Nuclear ribosomal DNA sequences characteristic of Sebacinaceae were detected by direct amplification of DNA from field collected ectomycorrhizal samples. A study was undertaken to confirm the formation of ectomycorrhizas by Sebacinaceae and to characterise representative samples genetically, morphologically and ultrastructurally. The investigated sebacinoid mycorrhizas were sufficiently characteristic to be identified morphologically. They are distinguished by a characteristic, clampless, hydrophilic extramatrical mycelium, which is very variable in diameter and in wall thickness, and by the presence of distinctive, y-shaped, inflated multibranchings. Differences in the mantle structure clearly discriminate the two investigated sebacinoid mycorrhizal types. Comparison of the D1/D2 domains of the nuclear large subunit pseudogene and the ITS1 and ITS2 regions identifies the fungal partner of one of the mycorrhizal samples as Sebacina incrustans. Ultrastructural investigations of the ectomycorrhizas show a doliporus/parenthesome architecture consistent with that of the Sebacinaceae. Recently published sequence data obtained from sebacinoid mycorrhizas are compared to our sequences and the complex trophic relationships in the Sebacinaceae are discussed. Observations on ectomycorrhizas and basidiomes suggest that species of Sebacinaceae are fairly common mycobionts in various ectomycorrhizal plant communities.

The entomopathogenic Beauveria bassiana is a potential biopesticide. Fungicide-tolerant isolates of this fungus would have an added advantage of being compatible with the conventional chemical methods of pest control. Therefore, 30 isolates of the fungus were screened for tolerance to bavistin a commonly used benzimidazole fungicide containing methyl benzimidazol-2-yl carbamate (MBC). Germination and growth bioassays in the presence of 0.05% bavistin were conducted for screening. Three tolerant isolates were identified, showing tolerance up to 2% bavistin. Mutation in the β-tubulin gene is known to confer tolerance to MBC; the nine known mutation sites in the gene involved were sequenced in the tolerant strains. The β-tubulin gene from codons 1–405 was amplified using two pairs of degenerate primers, designed for the conserved region of the β-tubulin gene and sequenced. From the sequence suitable primers were designed for the regions flanking the nine known sites involved in mutations conferring MBC tolerance. The amplified products with these primers from the MBC-tolerant isolates were sequenced and in two (ARSEF 1315 and ARSEF 1316) a mutation was detected in the 198 codon resulting in replacement of glutamic acid with lysine. In the third tolerant isolate (ITCC 913) no mutation could be detected in any of the nine known sites conferring tolerance to MBC. To complete the sequencing of the β-tubulin gene, the remaining part (from codon 405 onwards) of the gene was isolated by a three-prime gene walk. The gene sequence showed a close homology to fungal β-tubulin genes with four introns.

The fly agaric is a remarkable mushroom in many respects; these are its bearing, history, chemical components and the poisoning that it provokes when consumed. The ‘pantherina’ poisoning syndrome is characterized by central nervous system dysfunction. The main species responsible are Amanita muscaria and A. pantherina (Amanitaceae); however, some other species of the genus have been suspected for similar actions. Ibotenic acid and muscimol are the active components, and probably, some other substances detected in the latter species participate in the psychotropic effects. The use of the mushroom started in ancient times and is connected with mysticism. Current knowledge on the chemistry, toxicology, and biology relating to this mushroom is reviewed, together with distinctive features concerning this unique species.

Two-dimensional gel electrophoresis was used to analyse stage-specific proteins from Phytophthora palmivora, a pathogen of cocoa and other economically important tropical crops. Approximately 1% of proteins appeared to be specific for each of the mycelial, sporangial, zoospore, cyst and germinated cyst stages of the life-cycle. Three proteins excised from protein gels of P. palmivora were identified as isoforms of actin by database searches to public libraries of Phytophthora infestans. The protein profiles of parallel samples of P. palmivora and P. infestans demonstrated that 30% of proteins precisely co-migrated suggesting that proteomics may be used to examine changes in the specific stages in the life cycles of Phytophthora spp.

In order to investigate phylogenetic relationships of selected members of Peronosporaceae to the genera Pythium, Halophytophthora, Phytophthora, and Peronophythora, Bayesian analysis of partial sequences of the ITS1-5.8S-ITS2 region was performed. In addition, sequences of the complete ITS1-5.8S-ITS2 region were analysed for 101 collections belonging to the genera Peronospora, Hyaloperonospora, Perofascia, Pseudoperonospora, Phytophthora and Peronophythora, using Bayesian inference of phylogeny and maximum parsimony. The results confirm a close relationship of these genera. The strongly supported Peronosporaceae clade is located within the paraphyletic genus Phytophthora (including Peronophythora). Monophyly of the genera Pseudoperonospora and Hyaloperonospora are strongly supported, but monophyly of Peronospora s. str. can neither be confirmed nor rejected. Within Peronospora s. str., basic relationships often remain unclear; however, some groups form highly supported monophyletic clades. Peronospora species parasitising the same host families or orders are only partly resolved as monophyletic, indicating frequent host-jumping also between distantly related host families. All species inhabiting flowers of different host families form a strongly supported monophyletic group.

Two new genera, Neophaeosphaeria and Phaeosphaeriopsis, are described to accommodate species of Paraphaeosphaeria that are not congeneric based on morphological characters and results of 18S rDNA sequence analyses. Paraphaeosphaeria s. str. is restricted to species with two-septate ascospores and anamorphs that produce non-septate, smooth, pale brown conidia enteroblastically from phialides which have some periclinal thickening. Species in Neophaeosphaeria have 3–4-septate ascospores and anamorphs that produce ovoid to ellipsoid, non-septate, brown, verrucose or punctate conidia from percurrently proliferating conidiogenous cells. Paraphaeosphaeria barrii, P. conglomerata, P. filamentosa and P. quadriseptata are transferred to Neophaeosphaeria. At present all species in Neophaeosphaeria occur on Yucca (Agavaceae). Phaeosphaeriopsis is described for species that produce 4–5-septate ascospores. Known anamorphs produce cylindrical, 0–3-septate, brown, punctate conidia from percurrently proliferating conidiogenous cells or bacillar conidia from simple phialides. P. agavensis, P. glauco-punctata, P. nolinae and P. obtusispora are transferred to Phaeosphaeriopsis. P. amblyspora is described as a new species.

Phylogenetic analyses of the ITS1-5.8S-ITS2 region of the ribosomal RNA gene cluster were carried out with two short-cycled (aecial/telial) European rusts on Asteraceae, Puccinia distincta causing the current pan-European epidemic on Bellis perennis, and P. lagenophorae causing a similar disease on Senecio spp., as well as the macrocyclic P. obscura which alternates between B. perennis (pycnial and aecial host) and Luzula spp. (main host). All three species formed a well-resolved cluster when compared with the ITS sequences of a range of other rust fungi, using both parsimony and distance methods. The sequences of P. distincta and P. lagenophorae differed from each other at three positions whereas P. obscura differed from P. distincta at 37 points. Together with consistent morphological and epidemiological differences across Europe, these data support the recognition of P. distincta as a separate species from P. lagenophorae. Both may be derived from P. obscura, although the precise evolutionary history remains obscure.

The Amphisphaeriaceae is an important family of ascomycetes within the Xylariales. There has been, however, disagreement regarding the taxonomic placement of many genera within this family and whether it should be confined to ascomycetes producing Pestalotiopsis-like anamorphs. In this study, phylogenetic relationships among members of the Amphisphaeriaceae are investigated using partial sequences of the 28S rDNA. Molecular data provided further evidence to support the association of several coelomycetous genera with the ascomycetous Amphisphaeriaceae. Phylogenetic analyses also show that all ascomycetous genera possessing Pestalotiopsis-like anamorphs are monophyletic and confirm the anamorphic-teleomorphic connections of some. There is, however, insufficient evidence to support the restriction of Amphisphaeriaceae to genera, which produce Pestalotiopsis-like anamorphs, because the phylogenetic placement of Amphisphaeria umbrina is not fully resolved and its affinities with other members received low bootstrap support. The results also indicate that Iodosphaeria and Arecophila should be excluded from the Amphisphaeriaceae. The placement of Lanceispora in the Amphisphaeriaceae is doubtful. A broad concept of the family Amphisphaeriaceae is advocated until further data are available.

The production of cellulases on minimal medium in the edible mushroom Agaricus bisporus is regulated by the carbon source: induced by cellulose and repressed by glucose. In order to isolate cellulose-growth specific sequences, a cDNA library from A. bisporus using suppression subtractive hybridization (SSH) was constructed. Northern blot analysis indicated that a high level of enrichment was achieved; 183 clones were isolated. A preliminary screen with cellulose-specific genes of A. bisporus (cel1, cel2, cel3 and cel4) using Southern hybridization resulted in 28 clones to be cel3, and 5 clones were cel2. The remaining 144 clones were sequenced. Partial sequences of the following genes were found: a β-glucosidase homologue of the blvk gene of Kluyveromyces marxianus; a cellulase homologue of an endoglucanase (avicellase III) of Aspergillus aculeatus, four different xylanases homologue of the xyn genes of different fungi, and one hexose transporter homologue to the hxtA gene of Aspergillus parasiticus. The apparent full-length of two hydrophobins homologue to the abh3 gene of A. bisporus and one histone homologue to the h2a gene of Aspergillus niger were also found. The remaining sequences did not have homology to any known genes.

A Coniothyrium minitans strain (T3) co-transformed with the genes for β-glucuronidase (uidA) and hygromycin phosphotransferase (hph), the latter providing resistance to the antibiotic hygromycin B, was used to investigate the survival and infection of sclerotia of Sclerotinia sclerotiorum by C. minitans over time in four different soils. Infection of sclerotia was rapid in all cases, with the behaviour of transformant T3 and wild type parent A69 being similar. Differences were seen between the soils in the rate of infection of sclerotia by C. minitans and in their indigenous fungal populations. Amendment of agar with hygromycin B enabled the quantification of C. minitans in soil by dilution plating where there was a high background of other microorganisms. In Lincoln soil from New Zealand, which had a natural but low population of C. minitans, the hygromycin B resistance marker allowed the umambiguous discrimination of the applied transformed isolate from the indigenous hygromycin B sensitive one. In this soil, although the indigenous C. minitans population was detected from sclerotia, none were recovered on the dilution plates, indicating the increased sensitivity of C. minitans detection from soil using sclerotial baiting. C. minitans was a very efficient parasite, being able to infect a large proportion of sclerotia within a relatively short time from an initially low soil population. The addition of hygromycin B to agar also allowed the detection of C. minitans from decaying sclerotia by inhibiting secondary fungal colonisers. This is the first report to show that fungi colonising sclerotia already infected by C. minitans mask the detection of C. minitans from sclerotia rather than displacing the original parasite.

Thirty South American species of Cortinarius belonging to the subgenera Telamonia, Dermocybe, Myxacium, Phlegmacium, and Cystogenes were studied using an integrated approach that included morphological, anatomical, and ultrastructural data, and also molecular phylogenetic analysis of nuclear rDNA sequences. The micromorphology of the basidiomes was studied by light microscopy, and the principal structures were illustrated by line drawings. Basidiospore ornamentation was studied by scanning electron microscopy (SEM). Nuclear internal transcribed spacers (ITS, including the 5.8S gene) and the DNA coding for the D1/D2 domains of the large ribosomal subunit (LSU) were sequenced and analysed using a Bayesian Markov chain Monte Carlo method to estimate phylogenetic relationships between the studied Cortinarius species. Morphology and anatomy of the pileus surface and basidiome pigmentation appeared to be the most useful characters to delimit some natural groups, whereas microcharacters related to the structure of pileus context, hymenophoral and stipe trama were of little taxonomic value. Basidiospore morphology and cheilocystidia seem to be taxonomically relevant at the species level. The following five infrageneric groups were supported by the morphological, chemical and molecular data: (1) Telamonia characterized by wide hyaline hyphae of the veil and by small basidiomes; (2) Dermocybe spp. with an epicutis as the most external layer of the pileus, and skyrin and hypericin pigments; (3) Dermocybe spp. with a thin viscid layer on the pileus, and endocrocin and dermolutein pigments; (4) Phlegmacium spp. characterized by a long and radicating stipe; and (5) Phlegmacium spp. that overlap in some macrocharacters with Telamonia species. Our analyses suggest that classification concepts based mainly on macromorphological characters are likely to lead to artificial grouping, whereas certain microscopical and chemical characters seem to be useful in constructing a more natural classification system for Cortinarius.

Double-stranded RNA (dsRNA) has been isolated from Agaricus bisporus fruit bodies exhibiting a wide range of disease symptoms. The symptoms which occurred singularly or in combination included; bare cropping areas on commercial beds (primordia disruption), crop delay, premature veil opening, off- or brown-coloured mushrooms, sporophore malformations and loss of crop yield. All symptoms were associated with loss of yield and/or product quality. Collectively, these symptoms are described as mushroom virus X (MVX) disease. The dsRNA titre was much lower than that previously encountered with the La France viral disease of mushrooms and a modified cellulose CF11 protocol was used for their detection. A broad survey of cultivated mushrooms from the British industry identified dsRNA elements ranging between 640 bp and 20.2 kbp; the majority have not previously been described in A. bisporus. 26 dsRNA elements were identified with a maximum of 17, apparently non-encapsidated dsRNA elements, in any one sample. Three dsRNAs (16.2, 9.4 and 2.4 kbp) were routinely found in mushrooms asymptomatic for MVX. Previously, La France disease was effectively contained and controlled by minimising the on-farm production and spread of basidiospores. Our on-farm observations suggest that MVX could be spread by infected spores and/or mycelial fragments.

Previous studies of morphological variation in the homobasidiomycete Hyphoderma setigerum have lead to suspicions of a species complex. This study explores variation in DNA sequences from the nuclear ribosomal ITS region of 45 specimens from America, Asia, and Europe in a phylogeographic context. Based on molecular analysis, morphological studies, and crossing tests, nine preliminary taxa are shown to exist inside the species complex, and the two previously described segregate species H. subsetigerum and H. nudicephalum are confirmed. The molecular analysis shows evidence of allopatric differentiation over intercontinental distances. Only one of the nine well-supported clades has a geographic distribution spanning more than one continent, probably indicating the importance of vicariance in the evolution of this species complex. The basionym of H. setigerum, Thelephora setigera, is neotypified to fix the application of that name.