Lactarius rimosellus Peck-a rarely discussed species-in oak-pine mixed forests in the Guatemalan highlands and describe the relevant ectomycorrhizae formed with Quercus sp. by means of molecular and morpho-anatomical tools. On the phylogenetic trees constructed on the basis of the partial LSU sequence, sporocarp- and ectomycorrhizae-derived sequences formed a common, statistically supported clade. The structural features of the ectomycorrhizae of L. rimosellus were generally found to match those described on various hosts for other Lactarius species belonging to the subgenus Russularia, where L. rimosellus has been traditionally assigned. These mycorrhizae are characterized by a pseudoparenchymatous outer mantle layer, with epidermoid or angular hyphal cells, and a plectenchymatous inner mantle layer; lactifers are embedded either in the middle and/or inner mantle layer. In the framework of a more general, ongoing study of the ethnomycology of the Maya populations in the Guatemalan highlands, we also report on the traditional knowledge about Lactarius mushrooms and their uses among native people.

Phylogeny and character evolution in the jelly fungi (Tremellomycetes, Basidiomycota, Fungi).

The Tremellomycetes (Agaricomycotina, Basidiomycota, Fungi) are a nutritionally heterogeneous group comprising saprotrophs, animal parasites, and fungicolous species (fungal-inhabiting, including lichen-inhabiting). The relationships of many species, particularly those with a lichenicolous habit, have never been investigated by molecular methods. We present a phylogeny of the Tremellomycetes based on three nuclear DNA ribosomal markers (nSSU, 5.8S and nLSU), representing all main taxonomic groups and life forms, including lichenicolous taxa. The Cystofilobasidiales, Filobasidiales, Holtermanniales, and Tremellales (including the Trichosporonales) are recovered as monophyletic, but this is not the case for the Tremellomycetes. We suggest, however, that the Cystofilobasidiales tentatively continue to be included in the Tremellomycetes. As currently circumscribed, the Filobasidiaceae, Sirobasidiaceae, Syzygosporaceae and Tremellaceae are non-monophyletic. Cuniculitremaceae, Sirobasidiaceae and Tetragoniomycetaceae are nested within Tremellaceae. The lichenicolous species currently included within the Tremellomycetes belong in this group, distributed across the Filobasidiales and Tremellales. Lichen-inhabiting taxa do not form a monophyletic group; they are distributed in several clades and sometimes intermixed with taxa of other nutritional habits. Character state reconstruction indicates that two morphological traits claimed to characterize groups in the Tremellomycetes (the basidium habit and basidium septation) are highly homoplastic. Comparative phylogenetic methods suggest that the transitions between single and catenulate basidia in the Tremellales are consistent with a punctuational model of evolution whereas basidium septation is likely to have evolved under a graduational model in the clade comprising the Holtermanniales, Filobasidiales, and Tremellales.

Molecular phylogenetics of the Gloeophyllales and relative ages of clades of Agaricomycotina producing a brown rot.

The Gloeophyllales is a recently described order of Agaricomycotina containing a morphologically diverse array of polypores (Gloeophyllum), agarics (Neolentinus, Heliocybe) and resupinate fungi (Veluti-Veluticeps, Boreostereum, Chaetodermella), most of which have been demonstrated to produce a brown-rot mode of wood decay and are found preferentially on coniferous substrates. Multiple phylogenetic studies have included taxa of Gloeophyllales, but none have sampled the order thoroughly, and so far only ribosomal RNA genes have been used. Consequently the limits and higher level placement of the Gloeophyllales are obscure. We obtained sequence data for three protein-coding genes (rpb2, atp6, tef1) and three rRNA regions (nuc-ssu, nuc-lsu, 5.8S) in 19 species of Gloeophyllales representing seven genera and analyzed them together with a diverse set of Agaricomycotina, emphasizing Polyporales. Boreostereum, which is suspected to produce a white rot, is the sister group of the rest of the Gloeophyllales, all of which produce a brown rot. Gloeophyllum contains at least two independent clades, one of which might correspond to the genus Osmoporus. White rot and resupinate fruiting bodies appear to be plesiomorphic in Gloeophyllales. Relaxed molecular clock analyses suggest that the Gloeophyllales arose in the Cretaceous, after the origin of Pinaceae.