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Molecular systematics of the Hypocreales: A teleomorph gene phylogeny and the status of their anamorphs
Abstract
Phylogenetic relationships among 40 species in the Hypocreales and Clavicipitales were inferred from sequence data obtained from the nuclear large-subunit ribosomal DNA. Cladistic analysis of these data support the monophyly of the Hypocreales, with the Clavicipitales derived from within the Hypocreales. Four groupings were resolved and are informally designated as the Hypocrea, Claviceps, Bionectria, and Nectria groups. Phylogenetic placement of teleomorphs including Melanospora and cleistothecial taxa, such as Heleococcum, Mycoarachis, and Roumegueriella, demonstrate the facility of molecular phylogenies to accommodate taxa with highly modified morphologies. Similarly, the hypocrealean origins of the anamorph species Verticillium lecanii and Acremonium chrysogenum illustrate the potential of the molecular phylogenetic approach to accommodate anamorph isolates within the context of a teleomorph phylogeny. Together these results suggest that a comprehensive classification of the Hypocreales, inclusive of teleomorph and anamorph states, is attainable through a molecular phylogenetic approach. Key words: Ascomycotina, Clavicipitaceae, Hypocreales, phylogeny, pyrenomycete, rDNA.
... The amplification cycle was as follow: denaturation (94°C for 2 min); 35 cycles of denaturation (94°C for 10 sec), annealing (56°C for 10 sec) and elongation (72°C for 30 sec); final elongation (72°C for 8 min). Finally, a PCR targeting the large-subunit ribosomal RNA region (LSU-rRNA) was done using the Allin™ RedTaq Mastermix, 2x (highQu GmbH) and the primers LR0R-F (5'-ACCCGCTGAACTTAAGC-3') [31] and LR6-R (5'-CGCCAGTTCTGCTTACC-3') [32]. The amplification cycle was as follow: denaturation (95°C for 5 min); 35 cycles of denaturation (94°C for 30 sec), annealing (52°C for 30 sec) and elongation (72°C for 1 min); final elongation (72°C for 15 sec). ...
Progresses in geothermal energy and deep drilling technologies have opened a new window into the terrestrial subsurface. This provides direct access to deep geothermal fluids used to produce heat and electricity, creating an opportunity to isolate and characterize novel microbial strains from these extreme habitats. In this study, we report the co-isolation of two fungal strains. Penicillium citrinum (strain HEK1) was isolated first and thought to be axenic. However, upon exposure to stress (frost and ethanol), a second strain, corresponding to the dimorphic yeast Meyerozyma guilliermondii (strain HEK2), appeared in HEK1 cultures. Strain HEK2 appeared first in the cultures and was followed by the subsequent re-growth of strain HEK1, underscoring their close relationship. Moreover, strain HEK2, able to switch from yeast cells to pseudohyphae when growing alone, did not produce pseudohyphae when in direct contact with strain HEK1. Altogether, our results indicate an intricate interaction between these strains that may allow them to thrive in the deep subsurface. These two fungi represent the first fungal strains isolated from deep geothermal fluids. Their presence within the fluids was confirmed through molecular analysis. The isolation of these strains emphasizes the importance of considering fungi when investigating microbial diversity in subsurface geothermal environments.
Graphical abstract
Highlights
First fungal strains isolated from a geothermal power plant
The two fungal strains were co-isolated from a geothermal fluid used for heat production
Surprising isolation of the cell-switching yeast upon stress exposure of an apparently axenic culture of the filamentous fungus
Fungal strains with high resistance to stressors and no apparent competition for carbon sources
... Polymerase chain reaction (PCR) amplification was applied to amplify four gene fragments, including ITS, LSU, tef1, and tub2. The primer pairs were used: ITS1/ITS4 for ITS (White et al. 1990), LR0R/LR5 for LSU (Rehner and Samuels 1995), EF1-728F/EF2 for tef1 (O'Donnell et al, 1998;Carbone and Kohn 1999), and T1/Bt2b for tub2 (Glass and Donaldson 1995;O'Donnell and Cigelnik 1997). PCR program for ITS amplification was conducted with an initial denaturation at 95 °C for 3 min, followed by 35 cycles of 95 °C for 30 s, annealing at 58 °C for 30 s, extension at 72 °C for 1 min, and a final extension at 72 °C for 7 min. ...
Apiospora species are widely distributed fungi with diverse lifestyles, primarily functioning as plant pathogens, as well as exhibiting saprophytic and endophytic behaviors. This study reports the discovery of three new species of Apiospora, namely A. gongcheniae, A. paragongcheniae, and A. neogongcheniae, isolated from healthy Poaceae plants in China. These novel species were identified through a multi-gene phylogenetic analysis. The phylogenetic analysis of the combined ITS, LSU, tef1, and tub2 sequence data revealed that the three new species formed a robustly supported clade with A. garethjonesii, A. neogarethjonesii, A. setostroma, A. subrosea, A. mytilomorpha, and A. neobambusae. Detailed descriptions of the newly discovered species are provided and compared with closely related species to enhance our understanding of the genus Apiospora.
... We amplified and sequenced the partial small subunit nuc rDNA (18S) using primers pair N24 and NS1 (Gargas and Taylor 1992;White et al. 1990), nuc rDNA internal transcriber spacers (ITS = ITS1+5.8S+ITS2) and the partial nuc rDNA large subunit (28S) with primers ITS5 and LR5 (Vilgalys and Hester 1990;White et al. 1990), and translation elongation factor-1 alpha (tef-1) with primers EF1-2228R and EF1-1983F (Rehner and Samuels 1995). The amplification parameters and additional detailed information on polymerase chain reaction (PCR), visualization of amplicons, and preparation of samples can be found in Rodriguez Flakus and Printzen (2014) and Flakus et al. (2019b). ...
Acrospermales represent one of the least studied lineages of Dothideomycetes and are characterized by diverse ecological strategies, including saprotrophic, epiphytic, fungicolous, lichenicolous, and bryophilous lifestyles. The order is composed of two teleomorphic genera, Acrospermum and Oomyces, and five anamorphic genera of unclear relationships. The objectives of the study were to establish the phylogenetic position of Acrospermum species collected from lichens in the tropical forest of Bolivia and to infer the evolution of the lichenicolous lifestyle in Acrospermales. Our results reveal that the examined specimens from Bolivia represent a new species, A. bolivianum, which is well characterized by its phylogenetic distinctness, morphological characteristics, and host selection. The new species is the first lichenicolous member of Acrospermum and forms a well-supported clade sister to the bryophilous Acrospermum adeanum. The evolution of lifestyles, concluded by phylogenetic analyses and ancestral state reconstructions, indicated that the saprotrophic lifestyle is ancestral to Acrospermales. This corresponds to their close relationship to other saprotrophic lineages of Dothideomycetes and indicates that the wide spectrum of nutritional strategies, currently observed in Acrospermales, may be a result of more recent shifts in their ecology. Our results also suggest that the lichenicolous lifestyle in Acrospermales appeared independently at least two times. Lichenicolous species are represented in our data set by Acrospermum bolivianum and Gonatophragmium physciae, which evolved from lichenicolous and plant-parasite ancestors, respectively. The genus Oomyces, represented by O. carneoalbus, was included for the first time in the phylogenetic analysis and showed a sister relationship to the remaining taxa of Acrospermales.
... Six gene regions were amplified through PCR, which included: (i) the internal transcribed spacer 5.8S rDNA (ITS) with primers ITS1 & ITS4 (White et al. 1990); (ii) the nuclear 28S large ribosomal subunit (LSU) with primers LROR & LR5 (Vilgalys and Hester 1990;Rehner and Samuels 1995); (iii) the nuclear 18S small ribosomal subunit (SSU) with primer pairs NS1 & NS4 and NS3 & NS8 (White et al. 1990); (iv) the RNA polymerase II second largest subunit (RPB2) with primers RPB2-5f2 & RPB2-7cR (Liu et al. 1999;Sung et al. 2007); (v) the mini chromosome maintenance protein complex 7 (MCM7) with primers Cer-MCM7F & Cer-MCM7R (De Beer et al. 2014); and (vi) the translation elongation factor 1-alpha (TEF1) large intron with primers EF1-782F & EF-2 (O'Donnell et al. 1998;Carbone and Kohn 1999). PCRs were prepared in 25 µL reaction volumes and contained 5 µL 5X MyTaq ™ Reaction Buffer (Bioline, London, UK), 0.5 µL (1 µL for MCM7) of each primer (10 µM), 0.3 µL MyTaq ™ DNA Polymerase and 17.7 µL (16.7 µL for MCM7) sterile deionized water. ...
Araucaria araucana is an ancient conifer, native to the mountain ranges in Chile and Argentina. These trees host a large number of organisms, mainly insects, strongly or even exclusively associated with them. The recent emergence of a novel canker disease on A. araucana has emphasised the importance of fungi associated with these iconic trees and has resulted in the discovery of various new species. In this study, we considered the identity of an unknown calicioid fungus consistently found on resin on the branches of A. araucana. Preliminary phylogenetic analyses placed isolates in the recently described sub-class Cryptocaliciomycetidae, closest to Cryptocalicium blascoi. However, the morphology of the ascomata and its occurrence in a unique niche suggested that the closest relative could be Resinogalea humboldtensis (Bruceomycetaceae, incertae sedis), a fungus with similar sporing structures found on resin of Araucaria humboldtensis in New Caledonia. There are no living cultures or sequence data available for either R. humboldtensis or its supposed closest relative, Bruceomyces castoris, precluding sequence-based comparisons. Morphological comparisons of the sporing structures on A. araucana confirmed that the ascomatal morphology of our unknown calicioid fungus and R. humboldtensis are almost identical and resemble each other more so than B. castoris or Cr. blascoi. A phylogenetic analysis based on the small subunit (SSU), internal transcribed spacer (ITS) and large subunit (LSU) rDNA regions resolved our strains into two clades with Cr. blascoi as its closest relative. Further analyses applying the Genealogical Concordance Phylogenetic Species Recognition (GCPSR) based on ITS, mini chromosome maintenance protein complex (MCM7), RNA polymerase II second largest subunit (RPB2) and translation elongation factor 1-alpha (TEF) gene regions, confirmed that strains represent two new species. Based on our morphological observations and phylogenetic analyses, we introduce two new Resinogalea species, R. araucana and R. tapulicola, and reclassify the genus in the subclass Cryptocaliciomycetidae.
Soil structure and aggregation are crucial for soil functionality, particularly under drought conditions. Saprobic soil fungi, known for their resilience in low moisture conditions, are recognized for their influence on soil aggregate dynamics. In this study, we explored the potential of fungal amendments to enhance soil aggregation and hydrological properties across different moisture regimes. We used a selection of 29 fungal isolates, recovered from soils treated under drought conditions and varying in colony density and growth rate, for single‐strain inoculation into sterilized soil microcosms under either low or high moisture (≤−0.96 and –0.03 MPa, respectively). After 8 weeks, we assessed soil aggregate formation and stability, along with soil properties such as soil water content, water hydrophobicity, sorptivity, total fungal biomass and water potential. Our findings indicate that fungal inoculation altered soil hydrological properties and improved soil aggregation, with effects varying based on the fungal strains and soil moisture levels. We found a positive correlation between fungal biomass and enhanced soil aggregate formation and stabilization, achieved by connecting soil particles via hyphae and modifying soil aggregate sorptivity. The improvement in soil water potential was observed only when the initial moisture level was not critical for fungal activity. Overall, our results highlight the potential of using fungal inoculation to improve the structure of agricultural soil under drought conditions, thereby introducing new possibilities for soil management in the context of climate change.
The apothecial fungus Ramomarthamyces octomerus sp. nov. is described from specimens collected in Mediterranean climate regions in southern Portugal, Spain (Canary Islands), and the Dalmatian region of Croatia. Presumably saprobic, R. octomerus occurs on intact, decorticated wood of Laurus novocanariensis and Olea europaea. Ascospores are cylindric-ellipsoid and seven-septate. Surprisingly, in our four-locus phylogenetic analysis (nuSSU, ITS1-5.8S-ITS2, LSU, mtSSU), this fungus clusters among species of Cyclaneusma, Marthamyces, Naemacyclus, and Ramomarthamyces in a core Marthamycetaceae clade that circumscribes primarily leaf-inhabiting, filiform-spored species. In addition, the asci of R. octomerus possess an amyloid pore, but the reaction varies between specimens collected in the Canary Islands and those collected in Portugal and Croatia. The occurrence of an amyloid reaction in the asci of R. octomerus challenges the characterization of Marthamycetales taxa as possessing inamyloid asci. In our discussion we provide background and analysis of these notable observations.
Community assembly is influenced by environmental niche processes as well as stochastic processes that can be spatially dependent (e.g. dispersal limitation) or independent (e.g. priority effects). Here, we sampled senesced tree leaves as unit habitats to investigate fungal community assembly at two spatial scales: (i) small neighborhoods of overlapping leaves from differing tree species and (ii) forest stands of differing ecosystem types. Among forest stands, ecosystem type explained the most variation in community composition. Among adjacent leaves within stands, variability in fungal composition was surprisingly high. Leaf type was more important in stands with high soil fertility and dominated by differing tree mycorrhizal types (sugar maple vs. basswood or red oak), whereas distance decay was more important in oak-dominated forest stands with low soil fertility. Abundance of functional groups was explained by environmental factors, but predictors of taxonomic composition within differing functional groups were highly variable. These results suggest that fungal community assembly processes are clearest for functional group abundances and large spatial scales. Understanding fungal community assembly at smaller spatial scales will benefit from further study focusing on differences in drivers for different ecosystems and functional groups, as well as the importance of spatially independent factors such as priority effects.
Mycophilic or fungicolous fungi can be found wherever fungi exist since they are able to colonize other fungi, which occupy a diverse range of habitats. Some fungicolous species cause important diseases on basidiomycetes and thus, they are the main reason for the destruction of mushroom cultivations. Nonetheless, despite their ecological significance, their genomic data remain limited. Cladobotrum mycophilum is one of the most aggressive species of the genus, destroying the economically important Agaricus bisporus cultivations. The 40.7 Mb whole genome of the Greek isolate ATHUM6906 is assembled in 16 fragments, including the mitochondrial genome and two small circular mitochondrial plasmids, in this study. This genome includes a comprehensive set of 12,282 protein coding, 56 rRNA, and 273 tRNA genes. Transposable elements, CAZymes and pathogenicity related genes were also examined. The genome of C. mycophilum contained a diverse arsenal of genes involved in secondary metabolism, forming 106 Biosynthetic Gene Clusters, which renders this genome as one of the most BGC abundant among fungicolous species. Comparative analyses were performed for genomes of species of the family Hypocreaceae. Some BGCs identified in C. mycophilum genome exhibited similarities to clusters found in family Hypocreaceae, suggesting vertical heritage. In contrast, certain BGCs showed a scattered distribution among Hypocreaceae species or were solely found in Cladobotryum genomes. This work provides evidence of extensive BGC losses, Horizontal Gene Transfer events and formation of novel BGCs during evolution, potentially driven by neutral or even positive selection pressures. These events may increase Cladobotryum fitness under various environmental conditions and potentially during host-fungus interaction.
Here, we present the newly identified Inosperma macrocarpa and the first record of I. afro-melliolens from West Africa. Inosperma macrocarpa is nested in an Old World Tropical clade, based on a molecular phylogeny inferred from the sequences of ITS, LSU, RPB2, and TEF1. Complete descriptions and illustrations, including photographs and line drawings , of the new species are presented. Morphological and molecular analyses based on collections from Benin confirmed the presence of I. afromelliolens in West Africa. Toxicity analysis showed that neither species contained muscarine, which further supports the hypothesis that the ability to produce muscarine is a derived trait of Inosperma.
This study documents the morphology and phylogeny of ascomycetes collected from karst landscapes of Guizhou Province, China. Based on morphological characteristics in conjunction with DNA sequence data, 70 species are identified and distributed in two classes (Dothideomycetes and Sordariomycetes), 16 orders, 41 families and 60 genera. One order Planisphaeriales, four families Leptosphaerioidaceae, Neoleptosporellaceae, Planisphaeriaceae and Profundisphaeriaceae, ten genera Conicosphaeria, Karstiomyces, Leptosphaerioides, Neoceratosphaeria, Neodiaporthe, Neodictyospora, Planisphaeria, Profundisphaeria, Stellatus and Truncatascus, and 34 species (Amphisphaeria karsti, Anteaglonium hydei, Atractospora terrestris, Conicosphaeria vaginatispora, Corylicola hydei, Diaporthe cylindriformispora, Dictyosporium karsti, Hysterobrevium karsti, Karstiomyces guizhouensis, Leptosphaerioides guizhouensis, Lophiotrema karsti, Murispora hydei, Muyocopron karsti, Neoaquastroma guizhouense, Neoceratosphaeria karsti, Neodiaporthe reniformispora, Neodictyospora karsti, Neoheleiosa guizhouensis, Neoleptosporella fusiformispora, Neoophiobolus filiformisporum, Ophioceras guizhouensis, Ophiosphaerella karsti, Paraeutypella longiasca, Paraeutypella karsti, Patellaria guizhouensis, Planisphaeria karsti, Planisphaeria reniformispora, Poaceascoma herbaceum, Profundisphaeria fusiformispora, Pseudocoleophoma guizhouensis, Pseudopolyplosphaeria guizhouensis, Stellatus guizhouensis, Sulcatispora karsti and Truncatascus microsporus) are introduced as new to science. Moreover, 13 new geographical records for China are also reported, which are Acrocalymma medicaginis, Annulohypoxylon thailandicum, Astrosphaeriella bambusae, Diaporthe novem, Hypoxylon rubiginosum, Ophiosphaerella agrostidis, Ophiosphaerella chiangraiensis, Patellaria atrata, Polyplosphaeria fusca, Psiloglonium macrosporum, Sarimanas shirakamiense, Thyridaria broussonetiae and Tremateia chromolaenae. Additionally, the family Eriomycetaceae was resurrected as a non-lichenized family and accommodated within Monoblastiales. Detailed descriptions and illustrations of all these taxa are provided.
The characteristics of ascomal development in Melanospora zamiae conformed to the ascohymenial type of ontogeny, in which the ascoma, a perithecium, developed from a coiled ascomal initial, which was eventually enclosed by investing hyphae that formed the walls. The centrum was composed of two tissues: a centrum prosenchyma composed of several layers of flattened cells that line the ascomal wall, and the centrum pseudoparenchyma that filled the center of the ascoma both above and below the layer of asci. Both the suprahymenial and subhymenial pseudoparenchyma tissues disintegrated as the asci matured, whereas the centrum prosenchyma remained intact. No paraphyses were formed. The ascogenous system was differentiated from the ascogonial cell and the asci formed from binucleate ascus mother cells without the formation of croziers, resulting in asci of different ages in a single perithecium. Ascomal ontogeny in M. zamiae showed similarities to the Sordaria and Diaporthe types of ontogeny, but also differed from them in the lack of paraphyses.
Light and scanning electron microscopy showed that conidiogenesis is holoblastic in the Ephelis states of Balansia epichloë, B. henningsiana, B. pilulaeformis, B. strangulans, and Atkinsonella hypoxylon; phialidic in the Sphacelia states of Claviceps purpurea and Epichloë typhina, and of A. hypoxylon, which produces both conidial states. Balansia strangulans produces a continuous palisade of conidiophores over the surface of the stroma and beneath an evanescent peridial layer. In B. epichloë and B. henningsiana, sterile ridges separate the conidiophore palisade into flat, hysteriform cavities in the surface of the stroma. In a previously unreported Ephelis state of B. pilulaeformis (=Balansiopsis pilulaeformis), conidiophores are restricted to small discoid cavities in the stroma. Atkinsonella hypoxylon produces apothecioid conidiomata in specialized protuberances from the stroma, as does B. claviceps (anamorph = E. mexicana, type of the genus Ephelis). In the Sphacelia state of C. purpurea conidiophores line chambers in the stroma, but in the Sphacelia states of Epichloë typhina and A. hypoxylon they are borne on the surface of a smooth stroma.
Hypomyces papulasporae (synanamorphs = Papulaspora sp., Sibirina sp.), H. papulasporae var. americanus (synanamorphs = Papulaspora candida, Sibirina clavisedum), H. mycogones (anamorph = Mycogone-like) on Geoglossum spp. and Trichoglossum spp.; Hypomyces stephanomatis (synanamorphs = Stephanoma strigosum, Sibirina sp.) on the operculate discomycete Humaria hemisphaerica; Hypomyces leotiicola (synanamorphs = Sepedonium leotiarum, Sibirina sp.) on Leotia lubrica, and Nectria discophila (anamorph = ?Acremonium sp.) on Lachnum spp. are proposed. Nectria albidopilosa and N. discicola, occurring on unidentified, inoperculate Discomycetes, are also described. Hypomyces sepulcralis on soil and possibly on Peziza sp., and Nectria sepultariae, on Sepultaria arenicola, are redescribed. Stephanoma tetracoccum and its Sibirina-like synanamorph, both found on members of the Geoglossaceae, are illustrated and described briefly.
A brief review of the taxonomic history of the order Hypocreales (Fungi, Ascomycetes) is presented. A modified characterization of the Hypocreales with one family, Hypocreaceae, and the acceptance and characterization of the Clavicipitales with one family, Clavicipitaceae, are given together with a key to the orders of the Ascomycetes subclass Ascomycetidae, and keys to the genera of the Hypocreales and the Clavicipitales. A total of 104 genera are tentatively maintained in the Hypocreales; 33 additional genera are indicated in synonymy. A total of 23 genera are included in the Clavicipitales, an additional 8 are indicated in synonymy.
Ten species of Hypomyces (Ascomycotina, Hypocreales, Hypocreaceae) occur on members of the Boletaceae (Basidiomycotina, Agaricales). Five new species, viz. H. badius, H. boletiphagus, H. chlorinigenus, H. melanochlorus, and H. microspermus are described. One new combination, H. completes, is made. All species have a proven (five species) or associated (five species) anamorph referable to Sepedonium. The five species grown in culture from isolated ascospores produced hyphomycetous synanamorphs, and synanamorphs have been detected in collections of the other five species.
Thirteen species of Hypomyces occur on gilled fungi. Most are found on members of the Russulaceae; other hosts include Amanita spp., Crepidotus spp., Leptonia strigosissima, and Pholiota sp. Anamorphs have been proven only for the four species, H. armeniacus (Cladobotryum verticillatum), H. odoratus (C. mycophilum), H. succineus (Verticillium succineum comb, nov.), and H. tremellicola (Verticillium sp.). Anamorphs have been putatively linked to H. lateritius (C. tulasnei), H. lithuanicus (C. arnoldii), and H. petchii (Verticillium sp.).
Cladistic analysis of partial sequences (840 nucleotide positions) from the nuclear encoded small subunit ribosomal DNA was performed to infer the higher taxonomic placement of the Clavicipitales within unitunicate perithecial ascomycetes. Two major classifications exist concerning the placement of this order of ascomycetes; one places it as a sister group to the Hypocreales and the other, as a member of or a near relative to the Xylariales. A strict consensus of 10 equally most parsimonious trees was in agreement with the placement of the Clavicipitales as a monophyletic sister group to the Hypocreales; relationships within the Clavicipitales were not fully resolved in the strict consensus. The taxa sampled from the Hypocreales comprised a paraphyletic lineage in the strict consensus. Characters derived from anamorphs, stromata, centrum anatomy and nutritional modes were reviewed with respect to their level of congruence with the results inferred from the molecular data.
The ascogonium of Glomerella cingulata forms as a loop or coil from an assimilative hypha and becomes septate, each cell being uninucleate. Branches from the parent hypha envelop the ascogonium, become septate, branch further and produce the nonascogenous portions of the centrum as well as the ascomal wall. The ascogonium wraps around itself in various ways, elongates, and spreads into a flattened layer above the wall at the base of the young ascoma. Ascogenous hyphae, croziers, and asci arise from the ascogonium. Paraphyses form as vertical rows of expanded cells among the ascogenous hyphae. This tissue appears as an area of pseudoparenchyma in sectional view but the rows of cells are separable and, therefore, are equivalent to more typically filamentous paraphyses. The paraphyses are digested as the asci develop and are no longer evident when asci are mature. A small cavity forms just beyond the tips of the paraphyses when the neck and neck canal begin formation. The neck canal is lined with periphyses. Development of the centrum of G. cingulata is shown to be sordariaceous and especially similar to that in Triangularia backusii and Plectosphaerella cucumerina.
Development of the ascoma of Plectosphaerella cucumerina begins with the formation of a coiled ascogonium that becomes surrounded by branches from the parent hypha or from neighboring hyphae or both. Paraphyses develop as rows of cells with the lower ones laterally expanded to form an apparent pseudoparenchyma. Apical meristcmatic cells of the ascoma grow upward parallel to each other and form the neck and neck canal. Neck hyphae are septate, may be branched, and remain unpigmented until late in development of the ascoma. A cavity within the ascoma is present only near the entrance to the ostiole until late in development. Centrum development is the Sordaria-type. The ascogonium branches in a horizontal layer among the bases of the paraphyses and produces short ascogenous hyphae which give rise to asci by crozier formation. Meiosis followed by a mitotic division gives rise to eight nuclei, each of which becomes enclosed within an ascospore. Mitosis and septum formation produce bicellular ascospores with two uninucleate cells. The apical apparatus of the ascus is inamyloid and annellascous but is often obscure. The haploid chromosome number is four.