The ability of the maize pathogen Fusarium subglutinans to metabolize benzoxazolin-2(3H)-one (BOA) and 6-methoxybenzoxazolin- 2(3H)-one (MBOA), important allelochemicals of Zea mays, was studied. Growth of mycelium was completely inhibited by 10 mM BOA and reduced by lower concentrations. In vitro the fungus completely degraded 1 mM concentrations of BOA and MBOA within 2 d. N-(2-hydroxyphenyl)-malonamic acid and N-(2-hydroxy-4-methoxyphenyl)-malonamic acid were identified as the main degradation products of these allelochemicals.

The effectiveness of surface sterilization methods were compared using root tissue of barley (Hordeum vulgare) colonized by two Chaetomium species in an aseptic plant growth system. The reliability of different sterilants to inhibit ascospore germination was tested. Ascospores on nitrocellulose membranes were either treated directly or ascospores adhering to axenic barley seedlings were treated on plant tissue. Inhibition of ascospore germination on nitrocellulose membranes was achieved with lower concentrations of sterilants than when the spores were on plant surfaces. A 10% peracetic acid treatment was necessary if experiments were conducted ad planta. The sterilants penetrated into epidermal root tissue and caused damage to this area, as shown by the vital dye DiOC7(3). It was concluded that effective surface sterilization techniques for ascospores adhering superficially to plant tissue were not appropriate for the detection of fungi growing within the epidermis.

The likelihood of development of penconazole resistance in Venturia inaequalis was demonstrated using chemical mutagenesis of a wild isolate of the pathogen not previously exposed to penconazole. Mutant strains displayed a high degree of resistance which was stable. There was significant variation in the rate of total lipid and ergosterol biosyntheses between the resistant and sensitive strains. Low uptake of penconazole due to energy-dependent efflux was found to be the mechanism underlying penconazole resistance in the mutant strains, which also exhibited negatively correlated cross-resistance to mancozeb.

Cronartium flaccidum and Peridermium pini are rust fungi occurring on two-needle hard pines. According to previous molecular and morphological analyses, they are very closely related despite differences in their life-cycles. In this study we showed that although a low level of genetic differentiation occurs among populations of both P. pini and C. flaccidum, there is no overall differentiation between the two rusts, and in this respect they resemble a single taxon. We also observed evidence for linkage disequilibrium occurring between different alleles of separate loci of Peridermium pini suggesting that its population structure would be clonal. This suggests that strains of P. pini would have originated as asexual or self-fertilizing host range mutants of C. flaccidum.

We developed a method for separating physiologically active intraradical hyphae of arbuscular mycorrhizal (AM) fungi from mycorrhizal roots, allowing the hyphae to be used for physiological and biochemical experiments. In the present study, the phosphate efflux from the intraradical hyphae in vitro was examined in relation to hyphal phosphatase activity. Onion seedlings (Allium cepa) were planted in the soil inoculated with Gigaspora margarita. Six weeks after transplanting, the intraradical hyphae were isolated from the mycorrhizal roots using plant cell-wall digestion enzymes. The hyphae were incubated briefly at 25 °C in a buffer solution (50 mM Tris/HCl, pH 7.4), then incubated for 2 h and gently shaken with various inhibitors. Phosphate efflux, the amount of phosphate released to the buffer, was analysed by EnzChek phosphate assay kit. Hyphal phosphatase activity was stained histochemically and the proportion of phosphatase-active arbuscules was examined for each inhibitor. Phosphate effluxes were to some degree reduced by all inhibitors used, while the phosphatase inhibitor, BeSO4, greatly reduced the efflux. The degree of inhibition in the arbuscular phosphatase by each chemical was closely correlated to the decrease in the phosphate efflux. These results suggest that hyphal phosphatase may be partially involved in the phosphate efflux process from intraradical hyphae.

It is difficult to accurately identify Mycosphaerella species associated with leaf diseases of Eucalyptus based on morphological characters, as there is considerable overlap between very similar species and subspecies, and isolation from the host is not easy. Thus, a PCR and RFLP assay based on the ITS region of nr DNA was developed for the rapid detection and differentiation of M. nubilosa, M. cryptica and two non-sporing unidentified Mycosphaerella species isolated from the foliage of trees in resistant and susceptible families of E. globulus in a seed orchard at Kinglake West, Victoria, Australia. The M. nubilosa primer pair MNF/MNR was highly specific. A PCR-RFLP system based on the primer pair MCF/MCR, coupled with two restriction enzymes (DdeI and Tru1I), differentiated M. cryptica, M. nubilosa, M. tasmaniensis and M. aff. vespa. One of the unidentified field-isolated Mycosphaerella species was identified as M. grandis on the basis of ITS sequence data while the other species remains unidentified. A PCR-RFLP system based on the primer pair U1F/U1R, coupled with the restriction enzyme StyI, differentiated between the two unidentified species. Unexpectedly, unlike isolation and culture studies, these assays detected M. nubilosa, M. cryptica and M. grandis in all single lesions examined on both juvenile and adult leaves, and on both highly resistant and highly susceptible E. globulus trees at this site.

Changes in pre- and post-fire biomass of hypogeous fungal sporocarps remaining at locations where an endangered mycophagous marsupial, the northern bettong (Bettongia tropica), had foraged, were assessed in fire-prone sclerophyll forest in northeastern Australia. Low to medium intensity experimental fires were set during the late dry season in 1995 and 1996 and post-foraging biomass of sporocarps (expressed as biomass of sporocarps remaining at recent B. tropica diggings) was measured at unburnt and burnt sites at approximately six-week intervals for a period of 14 months. Post-foraging biomass was significantly higher at burnt sites immediately following fire compared with control sites, solely due to increased biomass of hypogeous species belonging to the family Mesophelliaceae. Several months after fire, post-foraging biomass was significantly higher on unburnt sites compared with very low biomass on burnt sites. Twelve months after fire, the biomass on burnt and unburnt sites was not significantly different, having returned to biomass observed pre-fire. All evidence points toward mesophellioid fungi being greatly more available to bettongs on recently burnt ground, but fire may make several other sporocarp taxa considerably less available several months following fire.

The dynamics of competitive colonisation of necrotic cyclamen tissue by the plant pathogenic fungus Botrytis cinerea and the saprophytic fungal antagonist Ulocladium atrum were studied immuno-histologically, while sporulation was studied macroscopically. The effect of different time intervals between inoculation of both fungi on resource capture by each species was explored. Colonisation and sporulation were used as indicators for competitive resource capture and the effectiveness of biological control of B. cinerea, using U. atrum. Mycelial biomass and sporulation showed logistical time courses in both species, in monocultures as well as in mixed cultures. Final colonisation and sporulation levels were lowered by competition, indicating competitive resource capture. Analysis of the extent to which sporulation of either fungus could be reduced by co-inoculation with the other fungus at different times, showed that B. cinerea can be completely excluded by ‘early’ pre-inoculation with U. atrum, but not vice versa, indicating that U. atrum can exploit resources in the leaf that are not accessible to B. cinerea. A model of competitive substrate colonisation and resource capture was developed on the basis of the experimental results. Model results confirm that competition for resources provides a sufficient biological explanation for the dynamic interactions between the fungi. The model provides a tool to optimise dose and timing of U. atrum applications providing effective biological control of B. cinerea.

To identify Stagonospora avenae genetically, 53 isolates from Europe and North America were assessed by RFLP analysis and internal transcribed spacer (ITS) sequence polymorphisms in comparison with isolates of S. nodorum, S. arenaria and Septoria tritici. Our results also provided insight on possible phylogenetic divergence within the species currently identified by characteristic conidial morphology as S. avenae. Isolates from Europe, Canada, Minnesota and North Dakota showed little genetic variation. In RFLP and ITS sequence analyses, S. avenae isolates from New York and ATCC 12277 (origin unknown) had a closer relationship with barley-biotype S. nodorum, and S. avenae f. sp. triticea isolates of ATCC 26370 and ATCC 26377 from Minnesota were closely associated with wheat-biotype S. nodorum. Enzyme restrictions of ITS PCR-amplified products could facilitate the identification of Stagonospora on cereals.

Pronase E and tunicamycin, a putative inhibitor of protein glycosylation, strongly reduced the frequency of germlings adhering to smooth polystyrene and completely inhibited appressorium formation of adhering germlings without inhibiting germ-tube growth of the pathogen. Additionally, α-mannosidase or α-glucosidase, but not β-glucosidase or lipase, partly inhibited adhesion to smooth polystyrene and substantially inhibited appressorium formation by germlings adhering to this substrate. Infection structure formation was also reduced by the addition of either lectins binding to mannose or glucose residues (ConA and LCA), or by the addition of IgG. The presence of ConA- and IgG-binding (glyco)proteins among proteins obtained from the germination fluid, or from differentiated germlings, has been confirmed on Western blots. Removal of proteinaceous surface material by pronase E inhibited appressorium formation on smooth polystyrene. After the protease was removed, newly formed ConA-binding glycoproteins have been detected at the germ-tube apex with FITC-conjugated ConA. The (re)appearance of these glycoproteins was correlated with the time period when appressorium formation was observed. Interestingly, removal of proteinaceous materials from the germ-tube surface by pronase E, and also the addition of IgG had no inhibitory effect on appressorium formation on furrows in polystyrene.

Our results suggest ConA-binding surface glycoproteins present at the germ-tube apex as essential factors for infection structure formation on smooth polystyrene. In contrast, these glycoproteins are unlikely to be involved in appressorium induction over grooves in the same substrate.

Sclerotinia sclerotiorum produced erythroascorbate at levels dependent on oxidative growth conditions, developmental stage and strain differentiating capacity. The transition of S. sclerotiorum from the undifferentiated to differentiated state under high oxidative stress growth conditions, was accompanied by a shift in the ratio of reduced/oxidized erythroascorbate towards its oxidized form. At low oxidative stress, the reduced form of erythroascorbate predominated. In a non-differentiating strain of S. sclerotiorum, reduced erythroascorbate predominated over the oxidized form during growth. The reduced/oxidized erythroascorbate ratio in this strain was 5-fold higher than the corresponding ratio in the differentiating strain. The level of oxidative stress (lipid peroxidation) in colonies of the differentiating strain (one day before they differentiated) was 3-fold higher than that formed in colonies of the non-differentiating strain. Exogenous ascorbate exhibited its antioxidant activity in the differentiating strain by causing a dose-dependent decrease in lipid peroxidation, and proportional inhibition of sclerotial development. It is suggested that during its initial growth stages, S. sclerotiorum cannot decrease lipid peroxidation to levels found in the non-differentiating strain, and then forms sclerotia. We propose that oxidative stress triggers differentiation in sclerotium-producing filamentous phytopathogenic fungi.

Mycosphaerella vespa sp nov. is described from diseased leaves of Eucalyptus from south-eastern Australia. It is associated with small, circular to irregular lesions that have red-brown margins and amphigenous pseudothecia. These characters, in combination with ascospore morphology and germination, distinguish M. vespa from previously described species of Mycosphaerella on eucalypt leaves. Lesions associated with this pathogen were often found to be harbouring wasps and this observation is discussed.

The entomopathogenic fungus Metarhizium anisopliae was physiologically manipulated by culturing in liquid culture under a range of water-stress conditions (0.98, 0.97 and 0.96 water activity, aw) imposed by either PEG 200, KCl or NaCl at 25 °C to optimise blastospore production and quality in terms of accumulation of endogenous compatible solutes and germinability. Optimum spore yield occurred at 0.98aw, when PEG 200 was the solute. Blastospores produced under different water-stress levels were subjected to hypo-osmotic and iso-osmotic washing treatments before quantification of endogenous sugar alcohols (polyols) and germinability studies. Endogenous amounts of the low molecular weight polyol erythritol were greatly affected by culture age, water-stress and hypotonic or isotonic washing treatment. Endogenous erythritol in blastospores produced under 0.98 and 0.97aw increased with culture age. Osmoprotection (washing with isotonic PEG 200 solution) of blastospores produced under all water-stress conditions resulted in retention of higher amounts of erythritol. Increased endogenous retention of the high molecular weight polyol mannitol was also observed with an isotonic washing treatment but only in blastospores produced under intermediate water-stress (0.98 and 0.97aw). Germination of all treatments was >90% on a medium with freely available water. Under water-stress conditions (0.96aw) only 10% of unmodified blastospores washed with water germinated. However, those grown under modified water availability conditions (0.98aw) and washed with isotonic solutions had significantly improved germination (38–90%). Blastospores produced at [les ]0.97aw washed with isotonic solutions had decreased germination. These results could have significant implications for conserving ecophysiological quality of fungal biocontrol propagules during harvesting and have an impact on storage and formulation.

Laccaria bicolor and L. laccata are ectomycorrhizal basidiomycetes which fruit abundantly over 1 or 2 months in autumn and sometimes in summer. We investigated the phenology of fruiting of the various Laccaria spp. genets in a Douglas fir plantation in the Nièvre (France), in which all genets had been identified by molecular methods. During the fruiting period of the population, the genets did not fruit synchronously: some fruited early, others later, and some over the whole period. For some genets that survived over 2–3 years (1995–97), the fruiting phenology remained similar from year to year, even when fruiting occurred in summer. We took advantage of the presence of a L. bicolor genet, which had been used to artificially inoculate Douglas firs in this plantation, to assess whether the timing of fruiting was influenced by microsite differences in the plantation (soil, host tree, etc.). In summer 1997, fruit bodies of this genet appeared synchronously everywhere at the beginning of the fruiting period. We conclude that the timing of Laccaria fruiting may be genetically determined, rather than explained by the size and position of the genet or the environmental conditions. The consequences for population studies are discussed.

Thirty-five isolates of Gaeumannomyces graminis were tested for ability to infect wheat, rye and oats, and for DNA polymorphisms using nuclear ribosomal DNA (rDNA) RFLP patterns and RAPD analysis. In general, the cereal-attacking isolates could be readily assigned to the rye-attacking (R) or non-rye-attacking (N) subgroups of var. tritici, or to var. avenae, on the basis of either of these molecular approaches. A small number of isolates gave anomalous rDNA RFLP patterns, but could nevertheless be assigned to one of the three groups by RAPD analysis. Two related G. graminis var. tritici isolates (T1–1 and T1–2) clearly grouped with the N rather than the R var. tritici subgroup on the basis of molecular analysis but were pathogenic to rye, indicating that the ability to infect this host may have arisen more than once. An earlier phylogenetic study of Gaeumannomyces involving DNA sequence analysis of the internal transcribed spacers of the rDNA indicated that several oat-infecting isolates originally classified as G. graminis var. tritici could be grouped with var. avenae isolates. From the rDNA RFLP and RAPD analysis described here, however, these isolates appear to be intermediate between var. avenae and var. tritici, although it is not clear whether they represent evidence of inter-varietal sexual hybridization.

Phenotypes of species hybrids created from in vitro fusion of zoospores from Phytophthora nicotianae and P. capsici were characterized and compared. The species hybrids were created as part of a study of sources of genetic variation in populations of the parent species that are pathogenic over a similar range of plants. Four isolates of species hybrids proved to be similar to both P. capsici and P. nicotianae in relation to vegetative and reproductive morphologies. As in a previous study, DNA of P. capsici was detected more readily than that of P. nicotianae in all hybrid isolates. In the present study, DNA of P. nicotianae was detected in three of four hybrids by hybridization of RAPD-PCR products with species-specific DNA from P. nicotianae. By thermal denaturation analyses, DNA melting temperatures and GC contents of parent species and species hybrids were similar. The mean GC content of 47·2% was similar to GC contents reported for other Phytophthora spp. Additionally, the distributions of GC-rich regions of hybrids were more similar to the distribution in P. capsici than in P. nicotianae. By these molecular analyses, the hybrids were shown to be more similar to P. capsici than to P. nicotianae. Even though interspecific somatic fusion is likely to occur rarely under natural conditions, it could contribute to the genetic diversity of heterothallic species of Phytophthora.

Zoospore cysts of four isolates of Aphanomyces euteiches with different host-specific pathogenicity germinated in response to peptone, pea root extract, CaCl2 or gum arabic but, in tests on one isolate, did not respond to several individual sugars, amino acids or polysaccharides. A. euteiches showed differential responses to substances applied at different times in the encystment process. Gum arabic triggered germination only when added to motile zoospores, which it caused to encyst. CaCl2 triggered germination when added to motile zoospores or during vortex-encystment of zoospores, but seldom immediately (<2 min) after encystment. Peptone and pea root extract triggered germination when added to motile zoospores, during vortex-encystment or immediately after encystment, but not ca 20 min after encystment. In comparative tests, Phytophthora parasitica was less stage-specific than A. euteiches, because it germinated in response to gum arabic applied to motile zoospores, during encystment and immediately after encystment.

Zoospore taxis, encystment and germination of A. euteiches and P. parasitica cysts were recorded on pea roots by video microscopy. The speed of germination and pattern of sporeling development on roots were most closely matched by treatment of motile spores with gum arabic in pure culture. The behaviour of A. euteiches conformed to a previous model for Pythium and Phytophthora spp., in which zoospores encyst by recognition of root surface components (simulated by gum arabic), germinate autonomously by a calcium-mediated process (simulated by calcium treatment during vortexing), and the germ-tube produces an assimilative hypha (simulated by peptone or pea root extract).

The fungal pathogen responsible for Colletotrichum leaf disease of Hevea, develops three different disease symptoms: raised spots, anthracnose and papery lesions. These diseases have been attributed to Colletotrichum gloeosporioides (teleomorph Glomerella cingulata) and the fungi involved are morphologically indistinguishable. Twenty-five Colletotrichum isolates, which originated from three different disease symptoms, were characterized initially using RAPD markers. Two major RAPD profiles were detected which were related to the type of disease symptom developed. Colletotrichum isolates causing raised spot symptom were easily distinguished from isolates originating from either anthracnose or papery lesions. Restriction analysis of the PCR amplified 5·8S ribosomal DNA (rDNA), including both the flanking internal transcribed spacers (ITS) of representative isolates from the three different disease symptoms, also revealed two distinct RFLP patterns reflecting the same groupings as detected through RAPDs. Both molecular approaches suggested that there were two species of Colletotrichum associated with Hevea inciting the development of three different symptoms: Colletotrichum acutatum causing raised spot symptom, and C. gloeosporioides causing both anthracnose and papery lesions. This is the first record of Colletotrichum acutatum on Hevea in India.

The composition and concentration of airborne fungal propagules are probably determined by many interrelated environmental factors, such as temperature, relative humidity, atmospheric pressure, wind speed, rainfall and gas pollutants. The importance of these variables was assessed in an outdoor sampling survey conducted at regular intervals for 12 months at a single site in Turin. Samples were collected with a single-stage volumetric sieve sampler on potato dextrose agar supplemented with 15 mg l−1 streptomycin and 50 mg l−1 chloramphenicol.

Canonical Correspondence Analysis showed that the four ordination axes accounted for 93·5% of the variance in relationships between fungal entities present in more than 20% of samples and the environmental variables. The Monte Carlo permutation test demonstrated that the ordination was highly significant (P=0·01). The community's qualitative and quantitative composition mainly depended on the factors that have the greatest influence on Turin's climate, i.e., in descending order, temperature, relative humidity and rainfall. The relative importance of these environmental variables on different groups of fungi was assessed. Wind speed positively correlated with the fungi producing conidia of larger sizes.

We report the first successful culturing and confirmation of identity (via sodium dodecyl sulphate polyacrylamide gel electrophoresis, SDS–PAGE) of Serpula lacrymans (the dry rot fungus) derived from basidiomes and mycelia growing in the ‘wild’. The fungus was found growing on well decayed coniferous wood within the Narkanda region of the Western Himalayas at between 2800 and 3100 m above sea level (a.s.l.). The ‘wild’ habitat of the fungus is described, as is the isolation of Serpula himantioides also found in these regions. Temperature, osmotic potential, initial pH and air-current influenced the average colony extension rate of ‘wild’ and a selection of building isolates on 2% malt extract agar. The ‘wild’ and building isolates behaved as two separate cohorts; the ‘wild’ isolates appeared to be less affected by extremes of temperature, whereas building isolates extended more rapidly at the moderate microenvironmental regimes.