Action spectra for fruiting of the mushroom Coprinus congregatus

Spectral sensitivity for photostimulated and photoinhibited phases of the development of Coprinus congregatus Bull. ex Fr. was determined from 405–730 nm. The cultures were exposed during 12 h to a constant number of incident quanta. Spectral sensitivity for photoinduction of primordia formation, for photoinhibition of primordia development and for photomaturation of primordia was quite similar, suggesting a common photoreceptor during fruit-body morphogenesis. A defined action spectrum for the photoinhibitory effect of a light break during the inductive night was determined from 407–690 nm. The most effective wavelengths were in the blue (445 nm). Wavelengths longer than 510 nm were ineffective. The general shape of this action spectrum was similar to those obtained for many blue light responses in which a flavoprotein was postulated to be the photoreceptor.     

Sequential production of enzymes and basidiospore formation in fruiting bodies of Coprinus macrorhizus

The production of NADP-dependent glutamate dehydrogenase was initiated at the stage of first meiotic prophase in pileus cells but not in stipe cells of dikaryotic and monokaryotic fruiting bodies in Coprinus macrorhizus. The production of chitinase and glucanases assayed with laminarin and lichenan was observed after the completion of meiosis only in pileus cells. The light conditions that were effective for the delay or inhibition of cellular events in the pileus cells were also effective for the delay or inhibition of enzyme production. But all sporeless mutants tested, which were defective at the various stages of basidiospore formation, produced the normal levels of these enzymes. The results indicate that the sequential production of enzymes and cellular events leading to basidiospore formation in pileus cells are independent from each other.Abbreviation GDH_NADP NADP-dependent glutamate dehydrogenase     

Reconstructing the evolution of agarics from nuclear gene sequences and basidiospore ultrastructure

Traditional classifications of agaric fungi involve gross morphology of their fruit bodies and meiospore print-colour. However, the phylogeny of these fungi and the evolution of their morphological and ecological traits are poorly understood. Phylogenetic analyses have already demonstrated that characters used in traditional classifications of basidiomycetes may be heavily affected by homoplasy, and that non-gilled taxa evolved within the agarics several times. By integrating molecular phylogenetic analyses including domains D1–D3 and D7–D8 of nucLSU rDNA and domains A–C of the RPB1 gene with morphological and chemical data from representative species of 88 genera, we were able to resolve higher groups of agarics. We found that the species with thick-walled and pigmented basidiospores constitute a derived group, and hypothesize that this specific combination of characters represents an evolutionary advantage by increasing the tolerance of the basidiospores to dehydration and solar radiation and so opened up new ecological niches, e.g. the colonization of dung substrates by enabling basidiospores to survive gut passages through herbivores. Our results confirm the validity of basidiospore morphology as a phylogenetic marker in the agarics.     

Intraspecific hybridization between Coprinus cinereus and Schizophyllum commune by PEG-induced protoplast fusion and electrofusion

Two irreversible inhibitors, iodoacetamide and diethylpyrocarbonate, were used to select intraspecific fusion products of two mushroom species, Coprinus cinereus and Schizophyllum commune. Iodoacetamide was the more suitable inhibitor because it gave a low breakage frequency and low survival rate of the cells in the inactivation experiments. Fusion-induced by polyethylene glycol and electro-fusion were compared and, under optimal conditions, gave fusion frequencies of 16.7% to 50.0% and 6.9% to 8.4%, respectively. All fusion progeny were heterokaryons (dikaryons) and had clamp connections. There were no differences in the morphology and fruiting ability of the fusion progeny and those of the heterokaryons generated from mating.     

Improved fruiting of Coprinus atramentarius using cold-shock treatment during growth

Coprinus atramentarius was grown on two commercial composts at a constant 20°C or with a cold shock (25°C20°C) after 10 days. Cold shock was required for it to form fruiting bodies.The authors are with the University of Western Sydney, Hawkesbury, School of Horticulture, Locked Bag 1, Richmond NSW 2753, Australia     

Microarray and real-time PCR analyses reveal mating type-dependent gene expression in a homothallic fungus

Sordaria macrospora is a homothallic ascomycete which is able to form fertile fruiting bodies without a mating partner. To analyze the molecular basis of homothallism and the role of mating products during fruiting body development, we have deleted the mating type gene Smta-1 encoding a high-mobility group domain (HMG) protein. The ΔSmta-1 deletion strain is morphologically wild type during vegetative growth, but it is unable to produce perithecia or ascospores. To identify genes expressed under control of Smta-1, we performed a cross-species microarray analysis using Neurospora crassa cDNA microarrays hybridized with S. macrospora targets. We identified 107 genes that are more than twofold up- or down-regulated in the mutant. Functional classification revealed that 81 genes have homologues with known or putative functions. Comparison of array data from ΔSmta-1 with those from three phenotypically similar mutants revealed that only a limited set of ten genes is deregulated in all mutants. Remarkably, the ppg2 gene encoding a putative lipopeptide pheromone is 500-fold down-regulated in the ΔSmta-1 mutant while in all other sterile mutants this gene is up-regulated. Electronic Supplementary Material Supplementary material is available for this article at and is accessible for authorized users.     

Identification of a mycorrhizal fungus in Epipogium roseum (Orchidaceae) from morphological characteristics of basidiomata

A mycorrhizal fungus, ME1-1, isolated from an achlorophyllous orchid, Epipogium roseum, was identified as Coprinellus disseminatus (≡ Coprinus disseminatus) based on characteristics of basidiomata that were artificially induced. Spawn of ME1-1 cultivated on a medium that consisted of sawdust and wheat, sandwiched between two mats of volcanic soils, which was incubated at 20.0° ± 0.5°C in 80.0% ± 0.5% relative humidity in the dark. The basidiomata were formed on the soil after 2 months. Morphological and anatomical characteristics of the basidioma mostly accorded with those of Coprinellus disseminatus. We therefore concluded that C. disseminatus is one of the mycobionts of E. roseum.     

Segregation of genetic markers in the two-spored secondarily homothallic basidiomycete Coprinus bilanatus

Summary Secondarily homothallic basidiomycetes, of which the cultivated mushroom Agaricus bisporus is an example, produce both self-fertile and non self-fertile spores. The random migration of nuclei from the basidia to give binucleate spores provides the simplest explanation for the regulation of breeding behaviour in this group of fungi. To test the predictions of the random migration hypothesis, the segregation of mating-type, auxotrophy and antimetabolite resistance has been determined in the secondarily homothallic ink-cap fungus, Coprinus bilanatus. In 41 of a total of 56 spore progenies tested, the segregation ratios conformed to the predictions of the random migration hypothesis. Poor fits to the predicted ratios were, in many instances, associated with an adenine auxotrophy. On the basis of the data reported, random migration can be regarded as the primary control of secondary homothallism.     

Secretional expression of a Bacillus subtilis xylanase gene in the Basidiomycete Coprinus cinereus

The Bacillus subtilis endo (β-1,4-) D-xylanase structural gene (xyn) was trimmed away from its signal sequence and then fused after the signal sequence of the basidiomycete Pleurotus ostreatus manganese(II) peroxidase cDNA. The resulting modified gene (xyn′) was inserted between the promoter and terminator of two chromosome-integrating, heterologous protein expression vectors. These recombinant plasmids were introduced into protoplasts of the monokaryotic Coprinus cinereus trp1 strain with the C. cinereus TRP1-containing plasmid. One Trp⁺ Xyn⁺ transformant for each of the recombinant plasmids was obtained, which showed a markedly high xylan-degrading activity as compared with the control Trp⁺ transformant.     

Removal of bisphenol A by polymerization and precipitation method using Coprinus cinereus peroxidase

Bisphenol A was efficiently removed by the polymerization and precipitation method using Coprinus cinereus peroxidase. The removal efficiency was optimal between pH 9–10 and at 40 °C with a molar ratio of H₂O₂ to bisphenol A of about 2. To remove 100 mg bisphenol A l^–1, peroxidase was required 5 U ml^–1 at pH 7 and 25 °C and 3 U ml^–1 at pH 10 and 40 °C.     

Differential resistance and the importance of antibiotic production in Acromyrmex echinatior leaf-cutting ant castes towards the entomopathogenic fungus Aspergillus nomius

Paired exocrine metapleural glands are present in almost all ants and produce compounds with antibiotic properties towards a variety of pathogenic fungi and bacteria. In Acromyrmex leaf-cutting ants, small workers have relatively large metapleural glands compared to large workers, and thus harbour approximately half the number of gland cells of large workers, despite being only one-fifteenth their body mass. Here we present results showing that when the two worker castes of A. echinatior are treated with spores of the pathogenic fungus Aspergillus nomius in doses that correspond to the difference in metapleural gland cell numbers they do not differ in survival. However, we also show, for the first time, that small workers survive significantly longer than large workers when both are challenged with a dose of spores that corresponds to their difference in body mass. Furthermore, the time until Aspergillus nomius hyphae and spores appear on the cadavers of workers dead from infection, is significantly increased in the small worker caste. In addition to supporting previous findings that the metapleural glands have an important defence function, the results of this study indicate that the relatively large glands in small workers makes this caste particularly well adapted to preventing pathogenic microorganisms from entering the colony. Received 23 January 2006; revised 7 April 2006; accepted 11 April 2006.     

PCR with degenerate primers amplifies a subgenomic DNA fragment from the endoglucanase gene(s) of Torula thermophila, a thermophilic fungus

The aim of this study was to enable the polymerase chain reaction (PCR) amplification of DNA fragments within endoglucanase gene(s) of Torula thermophila, by using degenerate primers so that the amplified fragment(s) could be used as homologous probe(s) for cloning of full-length endoglucanase gene(s). The design of the degenerate PCR primers was mainly based on the endoglucanase sequences of other fungi. The endoglucanase gene sequence of Humicola insolens was the only sequence from a thermophilic fungus publicly available in the literature. Therefore, the endoglucanase sequences of the two Trichoderma species, Trichoderma reesei and Trichoderma longibrachiatum, were used to generalize the primers. PCR amplification of T. thermophila genomic DNA with these primers resilied in a specific amplification. The specificity of the amplified fragment was shown by Southern hybridization analysis using egl3 gene of T. reesei as probe. This result suggested that the degenerate primers used in this study may be of value for studies aimed at cloning of endoglucanase genes from a range of related fungi.     

Construction of a bacterial artificial chromosome (BAC) library of Coprinus cinereus

A bacterial artificial chromosome (BAC) library of the genomic DNA of Coprinus cinereus strain MP#2 was constructed using the BAC vector pBACTZ, which carries the C. cinereus trp1 gene. The library consists of 1536 clones. Analysis of inserts in some of the clones suggested that the library covers five times the C. cinereus genome. Screening of the BAC clones using ten markers mapped on nine different chromosomes also indicated that the library is likely to cover the whole length of the genomic DNA. We show an example of transformation of C. cinereus with BACs containing inserts of longer than 170kb.     

Extracellular lipase production by a sapwood-staining fungus,Ophiostoma piceae

The extracellular lipase production of a sapwood-staining fungus, Ophiostoma piceae, grown in liquid media, was optimally active at pH 5.5 and 37°C. Although glucose, fructose, sucrose, starch and dextrin, as carbon sources for growth gave similar mycelial yields, which were higher than those obtained with arabinose, galactose or raffinose, the cells growing on those carbohydrates produced little extracellular lipase. However, both high biomass and lipase activity were obtained when plant oils (olive, soybean, corn, sunflower seed, sesame, cotton seed or peanut) were used as carbon sources. Among the nitrogen sources examined, Casamino acids gave the best growth, whereas (NH₄)₂SO₄ gave the best lipase production. The highest lipase productivity seen was obtained in a medium with olive oil as carbon source and a combination of (NH₄)₂SO₄and peptone as nitrogen source.The authors are with Forest Products Biotechnology, Department of Wood Science, Facully of Forestry, University of British Columbia, Vancouver, BC, V6T 1Z4, Canada