Cladistic analysis was performed on partial sequences from the nuclear-encoded small subunit ribosomal DNA. Four Halosarpheia spp., Lignincola laevis and Nais inornata were examined to determine the placement of these genera within the unitunicate ascomycetes. Two equally most parsimonious cladograms resulted from the analysis of the data. In a strict consensus of the two cladograms the taxa sampled did not form a monophyletic group, and Halosarpheia was shown to be polyphyletic. The marine Halosphaeriales formed a subclade with taxa from the Microascales. Data indicate that L. laevis and N. inornata may be congeneric, but other species of both genera require further study.

The phylogenetic relationship between Epicoccum nigrum and Phoma epicoccina has been assessed by means of sequencing of the ITS regions of rDNA. The analysis of the sequences from five isolates of E. nigrum and four of P. epicoccina suggests that both entities represent the same biological species.

Scytalidium thermophilum plays an important role in determining selectivity of compost produced for growing Agaricus bisporus. The objective of this study was to characterise S. thermophilum isolates by random amplified polymorphic DNA (RAPD) analysis and sequence analysis of internally transcribed spacer (ITS) regions of the rDNA, to assess the genetic variation exhibited by this species complex and to compare this with existing morphological and thermogravimetric data. RAPD analysis of 34 isolates from various parts of the world revealed two distinct groups, which could be separated on the basis of the differences in the banding patterns produced with five random primers. Nucleotide sequence analysis of the ITS region, which was ca 536 bp in length, revealed only very minor variation among S. thermophilum isolates examined. Several nucleotide base changes within this region demonstrated variation. Genetic distance values among type 1 and 2 S. thermophilum isolates, as determined by ITS sequence analysis, varied by a value of 0.005%. Molecular analyses carried out in the present study would suggest that isolates within this species complex exhibit genetic differences which correlate well with morphological variation and thermogravimetric data previously determined.

Growth in Neurospora proceeds by the highly polarised extension of hyphal tips and the formation of new tips (branches). While much attention has been devoted to tip growth, the mechanisms responsible for the decision to form a branch are unknown. One important issue is the degree to which the decision to branch is made independently by each tip, or is under the control of proximal regions of the colony. We address this question by comparing the lengths of branch intervals growing from one branch point. We find significant correlations between such intervals under nearly all circumstances tested. In contrast, we find no significant correlation between branch intervals adjacent in series. The strength of the correlation depends on the qualitative nature of the branches surveyed. Dichotomous/apical branching shows a high degree of correlation while lateral branching shows a weaker but still statistically significant correlation (this difference cannot be explained either by differences between the growth rates or distributions of lengths of branch intervals). We interpret the observed correlations to mean that the decision to form a branch is not independently controlled by the tip where the branch forms. Instead it is determined, at least in part, by factors at or close to the previous branch.

The effect of hydroxyl radical scavengers dimethyl sulphoxide, p-nitrosodimethylaniline, ethanol, benzoate, salicylate and thiourea was studied on sclerotial differentiation and growth of Sclerotinia sclerotiorum and Rhizoctonia solani. There is a correlation between both scavenger concentration and scavenger-hydroxyl radical reaction rate with delay and inhibition of differentiation at growth-noninhibiting scavenger concentrations. Growth-inhibiting scavenger concentrations further increased delay and inhibition of differentiation, and eventually stopped fungal growth, acting as antifungal antioxidant alternatives to traditional fungicides. p-nitrosodimethylaniline (with the highest hydroxyl radical reaction rate) was the most effective inhibitor of sclerotial differentiation and growth. The results strongly support the theory that oxygen free radicals induce differentiation in sclerotium-producing phytopathogenic fungi.

Four different primer pairs were designed for the preferential PCR amplification of fungal SSU rDNA directly from environmental samples. Most of the amplification products obtained from a reference collection of 46 wood-decomposing fungi could be separated by denaturing gradient gel electrophoresis (DGGE) using 1650 bp rDNA fragments produced by primer pair FR1+NS1. A relatively high level of resolution was also achieved using 390 bp products amplified with primer pair FR1+FF390. In contrast, the separation of amplification products obtained by the remaining two primer pairs (FR1+FF700 and FR1+FF1100; 700 bp and 1100 bp, respectively) was inadequate when applied to our fungal collection. Differentiation between all the species tested was achieved by combined analysis of the rDNA fragments produced by primer pairs FR1+NS1 and FR1+FF390. The DGGE analysis of environmental samples collected from Norway spruce stumps showed that the analysis of fungal DNA extracted directly from wood was usually in accordance with the investigation of mycelial cultures isolated from the same decay column. In some cases, however, disagreement was observed, which suggests that these two fundamentally different techniques present different views about fungal diversity. The investigation of fungal species profiles directly from environmental samples using DGGE analysis of PCR-amplified SSU rDNA molecules can be used to improve the detection of fungal groups that are difficult to cultivate.

275 isolates of Ophiostoma novo-ulmi, sampled across the southern Great Lakes region of North America from Wisconsin to Ohio in 1996, were analysed for vegetative compatibility (vc) types. Over 60% of the sample was a clonal vc component, comprising only two vc types (the AMSG and EUSG); the remainder of the sample was highly heterogeneous for vc types. Vc diversity was highest in Indiana and Michigan; close to where NAN O. novo-ulmi probably first appeared in North America. Each vc clone exhibited a uniform RAPD haplotype and a lower frequency of the A compared with the B mating type. This is consistent with the clones being mainly asexually spread, but with a potential for inbreeding, an unusual feature in a ‘genetic clone’. The heterogeneous component, however, exhibited diverse RAPD haplotypes and a near 1[ratio ]1 ratio of A[ratio ]B mating types, indicating that the mainly novel vc types of this component arise via sexual recombination. In Europe, dominant vc clones have been quickly replaced by novel vc types. In the Great Lakes region, however, vc clones appear to have survived for over 50 years despite a high potential for emergence of new vc types via sexual recombination. No differences were found between the clonal and heterogeneous components with regard to growth rate and pathogenicity, two important fitness parameters. Two other factors, a low level of selection imposed by deleterious d-factor viruses and a density dependent effect associated with vegetative incompatibility, may have favoured prolonged survival of the clones in North America.

The production and properties of Agaricus bisporus endoxylanase and β-xylosidase were studied in compost and liquid cultures. A. bisporus was grown on minimal medium with solka floc BW40, oat spelt xylan, Whatman CC41 microcrystalline cellulose, D-xylose and D-glucose as carbon sources. Endoxylanase activity and β-xylosidase activity were found during growth with complex carbon sources such as cellulose or xylan and no activity was present with sugars such as glucose or xylose. In compost culture endoxylanase activity increased during and after the button stage of fruit body development and decreased after fruit body harvest. The production of β-xylosidase differed in that it was only detectable during the button stage of development. Using activity staining and preparative IEF, three endoxylanase activities were separated from liquid cultures and three principal endoxylanase activities were separated from compost extract. The pH optima and IEF profiles of these activities were similar. Only one β-xylosidase activity was found in liquid culture, whereas two were found in compost culture; the IEF profiles of the compost activities differed markedly from those of the β-xylosidase found in liquid culture.

The genetics of somatic incompatibility in tetrapolar Collybia fusipes was studied using eight dikaryotic isolates collected from the wild and their experimentally derived progeny. Monokaryons from each isolate were all paired with the same unrelated monokaryon and also paired together in all combinations. The somatic compatibility of the two resulting sets of dikaryons was studied. Two different types of somatic incompatible interaction were observed, lightly or heavily pigmented lines developing between the two isolates. The dikaryons that had one nuclear type in common and one coming from sibling monokaryons were compatible in 7–27% of the cases, incompatible with a lightly pigmented interaction in 30–93% and incompatible with a heavily pigmented interaction in 0–53%. The results suggest that at least three to four loci control the somatic incompatibility in C. fusipes, one of them alone controlling the heavily pigmented interaction.

Colletotrichum acutatum is cosmopolitan and causes diseases in many crops including Hevea brasiliensis. It sporulated freely on PDA at 10–40 °C with peaks at around 15° and 25°. Self-inhibition of spore germination occurred at concentrations above 8×106 spores ml−1. Ultraviolet radiation (especially 254 nm) inactivated the spores. Spore germination was around 90% between 15–35°. Free water promoted spore germination but was not essential; high relative humidity (95%) was sufficient. Spores could also withstand temperatures up to 35°. It is concluded that the most favourable conditions for the spread of C. acutatum are prevalent in the major rubber growing areas in Sri Lanka throughout the monsoons.

Clonal isolates of Uncinula necator, the grapevine powdery mildew pathogen, were characterized by mating type, RFLP and PCR analyses. Five random genomic clones, consisting of three multi-copy and two low-copy sequences, were used as probes for hybridization. The oligonucleotides (CAC)5, (GACA)4, the core sequence of the M-13 minisatellite region and the consensus region from a plant intron splice junction (R1), were used as primers in PCR reactions. The DNA probes detected high levels of genetic diversity in U. necator, identifying 34 unique haplotypes among the 81 isolates analysed, with a Shannon diversity index of D′=0·85. In comparison, 15 haplotypes were resolved in PCR reactions with the four primers, resulting in a diversity index of D′=0·59. The overall diversity within the collection, by combining RFLP and PCR data, was D′=0·88, distinguishing 37 haplotypes. The RFLP analysis provided greater discrimination of individual haplotypes than did PCR. Phenetic analysis of the RFLP and PCR data separately and in combination revealed two broad phenetic groups. The average overall similarity between the two groups was 40%. Mating of isolates from the two phenetic groups resulted in viable ascospores. The combination of RFLP, PCR and mating techniques was effective in determining levels of diversity and provided insight into the genetic relatedness of U. necator isolates.

The hfa (high frequency of aneuploidy) mutants of Aspergillus nidulans carry conditional lethal (temperature-sensitive) defects which cause an increased frequency of aneuploids to be produced amongst their asexual progeny. When examined microscopically, most of the mutants grew and divided their nuclei at restrictive temperature, albeit more slowly than the wild-type, and aneuploidy was not attributable to an obvious cell cycle lesion. Exceptions were hfaB3 and hfaL1 which exhibited defects in nuclear division, although neither mutant arrested at a specific point in the cell cycle. Cells carrying hfaB3 contained only a single enlarged nucleus which was often transected (‘cut’) by the first septum and temperature-shift experiments showed that the mutation triggers aneuploidy by causing failure to properly exit mitosis. Although the hfaD1 mutant underwent nuclear division, it differed morphologically from wild-type by exhibiting a hyper-branching phenotype. The original hfaD1 isolate was shown also to carry a second unlinked mutation (designated hurA1) which confers resistance to hydroxyurea and partly alleviates the growth defects imposed by hfaD1.

The mechanisms responsible for controlling hyphal extension and branching are still poorly understood. We have investigated these processes by studying their dependence on temperature and nutrient concentration. Tip growth is highly responsive to temperature change, increasing linearly from 4 to 37 °C. Over this range of temperatures the branching pattern shows virtually no response. Likewise, varying nutrient concentration does not affect branch distribution. Colonies subjected to rapid extreme temperature downshifts (for example from 25 to 4°) display a strong and highly predictable branching response. There are three stages to this response. First there is an initial lag phase of growth without branching. After this, the growing tips display a series of tightly spaced, dichotomous branches dubbed starbursting. Following continued growth at 4°, tips enter a recovery phase, returning to normal branching frequencies with lateral branches. The strength of the response to cold is correlated with the ratio of growth rates before and after the downshift. The combined observations point to a homeostatic set point for branch distribution that compensates for temperature and nutrient concentration.

Evidence that chemical changes in litter exposed to elevated CO2 might alter the composition and function of fungal communities in soil is presented. Some potential effects of elevated atmospheric CO2 on the decomposition of Fraxinus excelsior leaf litter and the interactions of the colonizing fungi, growing singly or in simple associations in microcosms, were investigated. Fungal colonization was monitored by analysis of the ergosterol content of litter and specific PCR-amplified ribosomal DNA spacer products. After < 42 d, fungal colonization was less on litter with a higher C[ratio ]N ratio when obtained from seedlings grown in elevated CO2 (600 ppm). After triple-species inoculation percentage α-cellulose and, in some combinations, nitrogen content was reduced on litter from seedlings grown in elevated CO2.

The analysis of random amplified microsatellite (RAMS) markers of Phytophthora cactorum from strawberry showed that leather rot of strawberry fruits and crown rot are not caused by genetically different strains of this species. UPGMA-analysis showed that the populations of P. cactorum on strawberry differ between North America and Europe and that no clear genetic separation between isolates from different plants can be made among these populations. Slight morphological variation exists in the diameters of oogonia and oospores among the isolates but is insufficient for the identification of genetic groups or host specificity of P. cactorum isolates. Pathogenicity experiments proved that strains show a tendency towards host specialization.

The lipid and fatty acid compositions of Glomus intraradices and G. claroideum mycelia, extracted from quartz sand in a compartmentalized growth system, were analysed. The fungi were grown in association with Cucumis sativus and Trifolium subterraneum, respectively. For both fungi, the fatty acids 16[ratio ]1ω5 and 16[ratio ]0 dominated in the neutral lipid fraction, and 18[ratio ]1ω7 made up a significant part of the phospholipids. The fatty acids were used as estimators of the amount of neutral lipids and phospholipids of AM fungi as well as to calculate the biomass of different parts of their mycelium. The phospholipid content was higher in hyphae than in spores, whereas the opposite was observed for neutral lipids. In 3-mo-old G. intraradices mycelium, spores accounted for 90% of the external biomass, and calculations indicated that about 20% of the spore biomass consisted of neutral lipids. In both fungi the fatty acid compositions of hyphae and spores were similar regardless of the age of the mycelium.

Using the signature fatty acid 16[ratio ]1ω5 to calculate the distribution of AM biomass for a 2-mo-old mycelium of G. claroideum, we found that the fungal biomass was equally distributed between the external mycelium and the internal mycelium in the host root.

Over 600 basidiomycetes were isolated from indigenous forests and commercial Eucalyptus and Pinus plantations in South Africa. One hundred and twenty of the cultures were identified and biochemical tests were done to screen the cultures for characteristics that are favourable for biopulping. Most of the white-rot fungi previously associated with biopulping elsewhere in the world were also isolated in South Africa, as well as an isolate with uniquely regulated ligninolytic systems. Phanerochaete chrysosporium was found to be a natural coloniser of wood chip piles in South Africa.

A germination inhibitor in the spore matrix of Pestalotiopsis neglecta was strongly inhibitory to other mitosporic fungi, especially coelomycetes. Germination of conidia of P. neglecta was enhanced by an exogenous source of certain carbohydrates, especially pectin but also some sugars. Light was not important for germination or growth, and was not essential for reproduction, although the number of acervuli trebled when light intensity was increased from 210 μmol m−2 s−1 to 300 μmol m−2 s−1. The possible significance of these adaptations in the ecology of P. neglecta is discussed.

PCR-based methods to detect Verticillium chlamydosporium on infected plant roots were developed. Arbitrary ERIC primers and those based on rRNA genes, to identify fungi grown in pure culture, were unsuitable for DNA extracted from nematode-infested roots, because of interference by plant and nematode DNA. A novel method utilizing specific primers designed from an amplified and cloned fragment of the V. chlamydosporium β-tubulin gene was developed. Although it could not discriminate between different isolates of V. chlamydosporium, one primer set could identify the fungus on tomato roots infested with root-knot nematodes. The V. chlamydosporium β-tubulin sequence data showed close homology to sequences from plant endophytic Acremonium and Epichloë species and the saprotrophic Trichoderma viride.

Extracellular matrix (ECM) surrounding conidia and germlings of B. sorokiniana was studied using light microscopy (LM), scanning (SEM) and transmission electron microscopy (TEM). Conidial ECM surrounding dry-inoculated, ungerminated conidia was fluid-like and observed only using a cryo-preparation technique, suggesting that the material was water soluble. ECM enveloping germlings appeared fibrillar in LM, TEM and SEM but amorphous in cryo-SEM, indicating that the structure of the ECM is dependent on the water content of the matrix. Fibrillar ECM formed thread-like structures that extended over long distances on the substrate or towards neighbouring conidia and hyphae. TEM of germlings negatively stained with uranyl acetate revealed the presence of fungal fimbriae. The strong resemblance between the extending organization of fibrillar thread-like ECM structures and fimbriae suggested that fimbriae constitute a basic structural component of the ECM and serve as the aggregation centre for the other ECM components. Histochemical labelling revealed significant differences between ECM surrounding the fungus at different morphological stages. The germ tube ECM was labelled for both proteins and polysaccharides whereas germling ECM consisted of two layers: an inner rich in proteins and an outer composed mainly of polysaccharides. Furthermore, the newly released ECM localized on germ tubes and hyphal tips showed affinity for microspheres carrying any type of surface properties while hyphal ECM had affinity only for negatively charged microspheres. This together suggests that ECM after its release is subjected to structural changes.