Genetic variation of 42 isolates of Corynespora cassiicola, a destructive fungal pathogen of many economically important crop plants including rubber, was investigated using RAPD–PCR analysis. Five genetic groups were identified using RAPD–PCR profiles generated by eight random primers. Results indicate that there is a significant genetic variation among C. cassiicola isolates collected from different host plants. These results should facilitate the development of rubber clones with enhanced resistance against all genetic groups of C. cassiicola.

DNA sequence alignment of the nuclear 5·8S ribosomal RNA (rRNA) gene and internal transcribed spacers (ITS) from Beauveria bassiana demonstrated that 6·62% sequence variation existed between nine isolates. A higher level of mutation was observed within the ITS regions, where 8·39% divergence occurred. Polymerase chain reaction restriction fragment length polymorphism, PCR–RFLP, and DNA sequence alignment of the ITS1 and ITS2 regions identified seven polymorphic restriction endonuclease sites, Alu I, Hha I, Hinf I, Sin I, Tru 9AI and two Tha I restriction sites. The allelic frequency of each genetic marker was determined from 96 isolates. PCR–RFLP defined 24 B. bassiana genotypes within the sample set, from which eight phylogenetic clusters were predicted to exist. AMOVA and Fst (θ) indicated that no significant correlation existed between B. bassiana haplotype and insect host range as defined by insect order from which each isolate was derived.

Basidiomes of 43 samples of eight Amanita species were gathered from different habitats of Hungary. The mineral composition (22 elements) was analysed by the ICP method in three independent replications, and mineral compositions found as discussed and compared. The Amanita species analysed were very different in As-, Cd-, Cr-, Mo-, Mn-, Se- and mainly in V-content. Other elements (Al, B, Ba, Ca, Co, Cu, Fe, Mg, Na, Ni, Sr, Ti, Zn) occur in the basidiomes in balanced concentrations. The K and P contents have the lowest differences. Summarizing all the data (n=43), the average mineral status of species of Amanita can be deduced. The lowest variability measured was for K and P, and the highest for chromium, nickel and vanadium.

Specific, significant accumulation was found only for vanadium, due to the previously demonstrated occurrence of a binding molecule ‘amavadine’ in the basidiomes of A. muscaria. Remarkable Cd-levels were estimated in A. pantherina and A. muscaria (11.4 and 12.3 mg kg−1D.M., respectively). The higher contents of other elements (e.g. K, practically in all species; Se in A. strobiliformis) are analytical facts but, not accumulations.

The mineral compositions of the ectomycorrhizal genus Amanita, of litter decomposing Agaricus and of wood decaying Trametes were compared. Some significant differences were found (AsAmanita<AsAgaricus; CdAmanita<CdAgaricus; CuAmanita<CuAgaricus; PAmanita<PAgaricus; KAmanita>KTrametes; PAmanita>PTrametes) but it seems that the mineral composition of the basidiomes is practically independent of the ectomycorrhizal habit. The specificities of the fungi-tree symbiotic interactions are known, and well documented (higher uptake and transport of certain elements first of all of P), however, the differences found in the mineral components are due to other factors (e.g. substrates, accumulating ability) and not to the mycorrhizal status.

Neurospora and Gelasinospora are traditionally distinguished by the ornamentation pattern of the surface of their ascospores, which are ribbed in the former and pitted in the latter. However, a detailed examination of the morphology of numerous strains of most of the species of both genera confirm the hypothesis that there are not enough criteria to distinguish them from each other. The names Neurospora and Gelasinospora are synonymized and the circumscription of the genus Neurospora amended. Partial sequences of the 28S rDNA gene from 27 species of both genera were analysed to infer their phylogenetic relationships. Species of the two genera were interspersed in the different clades and confirmed that they are genetically very similar. The grouping obtained demonstrates that the morphology of the episporial-layer of the ascospores is an informative phylogenetic character. Two recent isolates from soils of Nigeria and Spain, which could not be classified as any known species of Neurospora are described, illustrated, and recognized as new: N. nigeriensis and N. uniporata spp. nov. A synopsis and key to the 49 species of Neurospora now recognized in the genus is presented, and the new genus Pseudogelasinospora described to accommodate P. amorphoporcata (syn. Gelasinospora amorphoporcata comb. nov.).

We conducted a series of experiments to determine if extraradical hyphae of VAM fungi acclimate to low temperatures. Blocks of field soil containing VAM fungi were either slowly cooled or held at room temperature prior to freezing. Infectivity of VAM fungi was greater in soil that was slowly cooled. We hypothesized that greater infectivity following freezing resulted from cold acclimation of extraradical hyphae. This hypothesis was tested using in vitro mycorrhizas cultured in two-compartment Petri plates, in which hyphae grew into a separate compartment. Metabolic activity of these hyphae following freezing was assessed using a vital stain. The majority of cultures that were slowly cooled prior to freezing contained active hyphae, whereas hyphal activity was almost completely eliminated by freezing in non-precooled cultures. Freezing temperature influenced survival and activity of hyphae. To our knowledge this is the first report of acclimation to cold temperatures by VAM fungi.

The morphology of two soil-borne Verticillium species, V. dahliae and V. tricorpus, was studied on two semi-selective agar media, in the absence and presence of soil. Morphology of the fungi differed considerably between the media, with respect to presence and shape of microsclerotia, dark hyphae (i.e. short melanised hyphae attached to the microsclerotia) and dark mycelium (i.e. melanised mycelium throughout the colony). On modified soil extract agar (MSEA), a pectate based agar, V. dahliae always had globose to elongate microsclerotia, without dark hyphae or dark mycelium, whereas V. tricorpus always had dark hyphae or dark mycelium, and microsclerotia, whenever present, were globose to irregular in shape. On ethanol agar (EA), V. dahliae had large microsclerotia and abundant dark hyphae, whereas V. tricorpus did not form microsclerotia, but always abundant dark mycelium. For the first time we observed the formation of dark hyphae by V. dahliae to a great extent. In the presence of soil, most characteristics were less pronounced, and V. dahliae microsclerotia were smaller, but V. tricorpus produced large microsclerotia, even when they were absent in pure culture. Morphological characteristics suitable for discrimination between the two species on MSEA plates in the presence of soil were selected and tested with fresh isolates from agricultural fields. The two fungi could be distinguished using qualitative characteristics and microsclerotial size. Molecular analysis and morphology on potato dextrose agar confirmed all identifications made on soil dilution plates.

Some Armillaria species cause major disease problems in both forestry and gardens. In order to determine routinely and rapidly the identity of an isolate, a PCR-based method was investigated. A fragment, approx. 900 bp, of the IGS region of the ribosomal DNA was amplified from 96 different isolates of six species of Armillaria collected from the U.K. and mainland Europe. Armillaria mellea, A. gallica and A. tabescens produced unique digestion patterns when the PCR fragment was restricted with Alu I. A. ostoyae could be distinguished from A. borealis and A. cepistipes when the PCR fragment was restricted with Bsm I. Previously unreported RFLP patterns were determined for A. mellea and A. gallica. The existence of A. gallica isolates which are putative heterozygotes for the rDNA cluster is also reported. Based on differences in banding patterns. A. mellea, A. gallica, A. cepistipes and A. borealis were subdivided further into groups which correlated with differences in cultural morphology.

A Melampsora lini β-tubulin gene and flanking sequence has been isolated and characterised. The gene contains 8 introns, is expressed in germinating urediospores and vegetative mycelia and encodes a 448 amino acid protein that is similar to other fungal β-tubulin genes. Evidence for a second β-tubulin gene within the M. lini genome is also presented. The predicted M. lini β-tubulin protein was compared with 34 other fungal β-tubulin proteins in a phylogenetic analysis and, in general, good agreement was observed between the derived β-tubulin based phylogeny and the current taxonomic status of these species. However, a second β-tubulin protein found in five fungal species is atypical when compared to other fungal β-tubulin proteins. Analysis of intron positions within 33 fungal β-tubulin genes showed that intron location was generally characteristic of the fungal class, with euascomycete fungi showing particularly striking conservation of intron position, although intron number was variable and intron sequences showed little conservation.

Amplified fragment length polymorphism (AFLP) analysis is based on selective PCR (polymerase chain reaction) amplification of genomic DNA restriction fragments. The present study was performed to: (1) assess the extent of AFLP variation among major cultivated strains of Lentinula edodes (the shii-take mushroom) in Japan; (2) evaluate the usefulness of AFLP as genetic marker for typing the cultivated strains; and (3) infer the genetic relatedness among them. Six AFLP primer pairs detected a total of 304 DNA fragments in a sample of 13 cultivated strains for wood log cultivation and 2 strains for sawdust cultivation, of which 179 DNA fragments (58·9% of detected fragments) were polymorphic between two or more strains. These polymorphic DNA fragments could differentiate all of the cultivated strains. Cluster analysis and principle coordinate analysis based on AFLP data showed two distinct groups; one group was composed of the strains in which fruiting occurred during the middle to low temperature period from autumn to next spring under outdoor wood log cultivation, and the other strains in which fruiting could be induced in the summer under outdoor cultivation and strains for sawdust cultivation under indoor air-controlled condition.

Genetic variability of 95 isolates of Beauveria bassiana from different geographical regions and hosts was assessed using RAPDs and ITS-RFLP molecular markers in order to characterize its genetic variability. Clusterings were obtained using UPGMA and Principal Coordinates Analysis. Four groups were identified with RAPDs. A 930 bp fragment was amplified with primers ITS1 and PN16, and polymorphisms were observed with AluI and MspI, generating two main clusters. Results indicate low genetic variability present in the Colombian B. bassiana population, no association to host type or geographic locality, and a population structure that is not clonal. This characterization sets criteria for determination of bio-insecticide potentiality, introduction of foreign strains and improvement of entomopathogens within an IPM program.

The Ophiostoma piceae species complex incorporates several economically important species, including serious tree pathogens and agents of bluestain. The species in the complex are morphologically similar, but can be distinguished from each other based on morphology, biology, mating type studies and molecular data. At present, all the species in the complex are considered to be native to the Northern Hemisphere, most of them with a very wide distribution. Only a few sporadic reports of members of the complex are available from the Southern Hemisphere, where they are believed to have been introduced, including New Zealand, Australia, Chile, Brazil and Uruguay. This study aims to confirm the identity of isolates resembling O. piceae originating from three Southern Hemisphere countries, using mating compatibility and rDNA sequencing. Our results show that O. quercus is widely distributed throughout South Africa on both native and exotic hardwoods. O. quercus is also reported for the first time from Brazil, again from a native host. O. floccosum is reported for the first time from South Africa, but from an exotic Pinus sp. These results suggest that species of the O. piceae complex are common in the Southern Hemisphere, and that current views on the origins of especially O. quercus need to be reconsidered.

Phylogenetic trees of 16 Claviceps species were constructed based on alignments of 5.8S rDNA and the adjacent ITS1 and ITS2 spacers. Two highly supported clades were found: (1) C. paspali, C. zizaniae, C. grohii, C. sulcata, C. fusiformis, and C. purpurea; and (2) C. citrina, C. phalaridis, two unidentified Claviceps spp. (isolates PM and SG), C. sorghicola, C. gigantea, C. sorghi, C. africana, C. viridis, and C. pusilla. No relationship was found between the species placement and its morphological markers. The probe from C. purpurea gene cpd1 for dimethylallyl tryptophan synthase, the first enzyme of alkaloid biosynthesis, was hybridized to Pst I digested genomes of the above species under non-stringent conditions. Hybridizing DNA was present in all species of clade 1, although the signal of the C. paspali gene was weaker. In clade 2, only C. africana, C. gigantea, and C. pusilla gave weak positive signals. Colorimetric detection found small amounts of alkaloids in cultures of Claviceps sp. SG and PM but despite that, no cpd1 hybridizing bands were found. The occurrence of two major clades of Claviceps and their biogeography suggests, that the genus originates from South America and that the evolution of its species was influenced by comigration with their hosts and with the global climatic changes that influenced spreading of grass subfamilies.

The ITS regions of 14 isolates of endophytic fungi obtained from the root systems of four Woollsia pungens plants were sequenced and compared to sequences of Hymenoscyphus ericae, Oidiodendron maius and to sequences from other fungal taxa currently available in the GenBank and EMBL nucleotide databases. Sequence data suggested that six distinct taxa were present in the root systems of the four plants and that up to four of these were present in the root system of a single plant. One of the endophytes was identified as an Oidiodendron species, while most other isolates probably belong to the Leotiales.

The temporal pattern of release and dispersal of inoculum of plant and insect pathogenic fungi play an important role in the spread of disease. Airborne concentrations of primary and secondary conidia of Erynia neoaphidis released from the rose-grain aphid Metopolophium dirhodum were monitored at the edge of two winter wheat crops on IACR-Rothamsted Experimental Farm between May and September in 1996 and 1997. Hourly average temperature and humidity were recorded at each spore trap site and daily totals of rain and sunshine hours and daily average wind speed recorded about 1.6 km from the monitoring sites. No airborne conidia were found in 1996, but large numbers were trapped at the two sites in 1997. They were present from mid-June until early August, reaching peak concentrations on 17–18 July. Concentrations were usually highest during the night and in the early morning (01:00–07:00 h GMT) and were generally low during the day. On the 3 days when significant numbers of conidia were caught in the afternoon, daytime relative humidity was high (about 89%) and day-time temperature low (about 16 °C). Night-time conditions nearly always favoured the production of conidia. This suggests day to day variation in airborne conidium concentrations may be affected more by underlying biological factors than environmental conditions.

In vitro, antibiotic effects of the culture filtrates of 29 species of aquatic hyphomycetes against Gram-negative and Gram-positive bacteria, yeasts and hyphomycetes were studied. The culture filtrates of 38% of isolates were active in at least one of our tests for biological activity. Fifteen species inhibited growth of bacteria and four demonstrated antifungal activity. Filtrates of 35% of the isolates were antibacterial. About 50% of the isolates, those with the largest in vitro inhibitory activities (Dimorphosphora foliicola, Flagellospora sp. and Mycocentrospora sp.) were obtained from wood. Filtrates of Articulospora tetracladia were not only active against Gram −ve bacteria, but were also antiviral against bacteriophage T4 and influenza virus.

Lactarius angiocarpus sp. nov. is described from miombo woodlands in Zambia. It is the third sequestrate representative of the Russulaceae described from tropical Africa. Morphological characters and DNA sequence data support its placement in Lactarius subgen. Plinthogali. Molecular kinship analyses confirm a multiple origin of sequestrate Russulaceae spp. As none of the previously sequenced sequestrate Russulaceae spp. has been assigned to the Plinthogali clade, the placement of L. angiocarpus indicates an additional point of origin of derivative sporocarp types within the Russulaceae. Within the same subgenus, another tropical African sequestrate species, L. dolichocaulis comb. nov. (syn. Arcangeliella dolichocaulis) is recognized.

36 fungal β-tubulin sequences were analysed to study the evolution of this gene and the phylogeny of basidiomycetes. The analysis comprises a representative selection of all major lineages of basidiomycetous fungi and some selected ascomycetes for comparison. Intron positions vary between the different lineages, but seem to be conserved in the Hymenomycetes and Ustilaginomycetes. The most conserved regions seem to be highly susceptible for introns. Splicing and branching sites of the introns are more variable in basidiomycetes than reported from other fungal groups so far.

Basidiomycete monophyly was confirmed with our data in respect to the ascomycetes studied. By analysing amino acid sequences, the Hymenomycetes and the Ustilaginomycetes were resolved as monophyletic groups. The phylogeny within these two groups is similar to that obtained with other genes. Based on β-tubulin data Naohidea sebacea, Chionosphaera apobasidialis, Jaculispora submersa, Platygloea pustulata, Platygloea disciformis and Melampsora lini, representing the Urediniomycetes, are not resolved in most analyses. The early radiation of major basidiomycetous lineages seems to be reflected in the highly conserved β-tubulin gene.

Gaeumannomyces graminis var. graminis produces 1,8-dihydroxynaphthalene (DHN) melanin in its hyphal and hyphopodial cell walls. We isolated G. graminis mutants that were affected in their melanin biosynthesis. One was unable to synthesize DHN-melanin and, because it accumulated 2-hydroxyjuglone, a DHN melanin pathway shunt product, it is most likely to be defective in the reductase that catalyzes the conversion of 1,3,8-trihydroxynaphthalene to vermelone, the penultimate reaction in DHN synthesis. Genetic crosses with our wild-type strain indicated that this trihydroxynaphthalene reductase deficiency was the result of a single mutation. Another mutant constitutively synthesized DHN melanin and genetic crosses with our wild-type strain suggested that this heavily melanized mutant had a single mutation responsible for its phenotype. This mutant produced more melanin than the wild-type strain as measured by Azure A binding to melanin. The wild type and constitutively melanized mutant hyphae were more hydrophobic and more resistant to lytic enzymes, benomyl, restrictive temperature, and uv light than the non-melanized mutant, which also autolysed more readily. The non-melanized mutant was not more sensitive to heavy metal than the melanized strains. In addition, the non-melanized mutant was unaltered in pathogenicity to rice, whereas the constitutively melanized mutant was less pathogenic. The constitutively melanized mutant produced less extracellular lytic enzymes than the wild-type and the non-melanized mutant, which may explain its reduced virulence.

The infection of round-leaved mallow (Malva pusilla) leaves by Colletotrichum gloeosporioides f. sp. malvae was studied using light and confocal microscopy. Conidia germinated and produced appressoria within 24 h after inoculation. An infection peg arose from the base of the appressorium and directly penetrated an epidermal cell. An intracellular infection vesicle appeared beneath the penetration site by 48 h after inoculation. Large primary hyphae (LPH, approx. 4 μM diam.) emerged from the vesicle and grew intracellularly through several adjacent epidermal cells and then intercellularly between mesophyll cells. Epidermal cells infected by LPH maintained their viability, as shown by their ability to plasmolyse and accumulate neutral red stain. No visible disease symptoms appeared during this biotrophic stage of infection. Thin secondary hyphae (TSH, approx. 2 μM diam.) developed from LPH in 4–5 days after inoculation and were associated with the appearance of necrotic lesions. Host cell wall maceration was visible only during the necrotrophic stage. The duration of the biotrophic stage decreased as mallow leaves became older or were senesced by placing them in the dark. TSH and host cell necrosis developed as soon as 48 h after inoculation of the most senescent leaves, and TSH were observed directly emerging from the infection vesicle. Application of thiol reagents, such as glutathione and dithiothreitol, lengthened the biotrophic stage and delayed symptom development, whereas an antioxidant, ascorbate, enhanced infection and promoted symptom development.

The illudins are a family of fungal sesquiterpenes that have been studied as anti-tumor agents, and they also have antibacterial activity. Over a four-year period, 25304 fungal isolates (approximately 97% ascomycetes and 3% basidiomycetes), were screened for antibacterial activity against methicillin-resistant Staphylococcus aureus. Illudin-like compounds with antibacterial and cytotoxic activity against tumor cell lines were observed in 10 basidiomycete strains. The isolates were recovered from different types of substrata using indirect methods and only formed sterile mycelia in pure culture. The isolates were genetically related but not identical, based on PCR-based fingerprinting techniques. DNA sequencing of the ITS1-5.8 S-ITS2 region of the strains revealed that nine had identical sequences, indicating that they were conspecific. The sequence of the remaining isolate was 96.34% similar, suggesting that it was a closely related species. The D1–D2 region of the 25 S rRNA gene of the two strain types was also sequenced. Both sequences were 99.39% similar, and Coprinopsis gonophylla (syn. Coprinus gonophyllus) was the closest match for both. Strains were grown in pure culture on a rice-based medium that allowed the development of basidiomata from one culture of the main strain type, which was identified as C. episcopalis, a close relative of C. gonophyllus. Both species (or strain types) produced different types of illudin-like compounds. Three novel illudins (I, I2 and J2) were found to be produced by the cultures identified as C. episcopalis, while only illudinic acid was produced by the other Coprinopsis sp. The taxonomical relationships of the Coprinopsis species identified in this study with other illudin producers previously reported in the literature are discussed.