The ability of the preferentially saprotrophic fungus Armillaria cepistipes and the pathogenic A. ostoyae to capture fresh stump resources was investigated in managed Alpine Norway spruce (Picea abies) forests where both species occur sympatrically. The incidence of Armillaria species and genets as rhizomorphs in the soil as well as external and internal (heart rot) stump colonisation were determined in three comparable 1 ha plots. The results indicate that A. cepistipes and A. ostoyae have a very similar strategy to capture fresh stump resources. Both species produce dense networks of rhizomorphs in the soil enveloping living trees. After the felling of the trees, the stumps are rapidly captured through spread of mycelial fans in the cambial zone. Our study suggests that A. ostoyae is slightly more competitive than A. cepistipes in primary resource capture.

Hebeloma strains of arctic and temperature origin, grown at 22° or 6°, were assayed for wall-bound and extracellular acid phosphomonoesterase (pNPPase) across a temperature range 2–37°. Only when grown at 6° was a cold active extracellular pNPPase induced in all the arctic strains and most of the temperature strains tested. Such enzymes are suggested to be an adaptation to low soil temperatures, and are discussed in the context of ectomycorrhizal access to soil PO4− monoesters at low temperature.

Datronia caperata, a basidiomycete fungus, is one of the dominant polypore species found in neotropical mangrove forest fragments, where it is locally specialized on Laguncularia racemosa. We examined the genetic structure of D. caperata populations from four Panamanian mangrove forests using AFLP markers. Using five primer pair combinations, 145 loci were detected, 98.6% of which were polymorphic. Each of the populations showed a high degree of genetic diversity (Nei's h ranging from 0.146 to 0.223). Results from minimum spanning trees and Mantel tests showed little evidence for small-scale spatial structure within sites. A significant amount of total genetic variation was partitioned among populations (ΦST=0.21) separated by 10s to 100s of km, a considerably greater amount than has been detected in other mushroom and wood-decaying fungi sampled at equal or greater geographic distances. These results suggest that despite production of copious basidiospores capable of long distance dispersal, some homobasidiomycete fungi may be susceptible to genetic isolation due to habitat fragmentation.

Beauveria bassiana is a popular biocontrol agent used as ‘green’ pesticide in crop insect pest management. Chromosome number has been variously reported as five, six, seven and eight in this species. The range of chromosome number and the minimum chromosome number in this economically important fungus were assessed through telomere fingerprint analysis of a sample of 17 isolates from different and similar hosts and distant and same geographic origin. Genomic DNA digested with EcoRI, which has no cutting site in the telomere repeat sequence arrays was probed with a radioisotope-labelled (5′-TTAGGG-3′)8 oligonucleotide. The probe-hybridised regions appeared as discrete bands – each representing a telomere. The number of bands in each lane was counted and halved to arrive at the chromosome number of that isolate. The chromosome number varied from 5 to 10 in the different isolates. The telomere probe hybridised bands were also scored for presence or absence in a 0–1 matrix and a dendrogram based on similarities between the isolates was constructed using the NTSYS-pc ver. 2.02i software. The isolates showed very little similarity; the overall similarity was 14%. Only two isolates which were of diverse host and geographic origin showed 100% similarity. Isolates from the same epizootic that showed 43% similarity in their telomere fingerprints had 96% similarity in their RAPD (Random amplified polymorphic DNA) fingerprints with 10 primers. The genetic distances computed from any one DNA fingerprinting method thus do not reflect the true genetic similarities of the isolates. The frequency distribution pattern of the pair-wise similarities computed from telomere fingerprints hinted at the occurrence of recombination in this fungus. Telomere fingerprinting proved very useful in typing isolates since each of them was found to have a unique fingerprint. Isolates with the same chromosome number neither showed a distinct morphology or virulence character nor a close similarity in telomere or RAPD fingerprints to merit their subgrouping into a taxonomically relevant or practically useful unit.

We report the cloning and sequencing of a DNA fragment encoding a putative C2H2 zinc finger protein from Aspergillus nidulans. The gene was isolated by complementation cloning of a salt sensitive phenotype of the A. nidulans sltA1 mutant. A 3·8 kb PstI fragment that restored wild type salt tolerance contained one large open reading frame of 2202 bp. The predicted protein (StzA) from this reading frame comprises 698 amino acids and has three Zinc fingers along with a putative transcriptional activation domain rich in acidic amino acids. The corresponding sequence from a sltA1 mutant contains a premature STOP codon resulting in loss of the putative transcriptional activator in the C-terminal region. The Zinc fingers show conserved motifs with a number of transcription factors including CreA from A. nidulans and the human Wilm's tumour susceptibility protein WT-1.

Previous phylogenetic studies based on ITS sequence data have shown that Ophiostoma species with Sporothrix anamorphs include several species complexes. Isolates from oak in Poland and Hungary, which have previously been referred to as O. stenoceras, as well as isolates morphologically similar to S. inflata formed the basis of this study. Identification was based on sequences for the ITS region of rDNA operon and partial β-tubulin gene. Analyses showed that isolates from Poland and Hungary reside in a well resolved clade, separate from those in the O. stenoceras-complex. The morphology of these isolates was compared with those of strains in the O. stenoceras complex and S. inflata. Morphological differences in teleomorph and anamorph structures were found between the isolates from Poland and Hungary and those in the O. stenoceras-complex. Growth characteristics and the presence of the teleomorph in culture could be used to separate this fungus from isolates in the S. inflata-complex. The fungus from Poland and Hungary is described here as O. dentifundum sp. nov. It is phylogenetically most closely related to isolates of S. inflata, which represent four well defined groups based on morphology and DNA sequence phylogeny.

A somatic incompatibility system was demonstrated in Rosellinia necatrix (anamorph Dematophora necatrix) through barrage formation in pairings among mass fungal isolates from infected avocado trees in commercial orchards in southern Spain, as well as among single ascospore isolates derived from perithecia formed in infected roots. Results revealed the occurrence of high diversity in somatic incompatibility in the R. necatrix population in the study. The occurrence of R. necatrix perithecia in naturally infected avocado roots in the field suggests that such a diversity might originate from sexual reproduction. However, the possibility that other parasexual mechanisms demonstrated in this fungus may contribute to the genetic diversity can not be discounted.

The 5′ end of the large subunit (LSU) region and the entire internal transcribed spacer (ITS) region of ribosomal DNA were sequenced from 11 species or special forms of Melampsora on Salix, and three species on Populus. For all the species, except for M. larici-epitea and M. coleosporioides, the sequences in both the examined regions were identical within a species. Within M. larici-epitea, f. sp. larici-epitea typica and f. sp. larici-retusae shared the same sequences which slightly differed from that of f. sp. larici-daphnoides. In the LSU region, M. larici-epitea, M. capraearum and the stem-infecting form on S. viminalis shared the same sequence and the Far-Eastern species M. epiphylla differed from them only slightly (p distance 0.006), indicating that they may share a common ancestral lineage. M. amygdalinae and M. coleosporioides formed a distict group (bootstrap value 100% for combined ITS and LSU data). M. larici-epitea and M. ribesii-purpurea, both belonging to the M. epitea complex, appeared to be distinct. The molecular data also suggest that the differences in certain characteristics, such as the thickness of teliospore walls and host specificity, may have evolved relatively recently.

Downy mildew of oilseed poppy (Papaver somniferum) has become a serious disease issue for the Tasmanian poppy industry since its first record in 1996. Previous reports have reported the pathogen as Peronospora arborescens, which is differentiated from the related species P. cristata, also known to infect Papaver spp., by conidium dimensions alone. This study investigated the taxonomic status of the downy mildew pathogen, using both morphological characters and molecular analysis of the internal transcribed spacer (ITS) region of the ribosomal DNA (rDNA). The inherent variability of conidium dimensions made differentiation of species difficult. Sequence homology and phylogenetic analyses of the ITS region showed the pathogen to be more closely related to P. cristata than P. arborescens. It is therefore proposed that downy mildew of oilseed poppy in Tasmania be reattributed to the pathogen P. cristata. In addition to this work, PCR primers have been developed for the specific detection of the downy mildew pathogen in Tasmania.

The arbuscular mycorrhizal (AM) fungus Scutellospora reticulata (CNPAB11) was characterized using morphological, ontogenetic and molecular approaches. Spore ontogenesis was studied using Ri T-DNA transformed carrot roots and observations were compared with those published for eight other, pot-cultured, Scutellospora species. The sporogenesis of S. reticulata exhibited an unreported pattern of outer spore wall differentiation. In addition, Denaturing Gradient Gel Electrophoresis (DGGE), targeting the V9 region of the SSU nrDNA, was used to differentiate S. reticulata from 16 other Scutellospora species and results were confirmed by sequencing analysis. Phylogenetic analyses, using nearly full length SSU nrDNA sequences, grouped S. reticulata in a cluster together with S. cerradensis and S. heterogama, species that share similar spore wall organization and also possess ornamented external walls. PCR-DGGE and sequence analysis revealed intragenomic SSU nrDNA polymorphisms in four out of six Scutellospora species tested, and demonstrated that SSU nrDNA intragenomic polymorphism could be used as a marker to differentiate several closely related Scutellospora species.

The interactions between Pythium oligandrum hyphae and two types of sclerotia, i.e. the plano-convexoid sclerotium of Botrytis cinerea and the tuberoid sclerotium of Sclerotinia minor, were investigated by ultrastructural and cytochemical experiments. In the mycoparasitism of P. oligandrum, some differences in relation to sclerotium anatomy and the role of the rind layer in preventing invasion are documented. Both types of sclerotia showed neither alterations of the heavily melanised rind walls, nor direct rind wall penetration by P. oligandrum. This oomycete successfully entered B. cinerea sclerotia only through breaches at the junction of rind cells and corresponding to gaps in melanin deposits. On the other hand, none of these breaches was observed in the small sclerotia of S. minor, and P. oligandrum ingress in the sclerotia stopped at the inner rind layer. After the penetration of B. cinerea sclerotia by the mycoparasite, it extensively colonised the cortical and medulla areas by intercellular growth. The invaded tissues displayed pronounced alterations as well as some disorganisation of tissue in places. Colonisation was associated with severe chitin degradations of all host walls, which occurred even at some distance from P. oligandrum hyphae. The observation of wall thickenings in some P. oligandrum–hyphae suggests that the sclerotial cells constitute a harsh environment unsuitable for survival of the mycoparasite. These wall thickenings could be interpreted as P. oligandrum defence-like reactions.

Restrictions patterns of the ITS rDNA and a partial fragment of the β-tubulin gene were shown to help identify Phaeoacremonium species associated with diseased grapevines. PCR-RFLP markers revealed a high frequency of P. parasiticum, together with P. aleophilum, in fungi isolated from stems and trunks of mature diseased vines showing ‘hoja de malvón’ disease, and from young declining plants in Argentina. This distribution is different from that in vineyards in other parts of the world, where P. aleophilum is dominant. Indeed, this is the first report associating P. parasiticum with diseased vines.

SSU rRNA gene sequences of Anamylopsora pulcherrima (Anamylopsoraceae), Placopsis gelida, Trapelia involuta and T. placodioides (Agyriaceae) were determined and aligned with the corresponding sequences of 39 other ascomycetes. Phylogenetic analysis (maximum parsimony, spectral analysis) of these sequences placed the Agyriineae outside the Lecanorales and suggested a sister group relationship to Ostropales/Pertusariales. The resurrection of the order Agyriales is proposed based on molecular and morphological data, such as faintly amyloid asci opening by dehiscence. The sequence data support the two families Agyriaceae and Anamylopsoraceae to be accommodated within the order Agyriales. The Agyriales, Ostropales and Pertusariales can be placed in the class Lecanoromycetes.

The diversity of ectomycorrhizal mycobionts of Pisonia grandis (Nyctaginaceae) from coral cays in the Capricorn-Bunker group, Great Barrier Reef, Australia, was examined. Only two ectomycorrhiza morphotypes (brown and black) were identified in soil from seven cays and DNA from both morphotypes was subjected to ITS-RFLP and sequence analysis. The brown morphotype was present in soil from all cays but the black morphotype was only observed in soil from three cays. ITS-RFLP analysis showed that the brown and black morphotypes were formed by different fungal taxa, with the RFLP pattern for the black morphotype being consistent with that of the culture previously obtained from black ectomycorrhizal roots on Heron Island. Comparison with the GenBank database revealed that closest matches to both morphotypes were sequences for various Thelephoraceae (Basidiomycota), but the brown and black morphotypes had only 80% sequence similarity to each other. Neighbour-joining analysis of these sequences with sequences for other Thelephoraceae grouped the brown and black morphotypes in a well-supported clade with several Tomentella species, suggesting that both belong to this genus. The data are discussed in relation to ectomycorrhizal fungal diversity and the coral cay habitat.

Intraspecific variation of 25 Metarhizium anisopliae var. anisopliae isolates from various insect hosts and geographical origins was studied using RFLP analysis of the rDNA gene complex and the mtDNA. Heterologous and homologous mitochondrial and ribosomal probes were used to produce RFLP profiles of the isolates. The rDNA analysis allowed the differentiation of only seven isolates but provided evidence for the possible presence of introns in different sites of the nuclear rRNA-gene-complex. PCR amplification of the ITS1-5.8S-ITS2 rDNA region and the mtDNA regions corresponding to the SrRNA and the LrRNA, followed by restriction analysis of the PCR products underlined the conserved nature of these regions. By contrast, digestion with any one of endonucleases Hae III, Cfo I, Hpa II and subsequent RFLP analysis of the mtDNA, allowed the classification of the 25 isolates in 10 distinct groups. Double digestions using the restriction endonucleases Kpn I and Sac I led to maximum isolate differentiation, producing unique patterns for 16 isolates and subdividing the remainder into four groups of two or three isolates. This indicates the potential of mtDNA RFLP analysis as an excellent tool for fingerprinting M. anisopliae isolates and for studying genetic polymorphism.

Botryosphaeria spp. from seeds of the native afromontane forest tree species, Podocarpus falcatus and Prunus africana, in Ethiopia have been identified. This is achieved by combining anamorph morphological characters and ITS sequence data. From a relatively small sample, four Botryosphaeria spp. were encountered. Two of the species from P. falcatus and the one from P. africana have not been previously described. The two species from P. falcatus were represented by only one isolate each and are not named here. The morphology of their Diplodia and Dothiorella anamorphs is, however, characterised. The anamorph of the other Botryosphaeria sp. from P. africana is described here as Diplodia rosulata sp. nov. Furthermore, B. parva is identified from P. falcatus based on ITS phylogeny. This species is also an important pathogen of various commercially important tree species in Ethiopia and elsewhere. This study highlights the ability of Botryosphaeria spp. to infect seeds and the possibility that they might be distributed in this way. The study also contributes to recent attempts to stabilize the taxonomy of Botryosphaeria anamorphs, especially regarding Diplodia, which is currently in taxonomic disarray.

Isozymes of five Australian Ganoderma species were studied using cellulose acetate gel electrophoresis (CAGE) and PAGE. Phenetic analysis supported the distinction between the five species, which were separated by unbiased genetic distances of between 0.532 and 3.330. Estimates of heterozygosity and polymorphisms identified considerable genetic variability within populations and species. The putative saprotrophic G. australe, G. incrassatum and G. cupreum had the lowest genetic variability while the pathogen Ganoderma sp. Group 6.3, morphologically similar to G. lucidum, had the highest, followed by G. weberianum. Isolates of G. adspersum, G. applanatum from Europe and G. australe from Australia differed at most loci, thus the commonly accepted synonymy between G. australe and G. adspersum was not supported. Pectic isozymes alone were sufficient to distinguish three laccate (possessing a cutis surface consisting of a palisade of inflated hyphal ends) Australian species, G. weberianum, Ganoderma sp. Group 6.3 and G. cupreum, but they did not distinguish between the non-laccate G. australe and G. incrassatum. The laccate group contains species of greatest economic importance. Australian species were resolved by a single isozyme (glucose-6-phosphate dehydrogenase) using CAGE, demonstrating the potential for this method as a rapid diagnostic tool for identifying species of Ganoderma. This is the first reported population genetics based study of Ganoderma using isozymes.

Ericoid mycorrhizal fungi increase the ability of their host plants to colonise soils polluted with heavy metal ions, although the mechanisms are not clearly understood. Two mycorrhizal mycelia of Oidiodendron maius isolated from contaminated soil were previously shown to tolerate high concentrations of heavy metal ions in the growth medium. The influence of zinc ions on the secretion and activity of polygalacturonase (PG), an extracellular enzyme that hydrolyses the pectin component of the plant cell walls, was investigated because of their significance during saprotrophic growth. Two major PG isoforms expressed both in the absence and in the presence of increasing concentrations of zinc ions were identified. The PG isoforms were purified from the tolerant Oidiodendron strains as well as from ericoid fungal isolates from non-polluted soils. Addition of increasing concentrations of Zn and Cd ions to the purified enzymes resulted in an increase of PG activity of both tolerant and non tolerant O. maius isolates at doses below 1 mM. By contrast, at the same metal ion concentrations PG activity of a sterile isolate from non-polluted soil was unaffected or slightly inhibited. We speculate that the response of PG to heavy metal ions may have been a pre-adaptive factor for the colonisation of polluted soils by O. maius.

Sequences of the internal transcribed spacer (ITS) region of the nuclear ribosomal DNA of ectomycorrhizal fungi were analysed and a specific PCR primer pair for Tricholoma matsutake was designed. Using the primer pair, part of the ITS region of T. matsutake (400 bp) was amplified, but no amplified fragment was detected for other ectomycorrhizal fungi. PCR was performed on DNA extracted from mycorrhizas of T. matsutake and Shiro soil (extramatrical mycelium of T. matsutake and soil complexes) and the 400 bp fragments were also specifically amplified. This indicates that presence of T. matsutake at any of its life stages can be easily confirmed by PCR typing.

The diversity and frequency of leaf endophytic fungi occurring in naturally regenerating and planted stands of manuka were investigated at several sites. A restricted and characteristic range of species was isolated from all sites, although marked differences were noted between natural and planted stands. In natural stands a Phyllosticta species was the single dominant endophyte. This fungus appears to be host species specific, as it was not isolated from the closely related kanuka (Kunzea ericoides) in mixed, even-aged stands of manuka and kanuka. The frequency of the Phyllosticta sp. fluctuated greatly between individual trees across single stands. No obvious environmental or other gradients were recognized which could explain this patchy distribution. The Phyllosticta sp., along with the two next most common species in natural stands, Diploceras leptospermi and an unidentified coelomycete, were absent from most of the planted stands sampled, and at very low frequencies when present. Although several of the planted stands were close to natural stands and had been established for several years, the difference between them has been maintained. The most common endophytes in planted stands were Botryosphaeria and Alternaria spp. These species, found at low levels in natural stands, are likely to be ‘opportunistic’ endophytes. They are probably also characteristic of the epiphytic mycota, invading only the living leaves of plants under stress. The difference in leaf endophyte communities between planted and natural stands of manuka may be an indication of wider differences in the microbial communities of the respective stands, and could prove significant for site restoration projects. Such projects using nursery-raised manuka plants as a pioneer species may be establishing stands lacking much of the natural diversity of manuka communities for extended periods following establishment.