Unitunicate Asci Research Articles

Portuguese marine fungi and the contribution of differential interference contrast (DIC) microscopy for their morphological identification

Marine fungi occur either in Open Ocean or in the intertidal zone of sandy beaches, salt marshes and mangroves, where their hosts and substrates are found. The development of morphological adaptations like appendages and sheaths of the spores are vital to the settlement and attachment to substrate surfaces, floatation and dispersion on seawater. The morphological features of these appendages and sheaths of spores also have an important role in the identification of marine fungi. Differential interference contrast (DIC) microscopy is an essential tool for the observation of these mucilaginous structures in marine fungi spores and was therefore applied to marine mycota from surveys along the Portuguese coast.Since 1991 Portuguese marine fungi have been studied and characterized based on morphological identifications. The studied environments included salt marshes, sandy beaches and marinas. On these environments different substrates were collected such as plants and baits of Spartina maritima, different categories of drift substrates and Fagus sylvatica and Pinus pinaster baits. These substrates, which had been exposed to different conditions of permanent and temporary submersion, were subjected to an initial examination under the stereoscope microscope in order to detect fruit bodies and spores of marine fungi. These structures were then observed in order to achieve the taxonomic identifications of these fungi, based on dichotomous keys for marine fungi.To observe and characterize the wide variety of appendages and mucilaginous sheaths of the ascospores, basidiospores and conidia often invisible in the bright field of the light microscope, the use of DIC microscopy was implemented, because three-dimensional images are produced, highlighting them. The identified fungi were microphotographed with a Leica Wild MPS 52 with Fujichrome RTP- 135, 64T Tungsten.The studies carried out with substrates subjected to conditions of permanent submersion highlighted the dominance of Ascomycota with unitunicate asci. The unitunicate asci are thin - walled, persistent or early deliquescing, favoring ascospores release on marine environmental conditions (fig 1a, 1b and 1c). On the other hand, Ascomycota with bitunicate asci, were mainly detected on temporary submersion conditions. These fungi presented asci with active spore discharge and mucilaginous sheaths that contributed to substrate attachment (fig. 2a and 2b). Ascospores and conidia presented morphological diversity on their appendages, which has particular importance on their success in marine environment (figs. 3, 4, 5, and 6).Marine Mycologists are in agreement that the best technique to observe the appendages and sheaths of spores, often invisible in the bright-field microscope is the use of differential interference or phase contrast microscopy. DIC microscopy was then applied to the observation of Portuguese marine fungi enabling to thoroughly characterize the structures essential to their morphological identifications.

Molecular systematics of Woswasia atropurpurea gen. et sp. nov. (Sordariomycetidae), a fungicolous ascomycete with globose ascospores and holoblastic conidiogenesis

The monotypic genus Woswasia is described for wood-inhabiting and mycotrophic fungi classified in the Sordariomycetidae. It is characterized by unicellular, hyaline, globose to ellipsoid, verruculose ascospores; unitunicate long-stipitate asci with an apical annulus staining blue in aqueous cotton blue and perithecia with a long neck immersed in a stroma exhibiting a conspicuous pH-dependent color reaction. In vitro, it produces branched subhyaline to hyaline conidiophores with terminally arranged sympodial conidiogenous cells and holoblastic hyaline conidia. The remarkable morphological similarity of Woswasia to Amplistroma of the Amplistromataceae, although suggestive of a close relationship, was not confirmed by molecular data. Phylogenetic analysis based on two functional ribosomal genes (large and small subunits of the nuclear rDNA) and one protein-coding gene (second largest subunit of RNA polymerase II) supports the placement of Woswasia in the Sordariomycetidae incertae sedis. Woswasia atropurpurea, the type and only species of the genus, groups within a large heterogeneous clade containing other small or monotypic genera of wood-inhabiting saprobic fungi, which are distantly related and of which the majority lacks an ordinal or familial affiliation. Within the clade a relationship of Woswasia to the freshwater genus Cyanoannulus is suggested.

Phylogenetic classification of Pleurothecium and Pleurotheciella gen. nov. and its dactylaria-like anamorph (Sordariomycetes) based on nuclear ribosomal and protein-coding genes

Two strains of an unidentified perithecial ascomycete with a dactylaria-like anamorph and another morphologically similar strain of a dactylaria-like fungus were collected on decaying wood submerged in freshwater. To study their phylogenetic relationships we (i) combined sequence data from the nuclear small and large subunits ribosomal DNA (nc18S and nc28S) and the second largest subunit of RNA polymerase II (RPB2) for a multigene phylogenetic analysis and (ii) used sequences of the internal transcribed spacer region (ITS) of the rRNA operon for a species-level analysis. The new genus Pleurotheciella is described for two new species, Pla. rivularia and Pla. centenaria, with nonstromatic perithecia, unitunicate asci, persistent paraphyses and hyaline, septate ascospores and dactylaria-like anamorphs characterized by holoblastic, denticulate conidiogenesis, subhyaline conidiophores and hyaline, septate conidia. Based on morphological and molecular data, Pleurotheciella is closely related to the genera Pleurothecium and Sterigmatobotrys. A key to the three genera and the known species is provided. In the three-gene inferred phylogeny, these genera grouped as a sister clade to the Savoryellales within a robust clade of uncertain higher rank affiliation. Phylogenetic study of the 12 strains that represent Pleurothecium recurvatum revealed four that grouped apart from the core of the species. Two of these strains, which form a monophyletic well supported clade in both phylogenies and share similar morphological characteristics, are described as a new species, Pleurothecium semifecundum.

Conlarium duplumascospora gen. et. sp. nov. and Jobellisia guangdongensis sp. nov. from freshwater habitats in China

Conlarium duplumascospora gen. et. sp. nov. and Jobellisia guangdongensis sp. nov. are described and illustrated from submerged wood collected from Guangdong Province, China. Conlarium duplumascospora is characterized by gregarious, coriaceous and beaked ascomata; cylindrical, unitunicate asci with a bipartite apical ring; biseriate, fusiform, hyaline, 0–5-septate ascospores with or without appendages; and anamorph with muriform conidia. Jobellisia guangdongensis is characterized by globose to subglobose, gregarious and papillate ascomata; three-layered peridium; cylindrical, unitunicate asci with a refractive apical ring; and one-septate, fusiform, greenish brown ascospores. Sequence analyses of partial nuclear large subunit ribosomal DNA (LSU rDNA) were performed to infer the phylogenetic affinities of these new taxa. A key to species of Jobellisia is provided.

Three new genera representing novel lineages of Sordariomycetidae (Sordariomycetes, Ascomycota) from tropical freshwater habitats in Costa Rica

Three new genera are established in the Sordariomycetidae based on morphological and molecular data (SSU and LSU nrDNA) to accommodate five ascomycete species collected from submerged woody debris in freshwater habitats from Costa Rica. The genus Bullimyces contains three new species, B. communis, B. costaricensis and B. aurisporus. Bullimyces is characterized by globose to subglobose, membranous, black, ostiolate ascomata; deliquescent, hyaline, globose cells that fill the center of the centrum; unitunicate asci that deliquesce early in some species; and septate, thick-walled ascospores with or without gelatinous sheaths or appendages. Bullimyces species form a well supported clade with 100% bootstrap support, but the position of the genus in the Sordariomycetidae remains unclear. The second genus, Riomyces, is represented by a single species, R. rotundus. Riomyces is characterized by globose to subglobose, membranous, black, ostiolate ascomata, unitunicate, cylindrical asci, hyaline, globose cells that fill the hamathecium and septate, thick-walled ascospores with a gelatinous sheath. Although Riomyces is morphologically similar to Bullimyces, the two genera did not group together with support in any analysis. The third genus, Hydromelitis, is represented by a single species, H. pulchella. Hydromelitis is characterized by pyriform, membranous, black, ostiolate ascomata, unitunicate asci lacking an apical structure, simple, thin-walled, septate paraphyses and hyaline to golden yellow, multiseptate, thick-walled ascospores with a gelatinous sheath. Bullimyces, Riomyces and Hydromelitis were nested within an unsupported clade consisting of members of the Ophiostomatales, Magnaporthales and freshwater Annulatacaceae sensu lato and sensu stricto.

Discovery of the teleomorph of the hyphomycete, Sterigmatobotrys macrocarpa, and epitypification of the genus to holomorphic status.

Sterigmatobotrys macrocarpa is a conspicuous, lignicolous, dematiaceous hyphomycete with macronematous, penicillate conidiophores with branches or metulae arising from the apex of the stipe, terminating with cylindrical, elongated conidiogenous cells producing conidia in a holoblastic manner. The discovery of its teleomorph is documented here based on perithecial ascomata associated with fertile conidiophores of S. macrocarpa on a specimen collected in the Czech Republic; an identical anamorph developed from ascospores isolated in axenic culture. The teleomorph is morphologically similar to species of the genera Carpoligna and Chaetosphaeria, especially in its nonstromatic perithecia, hyaline, cylindrical to fusiform ascospores, unitunicate asci with a distinct apical annulus, and tapering paraphyses. Identical perithecia were later observed on a herbarium specimen of S. macrocarpa originating in New Zealand. Sterigmatobotrys includes two species, S. macrocarpa, a taxonomic synonym of the type species, S. elata, and S. uniseptata. Because no teleomorph was described in the protologue of Sterigmatobotrys, we apply Article 59.7 of the International Code of Botanical Nomenclature. We epitypify (teleotypify) both Sterigmatobotrys elata and S. macrocarpa to give the genus holomorphic status, and the name S. macrocarpa is adopted for the holomorph. To evaluate the ordinal and familial affinities of Sterigmatobotrys and its relationships with the morphologically similar genera Carpoligna and Chaetosphaeria, phylogenetic relationships were inferred based on aligned sequences of the large subunit nuclear ribosomal DNA (ncLSU rDNA).

Studies onMicrothyriaceae: Placement ofActinomyxa,Asteritea,Cirsosina,PolystomellinaandStegothyrium

Abstract Actinomyxa, Asteritea, Cirsosina, Polystomellina and Stegothyrium are genera presently included in Microthyriaceae. We have examined the generic types and consider that their characters differ from those of Microthyriaceae. In Actinomyxa ascoma dissolve as a slimy mass in water and ostioles are star-like, thus the genus would be better placed in Stictidaceae. Asteritea has flattened ascomata with a star-like opening and superficial mycelium with hyphopodia and is better placed in Asterinaceae. Cirsosina has superficial hyphae without hyphopodia and elongate ascomata opening by a longitudinal split and is better placed in Aulographaceae. In Polystomellina ascomata also open by a longitudinal slit and have superficial hyphae and is better placed in Asterinaceae. Stegothyrium is a very unusual genus as it has thick-walled asci that appear to be bitunicate, but there is a J+ apical ring which is indicative of unitunicate asci. We suggest that Stegothyrium is excluded from Microthyriaceae and placed i...

Verrucostoma, a new genus in the Bionectriaceae from the Bonin Islands, Japan

Verrucostoma freycinetiae gen. et sp. nov. is described and illustrated from specimens on dead leaves of Freycinetia boninensis (Pandanaceae) collected in Hahajima, Bonin (Ogasawara) Islands, Japan. The genus is characterized by pale orange perithecia with protuberances around the perithecial apex, no color change in 3% potassium hydroxide and lactic acid, unitunicate asci, spinulose ascospores and an Acremonium-like anamorph. Morphological characters were compared with other genera in the Bionectriaceae and Nectriaceae (Hypocreales). Verrucostoma is morphologically similar to Bionectria (Bionectriaceae) from which it differs in the formation of conspicuous protuberances around the perithecial apex and the Acremonium-like anamorph. Moreover molecular analyses of Verrucostoma and other members of the Bionectriaceae and Nectriaceae based on α-actin, large subunit nuclear ribosomal DNA and RNA polymerase II subunit 1 sequences support the conclusions based on morphological data. Our results confirm that V. freycinetiae is distinct from other genera among the Nectria-like fungi and represents a new genus belonging to the Bionectriaceae.

Sedecimiella taiwanensis gen. et sp. nov., a marine mangrove fungus in the Hypocreales (Hypocreomycetidae, Ascomycota)

In an ongoing survey of marine mangrove fungi in Taiwan, a fungus with light-coloured ascomata, resembling members of the Hypocreales, was found on twigs of Kandelia obovata. Based on morphological characteristics and partial nuclear small and large subunit ribosomal DNA data, the fungus is described as a new genus and species, Sedecimiella taiwanensis, and placed in the Hypocreales (Hypocreomycetidae, Ascomycota). Phylogenetic analyses based on LSU/SSU sequences did not resolve the familial placement of S. tawianensis within the Hypcreales, but it may be related to the Niessliaceae. The characteristic features of S. tawianensis are orange to dark-brown colour, soft-texture and perithecial ascomata with the inner wall layer of the neck extending into the upper part of the centrum; cylindrical, 16-spored, unitunicate asci without an apical pore; and hyaline, smooth-walled and globose ascospores.

New fungal genera, Tectonidula gen. nov. for Calosphaeria-like fungi with holoblastic-denticulate conidiogenesis and Natantiella gen. nov. for three species segregated from Ceratostomella

Two morphologically similar groups of ascomycetes with globose to subglobose perithecia, elongate necks, unitunicate asci floating freely at maturity, and hyaline ascospores currently placed in Calosphaeria s. lat. and Ceratostomella s. lat., respectively, are studied. The Calosphaeria-like fungi have groups of perithecia growing between cortex and wood, arranged in circular groups with converging necks and piercing the cortex in a common point; the asci with a shallow apical ring and U- to horseshoe-shaped hyaline ascospores are compared with Calosphaeria pulchella, the type species of the genus. Conidiogenesis of the investigated Calosphaeria-like fungi is holoblastic-denticulate; ramichloridium-like and sporothrix-like conidiophores and conidia were formed in vitro. Ascospore and ascus morphology, structure of the ascal apex, ascogenous system, mode of conidiogenesis and the large subunit rRNA sequences of this group differ considerably from C. pulchella and both groups are unrelated. Thus a new genus, Tectonidula, is described with two accepted species, T. hippocrepida and T. fagi; they are separated by ascospore and ascus morphology and holoblastic-denticulate conidiogenesis from the core species of Calosphaeria. The placement of Tectonidula among perithecial ascomycetes is discussed. The relationship of Tectonidula with Barbatosphaeria and two ramichloridium-like hyphomycetous genera Rhodoveronaea and Myrmecridium is investigated. Three species formerly attributed to Ceratostomella are studied. The revision of the herbarium type specimen and fresh material of Ceratostomella ligneola revealed that it is conspecific with Ceratostomella ampullasca and Ceratostomella similis. The LSU phylogeny clearly separated C. ligneola from Ceratostomella s. str. and morphologically similar Lentomitella. On the basis of molecular sequence data and detailed comparison of morphology of asci, ascospores and ascogenous system the genus Natantiella is described for C. ligneola with C. ampullasca and C. similis as its synonyms. Natantiella produced sterile mycelium in vitro.

Bellojisia, a new sordariaceous genus for Jobellisia rhynchostoma and a description of Jobellisiaceae fam. nov.

The phylogenetic analyses of partial nucLSU rDNA sequence data of three Jobellisia species indicate that J. rhynchostoma is distinct from the core species of Jobellisia. Jobellisia luteola, the type species of the genus, and J. fraterna reside as a strongly supported monophyletic clade in a basal position in a grouping containing the Diaporthales, the Calosphaeriales and the Togniniaceae, while all phylogenies confirm the placement of J. rhynchostoma within the Sordariales. The new family Jobellisiaceae (incertae sedis) is described for Jobellisia. A new perithecial ascomycete genus, Bellojisia (Lasiosphaeriaceae, Sordariales), is introduced for J. rhynchostoma. The fungus produces nonstromatic, long-necked perithecia with a superficial to semi-immersed pyriform venter and carbonaceous three-layered perithecial wall, 1-septate, hyaline, later brown, reniform to navicular ascospores with a polar germ pore formed in unitunicate asci. The fungus was not observed to produce a conidial anamorph in vitro. Both morphological and molecular data suggest Corylomyces selenosporus of the Sordariales is the closest relative of J. rhynchostoma. The other relatives of Bellojisia (viz. Cercophora, Lasiosphaeria and Podospora) recruit from the Lasiosphaeriaceae (Sordariales). Cercophora and Podospora are shown as polyphyletic within the Sordariales, which is in agreement with previous molecular studies.

Two new ascomycetes from rainforest litter in Costa Rica

Two new ascomycetes, Boerlagiomyces costaricensis (Pleosporales) and Scopinella musciformis (Sordariales sensu lato), from litter samples collected in rainforests of Costa Rica, are described and illustrated. Boerlagiomyces costaricensis has globose, ostiolate ascomata covered by numerous setae-like hairs; cylindrical, fissitunicate asci without apical structures; and large, fusiform, muriform, hyaline to pale brown ascospores. Scopinella musciformis is characterized by ostiolate ascomata with a few compact clusters of hypha-like hairs distributed on the peridial surface and a long neck; ovate to ellipsoidal unitunicate asci; and small quadrangular ascospores with diagonal germ slits.

Two new ascomycetes from rainforest litter in Costa Rica

Two new ascomycetes, Boerlagiomyces costaricensis (Pleosporales) and Scopinella musciformis (Sordariales sensu lato), from litter samples collected in rainforests of Costa Rica, are described and illustrated. Boerlagiomyces costaricensis has globose, ostiolate ascomata covered by numerous setae-like hairs; cylindrical, fissitunicate asci without apical structures; and large, fusiform, muriform, hyaline to pale brown ascospores. Scopinella musciformis is characterized by ostiolate ascomata with a few compact clusters of hypha-like hairs distributed on the peridial surface and a long neck; ovate to ellipsoidal unitunicate asci; and small quadrangular ascospores with diagonal germ slits.

Dyrithiopsis lakefuxianensis gen. et sp. nov. from Fuxian Lake, Yunnan, China, and notes on the taxonomic confusion surrounding Dyrithium

A new taxon with Dyrithium-like characteristics was collected from Lake Fuxian in China. The taxon is typical of the Amphisphaeriaceae in that it has relatively large, ostiolate, immersed ascomata, unitunicate asci with a J+ subapical ring, and brown ascospores. It is similar to Dyrithium in that it has muriform ascospores, but considerable confusion surrounds this genus. In Dyrithium asci are bitunicate and lack a J+ subapical ring, while this was not true of our species. A new genus, Dyrithiopsis, therefore is established to accommodate this new taxon. Details of its anamorph also are provided, based on cultural studies. Parsimony analyses of part of the large-subunit rDNA provide further evidence to support the familial placement of this new genus in the Amphisphaeriaceae. The taxonomic position of Dyrithium also is discussed.

Dyrithiopsis lakefuxianensis gen. et sp. nov. from Fuxian Lake, Yunnan, China, and Notes on the Taxonomic Confusion Surrounding Dyrithium

A new taxon with Dyrithium-like characteristics was collected from Lake Fuxian in China. The taxon is typical of the Amphisphaeriaceae in that it has relatively large, ostiolate, immersed ascomata, unitunicate asci with a J+ subapical ring, and brown ascospores. It is similar to Dyrithium in that it has muriform ascospores, but considerable confusion surrounds this genus. In Dyrithium asci are bitunicate and lack a J+ subapical ring, while this was not true of our species. A new genus, Dyrithiopsis, therefore is established to accommodate this new taxon. Details of its anamorph also are provided, based on cultural studies. Parsimony analyses of part of the large-subunit rDNA provide further evidence to support the familial placement of this new genus in the Amphisphaeriaceae. The taxonomic position of Dyrithium also is discussed.

First report of powdery mildew on mulberry caused by Phyllactinia guttata in the eastern Mediterranean region of Turkey

Mulberry (Morus spp.) is an increasingly importantcrop that is widely planted in the Hatay province,Turkey as a food source for silkworms. Production ofsilk products is the main source of income in thisprovince. Serious damage caused by powdery mildewwas discovered in the main mulberry production areasof Samandag and Harbiye regions in 2000. A diseaseincidence of 81–95% was recorded in affected orch-ards. Symptoms included white superficial myceliumwith abundant sporulation on lower leaf surfaces, andchlorotic spots on upper leaf surfaces. Mycelial growthwas observed on leaves, petioles and pedicels, but noton fruit and branches. Symptoms were apparent on eachplant, with disease severity ranging from a fewindividual spots to numerous lesions almost coveringwhole leaves.The white mycelium had lobed appressoria. Conidio-phores were straight and cylindrical. Single-celled, club-shaped conidia occurred usually singly on unbranchedconidiophores. Conidia ranged in length from 55 to85 mm (mean 68·25 ^2·88 mm), and in width from 15to 20 mm (mean 16·65 ^0·7 mm). Cleistothecia withtapering appendages and a bulbous base were yellowwhen young and turned black at maturity. Eachcleistothecium contained nine to 13 unitunicate asci,broadly oval to ellipsoid, short-stalked, and containingtwo ascospores. The size of cleistothecia ranged from225 to 255 mm in diameter (mean 235 ^3·27 mm).Asci were 68–95 mm long (mean 78·5 ^2·1 mm) and35–40 mm wide (mean 37·5 ^0·9 mm). Ascosporeswere 35–45 mm long (mean 40 ^1·29 mm) and 12·5–17·5 mm wide (mean 16 ^0·7 mm). The length ofappendages on cleistothecia ranged from 180 to285 mm (mean 242·6 ^6·7 mm).Pathogenicity tests were conducted on 1-year-old,healthy mulberry seedlings. These plants were inocu-lated by shaking conidia from powdery mildew-infectedleaves onto the leaves of healthy plants. Plants wereincubated in a growth chamber at 26 ^28C with a 16 hphotoperiod. Inoculated plants developed powderymildew symptoms after 9 days. The symptoms oninoculated plants and the morphological characteristicsof the pathogen were similar to those for naturallyinfected plants.Based on the characteristics of sexual and asexualstates and host specialization, the fungus was identifiedas Phyllactinia guttata (Webster, 1979; Hanlin, 1992).This is the first report of powdery mildew on mulberryin the eastern Mediterranean region of Turkey. Acombination of high inoculum pressure, low humidityand an unusually hot, dry summer during 2000favoured fungal infection and disease development.The disease is important as severe foliar lesions renderplants unacceptable for silkworm production.

Annulatascus fusiformis sp. nov., a new freshwater ascomycete from the Philippines

Annulatascus fusiformis sp. nov. is described based on a specimen collected on submerged wood from a river in the Philippines. The species possesses globose to subglobose, black, immersed ascomata with long hyaline necks; wide, septate, tapering paraphyses, and cylindrical unitunicate asci with a relatively massive refractive apical ring. In this respect it is similar to species of Annulatascus, however, A. fusiformis differs from other species in the genus, as ascospores are provided with bipolar mucilaginous pad-like appendages. The species is illustrated with light, scanning and electron micrographs.

Annulatascus fusiformis sp. nov., a New Freshwater Ascomycete from the Philippines

Annulatascus fusiformis sp. nov. is described based on a specimen collected on submerged wood from a river in the Philippines. The species possesses globose to subglobose, black, immersed ascomata with long hyaline necks; wide, septate, tapering paraphyses, and cylindrical unitunicate asci with a relatively massive refractive apical ring. In this respect it is similar to species of Annulatascus, however, A. fusiformis differs from other species in the genus, as ascospores are provided with bipolar mucilaginous pad-like appendages. The species is illustrated with light, scanning and electron micrographs.

Studies on Amphisphaeriales: The Cainiaceae

The Cainiaceae is revived to accommodate Atrotorquata, Arecophila, Cainia, Ceriophora and Reticulosphaeria , all of which are described and illustrated. Ommatomyces is also included in the family and discussed. The essential characteristics of the family are ascomata which are usually immersed under a clypeus, unitunicate asci with a complex, J+, apical apparatus comprising a series of rings, and brown ascospores which may have longitudinal germ slits, striations, reticulate ornamentations and/or germ pores.

Cannonia gen. nov., from palms in the Southern Hemisphere

Cannonia gen. nov. is introduced to accommodate three collections of an unusual taxon, originally described as Ceratostoma australe. Cannonia is thought to be xylariaceous, due to its globose immersed ascomata with the stroma reduced to a clypeus, well-developed paraphyses, unitunicate asci lacking an apical apparatus, and dark brown, aseptate ascospores with a germ slit. Cannonia is compared with other possibly related genera and illustrated with interference contrast micrographs.