Techniques for selective isolation, semiquantification and rapid virulence testing are described for Catenaria anguillulae, an important endoparasitic fungus of nematodes. Selective isolation was achieved by using second-stage juveniles of Anguina tritici as bait in soil aliquots for 48 h. Semiquantitative estimation of the population of the fungus was made by calculating the infection percentage of inoculated juveniles. Rapid virulence testing was done by using second-stage juveniles of Heterodera cajani, Meloidogyne incognita and A. tritici.

A taxonomic review and revision of the dematiaceous hyphomycete genus Acrogenospora is presented. Species in this genus produce acrogenous, solitary, nonseptate conidia from mononematous, simple conidiophores, with multiple percurrent proliferations. Acrogenospora altissima comb. nov. and A. megalospora comb. nov. are proposed for the anamorphic states of two Farlowiella species. A. ovalia and A. subprolata spp. nov. are described from submerged wood in Hong Kong. They differ from other Acrogenospora species in shape, colour, and size of their conidia. A taxonomic key to species in the genus is provided.

An evaluation was made of the genotypic diversity (DNA fingerprinting) of 269 A. flavus strains, including subpopulations isolated from grain sampled at harvest (91 unique ‘fingerprints’ or genotypes/128 strains), field soil (26 genotypes/31 strains), maize insects (49 genotypes/52 strains) and air-spora (56 genotypes/58 strains), from a maize field near Kilbourne, Illinois. Eight A. flavus genotypes were isolated from grain samples harvested in different years (1988–1991). Genotype 36, isolated from three maize samples, matched the DNA fingerprint of a K. E. Papa strain NRRL 19997, isolated from maize grown in Georgia. Ninety-eight percent of the A. flavus genotypes produced sclerotia and 53% produced aflatoxin. Contrasts of DNA fingerprints revealed two matches involving subpopulations from grain and soil, one match for grain and maize insects, and no matches for grain and air-spora. The high genotypic diversity recorded for each subpopulation, in addition to a limited sample size, precluded any assessment of the relative importance of these subpopulations as sources of A. flavus infective inoculum. Aspergillus parasiticus was routinely isolated from soil samples.

Isolates of Tiarosporella parca from needles of Picea abies growing in Lapland, southern Finland, southern Norway and Switzerland were compared with each other by using their combined fatty acid and sterol profiles (FAST) and randomly amplified microsatellite (RAMS) markers.

Nineteen fatty acids and four sterols were recorded, nine of which appeared only at low amounts (<1% of the total amount of extractives). In a discriminant analysis of their FAST-profiles the isolates could be separated into three non-overlapping groups: (i) Lappish and southern Norwegian, (ii) Southern Finnish and (iii) Swiss isolates. The Swiss isolates form the most distinct group with 11 out of 23 extractives significantly different from those of the other geographical populations.

In RAMS analysis two out of five primers tested with ten isolates resulted in polymorphic banding patterns. Using these two primers 26 variable markers in eight size areas were observed which allowed separation of the 39 isolates investigated into 35 different haplotypes. Thus, only four pairs of identical isolates were observed. Determination of the genetic diversity among the isolates revealed that 59% of variation was due to variance within populations, 27% was due to variance between populations within geographical regions and 13% was due to variation between regions.

Hence, both methods show a differentiation of the European T. parca population according to geographical origin.

Two double-stranded ribonucleic acid (dsRNA) elements were detected in an isolate of the clavicipitaceous grass endophyte Epichloë festucae obtained from an asymptomatic Festuca rubra plant. No dsRNA was detected in three other isolates of E. festucae. The size of the two dsRNA elements isolated from mycelium was of 5·2 and 3·2 kbp. Hybridization experiments with a cDNA probe complementary to the 5·2 kbp dsRNA showed that there was no sequence similarity between the dsRNA elements. Isometric virus-like particles of about 50 nm in diameter were observed by electron microscopy in sucrose-gradient purified virus preparations from the isolate containing dsRNA. The 5·2 kbp dsRNA element was present in the partially purified virus preparations, suggesting that this element is encapsidated. There was no difference with respect to growth rate or colony morphology between the dsRNA containing and dsRNA-free isolates. The presence of dsRNA and virus-like particles suggests the presence of a mycovirus in the Vit5 strain of E. festucae.

Epichloë species are endophytic symbionts of grasses which may differ in the relative importance of their sexual or asexual life cycles. Sexual reproduction of the fungus by stroma-formation prevents host flowering (choke) and thus is highly antagonistic, whereas asexual reproduction through clonal propagation in host seeds does not affect host fitness. Stroma-forming Epichloë endophytes from Bromus erectus and non-stromal strains from B. benekenii and B. ramosus were recognized as a distinct mating population (MP) based on complete sexual compatibility among strains and intersterility between other MPs established by mating tests. This biological species represents the only documented case of highly antagonistic strains interfertile with highly mutualistic strains. A second distinct MP of Epichloë was evident on Brachypodium sylvaticum including both stroma-forming and non-stromal isolates. These two MPs were further characterized by distinct morphologies of fruiting structures, allozyme divergence, β-tubulin gene phylogeny, and host preferences, and were described as new species: E. bromicola associated with Bromus spp. and E. sylvatica with Bp. sylvaticum. Additional mating tests among Epichloë from several, previously unexamined, hosts including Brachypodium pinnatum, Calamagrostis villosa, Festuca spp., Phleum pratense, and Poa spp., expanded the known host ranges of three other European species, E. typhina, E. festucae and E. baconii. Genetic variability of all five European species and gene diversity of host subpopulations were analysed based on allozyme data from a total of 497 Epichloë isolates. Average gene diversity (Hs) within MPs ranged from 0·09 to 0·36 with E. typhina being the most diverse, and GST values, a measure for between subpopulation differentiation, ranged from 0·73 to 0·90 indicating that genetic isolation of endophytes on many host grasses is likely.

As part of an inventory of mycodiversity in the Brazilian Cerrado new coelomycetes were collected and are herein described: Phyllosticta xylopiae-sericeae sp. nov., Dinemasporium duguetiae sp. nov., Harknessia salvertiana sp. nov., Harknessia qualeae sp. nov. on living leaves of Xylopia sericea, Duguetia furfuracea, Salvertia convallariodora, and Qualea grandiflora, respectively. Pseudothiopsella hirtellae is reported on a new host, Hirtella gracilipes, and for the first time illustrated in detail.

Selenate-resistant (SelR) mutants of Botrytis cinerea were selected by plating conidia or mycelial plugs onto minimal medium amended with selenate and taurine. Ultraviolet irradiation of conidia to give 5% survival increased mutant yield twenty-fold. Mutants could be divided into three classes based on growth in the presence of selenate or chromate and on response to taurine in minimal media. Nitrate non-utilizing (Nit) mutants, generated as spontaneous sectors on minimal media amended with chlorate, behaved as nit1 mutants in growth tests (i.e. putatively defective in nitrate reductase apoenzyme). When nit1 mutants were paired on medium with nitrate as sole nitrogen source some pairings complemented, behaviour attributed to intragenic complementation. Selected crosses of SelR and nit1 mutants with wild-type strains gave 1[ratio ]1 segregation of both phenotypes and no evidence of linkage to either Mbc1 (benzimidazole resistance) or Daf1 (dicarboximide resistance) markers; loose linkage was confirmed between Mbc1 and Daf1. Strains showing the SelR phenotype may result from mutations in different genes; the genotypic symbol Sel 1 is allocated to one of these mutations. Four-point crosses involving the markers at Sel 1, nit1, Mbc1 and Daf1 generated progeny with all 16 genotypes. Both Sel1R and nit1 mutants were stable following subculture and retained pathogenicity in a bean seedling assay. Complementation was demonstrated between SelR and nit1 mutants selected from the same parent.

Antrodia lalashana sp. nov. and Antrodiella formosana sp. nov. found growing saprotrophically on the decayed stems of Persea zuilhoensis and Abies kawakamii, respectively, in Taiwan, are described and illustrated. Cultural characters are described for the two species.

The relation between the pattern of fumonisin production and the mating population and mating type of Fusarium isolates from maize in Argentina at different maturity stages has been evaluated. Fifty-one isolates of Fusarium species belonging to the Liseola section were identified to mating population and tested for their ability to produce fumonisins (FB1, FB2 and FB3). Only mating populations associated with maize, A (Fusarium moniliforme, 23 isolates), D (F. proliferatum, 24 isolates) and E (F. subglutinans, 4 isolates) were found. All but two isolates of populations A and D produced, when grown on maize substrate, high levels of fumonisins ranging from 0·01 to 3·99 mg g−1 (mean 2·00 mg g−1 and 1·69 mg g−1, respectively), whereas isolates of population E yielded less than 0·02 mg g−1 (mean 0·01 mg g−1). Five isolates of F. proliferatum, all belonging to mating type D−, produced more FB2, than FB1, which is consistent with finding a relatively large amount of FB2 (0·01 mg g−1; FB2/FB1=2·27) in one of the maize samples from which these isolates were obtained. Amounts of FB3 were similar to FB2 in cultures of mating population A (mean FB3/FB2=0·89), but much lower than FB2 in cultures of mating population D (mean FB3/FB2=0·21).

The occurrence of Lophodermium on fallen needles and Leptostroma, the mitosporic state of Lophodermium, on living needles of Japanese black pine trees treated with simulated acid rain (SAR, pH 3) or tap water (control, pH 6·3) were studied. The amount of fallen needles was also examined. Significantly more needles fell from SAR-treated trees than from control trees, though there were no visible symptoms in the needles of SAR-treated trees. The detection frequency of Lophodermium on fallen needles, and that of endophytic Leptostroma on 2 year old and 3 year old needles, was lower in SAR-treated than control trees. The reduction in the detection of Lophodermium/Leptostroma on needles implied the reduction in the density of endophytic fungi with air pollution.

Cladistic analysis of characters derived from nuclear-encoded 18S rDNA sequences was used to infer the phylogenetic placement of ambrosial species of the anamorph genus Raffaelea among perithecial ascomycetes. Of eight species in the genus investigated, seven resolve as a monophyletic lineage that forms a sister group to the sexual genus Ophiostoma. Raffaelea hennebertii, the single species excluded from this lineage, appears to be allied phylogenetically with species of Melanospora. These data are discussed in relation to the monophyly of Raffaelea and its relationship with other sexual and asexual fungal associates of wood-boring Coleoptera. In addition, we report the presence of significant length variation in PCR-amplified 18S rDNA fragments for three of the Raffaelea species studied.

Phylogeny and biogeography of Lentinula, which includes cultivated shiitake mushrooms, were investigated using parsimony analyses of an expanded nuclear ribosomal DNA dataset. Lentinula occurs in the New World as well as Asia and Australasia. The Asian–Australasian Lentinula populations appear to form a clade, but species limits within this group are controversial. We refer to the entire Asian–Australasian Lentinula clade as shiitake. Thirty-seven wild-collected isolates of shiitake were examined, representing Australia, Borneo, China, Japan, Korea, Nepal, New Zealand, Papua New Guinea (PNG), Tasmania and Thailand. Five isolates of the New World species, L. boryana, were included for rooting purposes. Levels of sequence divergence between North and Central American L. boryana isolates are higher than those between the most divergent shiitake isolates. In shiitake, five independent lineages of rDNA were identified, which we call groups I–V, but relationships among these lineages are not well resolved. Group I includes populations from northeast Asia to the South Pacific. Group II includes populations from PNG, Australia and Tasmania. Group III is limited to New Zealand. Group IV is from PNG. Finally, group V is from eastern China and Nepal. The distribution of rDNA lineages suggests a complex biogeographic history. Although many areas remain unsampled, our results suggest that certain areas have particularly high levels of diversity and should be targeted for further study and conservation.

Variation within the internal transcribed spacer (ITS) and the intergenic spacer (IGS) of the ribosomal RNA gene of isolates representing seven European species of Armillaria was examined by PCR, coupled with RFLP analysis and partial sequencing of the ITS region.

The amplified-ITS region was about 840 bp long and uniform in length among the European isolates, while the amplified-IGS region showed four different lengths which corresponded to A. mellea, A. tabescens, A. ectypa and a group including the four other species. A. borealis, A. ostoyae, A. cepistipes and A. gallica.

Restriction digestions of the ITS and IGS regions by Alu I and Rsa I, respectively, gave rise to the same four polymorphic groups. However, A. borealis and A. ostoyae were easily separated from A. cepistipes and A. gallica by digestion of the two rDNA spacers with Hinf I and Taq I. Nde II digests of the amplified ITS could distinguish A. borealis from A. ostoyae and each of the seven species were separated by Alu I digestion of the IGS region. A North American A. mellea isolate, partly examined in this work, was found to be different from the European A. mellea isolates, while there was a close similarity between the European A. ostoyae and an isolate of the same species isolated from the U.S.A.

Cluster analysis based on the presence or absence of comigrating restriction fragments indicated more than 80% similarity between A. borealis and A. ostoyae and between A. cepistipes and A. gallica, while A. mellea, A. tabescens and A. ectypa were found in separate clusters exhibiting, respectively, about 40, 38 and 32% average similarity with the other species.

Although little intraspecific variation was observed in many species, A. gallica and A. cepistipes were found to be heterogeneous. In view of recent results suggesting several groups in A. borealis and A. cepistipes, the Alu I restriction patterns obtained in this work would identify the former species as type B, characterized by an Alu I pattern different from that of A. ostoyae, and the latter species as composed of one isolate (C4) of type B and three isolates (C1, C2 and C3) of type A.

Nucleotide sequences of the rDNA internally transcribed spacer 1 (ITS1) of one isolate of each species showed that A. mellea and A. ectypa differed from the other species by several insertions and point mutations giving rise to a level of similarity ranging from 66 to 79%. This DNA region was highly conserved within the other species which revealed a similarity of 97 to 100%.

These results demonstrate that analysis of rDNA spacers provides appropriate data to circumscribe taxonomic entities within the Armillaria complex and that the phylogenic relationships among species deduced from the present study are consistent with previous analyses based on pairing tests as well as morphological and physiological characters.

A total of 34 isolates of Cylindrobasidium evolvens were obtained from wounded Picea abies in five sample plots in Sweden and Lithuania. Population structure and genetic variation within the species were assessed by somatic incompatibility and arbitrary primed DNA (AP-DNA) analysis, using the M13 core sequence as a primer. All somatic incompatibility pairings between different C. evolvens isolates resulted in weak mycelial antagonism. The AP-DNA profiles indicate genetic variation was high in the species with no differentiation among isolates being found either on a large (more than 700 km) (R2=0·108) or a fine scale (200 m) (R2=0·024). In a plot, genetic similarity was significantly higher (P<0·02) among isolates from wounds infected during the previous 2 years than among the rest of the same population.

Faecal samples collected from the Mountain brushtail possum (Trichosurus caninus) at several sites in eastern Australia showed the presence of spores from at least 21 hypogeous fungi, and several epigeous species. Most were found in faeces of animals at one site, where samples were collected in different seasons. Further collections are needed from the other sites at different times of the year to clarify how widespread the mycophagous habits of T. caninus are. Many of the hypogeous fungi identified are either known or presumed to form mycorrhizal relationships with the roots of forest trees and shrubs and T. caninus may disperse their spores in its faeces.

Podosphaera clandestina var. cydonia var. nov., Setoerysiphe kashmiriensis sp. nov. and Brasiliomyces kumaonensis sp. nov., collected from Northern India on Cydonia oblonga, Sambucus wightiana and Quercus sp., respectively, are described and illustrated.

Using the mating type genes (MAT-1 and MAT-2) from Cochliobolus heterostrophus (a pathogen of maize) as probes, MAT genes from C. carbonum (also a maize pathogen) and C. victoriae (an oat pathogen) were cloned. Sequence analyses and functional tests showed that C. carbonum MAT-1 and C. victoriae MAT-2 are structurally similar to their C. heterostrophus homologues, and all three genes function comparably when expressed in C. heterostrophus. Gel blot analyses of DNAs from C. heterostrophus or C. carbonum, probed with C. heterostrophus MAT-specific fragments, revealed that field isolates from diverse geographical locations contained either MAT-1 or MAT-2, but never both. The entire currently existing collection of C. victoriae isolates, however, contained only MAT-2, and all members of the collection were found to be either female sterile or completely infertile. Thus, crosses between isolates of C. victoriae were not possible, although crosses between C. victoriae and C. carbonum (but not C. heterostrophus) were readily made, confirming previous reports that these two species are interfertile. These observations, combined with careful scrutiny of the literature, have led to the hypothesis that MAT-1 female fertile isolates of C. victoriae have never existed. All available evidence is consistent with the notion that the entire C. victoriae field population was derived from a single progenitor strain, which arose as the result of horizontal transfer of genes for pathogenicity to oats into a strain of C. carbonum that was MAT-2 and female sterile.

The internal transcribed spacers, ITS1 and ITS2, and 5.8S region of ribosomal DNA (rDNA) from 20 species of Ascosphaera were amplified by PCR, and their sequences determined and compared. No variation was detected in the sequences from 23 widely distributed isolates of A. apis, in sequences from 11 widely distributed A. atra isolates, in four A. aggregata isolates, or in sequences from two isolates each of A. acerosa, A. duoformis, A. flava, A. larvis, A. pollenicola and A. proliperda. However, the ribosomal sequences from each of these nine species, and from another 11 species of which only a single isolate was examined, differed from one another by 0·18–30·9%. Thus these sequences provide a rapid method for identifying species, and searches using them showed that the sequence of A. apis rDNA recorded in the international databases is, in fact, that of A. atra.

The rDNA sequences also provided data for assessing the relationships of these fungi. Of the rDNA sequences in current international databases, that of Eremascus albus was very close to, but distinct from, those of the Ascosphaera species. Comparisons of the Ascosphaera sequences showed that most formed consistent clusters irrespective of the method of comparison used (distance matrix and parsimony), or whether the ITS1 or ITS2 portions were used; A. acerosa with A. asterophora, A. atra with A. duoformis, A. colubrina closely with A. flava, A. larvis, A. major, A. variegata and more distantly with A. apis and A. celerrima, and, also, A. aggregata with A. subcuticulata, A. proliperda and more distantly with A. solina. The apparent relationships of these clusters were inconsistent, depended on the alignment of several regions of ‘indels’, and could not be resolved. The A. fusiformis, A. naganensis and A. osmophila sequences showed inconsistent relationships with the others, especially that of A. osmophila, which had an A. solina-like ITS2 region, but an atypical ITS1 sequence with a large unique repetitive insertion. The clusters based on gene sequence comparisons clearly correlated with groupings based on ascospore morphology and other characters.

Agaricus bisporus browning is a common and economically detrimental phenomenon, in which melanogenic phenols are enzymically processed into quinones, which evolve eventually to melanins. This review deals with the two fundamental sides of this process, enzyme(s) and phenolic substrates. Mushroom tyrosinase, the main polyphenol oxidase encountered in the A. bisporus sporophore, is treated in the first part. Its overall molecular architecture, isoforms, primary sequence and genetic background are considered. The presentation of tyrosinase catalytic features, including enzyme assays, kinetic properties, substrates and inhibitors, is followed by a comprehensive description of the active site and reaction mechanisms. Because of their relevance for studies of mushroom browning during development and post-harvest storage, the occurrence and properties of latent enzyme forms, as well as the location of tyrosinase and variations of its activity during the A. bisporus life cycle, are also reviewed. The second part deals with the substrates, particularly γ-L-glutaminyl-4-hydroxybenzene (GHB) and its derivatives. Main data concerning the nature, obtention (by extraction or synthesis), spectrometric and chromatographic characteristics, chemical stabilities and biological properties of these typical Agaricaceae compounds are presented. Their distribution and levels according to the strains and flushes are described, as well as their variations during storage. Thirdly, the relationship between browning and the natural or pathogenic discolouration intensity is developed.