Tuber eburneum and Tuber mujicii: New pine-associated Tuber species from eastern North America.

The ectomycorrhizal truffle genus Tuber is widespread and diverse. Recent sampling of ascomata, ectomycorrhizal root tips, and environmental sequences has resulted in the identification of many Tuber species that cannot be assigned to described species and require formal description. Using morphological and molecular phylogenetic analysis, we describe two North American Tuber species associated with pines (Pinus spp.). Tuber eburneum, sp. nov., is an early-diverging taxon in the Melanosporum clade that differs substantially from all other taxa in that clade due to its light-colored peridium and gleba, lack of peridial warts, and peridial hairs that are ornamented with small, irregular protrusions. Tuber mujicii, sp. nov., is a whitish truffle species in the Puberulum clade. Although T. mujicii is morphologically similar to many related taxa, it can be distinguished by a combination of characters, including peridium color, spore size, number of ascospores per ascus, and number of reticulations across the spore surface.

Reappraisal of the Genus Exsudoporus (Boletaceae) Worldwide Based on Multi-Gene Phylogeny, Morphology and Biogeography, and Insights on Amoenoboletus.

The boletoid genera Butyriboletus and Exsudoporus have recently been suggested by some researchers to constitute a single genus, and Exsudoporus was merged into Butyriboletus as a later synonym. However, no convincing arguments have yet provided significant evidence for this congeneric placement. In this study, we analyze material from Exsudoporus species and closely related taxa to assess taxonomic and phylogenetic boundaries between these genera and to clarify species delimitation within Exsudoporus. Outcomes from a multilocus phylogenetic analysis (ITS, nrLSU, tef1-α and rpb2) clearly resolve Exsudoporus as a monophyletic, homogenous and independent genus that is sister to Butyriboletus. An accurate morphological description, comprehensive sampling, type studies, line drawings and a historical overview on the nomenclatural issues of the type species E. permagnificus are provided. Furthermore, this species is documented for the first time from Israel in association with Quercus calliprinos. The previously described North American species Exsudoporus frostii and E. floridanus are molecularly confirmed as representatives of Exsudoporus, and E. floridanus is epitypified. The eastern Asian species Leccinum rubrum is assigned here to Exsudoporus based on molecular evidence, and a new combination is proposed. Sequence data from the original material of the Japanese Boletus kermesinus were generated, and its conspecificity with L. rubrum is inferred as formerly presumed based on morphology. Four additional cryptic species from North and Central America previously misdetermined as either B. frostii or B. floridanus are phylogenetically placed but remain undescribed due to the paucity of available material. Boletus weberi (syn. B. pseudofrostii) and Xerocomus cf. mcrobbii cluster outside of Exsudoporus and are herein assigned to the recently described genus Amoenoboletus. Biogeographic distribution patterns are elucidated, and a dichotomous key to all known species of Exsudoporus worldwide is presented.

Greetings from belowground: two new species of truffles in the genus Pachyphlodes (Pezizaceae, Pezizales) from México.

Pachyphlodes is a lineage of ectomycorrhizal, hypogeous, sequestrate ascomycete fungi native to temperate and subtropical forests in the Northern Hemisphere. Pachyphlodes species form ectomycorrhizae mainly with Fagales hosts. Here we describe two new species of Pachyphlodes, P.brunnea, and P.coalescens, based on morphological and phylogenetic analysis. Pachyphlodesbrunnea is distributed in the states of Tamaulipas and Nuevo León in northern México, occurring with Quercus and Juglans species. It is characterized by its dark brown peridium, white gleba, and spores with capitate columns. Pachyphlodescoalescens is distributed in the states of Michoacán and Tlaxcala in central and southwestern México co-occurring with Quercus and is distinguished by its reddish-brown peridium, light yellow gleba, and spore ornamentation. Both species, along with P.marronina, constitute the Marronina clade. This clade contains North American species characterized by a brown peridium and spores ornamented with capitate spines to coalesced spine tips that form a partial perispore.

Molecular systematics and taxonomic overview of the bird's nest fungi (Nidulariaceae).

Fungi in the Nidulariaceae, otherwise known as 'bird's nest fungi', are among the least studied groups of Agaricomycetes (Basidiomycota). Bird's nest fungi are globally distributed and typically grow on woody debris or animal dung as saprotrophs. This group of fungi is morphologically diverse with ca. 200 described species. Phylogenetic relationships of bird's nest fungi were investigated with four commonly used loci (ITS, LSU, tef, and rpb2). The family was resolved as a monophyletic group with Squamanitaceae as a potential sister taxon. Cyathus and Crucibulum each formed its own independent and well-supported clade. Nidula and Nidularia formed a clade together, but each genus is polyphyletic. Two Mycocalia species included in our analyses were on their own separate branches, indicating that this genus is also polyphyletic. Misidentifications were detected in most genera, suggesting that species concepts need to be revisited and refined throughout Nidulariaceae. Several bird's nest fungi species have global geographical distributions whereas others may have more limited ranges. Basic morphological characters of bird's nest fungi have likely been lost or gained multiple times. The phylogenetic placement of Crucibulum is unclear and the sister lineage of bird's nest fungi is not conclusive. Further studies with data from rare species and additional informative genes are needed to fully resolve the topology of Nidulariaceae and identify its sister group with more certainty.

Systematic study of truffles in the genus Ruhlandiella, with the description of two new species from Patagonia.

Ruhlandiella is a genus of exothecial, ectomycorrhizal fungi in the order Pezizales. Ascomata of exothecial fungi typically lack a peridium and are covered with a hymenial layer instead. Ruhlandiella species have nonoperculate asci and highly ornamented ascospores. The genus was first described by Hennings in 1903 to include the single species, R. berolinensis. Since then, mycologists have uncovered Ruhlandiella species in many locations around the globe, including Australia, Spain, Italy, and the USA. Currently, there are four recognized species: R. berolinensis, R. peregrina, R. reticulata, and R. truncata. All were found near Eucalyptus or Melaleuca trees of Australasian origin. Recently, we discovered two new species of Ruhlandiella in Nothofagaceae forests in South America. They regularly form mitotic spore mats directly on soil in the forests of Patagonia. Here, we formally describe these new species and construct the phylogeny of Ruhlandiella and related genera using a multilocus phylogenetic analysis. We also revise the taxonomy of Ruhlandiella and provide an identification key to accepted species of Ruhlandiella.

A molecular and morphological re-examination of the generic limits of truffles in the tarzetta-geopyxis lineage - Densocarpa, Hydnocystis, and Paurocotylis.

Truffle species within the /tarzetta-geopyxis lineage share smooth, globose, hyaline spores, but differ in the amount of convolution of hymenia in ascomata. The relationships among truffle species in this lineage have historically been confused. Phylogenetic analyses of the ITS and 28S nuclear ribosomal DNA from recently collected members of the /tarzetta-geopyxis lineage from Asia, Austral Asia, North America, and South America prompted a reinvestigation of species and generic limits in the truffle genera Hydnocystis, Paurocotylis, and Stephensia. Our analyses support emendations of Hydnocystis and Paurocotylis, abandonment of Stephensia and the resurrection of the genus Densocarpa. Nomenclatural changes include the transfer of Stephensia bombycina to Hydnocystis, the transfer of Hydnocystis singeri and Stephensia bynumii to Paurocotylis, the reinstatement of Densocarpa for Stephensia shanori and transfer of Stephensia crocea to Densocarpa. This is the first detection of the genus Paurocotylis in the Americas. We describe three new species, Hydnocystis transitoria from North America, Paurocotylis patagonica from South America, and Paurocotylis watlingii from Australia. Our work highlights the unexplored diversity, morphological plasticity, and remaining taxonomic problems among truffles in the /tarzetta-geopyxis lineage.

Mycorrhizal detection of native and non-native truffles in a historic arboretum and the discovery of a new North American species, Tuber arnoldianum sp. nov.

During a study comparing the ectomycorrhizal root communities in a native forest with those at the Arnold Arboretum in Massachusetts (USA), the European species Tuber borchii was detected on the roots of a native red oak in the arboretum over two successive years. Since T. borchii is an economically important edible truffle native to Europe, we conducted a search of other roots in the arboretum to determine the extent of colonization. We also wanted to determine whether other non-native Tuber species had been inadvertently introduced into this 140-year-old Arboretum because many trees were imported into the site with intact soil and roots prior to the 1921 USDA ban on these horticultural practices in the USA. While T. borchii was not found on other trees, seven other native and exotic Tuber species were detected. Among the North American Tuber species detected from ectomycorrhizae, we also collected ascomata of a previously unknown species described here as Tuber arnoldianum. This new species was found colonizing both native and non-native tree roots. Other ectomycorrhizal taxa that were detected included basidiomycetes in the genera Amanita, Russula, Tomentella, and ascomycetes belonging to Pachyphlodes, Helvella, Genea, and Trichophaea. We clarify the phylogenetic relationships of each of the Tuber species detected in this study, and we discuss their distribution on both native and non-native host trees.

Cultural and cytological characterization of Dacryopinax primogenitus, a new species in the Dacrymycetes with a fully sequenced genome.

A Dacryopinax species that was cultured in Costa Rica and fruited in the laboratory provided DNA for the first sequenced genome for the Dacrymycetes. Here we characterize the isolate morphologically and cytologically and name it D. primogenitus Molecular sequences from the nuclear large subunit gene and internal transcribed spacer indicated that it is closely related to the South American D. indacocheae with which it agrees structurally. Both species form conidia on the basidiocarp, and D primogenitus also forms them on the mycelium. Unlike previous reports for the Dacrymycetales postmeiotic nuclear division results in uninucleate basidiospores and six residual nuclei in the basidium after basidiospore discharge. Ultrastructural analysis shows the characteristic septal-pore apparatus for the class and endogenous origin of the epibasidia/sterigmata, which may be a common occurrence in Dacrymycetes and the early diverging orders of its sister class, the Agaricomycetes.

A novel orellanine containing mushroom Cortinarius armillatus.

Orellanine (3,3',4,4'-tetrahydroxy-2,2'-bipyridine-1,1'-dioxide) is a tetrahydroxylated di-N-oxidized bipyridine compound. The toxin, present in certain species of Cortinarius mushrooms, is structurally similar to herbicides Paraquat and Diquat. Cortinarius orellanus and Cortinarius rubellus are the major orellanine-containing mushrooms. Cortinarius mushrooms are widely reported in Europe where they have caused human poisoning and deaths through accidental ingestion of the poisonous species mistaken for the edible ones. In North America, Cortinarius orellanosus mushroom poisoning was recently reported to cause renal failure in a Michigan patient. Cortinarius mushroom poisoning is characterized by delayed acute renal failure, with some cases progressing to end-stage kidney disease. There is debate whether other Cortinarius mushroom contain orellanine or not, especially in North America. Currently, there are no veterinary diagnostic laboratories in North America with established test methods for detection and quantitation of orellanine. We have developed two diagnostic test methods based on HPLC and LC-MSMS for identification and quantitation of orellanine in mushrooms. Using these methods, we have identified Cortinarius armillatus as a novel orellanine-containing mushroom in North America. The mean toxin concentration of 145 ug/g was <1% of that of the more toxic C. rubellus. The HPLC method can detect orellanine at 17 µg g(-1) while the LC-MSMS method is almost 2000 times more sensitive and can detect orellanine at 30 ng g(-1). Both tests are quantitative, selective and are now available for veterinary diagnostic applications.

Evolution of zygomycetous spindle pole bodies: Evidence from Coemansia reversa mitosis.

PREMISE OF THE STUDY: The earliest eukaryotes were likely flagellates with a centriole that nucleates the centrosome, the microtubule-organizing center (MTOC) for nuclear division. The MTOC in higher fungi, which lack flagella, is the spindle pole body (SPB). Can we detect stages in centrosome evolution leading to the diversity of SPB forms observed in terrestrial fungi? Zygomycetous fungi, which consist of saprobes, symbionts, and parasites of animals and plants, are critical in answering the question, but nuclear division has been studied in only two of six clades. METHODS: Ultrastructure of mitosis was studied in Coemansia reversa (Kickxellomycotina) germlings using cryofixation or chemical fixation. Character evolution was assessed by parsimony analysis, using a phylogenetic tree assembled from multigene analyses. KEY RESULTS: At interphase the SPB consisted of two components: a cytoplasmic, electron-dense sphere containing a cylindrical structure with microtubules oriented nearly perpendicular to the nucleus and an intranuclear component appressed to the nuclear envelope. Markham's rotation was used to reinforce the image of the cylindrical structure and determine the probable number of microtubules as nine. The SPB duplicated early in mitosis and separated on the intact nuclear envelope. Nuclear division appears to be intranuclear with spindle and kinetochore microtubules interspersed with condensed chromatin. CONCLUSIONS: This is the sixth type of zygomycetous SPB, and the third type that suggests a modified centriolar component. Coemansia reversa retains SPB character states from an ancestral centriole intermediate between those of fungi with motile cells and other zygomycetous fungi and Dikarya.

Functional and phylogenetic implications of septal pore ultrastructure in the ascoma of Neolecta vitellina.

Neolecta represents the earliest derived extant ascomycete lineage (Taphrinomycotina) to produce ascomata. For this reason the genus has been of interest with regard to ascoma evolution in ascomycetes. However, the evidence is equivocal regarding whether the Neolecta ascoma is homologous or analogous to ascomata produced in the later derived ascomycete lineages (Pezizomycotina). We investigated phylogenetically informative septal pore ultrastructure of Neolecta vitellina to compare with Pezizomycotina. We found that crystalline bodies that block nonascogenous septal pores in Neolecta differ from Woronin bodies, a synapomorphy for the Pezizomycotina, in three ways: (i) vacuolar origin, (ii) associated material and (iii) being loosely membrane bound. We also observed a unique type of membranous material within the septal pore, as well as distant from the septal pore, that appears to be associated with the endoplasmic reticulum. The vacuolar crystals and membranous material might have a function analagous to septal pore structures (e.g. Woronin bodies, lamellate structures) in the Pezizomycotina. Morphological evidence from our study supports an independently derived septal pore-occluding structure in the Neolecta lineage.

Historical biogeography and diversification of truffles in the Tuberaceae and their newly identified southern hemisphere sister lineage.

Truffles have evolved from epigeous (aboveground) ancestors in nearly every major lineage of fleshy fungi. Because accelerated rates of morphological evolution accompany the transition to the truffle form, closely related epigeous ancestors remain unknown for most truffle lineages. This is the case for the quintessential truffle genus Tuber, which includes species with socio-economic importance and esteemed culinary attributes. Ecologically, Tuber spp. form obligate mycorrhizal symbioses with diverse species of plant hosts including pines, oaks, poplars, orchids, and commercially important trees such as hazelnut and pecan. Unfortunately, limited geographic sampling and inconclusive phylogenetic relationships have obscured our understanding of their origin, biogeography, and diversification. To address this problem, we present a global sampling of Tuberaceae based on DNA sequence data from four loci for phylogenetic inference and molecular dating. Our well-resolved Tuberaceae phylogeny shows high levels of regional and continental endemism. We also identify a previously unknown epigeous member of the Tuberaceae--the South American cup-fungus Nothojafnea thaxteri (E.K. Cash) Gamundí. Phylogenetic resolution was further improved through the inclusion of a previously unrecognized Southern hemisphere sister group of the Tuberaceae. This morphologically diverse assemblage of species includes truffle (e.g. Gymnohydnotrya spp.) and non-truffle forms that are endemic to Australia and South America. Southern hemisphere taxa appear to have diverged more recently than the Northern hemisphere lineages. Our analysis of the Tuberaceae suggests that Tuber evolved from an epigeous ancestor. Molecular dating estimates Tuberaceae divergence in the late Jurassic (~156 million years ago), with subsequent radiations in the Cretaceous and Paleogene. Intra-continental diversification, limited long-distance dispersal, and ecological adaptations help to explain patterns of truffle evolution and biodiversity.

New North American truffles (Tuber spp.) and their ectomycorrhizal associations.

Recent surveys of belowground fungal biodiversity in México and USA have revealed many undescribed truffle species, including many in the genus Tuber. Here we describe seven new species: Tuber beyerlei, T. castilloi, T. guevarai, T. lauryi, T. mexiusanum, T. miquihuanense and T. walkeri. Phylogenetic analyses place these species within the Maculatum group, an understudied clade of small truffles with little apparent economic value. These species are among the more taxonomically challenge-ing in the genus. We collected Tuber castilloi, T. mexiusanum and T. guevarai as fruit bodies and ectomycorrhizae on Quercus spp. in forests of eastern México. Tuber mexiusanum has a particularly broad geographic range, being collected in eastern USA under Populus deltoides and in Minnesota and Iowa in mixed hardwood forests. T. walkeri is described from the upper midwestern USA, and T. lauryi and T. beyerlei occur in the western USA.

Terfezia disappears from the American truffle mycota as two new genera and Mattirolomyces species emerge.

Reexamination and molecular phylogenetic analyses of American Terfezia species and Mattirolomyces tiffanyae revealed that their generic assignments were wrong. Therefore we here propose these combinations: Mattirolomyces spinosus comb. nov. (≡ Terfezia spinosa), Stouffera longii gen. & comb. nov. (≡ Terfezia longii) and Temperantia tiffanyae gen. & comb. nov. (≡ Mattirolomyces tiffanyae). In addition we describe a new species, Mattirolomyces mexicanus spec. nov. All species belong to the Pezizaceae. Based on these results Terfezia is not known from North America, Mattirolomyces is represented by two species and two new monotypic genera are present.

Otidea subterranea sp. nov.: Otidea goes below ground.

Evidence suggests that truffle-like sporocarp forms have evolved many times in the Pezizales, but primarily from epigeous ancestors within ectomycorrhizal clades. There are several ectomycorrhizal clades, however, that contain no known hypogeous species. We collected specimens of an unusual unidentified truffle from mixed oak woodlands in Iowa. Although clearly a member of the Pezizales (Ascomycota), this hypogeous species did not belong to any of the described truffle genera. Based on a combination of ecological, phylogenetic, and morphological evidence we determined that this new truffle is a hypogeous member of the genus Otidea (Pyronemataceae), a lineage with no described truffle species. We describe it here as a new species, Otidea subterranea.