Development of specific DNA probes targeting rDNA internal transcribed spacer (ITS-1 or -2) sequences is described for the detection of strains representing uninucleate and binucleate Rhizoctonia spp. and Suillus bovinus. Discriminatory taxon/species-specific target sequences were identified following full length ITS sequence alignment of the test fungal sequences and those of other root associated pathogenic or mycorrhizal fungi. Both long (124–151 bp) and shorter (20–25 bp) probes were generated for assessment in Southern dot blot and liquid hybridization ITS capture-fragment length polymorphism assays. Fungal genomic DNA was presented as the target in dot blot protocols using the longer DIG (digoxigenin) labelled probes whilst the shorter 3′ biotin-labelled oligonucleotide probes were hybridized to PCR amplified full length ITS (ITS1-5.8S-ITS2) in both dot blot and liquid hybridization assays. The optimal hybridization temperatures for dot blot detection also produced maximal target specific signals in the liquid hybridization protocol. The latter protocol was found to be more discriminatory as target fungi were detected on the basis of combined probe hybridization-ITS capture and 5′ Cy-5 labelled ITS length polymorphism analysis (±5 bp) following denaturing sequencing gel electrophoresis in a ALFexpress DNA sequencer.

The vegetative life-cycle of Cymadothea trifolii (anamorph Polythrincium trifolii), causing sooty blotch of clover, is described using chemically as well as cryofixed and freeze-substituted samples. The pathogen enters the leaf through stomata and proliferates intercellularly. Nutrients are assumedly obtained via an interaction apparatus produced within the pathogen's hyphae, opposite to which the host cell is triggered to invaginate its plasmalemma. Rare attempts of ‘self-parasitism’ were also seen. Entering the conidial stage, stromata are laid down under the lower epidermis. The dying tissue above may explain the necrotic spots observed on infected leaflets. Foot cells in the conidial stromata produce thick-walled conidiophores, which grow sympodially. New conidiophores may grow into empty shells of old ones. Conidia are detached after pores between them and conidiophores have become plugged by organelles resembling Woronin bodies. Conidia are usually two-celled and their walls contain chitin and β-1,3-glucans as indicated by labelling with gold-conjugated wheat germ agglutinin and anti-β-1,3-glucan antibodies. Both conidiophores and conidia contain a structure which we regard as a new organelle with as yet unknown function.

Genetic variation was found between isolates of Botrytis elliptica from the USA and Taiwan based on 22 polymorphic markers from five RAPD primers. Among these 69 isolates, there were 43 different genotypes. One clonal genotype was found spanning Oregon and Washington, USA, and in a population from Hsinshe, Taiwan, clonal genotypes were frequent, demonstrating the importance of asexual spread of this pathogen. However, when only unique genotypes were considered, gametic disequilibrium among putative RAPD loci of the genotypes from Taiwan (33) or Hsinshe (23) was not detected, suggesting an underlying sexual recombination system for this population of B. elliptica. A weak level of cultivar specialization was detected among B. elliptica isolates from the Hsinshe population based on analysis of genetic distances between isolates.

Lewia avenicola sp. nov. (anamorph Alternaria sp.) is described. The fungus was isolated from oat grain in Norway. The fungus produced conidia within a month, and ascomata with fully mature ascospores within 8 months following inoculation, when stored on slants of synthetic nutrient agar (SNA) at 4 °C in darkness. A comparison with other known Lewia species is made, and a key to the five known Lewia species is included.

Candelabrum desmidiaceum sp. nov. and C. clathrosphaeroides sp. nov. are illustrated and described. Morphologically, they represent a new type of conidium development previously unknown in Candelabrum: the formation of a basal plate by dicho- or trichotomous branching of the conidial body in combination with branches growing upwards at the outer margin of the basal plate which are distinctly verrucose outside, forming a hollow sphere enclosing an air bubble in the centre. In Austria, both species were found mainly on submerged beech leaves, but in the U.S.A., C. clathrosphaeroides was isolated from submerged leaves of many different deciduous species, indicating a wider host range. Both species are aero-aquatic fungi, and C. clathrosphaeroides, at least, is widely distributed in temperate climates. A key to the species of Candelabrum is provided.

The utilization profile of 49 carbohydrates, based on API 50 CH biochemical tests, was used for the identification and the discrimination of 75 isolates of Beauveria and Tolypocladium. The data were used to construct a most parsimonious tree in which Beauveria and Tolypocladium were separated into two main branches. Although the Beauveria isolates could not be separated at the species level, eight different clusters corresponding to the eight Tolypocladium species studied were revealed. The reproducible carbohydrate utilization profiles of B. bassiana, B. brongniartii, T. cylindrosporum, T. niveum, T. geodes and T. tundrense have permitted the construction of an identification key that could serve as a tool to complement the morphological description of selected Beauveria and Tolypocladium isolates for their correct identification at the species level. The API 50 CH method is time-saving and can complement effectively more classical techniques for the discrimination between Beauveria and Tolypocladium and for the taxonomy of Tolypocladium at the species level.

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 South African Cape Floral Kingdom is the world’s smallest and most diverse plant Kingdom, but is greatly under threat from urbanisation and agriculture. It covers only 90000 km2 and has a Mediterranean climate. This area has a long history of botanical studies, but little research has been undertaken on the associated fungi. Therefore, a biodiversity study of the fungi has been initiated. The catalyst for this study has been the diversity of unusual fungi encountered whilst studying the fungal pathogens of Proteaceae. These pathogens include species, or entire genera of fungi that are specific to genera of Proteaceae. The diversity and uniqueness of the Proteaceae pathogens may indicate a similar diversity in saprobic fungi in this unusual habitat. Therefore, microfungi associated with genera of Proteaceae in various habitats are being systematically sampled over a two-year period. The initial results of this study are presented.

Colletotrichum dematium is the cause of a new fungal stem disease on cowpea in South Africa. The pre-penetration and infection process of this fungus on cowpea stems was studied by light microscopy and SEM. Conidia began germinating at 6 h post inoculation (hpi), forming appressoria directly or at the ends of germ-tubes. By 14 hpi appressoria had melanized and direct penetration of host tissue had begun. At 20 hpi infection vesicles formed in epidermal cells. Thick, knotted primary hyphae formed from these vesicles and entered adjacent cells. At ±40 hpi C. dematium produced secondary hyphae which were highly branched, and grew extensively inter- and intracellularly. After approximately 48 hpi light brown lesions appeared on the stem, associated with the invasion of secondary hyphae into cells and cell necrosis. Acervuli with one or two melanized setae were visible on lesions by 70 hpi.

A new species of Chaetomium, C. cuyabenoensis, is described from the Ecuador rain forest. It is similar to Chaetomium longicolleum but differs by having smaller and, in face view, strongly limonifom to quadrangular ascospores.

The electrophoretic karyotype (EK) of 24 strains of Botryotinia fuckeliana was resolved to determine the extent of chromosomal polymorphism among them. Based on EK variation, nine different profiles with a number of bands ranging from 5 to 8 were found. Those profiles were not related to the origins of the strains, hosts, or year of isolation. Inheritance of chromosomal bands was studied by analyzing the EK of the progeny from crosses between sexually compatible B. fuckeliana strains that had different karyotype. The EKs of 44 monoascospore strains showed 12 new EK profiles, suggesting that chromosomal rearrangements generated after meiotic recombination provides EK variability in this fungus. Also, we report here that the degree of EK variability indicates that sexual reproduction under natural conditions might be quite important in the life cycle of B. fuckeliana. Hybridization analysis showed that, in most strains, the rDNA is located in both a chromosomal band of high molecular weight and different size depending on the strain and a band of low molecular weight, revealing evidence for chromosome rearrangements in this plant pathogen.

Phylogenetic analyses of nucleotide data from the ITS and 5.8S rDNA gene region were performed on 37 strains of Onygenales representing all the accepted species of Auxarthron (9 strains), several species of Amauroascus (16 strains), and morphologically similar genera (3 strains of Amaurascopsis, 1 of Apinisia, 1 of Shanorella, 1 of Mallochia, 2 of Nannizziopsis, 1 of Pectinotrichum, and 3 of Uncinocarpus). The inferred phylogeny provided strong support for the monophyly of Auxarthron. We propose the new combinations Auxarthron kuehnii and A. pseudoreticulatum combs nov. and provide a key to the accepted species of Auxarthron. Although this study demonstrated that Amauroascus is polyphyletic, relationships among the members of that genus could not be resolved in phylogenies inferred from the analyses of ITS sequences; the new combination A. queenslandicus comb. nov. is also proposed. Molecular data suggested that Amaurascopsis reticulatus, characterised by the absence of discrete ascomata and prominently echinulate ascospores, can be accomodated in Mallochia.

Unidentified isolates of Cylindrocladium with 1–3-septate conidia and clavate vesicles were compared with C. theae, C. colhounii var. colhounii, C. colhounii var. macroconidiale, C. gracile, C. pteridis and Calonectria gracilis. Ribosomal DNA (rDNA) of these isolates were digested with the restriction enzymes EcoR I, Hind III and Xho I, and Southern analysis performed with the 6·3-kb rDNA repeat unit of Neurospora crassa as DNA probe. Based on differences in general morphology, supported by their rDNA restriction fragment length polymorphisms, isolates of all species and varieties could be distinguished. Furthermore, the Cylindrocladium anamorph of Calonectria gracilis was shown to be distinct from Cylindrocladium pteridis and C. gracile. The name Cylindrocladium pseudogracile sp. nov. is, therefore, proposed for the undescribed anamorph of Calonectria gracilis.

Specimens of Daldinia (Xylariaceae) from around the world possessing large ascospores were studied for teleomorphic and anamorphic morphological characters and compared with authentic material of D. grandis. A culture made from a specimen of D. grandis collected from Ecuador produced stromatic structures, but no conidiogenous structures referable to known xylariaceous anamorphs were observed. D. grandis is reconsidered and viewed as a species of warmer climates in the Americas. Three new species are recognised from new combinations of anamorphic and teleomorphic characters: (1) D. novaezelandiae sp. nov. from New Zealand, a fungus obviously related to D. bakeri, which is also reported from that country; (2) D. dennisii sp. nov., from Australia and New Zealand, of which two varieties are erected; and (3) D. loculatoides sp. nov., from the UK and Canada with affinities to D. loculata. The type of Sphaeria durissima was identified as D. loculata. Notes on further Daldinia spp. are included.

A study was designed to isolate and identify the endophytic mycobiota in leaves and twigs of Rubus spectabilis and R. parviflorus, the long-term goal being biological control of these plants with endophytes. Endophytic fungi were isolated from healthy-looking leaves and twigs collected from each species on Vancouver Island, British Columbia, Canada. More than 90% of the leaves of both species were colonised by endophytes. In addition, one third of the twig samples of each species were also colonised. Most isolates belonged to Phomopsis, and five species were identified and characterised according to morphology of the cultures and fructification structures. All Phomopsis species occurred on both Rubus species, but, one primarily colonised R. parviflorus, whereas another preferentially colonised R. spectabilis. The suitability of the isolated endophytes to inhibit growth of R. spectabilis and R. parviflorus, which compete with conifers in reforestation sites, is discussed.

A thermostable intracellular lipoxygenase from Thermomyces lanuginosus was purified to sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) homogeneity by ion-exchange chromatography on diethylaminoethyl (DEAE)-Sepharose, ion-exchange chromatography on carboxymethyl (CM)-Sepharose and Phenyl-Sepharose hydrophobic interaction chromatography. The molecular weight of the enzyme was estimated to be 100 kDa by SDS-PAGE. The lipoxygenase exhibited maximal activities at pH 6.0. The optimum temperature for the activity was 55 °C. The enzyme was thermostable at 50 ° with half-lives at 60 ° of 20 min and 70 ° of about 7 min. The lipoxygenase activity was completely lost by heating at 80 ° for 5 min. The lipoxygenase catalyzed the oxidation of linoleic acid into 13-L-hydroperoxy-9,11(Z,E)-octadecadienoic acid(13-HPOD) as the major product. The enzyme had an apparent Km of 8.5 μM and Vmax of 10.8 μmol mg−1 min−1 with linoleic acid as substrate.

Penicillium verrucosum is one of the main producers of ochratoxin A (OA) in agricultural commodities. To forecast the risk of OA contamination, there is a need to improve our knowledge on the ecology of P. verrucosum in the field. Dilution plating on ‘dichloran yeast extract sucrose 18% glycerol agar’ (DYSG) offers a simple and very sensitive method of detecting P. verrucosum propagules in soil. The properties of DYSG are illustrated in a suspension mixture experiment and confirmed in a soil mixture experiment. In the latter, P. verrucosum could be detected in conidial concentrations below 200 colony forming units (cfu) g−1 soil even when it constituted no more than 0·3% of the cfu. Furthermore, the DYSG method can be used to estimate the abundance of P. verrucosum propagules in soil with good precision. In some of the analysed cases, however, it was necessary to use appropriate mathematical models to treat results with high numbers of cfu on the Petri dishes.

The geographical distribution and ecological habitats of the Pleurotus eryngii complex in Israel are described from a study of 60 genotypes from 10 populations. Both MEA and PDA media were suitable for growth of all genotypes at all temperatures tested (4–37 °C). There was a high correlation between a coefficient of growth and the mean colony diameter growth rate. All genotypes reached the maximum growth at 27°. Growth rate at 30° was greater than at 19°. We found highly significant (P < 0.0001) variability among growth rates of Israeli genotypes, indicating ecogeographical differences among populations. The growth rate of 12 European genotypes from the Ukraine and Slovakia was higher than most of the Israeli genotypes when grown at 27° (non-stress conditions), but Israeli genotypes tolerated 37° (stress conditions) better, and recovered faster than European populations when returned to 27°. We conclude that Israeli genotypes are better adapted to hot and dry climates than European genotypes. Based on multiple regression analyses, strong correlations were found between relative humidity and rainfall parameters, but not temperature, of the locations from which Israeli isolates were collected, and their growth rates. This indicates that, paradoxically, relative humidity and rainfall have a stronger effect than temperature on the adaptability of this complex to different environments.

We determined the occurrence of birth rust fungus (Melampsoridium betulinum) within mature mountain birch (Betula pubescens var. czerepanovii) trees and compared the airborne urediniospore concentrations with overall rust infection levels over five successive years. Fifteen groups of three trees 3–5 m high were chosen from a homogenous mountain birch stand. Each group contained a tree with ‘low’, ‘moderate’ and ‘high’ infection covering the natural range of infection levels. The difference in rust classes remained constant throughout the study; trees in class ‘high’ consistently had always highest infection levels and trees in class ‘low’ always the lowest infection levels. Infection levels remained constant between years in ‘low’ class, while the infection levels varied significantly between years in ‘moderate’ and ‘high’ classes. The incidence of disease caused by M. betulinum varied from year to year and was correlated with the total airborne urediniospore counts. The data indicate that trees differ in resistance to birch rust and that infection levels of susceptible compared to resistant trees are more dependent on environmental conditions.