Fungi from the roots of the common terrestrial orchid Gymnadenia conopsea

The fungal community associated with the terrestrial photosynthetic orchid Gymnadenia conopsea was characterized through PCR-amplification directly from root extracted DNA and cloning of the PCR products. Six populations in two geographically distinct regions in Germany were investigated. New ITS-primers amplifying a wide taxonomic range including Basidiomycetes and Ascomycetes revealed a high taxonomic and ecological diversity of fungal associates, including typical orchid mycorrhizas of the Tulasnellaceae and Ceratobasidiaceae as well as several ectomycorrhizal taxa of the Pezizales. The wide spectrum of potential mycorrhizal partners may contribute to this orchid's ability to colonize different habitat types with their characteristic microbial communities. The fungal community of G. conopsea showed a clear spatial structure. With 43 % shared taxa the species composition of the two regions showed only little overlap. Regardless of regions, populations were highly variable concerning taxon richness, varying between 5 and 14 taxa per population. The spatial structure and the continuous presence of mycorrhizal taxa on the one hand and the low specificity towards certain fungal taxa on the other hand suggest that the fungal community associated with G. conopsea is determined by multiple factors. In this context, germination as well as pronounced morphological and genetic differentiation within G. conopsea deserve attention as potential factors affecting the composition of the fungal community.     

Deoxyribonucleic acid base composition and taxonomy in the genus Geotrichum Link

The molar percentage of guanine + cytosine (% GC) in the DNA from 25 yeast-like fungi related to the genus Geotrichum has been measured. This criterion together with the biochemical characteristics allow the division of the species into 3 groups: one group resembling G. candidum (% GC: 31.5–42) and containing 8 imperfect and 6 perfect forms. In this group only G. capitatum has a lower % GC (31.5). A second group of 6 species resembling the genus Trichosporon (57–60%) and a third intermediate category of 4 species (42–53.5%) were observed. The GC-values correlate well with biochemical characters, e.g. the presence of urease, the number of organic substrates assimilated, and also with the presence of characters typical of the Ascomycetes or the Basidiomycetes.This work was supported by I.N.S.E.R.M. (Institut National de la Santé et de la Recherche Médicale) C.R.L. n^o 76 1 153 8.     

Comparative phylogenies and host specialization in the alder ectomycorrhizal fungi Alnicola, Alpova and Lactarius (Basidiomycota) in Europe

Background  

Mycorrhizal fungi form intimate associations with their host plants that constitute their carbon resource and habitat. Alnus spp. (Betulaceae) are known to host an exceptional species-poor and specialized ectomycorrhizal (ECM) fungal community compared to other tree species, but the host-specificity pattern and its significance in terms of fungal diversification and speciation remain poorly documented. The degree of parallel speciation, host switching, and patterns of biogeography were explored in the historical associations between alders and three ECM taxa of Basidiomycetes: Alnicola (Agaricales), Alpova (Boletales), and Lactarius (Russulales). The aim was to develop an evolutionary framework on host specificity and diversification of Basidiomycetes in this highly specialized plant-fungus symbiosis.     

Relatedness Among Agronomically Important Rusts Based on Mitochondrial Cytochrome b Gene and Ribosomal ITS Sequences

A fragment of the mitochondrial cytochrome b (cyt b) gene of 13 agronomically important plant pathogenic Basidiomycetes was sequenced, including several Puccinia spp., Uromyces appendiculatus, Phakopsora pachyrhizi, Hemileia vastatrix and Rhizoctonia solani. The deduced amino acid sequences (residues 142–266) were used to study the relatedness of these pathogens as compared to other species of the Basidiomycetes, Ascomycetes and Oomycetes. The relatedness was also studied at nuclear level using the Internal Transcribed Spacers (ITS) in the ribosomal DNA. Phylogenic trees were constructed with the maximum parsimony (MP) and the neighbour‐joining (NJ) methods. On the basis of both cytochrome b and ITS sequences, the Puccinia species pathogenic to graminaceous crop plants, such as Puccinia recondita f. sp. tritici, Puccinia graminis f. sp. tritici, Puccinia striiformis f. sp. tritici, Puccinia coronata f. sp. avenae, Puccinia hordei, P. recondita f. sp. secalis and Puccinia sorghi, together with Puccinia horiana from Chrysanthemum, were very closely related to each other, whereas Puccinia arachidis (from peanut) was closely related to U. appendiculatus (from beans) but more distant from the other Puccinia species. Both rusts on soybean (P. pachyrhizi) and coffee (H. vastatrix) were outside the Puccinia cluster. All rusts were separated from other Basidiomycetes such as R. solani and the strobilurin producing species Strobilurus tenacellus and Mycena galopoda. Our results demonstrate that the amino acid sequence of the mitochondrial cytochrome b is a valid tool to study phylogenic relatedness among plant pathogenic Basidiomycetes and supports taxonomic grouping based on morphological structures and host specificity. Because of their high variability, ITS sequences were able to discriminate Puccinia species, which were identical on the basis of the cytochrome b amino acid sequence. Thus, ITS sequences could better show differences among species or within a species, whereas cytochrome b is more suitable than ITS for phyologenic inference at family or genus level. In addition, the sequence data obtained during this study represent essential information for easy isolation of the cyt b gene and detection of point mutations conferring resistance to Qo Inhibitor fungicides that eventually may evolve.     

The potential of Pseudozyma yeastlike epiphytes for the production of heterologous recombinant proteins

Although Basidiomycetes represent the most evolved class of fungi, they have been neglected with regard to recombinant gene expression. In this work, basidiomycetous yeasts belonging to Pseudozyma spp. were studied with respect to their amenability to heterologous protein production. Single plasmid or cotransformation experiments routinely afforded 100 to 200 independent transformants for the two tested species of Pseudozyma. Green fluorescent protein (GFP) was expressed in the correctly folded conformation, as demonstrated by fluorescence microscopy, and hen egg white lysozyme (HEWL) was expressed in its active form, as revealed by its lytic activity on Micrococcus lysodeikticus cells. Protease analysis established that Pseudozyma spp. contained equivalent or less extracellular protease activity than yeasts and far less protease activity than ascomycetous filamentous fungi in similar culture conditions. This proteolytic activity was inhibited by over 97% with a combination of PMSF and Pepstatin A. N-glycosylation patterns of native Pseudozyma flocculosa secreted proteins were comprised of one or a few short glycan chains that possess a classic eukaryotic structure typical of higher fungi and animal cells. This is the first report of a Basidiomycete that possesses multiple intrinsic characteristics necessary for use as a heterologous gene expression system.     

The Colacosomes: New Structures at the Host-parasite Interface of a Mycoparasitic Basidiomycete*

The ultrastructure of mycoparasitic interactions in heterobasidiomycetes is diverse. A previously unknown cell structure, involved in a specific parasitic interaction, is described and illustrated. Cells of the parasite Platygloea peniophorae (Auriculariales s.l., Basidiomycetes) which attach to host cells of Hyphoderma praetermissum (Aphyllophorales, Basidiomycetes) form distinct marginal vesicular bodies with electron-opaque cores and electron transparent sheaths. Finally, the vesicular content projects through the cell wall of the parasite and then interacts with the cell wall of the host. Ontogenetic stages are described and illustrated in detail.     

THE SEPTAL PORES OF COPFINUS LAGOPUS IN RELATION TO NUCLEAR MIGRATION

The septal pores of homokaryotic and dikaryotic mycelia of Coprinus lagopus have the complex structure characteristic of certain Basidiomycetes. The edge of the pore is thickened and covered on either side by a dome-shaped, perforated, membranous cap. However, in heterokaryotic mycelia in which nuclear migration was believed to be taking place, in addition to the complex pores, simple pores were also observed as well as various intermediate stages interpreted as steps in the breakdown of complex to simple pores. The simple pores may facilitate nuclear migration.     

FINE STRUCTURE OF MYCOTA. 7. OBSERVATIONS ON SEPTA OF ASCOMYCETES AND BASIDIOMYCETES

Moore , R. T., and J. H. Mc Alear . (Cornell U., Ithaca, N. Y.) Fine structure of mycota. 7. Observations on septa of Ascomycetes and Basidiomycetes. Amer. Jour. Bot. 49(1): 86–94. Illus. 1962.—Electron microscopy of 13 carpomycete species (5 Ascomycetes, 5 Basidiomycetes, and 3 Deuteromycetes) presents evidence that suggests there is an Ascomycete- and a Basidiomycete-type septum. The observed Ascomycete septa taper slightly or not at all toward a simple, clear-channel, central pore, while the observed Basidiomycete septa flare sharply and broadly at the center to produce what is termed a dolipore septum. On each side of this septum is a double membrane structure that is crescent-shaped in section. This is given the name parenthesome. These dolipore-parenthesome septa were found to be characteristic of basidiocarpic, dikaryotic, hyphae. Interpretative drawings of both types of septa are presented. From these and the micrographs, it is not difficult to see how materials, including nuclei, could pass through the Ascomycete-type septum, but it would appear that the modifications of the Basidiomycete-type septum while maintaining cytoplasmic continuity would prohibit nuclear migration in dikaryotic hyphae. It is postulated that as the plant and animal phylogenetic lines have developed actual diploidism, the terminal group of the fungi may have achieved functional diploidism. Of the 3 species of Deuteromycetes examined, 2, Aspergillus variecolor and Stilbum zacalloxanthum, have typical Ascomycete-type septa; the third, QM 7739, has Basidiomycete-type septa. This fungus is a species of Sclerotium, but the present confusion and lack of definitive information regarding the taxonomy, genetics, and perfect stage of this genus prohibit the enlarging at present on the possible significance of this observation.     

Yeasts in high Arctic glaciers: the discovery of a new habitat for eukaryotic microorganisms

Recently a new habitat for microbial life has been discovered at the base of polythermal glaciers. In ice from these subglacial environments so far only non-photosynthetic bacterial communities were discovered, but no eukaryotic microorganisms. We found high numbers of yeast cells, amounting to a maximum of 4,000 CFU ml⁻¹ of melt ice, in four different high Arctic glaciers. Twenty-two distinct species were isolated, including two new yeast species. Basidiomycetes predominated, among which Cryptococcus liquefaciens was the dominant species (ca. 90% of total). Other frequently occurring species were Cryptococcus albidus, Cryptococcus magnus, Cryptococcus saitoi and Rhodotorula mucilaginosa. The dominant yeast species were psychrotolerant, halotolerant, freeze-thaw resistant, unable to form mycelium, relatively small-sized and able to utilize a wide range of carbon and nitrogen sources. This is the first report on the presence of yeast populations in subglacial ice.     

Fungal community diversity and soil health in intensive potato cropping systems of the east Po valley, northern Italy

An ecological approach was used to investigate the relationship between diversity of soil fungal communities and soil‐borne pathogen inoculum in a potato growing area of northern Italy affected by yield decline. The study was performed in 14 sites with the same tillage management practices: 10 named ‘potato sites’, that for many years had been intensely cultivated with potatoes, and 4 named ‘rotation sites’, subject to a 4‐year rotation without potatoes or any recurrent crop for many years. Fungal communities were recorded using conventional (soil fungi by plate count and endophytic fungi as infection frequency on pot‐grown potato plant roots in soil samples) and molecular approaches [Basidiomycetes and Ascomycetes with specific and denaturing gradient gel electrophoresis (DGGE) analysis]. Diversity of fungal communities in potato sites was significantly lower than that in rotation sites. In addition, fungal communities in rotation sites showed lower Berger–Parker dominance than those in the potato sites, suggesting that rotation sites had a higher diversity as well as a better fungal community balance than potato sites. The ANalysis Of SIMilarity test of soil fungi and root endophytic fungi revealed that the two cropping systems differed significantly for species composition. Root endophytic fungal communities showed a greater ability to colonise potato roots in soil samples from potato sites than those from rotation sites. Moreover, the majority of endophytic root fungal community species in potato sites belonged to the potato root rot complex and storage disease (Colletotrichum coccodes, Fusarium solani and Fusarium oxysporum), while those in rotation sites were mainly ubiquitous or saprobic fungi. Soil rDNA analyses showed that Ascomycetes were much more frequent than Basidiomycetes in all the soils examined. DGGE analysis, with the Ascomycete‐specific primer (ITS1F/ITS4A), did not reveal distinctions between the communities found at the potato and rotation sites, although the same analysis showed differences between the communities of Basidiomycetes (specific primer ITS1F/ITS4B). These findings showed that recurrent potato cropping affected diversity and composition of soil fungal communities and induced a shift in specialisation of the endophytic fungi towards potato.     

Endophytic fungi from Melia azedarach

A collection of endophytic fungi was obtained from living apparently symptomless roots, stems, leaves and fruits of Melia azedarach. This is an exotic tree introduced into Brazil from Asia and is a producer of insecticidal compounds. It grows very well and is apparently resistant to many pathogens. Each segment from the plant was sampled during two periods. A total of 55 fungal isolates were recovered. Hyphomycetes were more prevalent than Ascomycetes, Coelomycetes and Basidiomycetes. The genera Aspergillus and Penicillium were the most common in the plant.     

Anthraquinone production by a fungus causing black heartwood stain in yellow cedar

Summary A yellow crystalline fungal metabolite was obtained from cultures of sterile, darkly pigmented fungi which had been isolated from the heartwood of yellow cedar. This chemical was identified as 1-hydroxy-3-methylanthraquinone and showed negligible toxicity against a range of wood-destroying Basidiomycetes and moulds. It also greatly reduced conidial formation inT. virgatum.     

Literature on Basic Palynology

The occurrence of airborne fungi at an urban coastal site was compared with that at a rural inland site in the South of Sardinia. Spore concentrations at the two sites differed markedly with lower concentrations at the urban site but the spore types found were similar. The general trend of the fungal airspores was also very similar at both sites as a result of the similar climatic conditions.

Mould spore concentrations were greatest in late spring (May-June) and autumn (October-November) with a decrease during summer months at both sites sampled. Cladosporium was the most common spore type identified followed by Basidiomycetes, Alternaria, Fusarium. These taxa are well known as allergens. It is recommended that further aeromycological surveys should be performed in different areas to determine the clinical importance of fungi as allergens in Sardinia.     

<Emphasis Type="Italic">Monotropastrum humile</Emphasis> var. <Emphasis Type="Italic">humile</Emphasis> is associated with diverse ectomycorrhizal Russulaceae fungi in Japanese forests

Monotropastrum humile is nearly lacking in chlorophyll and obtains its nutrients, including carbon sources, from associated mycorrhizal fungi. We analyzed the mycorrhizal fungal affinity and species diversity of M. humile var. humile mycorrhizae to clarify how the plant population survives in Japanese forest ecosystems. We classified 78 samples of adult M. humile var. humile individuals from Hokkaido, Honshu, and Kyusyu Islands into 37 root mycorrhizal morphotypes. Of these, we identified 24 types as Russula or Lactarius fungal taxa in the Russulaceae, Basidiomycetes, but we could not identify the remaining 13 types as to their genus in the Basidiomycetes. The number of fungal species on M. humile var. humile was the highest in the plant subfamily. The diversity of fungal species revealed its increased trends in natural forests at the stand level, fagaceous vegetation, and cool-temperate climate. The most frequently observed fungus colonized mainly samples collected from sub-alpine forests; the second most frequently observed fungus colonized samples collected from sub-alpine to warm-temperate forests. These results suggest that Japanese M. humile populations are associated with specific but diverse fungi that are common ectomycorrhizal symbionts of various forest canopy trees, indicating a tripartite mycorrhizal relationship in the forest ecosystem.     

Necrotrophic mycoparasitism of Botrytis cinerea by cellulolytic and ligninocellulolytic Basidiomycetes

Twenty-six isolates representing 17 species of aphyllophoraceous, wood-decaying Basidiomycetes and five species of agaricoid, turf-borne, thatch-decaying Basidiomycetes were screened for their abilities to degrade cellulose, lignin, and melanin by using colorimetric degradation assays on agar media. Selected ligninocellulolytic Basidiomycetes capable of degrading melanin were screened for antagonism of Botrytis cinerea Per.:Fr. The greatest inhibition of Botrytis colony and hyphal growth in vitro was observed in confrontations with Irpex lacteus (Fr.) Fr., Trametes versicolor (L.:Fr.) Pilat, and Chondrostereum purpureum (Pers.:Fr.) Pouzar. Hyphal interference and necrotrophic mycoparasitism by these ligninocellulolytic Basidiomycetes were recognized microscopically as coagulation and degeneration of Botrytis cytoplasm and as coiling and invasion of hyphae, conidiophores, and conidia, respectively. Sclerotia of B. cinerea were killed and parasitized in agar media, straw mulch, or moist sand infested separately with these three mycoparasites.     

Properties of calcium-dependent regulatory proteins from fungi and yeast

Calmodulins were isolated from vegetative mycelia of Basidiomycetes fungi, Agaricus campestris and Coprinus lagopus. These calmodulins showed similar mobilities to those of animal calmodulins on nondenaturing polyacrylamide gel electrophoresis in the presence or absence of Ca2+. The molecular weights of both calmodulins were determined to be 16,000. Agaricus calmodulin consisted of 148 amino acids including epsilon-N-trimethyllysine and cysteine. The UV-absorption spectrum showed the relatively high content of phenylalanine in Basidiomycetes calmodulins. The difference UV-absorption spectrum due to the blue shift by Ca2+ was observed. Both calmodulins activated muscle myosin light chain kinase and pea NAD+ kinase in a Ca2+-dependent manner, and the activities were inhibited by trifluoperazine or chlorpromazine. A calmodulin-like protein was partially purified from baker's yeast, Saccharomyces cerevisiae. However, detection of a calmodulin-like protein in prokaryotes was not successful.     

Biotransformations of aryl alkyl sulfides by whole cells of white-rot Basidiomycetes

White-rot Basidiomycetes promoted the oxidation of aromatic pro-chiral sulfides into sulfoxides with good enantioselectivity, conversion and a small production of sulfones. The reactions were carried out using whole cells of Irpex lacteus, Pycnoporus sanguineus, Trichaptum byssogenum, Trametes rigida, Trametes versicolor and Trametes villosa. The enantioselectivity for all the aryl alkyl sulfoxides was in favor of (S)-enantiomers. Oxidation of (phenylpropyl)sulfide produced (S)-(phenylpropyl)sulfoxide with high enantiomeric excess (e.e. ≥99%) by all the Basidiomycetes employed. Basidiomycetes isolated from Brazilian biomes were used as biocatalysts in the oxidation reaction.     

Free-Radical Scavenging Activity of Submerged Mycelium Extracts from Higher Basidiomycetes Mushrooms

Twenty-four Basidiomycetes strains were evaluated to determine their free-radical scavenging capacity in submerged cultivation. The scavenging capacity of the extracts varied from 1 to 85% depending on the mushroom species, solvent used, and concentration. A calculation of EC₅₀ of extracts from several wood-rotting Basidiomycetes showed high scavenging abilities at low effective concentration.     

New records of <Emphasis Type="Italic">Ardhachandra</Emphasis>, <Emphasis Type="Italic">Dicyma</Emphasis>, and <Emphasis Type="Italic">Sibirina</Emphasis> species from basidiomata of Aphyllophorales (Basidiomycetes) in Japan

Three mitosporic fungi, i.e., Ardhachandra cristaspora, Dicyma pulvinata, and Sibirina gamsii from basidiomata of Aphyllophorales (Basidiomycetes), are described and illustrated. These fungi have not been previously reported in Japan.     

The β-xylosidase production by yeastCryptococcus podzolicus

In studying the β-xylosidase production by yeastCryptococcus podzolicus (Basidiomycetes), it was shown to be an inducible secretory enzyme. Xylooligosaccharides generated from xylan and methyl β-xyloside were found to induce the β-xylosidase synthesis. The enzyme activity in the medium containing xylan or methyl β-xyloside was 1.0 and 1.5 U/ml, respectively; this production level is similar to that achievable at the β-xylosidase production by mycelial fungi.