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.

Until now gametangia have not been obtained between paired European A1 and American A2 isolates of Phytopthora ramorum in vitro. Their production in artificial culture relies on interspecific pairings. Using P. drechsleri and P. cambivora testers, 51 of 110 P. ramorum isolates from across Europe were all shown to be A1s; while 32 of 38 American isolates from across California and southwest Oregon were shown to be A2s. However, these interspecific pairings are complex, unusually slow and unpredictable. A range of culture media and conditions are described that were tested, unsuccessfully, with a view to enhancing the efficiency of the interspecific pairings. In further tests, gametangia were obtained between A1 and A2 isolates of P. ramorum when juvenile, pre-chlamydospore producing mycelia were mixed together on carrot agar. The gametangia formed in 3–10 d, sparsely to frequently, initially only within the boundaries of the mixed inocula but subsequently in the extended mycelial growth. Chlamydospores were also produced. This inoculum-mixing method, though again sometimes unpredictable, should enhance efficiency of testing for compatibility types and facilitate further studies on whether the sexual outcrossing system of P. ramorum is functional. Differences between sexual reproduction of P. ramorum and that of other heterothallic Phytophthora species are discussed.

A considerable number of species of lichen-forming fungi have wide geographical distributions, but studies of their genetic variability are minimal. ITS rDNA sequences of 32 populations of Parmelia saxatilis from five continents revealed two monophyletic groups. β-tublin gene sequences from a subset of nine collections supported these conclusions. While the number of collections sequenced is limited, one monophyletic group (the Atlantic Population. AtP) was recognized as occurring in Arctic and Antarctic regions and also included collections from more atlantic sites. Samples from more mesic environments in the Mediterranean region belonged to a second monophyletic group (the Mediterranean Population, MeP). In addition, four subgroups were distinguishable within the Atlantic Population. Norstictic and protocetraric acids are reported from the species for the first time, the norstictic acid only being found in the Atlantic Population. Living thalli from the Atlantic Population were provenance-tested; specimens transported from the UK to central Spain where the Mediterranean Population occurs showed adverse symptoms after six months. These results demonstrate that there can be substantial large-scale genotypic variability within widespread lichen phenospecies, something which has implications for comparative ecological, physiological, and air pollution sensitivity studies as well as for lichen conservation.

RFLP, DNA fingerprinting and VCG methods were used to characterize fourteen Verticillium dahliae isolates collected from southwestern Ontario, Canada. The isolates were typed as not pathogenic to tomato (NP), race 1 (avirulent on cvs carrying the Ve resistance gene) or race 2 (virulent on Ve cvs). On the basis of RFLPs, RAPDs, and DNA fingerprints detected by hybridization to a dispersed, repetitive genomic DNA probe, the isolates were classified into five DNA types. Type I included two NP isolates. Type II included four race 2, and three NP isolates. Types III and V were represented by single race 2 and race 1 isolates, respectively. Type IV included one race 2, and two race 1 isolates. Vegetative compatibility was determined for selected NP, race 1, and race 2 isolates of each race type/DNA type combination. Isolates of the same DNA type were compatible, as were type II and III isolates (VCG 4B), and type IV (VCG 2A) and V isolates (VCG 2B). These data show a level of genetic diversity not previously identified in the V. dahliae tomato pathogen population, and suggest multiple origins for the Ontario race 2 pathotype.

Chaenothecopsis bitterfeldensis sp. nov. is described and illustrated from Bitterfeld amber dating back to at least 20 million years ago. There has been no previous report of the sporing perfect stage of any Ascomycota in amber. The fungus also represents the first fossil record of resinicolous fungi and of the Mycocaliciaceae. The fossilised specimen contains over 20 fruiting bodies arising from an extensive mycelium. There are also hundreds of detached ascospores, some of which had germinated before the resin was transformed into amber. The taxonomy and palaeontology of the fossil fungus is discussed and stages in its preservation are described. Also some general aspects in the biology of resinicolous Mycocaliciaceae are reviewed. The striking similarity of C. bitterfeldensis to some extant species from East Asia suggests that the present distribution of these fungi is a relict of an ancient Laurasian range.

A particular group of saprotrophic and ectomycorrhizal fungi produced reproductive structures successively on the forest floor after treatment with some nitrogenous compounds (N compounds) such as urea and aqua ammonia. These fungi were also observed after the decomposition of dead bodies or faeces of animals. Their growth in pure culture was examined in liquid media containing ten different N sources (four inorganic and six organic). The saprotrophic Amblyosporium botrytis, Tephrocybe tesquorum, Coprinus echinosporus and C. phlyctidosporus grew well on ammonium, some amino acids (asparagine, glycine), urea and bovine serum albumin, although A. botrytis did not grow on bovine serum albumin. Coprinus phlyctidosporus grew on nitrate as well as on the above N compounds. Peziza urinophila and Pseudombrophila deerata did not grow on any of the N compounds examined. The ectomycorrhizal Laccaria bicolor, Hebeloma vinosophyllum and Hebeloma sp. showed different patterns of nitrogen utilization: L. bicolor grew well on ammonium, nitrate and urea; Hebeloma spp. grew well on ammonium, nitrate, amino acids, urea and bovine serum albumin. None of the fungi tested grew on ethylenediamine or putrescine. The patterns of growth in relation to the nature of nitrogen supply were discussed in relation to the conditions of soil where these fungi occur.

Pestalotiopsis microspora is an endophyte associated with many plants including Taxus wallachiana. Isolates commonly produce taxol in liquid culture. Defining culture amendments to optimize taxol production by P. microspora is a critical step toward the realization of fungal taxol for treating human cancers. The lowering of phosphate and the addition of sodium benzoate in the medium increased taxol production. Sterol biosynthesis inhibitors, such as tebuconazole and triadimefon, dramatically increased taxol yields.

Serine proteinase activity was assayed during sporophore maturation and postharvest senescence in Agaricus bisporus. Serine proteinase accumulated largely in stipe tissue early during postharvest storage and in the later stages of maturation of unpicked sporophores. Western blot staining correlated with enzyme activity indicating de novo synthesis. The enzyme could not be detected by Western blotting in freshly harvested sporophores at an early maturation stage. Small increases in activity were detected in the cap tissue during the final stages of senescence, postharvest and in unpicked sporophores. The distribution of the serine proteinase in senescent sporophore tissues (high in stipe, low in cap) was confirmed by histochemical and immunologically stained tissue prints.

Intercellular hyphae of Cladorrhinum foecundissimum were observed in a dense layer surrounding the stele of cotton roots and were distinguished inside root hairs of plants grown in substrate containing the endophyte. No damage was observed in the epidermis of roots from infested substrate. At the blossom stage height of plants grown in substrate with the endophyte was 50% greater than in the controls. In the same group of plants growing in phosphorus-deficient substrate the endophyte doubled the phosphorus concentration compared to the control plants. In 25 day-old plants growing in substrate with high phosphorus content, the use of 32P showed a L value [available P (mg P 100 g−1)] 29% greater in soil colonized by the endophyte than in the non-infested substrate, but dry matter and phosphorus content in plants were not significantly different.

A region of the nuclear ribosomal DNA containing the internal transcribed spacer 1 (ITS1), the 5·8S DNA and the internal transcribed spacer 2 (ITS2) was sequenced in 12 Polymyxa graminis and P. betae isolates, with particular emphasis on P. graminis from peanut clump-infested areas of the Indian subcontinent and West-Africa. Four different sequences were obtained from the seven isolates on sorghum or pearl millet, which differed from four sequences previously published for Polymyxa species and obtained for P. graminis isolates on barley, oat and wheat originating from temperate areas in Europe and America (two distinct sequences), for isolates on rice from Colombia and for P. betae isolates on sugar beet from several origins. The sequence variations concerned mainly the composition and length of ITS1 and ITS2 regions. Phylogenetic trees built with the distinct sequences currently known for Polymyxa spp. using parsimony, maximum likelihood and neighbour-joining methods separated P. betae from P. graminis. Within P. graminis, the hierarchy of the clustering partially matched the host range and geographical origin of the isolates. These results confirm the great diversity within P. graminis that has already been observed in host range and temperature requirements studies, and provide new arguments for considering several taxa within the species. On the basis of the ecological requirements and rDNA sequences of distinct P. graminis isolates, five special forms are proposed: P. graminis f. sp. temperata, P. graminis f. sp. tepida, P. graminis f. sp. subtropicalis, P. graminis f. sp. tropicalis and P. graminis f. sp. colombiana.

Internal transcribed regions (ITS1 and ITS2) of 11 different fungi, i.e. 8 species of Alternaria, Nimbya gomphrenae, Stemphylium vesicarium, and Ulocladium botrytis, were sequenced. Their phylogenetic relationships with another 36 species in Pleosporaceae in GenBank were analyzed by parsimony and distance methods. These two methods positioned filament-beaked Alternaria as a monophyletic group discrete from the other members in genus Alternaria. A hypothesis proposing that the filament-beaked speices of Alternaria evolved a particular evolutionary route distinguishable from other species based on molecular evidence, the unique traits of morphological adaptation for liberation and their subsistent strategies has been discussed.

The genus Pilidiella, including its teleomorphs in Schizoparme, has a cosmopolitan distribution and is associated with disease symptoms on many plants. In the past, conidial pigmentation has been used as a character to separate Pilidiella (hyaline to pale brown conidia) from Coniella (dark brown conidia). In recent years, however, the two genera have been regarded as synonymous, the older name Coniella having priority. To address the generic question, sequences of the internal transcribed spacer region (ITS1, ITS2), 5.8S gene, large subunit (LSU) and elongation factor 1-α gene (EF 1-α) were analysed to compare the type species of Pilidiella and Coniella. All three gene regions supported the separation of Coniella from Pilidiella, with the majority of taxa residing in Pilidiella. Pilidiella is characterised by having species with hyaline to pale brown conidia (avg. length[ratio ]width >1.5), in contrast to the dark brown conidia of Coniella (avg. length[ratio ]width [les ]1.5). Pilidiella diplodiella, which is a pathogen associated with white rot of grapevines, was shown to be an older name for C. petrakii. To delineate species in the P. diplodiella species complex, isolates were also compared based on histone (H3) gene sequences. Analyses derived from these sequence data separated P. diplodiella from a newly described species, P. diplodiopsis. The new species P. eucalyptorum sp. nov. is proposed for isolates formerly treated as C. fragariae and associated with leaf spots of Eucalyptus spp. This species clustered basal to Pilidiella, and may represent yet a third genus within this complex. Pilidiella destruens sp. nov. is newly described as anamorph of Schizoparme destruens, which is associated with twig dieback of Eucalyptus spp. in Hawaii. A key based on morphological characteristics is provided to separate the taxa treated in this study.

Different isolates of a foliar endophytic species of Phyllosticta were isolated from different tropical tree species in India to examine genetic variation among the isolates. Internal transcribed spacer-restriction fragment length polymorphism (ITS-RFLP) analysis did not detect any variation among the isolates, suggesting that they all belong to the same species. Sequence data of the ITS region of ribosomal DNA (ITS1 and ITS2, including 5.8S rDNA) supported the identity of the present fungal isolates as P. capitalensis. These results show that P. capitalensis (teleomorph Guignardia endophyllicola?) is an ubiquitous foliar endophyte that can infect tree hosts from different families and habitats.

Morphological characters and genetic relationships were studied among 36 Verticillium isolates derived from different host species. Conidial length varied from 3.5 μm to 12 μm and the DNA content ranged from 0.028 pg to 0.078 pg with a bias towards large conidia and higher DNA content among the Swedish and German isolates from Brassica napus. Ability to produce nitrate nonutilising (nit) mutants was also assayed. Of all 36 isolates tested, eight V. dahliae isolates from non-Brassica hosts and two from B. napus, V. nigrescens from flax and V. albo-atrum from tomato generated nit mutants. To facilitate disease diagnosis, a species-specific PCR primer pair was developed, aiming towards a PCR-based analysis to confirm Verticillium wilt infection in Brassica plants. To identify useful DNA templates for the PCR primer design, random genomic clones from isolates VD12 and VD17 were used as probes in RFLP analysis. The RFLP results were also utilized in phenetic analysis and data from 29 isolates revealed three major groups. The first group consisted of seven non-Brassica V. dahliae isolates and the V. nigrescens isolate. The second group contained the two V. albo-atrum isolates and the V. dahliae isolate from red clover. Finally, the third group contained all the 18 isolates of Verticillium from B. napus. In the third group, two subgroups were identified, separating the three French isolates from the German and Swedish isolates. The dendrogram indicated that the isolates in the latter subgroup were closely related to each other. Based on the above mentioned criteria, we confirm that the isolates of Verticillium from B. napus in Sweden and Germany should be classified as V. longisporum.

Accumulation and transformation of tellurite (TeO32−) by a species of Fusarium and Penicillium citrinum was examined using both solid and liquid Czapek Dox medium. In liquid medium, tellurite partitioned into soluble and insoluble species, and in tellurite-containing (1 mM) liquid medium at pH 6, approx. 60% of added tellurite precipitated after 48 h. Experiments showed that in liquid medium containing 1 mM sodium tellurite, the Fusarium sp. accumulated a maximum of ∼0·6 μmol Te (mg D.W.)−1 after 48 h. P. citrinum accumulated a much lower amount of tellurium, ∼0·07 μmol (mg D.W.)−1 falling to <0·02 μmol (mg D.W.)−1 after 48 h. Both organisms showed marked differences in the pattern of pH change of the medium, with the pH increasing during growth of the Fusarium sp. in 1 mM tellurite to ∼pH 6·7 after 2 wk. In contrast, the pH decreased during growth of P. citrinum in 1 mM tellurite, to a level of ∼pH 2·7 after 2 wk. On agar medium, test fungi exhibited tolerance to high levels of tellurite (up to 100 mM Na2TeO3) and this was associated with blackening of the growing colonies as well as the surrounding agar. TEM revealed the deposition of large black granules, apparently in vacuoles, which corresponded with the reduction of tellurite to amorphous elemental tellurium. Precipitation of amorphous tellurium on and around the biomass was also observed and confirmed by energy-dispersive X-ray microprobe analysis. In addition to the reductive transformation of tellurite, Fusarium sp. also displayed transformation of tellurite into a volatile form. The production of volatile tellurium by Fusarium sp. occurred over the whole growth period and amounted to an average value of 7·8 μmol Te (∼0·16%) from 51 growth medium with an initial concentration of 1 mM Na2TeO3. Although P. citrinum also transformed tellurite to elemental tellurium, volatilization of tellurium was not detected. It is concluded that different mechanisms of tellurium transformation are species-dependent and can be influenced by physico-chemical changes in the medium, e.g. pH, which can affect tellurium speciation into soluble and insoluble forms and bioaccumulation. In view of the extremely small amounts of Te volatilized by the Fusarium sp., this process cannot be considered to be an important detoxification mechanism.

The status of anaerobic zoosporic (Chytridiomycota) fungi along the entire digestive tract of sheep was assessed both analytically and microscopically. Digest samples were taken from different segments of the digestive tracts of three newly killed sheep that previously had been used in experimental dietary studies. These digest samples were tested for the presence of rumen fungi by assessing the recovery of live fungi from the samples, direct observation of digested plant remains under the scanning electron microscope (SEM), and using a chitin assay as an estimation of fungal biomass. Live anaerobic fungi were recovered from the abomasum, small and large intestine, caecum and faeces of sheep, but not from the digest samples of rumen and omasum. However, SEM examination of the samples confirmed the presence of fungal structures from all of these organs. In the large intestine and caecum samples the observed sporangial structures were rounded and showed conspicuous surface pitting. Results of the chitin assay indicated that the anaerobic fungi might account for up to 20% of the total microbial biomass in the rumen of sheep. The results of this study support the view that anaerobic fungi may be present as a resistant stage in the lower reaches of the digestive tract.

Candida sake was cultured on nutrient yeast dextrose broth, which was diluted and/or modified by the addition of either glycerol, glucose to 0·96 or trehalose to 0·97 water activity (aw) to modify endogenous sugar alcohol and sugar content. Sugar alcohols (glycerol, erythritol, arabitol and mannitol) and sugars (trehalose and glucose) were extracted from the yeast cells and quantified using HPLC. Total polyol and sugar content varied significantly between treatments. The total intracellular concentrations in NYDB medium were significantly increased in NYDB+glucose media. The major intracellular polyols/sugars in cells grown on unmodified NYDB were arabitol, trehalose and glucose with small amounts of glycerol and erythritol. This was changed by reducing aw of the growth medium, particularly with glucose or glycerol. The major polyols in C. sake cells grown on glucose-modified media were arabitol and the low mol wt polyol glycerol, with smaller amounts of glucose. In glycerol-amended full strength normal and diluted media, glycerol was the major intracellular polyol with lower amounts of the other polyols and sugars. The viability of the yeast cells with modified polyols/sugars was significantly improved at lowered aw level (0·935 and 0·95) when compared to unmodified yeast cells. Improvements in water stress tolerance was better in yeast cells grown for 48 than 24 h, especially in those from NYDB modified with either glucose, glycerol or trehalose. Such modifications of endogenous reserves were also shown to preserve or improve the biocontrol potential of the yeast against Penicillium expansum rot of apples. Thus physiological manipulation of intracellular polyol and sugar content may provide a means for developing biocontrol agents with improved ecological fitness in field environments.

An assay based on the competitive polymerase chain reaction (PCR) was developed to quantify Glomus mosseae, an arbuscular mycorrhizal (AM) fungus, within plant roots. Using previously designed G. mosseae specific primers, a heterologous internal standard was constructed by amplifying Pseudomonas DNA under low stringency annealing conditions. Co-amplification of G. mosseae and internal standard DNA within leek root extracts provided accurate quantification of target DNA. Colonization of leek roots by G. mosseae was monitored in a comparative study by competitive PCR and microscopy, a conventional method of quantification. These two methods gave closely parallel data for G. mosseae colonization from three different inoculum levels over a 6 week period. Results indicate that competitive PCR is a sensitive and accurate method of quantification. The major advantage of competitive PCR over microscopy is that it can quantify specific AM fungi.

The ascomycetous genus Cordyceps accommodates endoparasitic species that attack arthropods or other fungi. Analyses of ITS nrDNA sequence data of 72 taxa from the teleomorph genera Cordyceps, Claviceps, Epichloë, and the anamorph genera Akanthomyces, Beauveria, Metarhizium, Hirsutella, Hymenostilbe, Paecilomyces, Polycephalomyces, and Tolypocladium assigned the taxa to four main evolutionary lineages not reflected in the current classification of Cordyceps. Ten subclades were recognized from separate analyses of data subsets. Judged from the ITS phylogenies, Cordyceps spp. with branched stromata were highly supported as a divergent lineage. Host specificity was found to be of limited phylogenetic significance, and several host shifts are suggested to have occurred during the evolution of Cordyceps. Similar ascospore morphology was not reflected in the phyletic groups, and closely related taxa showed large interspecific variation with respect to the number of segments in which the ascospores are divided. However, combinations of selected characters were found to delimitate some lineages, e.g. all Cordyceps spp. that attack hosts in the insect orders Coleoptera and Lepidoptera, and with non-immersed perithecia and clavate to acicular, brightly yellowish to reddish stromata, constituted a separate clade. Furthermore, all Cordyceps spp. with perithecia obliquely immersed in the stroma were recognized as a distinct monophyletic group. This clade is additionally characterized by the formation of anamorphs ascribable to the genus Hymenostilbe. The mycogenous Cordyceps spp. grouped in a separate subclade, interspersed by two cicadaen parasites and all Tolypocladium spp. except T. parasiticum. Tolypocladium and Beauveria were found to be polyphyletic. The included Claviceps and Epichloë taxa appeared to be derived within Cordyceps, thus making Cordyceps paraphyletic as suggested in other studies.