This issue of Mycological Research News features: In this issue; and Changing the biological clock in Neurospora crassa.

Original papers in this number include morphological and molecular investigations on: Endophyllum species on asteraceous plants; Melampsora rusts on Salicaceae; myxotoxin producing Fusarium chemotypes; host specificity and nutrient uptake in Eudarluca caricis; Ampelomyces populations on powdery mildews; salt marsh populations of Claviceps purpurea; the classification of Omphalina foliacea; and endophytes in different genera of native Australian orchids.

A gene encoding a malic enzyme involved in the anaerobic growth of Mucor circinelloides is characterized. In the ericoid mycorrhizal Hymenoscyphus ericae, one paper shows how carbon availability affects nitrogen source utilisation, and another the effects of copper toxicity in relation to phosphatase activity.

The colonization of wood by airborne spores has been studied in experimental forest plots, and the climatic and altitudinal factors correlated with spore catches of Alternaria and Cladosporium determined.

Early development of Morchella fruit bodies is described and documented by scanning electron microscopy.

The following new scientific names are introduced in this issue: Endophyllum dimorphothecae sp. nov.; E. elytropappi (syn. Aecidium elytropappi) comb. nov.

Endophyllum osteospermi is an autoecious, endocyclic rust fungus, which has only been recorded on Chrysanthemoides monilifera ssp. monilifera (Asteraceae, Calendulae), a perennial woody shrub. Both organisms are indigenous to South Africa. Because E. osteospermi is being considered for release in Australia as a biocontrol agent against C. monilifera ssp. monilifera, it was necessary to determine its host range and natural distribution in South Africa. To address this, natural stands of Chrysanthemoides species, as well as other South African asteraceous plants, were monitored for E. osteospermi between 1992 and 2003. A morphological and molecular comparison of specimens referable to Endophyllum was undertaken. Based on these results, E. osteospermi was recorded on C. monilifera sspp. monilifera, pisifera, rotundata, canescens, and subcanescens, C. incana, and an undescribed taxon. E. osteospermi was also recorded on Osteospermum ciliatum, O. polygaloides, and O. potbergense. Furthermore, a closely related but previously undescribed species, E. dimorphothecae sp. nov. is described on Dimorphotheca cuneata. Aecidium elytropappi is transferred to Endophyllum as E. elytropappi comb. nov., being recorded on Elytropappus rhinocerostis and Stoebe plumosa. This study shows that in South Africa E. osteospermi is restricted to a small group of related plant species in the Calenduleae. This rust is therefore considered suitable as a candidate agent for the biocontrol of C. monilifera ssp. monilifera, and pending the results of host specificity testing, would most likely be safe to introduce into Australia.

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.

Fusarium graminearum and F. pseudograminearum are important plant pathogens in New Zealand and around the world. Headblight and crown rot diseases of cereals caused by these species are responsible for large economic losses due to reduction in seed quality and contamination of grain with tricothecene mycotoxins. In the current study we have used two different molecular phylogenetic approaches, AFLPs and gene genealogies, to gain insight into the evolutionary relationships between F. graminearum, and F. pseudograminearum in New Zealand. The worldwide genetic diversity of F. graminearum clade is represented by at least eight biogeographically distinct species (previously designated as lineages of F. graminearum). Our analysis demonstrated that this clade is represented by F. graminearum (=F. graminearum Lineage 7) and F. cortaderiae (=F. graminearum Lineage 8) in New Zealand. Through our analysis we also confirm the presence of F. pseudograminearum in New Zealand as a first record for this organism. Information on species is necessary for preventing the inadvertent intercontinental introduction of genetically unique foreign pathogens associated with world trade. The ability to place species information into a worldwide context enabled postulation that the New Zealand representatives of F. graminearum clade originated from at least two regions, and probably on at least two hosts. Correlation of species descriptions with biogeographical and host information revealed evidence for co-localisation of F. graminearum clade species with potential for genetic outcrossing in the field. Mycotoxin analysis showed F. graminearum (=lineage 7) isolates produce either nivalenol (NIV) or deoxnivalenol (DON). In contrast, F. cortaderiae isolates produced only NIV. These findings support earlier observations that mycotoxin production in the F. graminearum clade is not species specific, but suggest maintenance of chemotype diversity through speciation may have been restricted to a subset of species.

Eudarluca caricis is assumed to be a nonspecific mycoparasite of rust fungi. The evidence for its mycoparasitism has rested on constant association with uredinia. In this study, stable isotopes provided additional evidence of mycoparasitism, as E. caricis was enriched with 15N relative to its associated rust fungus, as were parasites and mycoparasites generally with respect to their hosts. Host specificity was directly tested in inoculations in the greenhouse. Isolates of E. caricis from Puccinia on two Eurasian grasses (i.e. Holcus lanatus and Phalaris arundinacaea) did not infect Melampsora on Populus that, in contrast, was successfully infected by a poplar isolate of E. caricis. An isolate from M. medusae on P. deltoides infected a significantly greater percentage of uredinia of M. medusae on P. deltoides than uredinia of M. occidentalis on P. trichocarpa. The host specificity of the three isolates was reflected in their divergence in a phylogenetic analysis based on ITS sequences. Interestingly, the analysis revealed that mycoparasites of rust and powdery mildew fungi have evolved from a common ancestor.

Pycnidial fungi belonging to the genus Ampelomyces are the most common natural antagonists of powdery mildews worldwide. During a study of the interactions between apple powdery mildew (Podosphaera leucotricha) and Ampelomyces mycoparasites, 52 new Ampelomyces isolates were obtained from P. leucotricha and, in addition, 13 new isolates from other species of the Erysiphaceae in four European countries. Their genetic diversity was screened using single-stranded conformation polymorphism (SSCP) analysis of the internal transcribed spacer (ITS) region of the ribosomal DNA (rDNA). For comparison, 24 isolates obtained from genetic resource collections or other sources were included in this study. Based on the ITS-SSCP patterns, the isolates were placed in eight groups. The isolates belonged to two types based on their growth in culture. The faster-growing and the slower-growing isolates were included in different SSCP groups. A phylogenetic analysis of the ITS sequences of representatives of these groups confirmed the results obtained with the SSCP method, and showed that the faster-growing isolates do not belong to Ampelomyces as suggested by earlier studies. All the isolates from P. leucotricha fell into a distinct SSCP group of genetically homogeneous isolates. This suggests that Ampelomyces mycoparasites which occur in apple powdery mildew are slightly different from the other Ampelomyces groups which contain mycoparasites from various powdery mildew species. This may be because the main growth period of Ampelomyces mycoparasites in apple powdery mildew is isolated in time from that of Ampelomyces isolates that occur in other species of the Erysiphaceae. P. leucotricha starts its life-cycle early in the season, usually in March–April, while most powdery mildews are active in the same environments only late in the year.

Claviceps purpurea specific to grasses in salt marsh habitats (Group G3) has previously been identified on Spartina spp. in two locations: New Jersey, USA and southern England. We have identified this subgroup of C. purpurea (G3) in 11 distinct populations including western Europe, South America, and along the Atlantic and Pacific Coasts of the USA. In addition, G3 C. purpurea was discovered on a new host grass genus, Distichlis. Unweighted pair group mean analyses of AFLP and RAPD data reveal distinct structure in G3 C. purpurea populations. Pacific coast populations show little diversity, suggesting they may have been introduced recently in that region. 43 isolates, representing 11 populations were identified as G3 based on the presence of an EcoRI restriction site in the 5.8S ribosomal DNA, and a clear genetic separation from isolates representing the other two C. purpurea subgroups (G1 and G2). In addition, all isolates originating from Spartina densiflora, S. foliosa, S. alterniflora, and S. anglica were identified as belonging to G3. RAPD and AFLP analyses supported the recognition of three discrete groups within C. purpurea and revealed high genetic variability between groups, with only 1.8% of polymorphic markers shared across all isolates. Similarly, analysis of molecular variation (AMOVA) revealed that genetic variability was mainly due to variations between groups (63.5%) rather than within groups (28.5%) or within populations (7.96%). G3 isolates were 35% similar, Pacific coast isolates 83% similar. Ninety percent similarity among isolates from the San Francisco Bay Area suggests this is a recently introduced population.

We examined the phylogenetic position of Omphalina foliacea and its putative closest relative, the lichen-forming basidiomycete Lichenomphalia lobata. Both species are restricted to specific habitats in the high andine regions of tropical America. We generated nuclear ITS and LSU rDNA sequences of two collections of each species and analysed the data together with sequences of omphalinoid basidiomycetes retrieved from GenBank using a Bayesian and a maximum parsimony approach. Our analyses show that both, O. foliacea and L. lobata are lichenized basidiomycetes. L. lobata is related to other species of the genus Lichenomphalia, whereas O. foliacea is not closely related to other Omphalina or Lichenomphalia species, but probably belongs in the hymenochaetoid clade. The analysis of a nuclear LSU data set of a broader sampling of hymenochaetoid basidiomycetes supported the placement of O. foliacea in this clade, but did not reveal any close relative.

We used ITS-RFLP and sequence analysis to determine the identities of the fungal endophytes of six terrestrial orchid species from southeastern Queensland, a region previously unexplored in this context. Pure cultures of orchid – colonising fungi were obtained and fungal identities were assessed by means of ITS-PCR, RFLP analysis, sequence comparison, and protocorm colonisation tests. ITS-PCR and RFLP analysis resulted in five main groupings. Sequencing and GenBank comparison of these five groups showed that the fungal endophytes isolated from the three Pterostylis species were probably Thanatephorus species. There was close sequence identity (90%) of the fungus isolated from Acianthus spp. to Epulorhiza repens, suggesting these may be the same fungal species. However, that only E. repens succeeded in colonising protocorms of Thelymitra pauciflora suggests these may be different species of Epulorhiza. Analysis of the ITS and LSU sequences of the fungus isolated from Caladenia carnea showed high identities with a sequence from a Sebacina vermifera originally isolated from Caladenia dilatata. These results show that there is specificity for fungal partners within the orchid genera Acianthus, Caladenia and Pterostylis.

A 3193 bp contiguous sequence has been cloned from the oleaginous fungus Mucor circinelloides, that contains a full-length gene encoding a putative NADP+[ratio ]dependent malic enzyme (EC. 1.1.1.40). The cloned DNA contains a 2154 bp putative open reading frame containing five introns and encoding a protein of 616 amino acids. The gene encoded what appeared to be an anaerobic isoform of malic enzyme (isoform II); this conclusion is supported by transcript analysis and by the fact that the ORF contains an N-terminal mitochondrial target sequence (a similar cellular location was identified for the anaerobic malic enzyme in Saccharomyces cerevisiae; Boles et al. 1998). The cloned gene did not encode either isoform III (the isoform associated with active growth) or isoform IV (associated with lipid accumulation) previously identified. M. circinelloides therefore must possess (at least) two structural genes for malic enzyme.

We compared the ability of five strains of the ericoid mycorrhizal fungus Hymenoscyphus ericae to utilise glutamine, ammonium or nitrate at high or low carbon (C) availability. The pattern of intraspecific variation in growth was affected by C availability. When C supply was high, growth differences between strains were explained by the total amount of nitrogen (N) taken up, suggesting variation in uptake kinetics. Under C-limiting conditions, strain differences were linked with their nitrogen use efficiency, implying intraspecific differences in N metabolism. The relationship between growth on glutamine and pH shifts in the media indicated that there was intraspecific variation in glutamine transporters. In addition, the correlation between pH changes and the amount of glutamine-N recovered as ammonium in the media indicated that there were intraspecific variations within the enzymatic pathways involved in glutamine metabolism. Our findings, compared with those of a previous study involving the same ericoid strains, draw attention to the temporal variation in nitrogen source utilisation by ericoid mycorrhizal fungi when maintained in axenic culture.

Ericoid endomycorrhizal fungi (two isolates of Hymenoscyphus ericae obtained from unpolluted heathlands and two H. ericae-type endophytes isolated from Calluna vulgaris growing on Cu-contaminated mine spoil) were grown for 14 d on 10% Rorison's solution containing sodium phytate as the sole P source and either trace (0.16 μM) or elevated (0.25 mM) concentrations of Cu. The elevated levels of Cu in the medium had no effect on the growth of the two H. ericae-type endophytes from mine spoil sites but caused a significant reduction in growth of the two H. ericae isolates from unpolluted sites. Wall, cytoplasmic and extracellular fractions were assayed for phosphomonoesterase (PMEase) and phosphodiesterase (PDEase) activity. Km and Vmax values varied between the different endophytes and both were highest in the wall fractions. Wall-bound phosphatase activity, excluding PDEase of one H. ericae-type endophyte, was generally unaffected after the isolates had been grown on medium containing 0.25 mM Cu. Extracellular PDEase of the two H. ericae-type endophytes from mine spoil sites was stimulated by 0.25 mM Cu in the growth medium. Cu concentrations up to 5.0 mM in the assay medium did not inhibit wall-bound phosphatase activity whereas three of the isolates showed a stimulation of extracellular activity with increasing Cu. The results are discussed in relation to phosphatase activity of ericoid endophytes on Cu-contaminated substrates.

Coarse woody debris is important for mycodiversity in forest ecosystems, but its availability in managed stands is reduced. Leaving dead wood during felling is suggested as an option to sustain and restore the diversity. However, little is known what fungi would colonise freshly cut wood left on managed sites, and how the colonisation process is influenced by ecological factors. During summer and autumn, 120 freshly cut Picea abies stem sections over 8 cm in diameter were placed upright in mapped locations over two discrete plots separated by 100 m in a north-temperate forest. After seven weeks the sections were collected, and isolation and identification of fungi was done from their upper surfaces. In all 943 fungal strains were isolated, representing 97 species. Species richness in the summer survey was 42.5% higher than during the autumn survey. Low species similarity characterized the different seasons (Sorensen indices: SS=0.36 and SN=0.34) and for 21 species (22%) observation frequency was significantly affected by season. As a result, community structures in summer and autumn differed notably (z-test; P<0.001). Species richness between the two plots differed by less than 10%, but there were 64 species (66%) found only in one of them, thus qualitative similarity was low (SS=0.49). Quantitative similarity was higher (SN=0.63), indicating that the dominant species colonised wood to a similar extent in both areas. Fungal community structure differed significantly among the two plots (z-test; P<0.001). Our data showed that freshly cut CWD contributed to mycodiversity in managed north-temperate forest, providing habitats for numerous individuals from over 100 species. The fungal community within a single stand differed markedly both across small distances and over the seasons. In order to sustain and enhance mycodiversity in managed stands, coarse wood should always be left during harvesting. This study also demonstrates the importance of molecular identification and ITS sequence databases for exploring fungal diversity in natural communities.

The study of mould spores is of major importance as many fungi can cause considerable economic losses worldwide acting as plant pathogens or triggering respiratory diseases and allergenic processes in humans. Knowledge of spore production relationships to different altitudes or weather patterns can be applied in a more efficient and reliable use of pesticides or improving diagnosis and treatment of respiratory allergic diseases. In this way monitoring of Cladosporium cladosporioides, C. herbarum and Alternaria spp. airborne spores during 2002 was carried out by means of three LANZONI VPPS 2000 pollen traps located in areas of north-west Spain at various altitudes and with various weather patterns.

High spore counts were recorded in the late summer and early autumn, with a fairly similar hourly spore-count pattern, increasing the concentrations in the late evening (7–10 p.m.). High spore concentrations were detected in inland rural areas in front of coastal ones. As the continentality index increased, C. cladosporioides spore concentrations rose and Alternaria declined. C. herbarum concentrations increased with increasing height above sea level. The weather factor displaying the strongest positive correlation with mean daily spore counts was temperature. The optimal conditions for high airborne spore concentrations were recorded at temperatures ranging from 23–29 °C and RH values of around 80%, followed rapidly by rainfall in Vigo and Ourense and preceded by heavy rain two days prior to recording peak values in Trives.

The external morphological changes occurring during initiation and early stages of fruit body development of a Morchella sp., before the development of asci, were examined by scanning electron microscopy. Four stages of primordial development were distinguished. First, disk-shaped knots of 0.5–1.5 mm were observed on the surface of the substrate. Next, the knot inflated and a primordial stem emerged from its centre. Afterward, the stem lengthened, oriented upward, and two types of hyphal elements developed: long, straight and smooth basal hairy hyphae and short stem hyphae, some of which were inflated and projected out of a cohesive layer of tightly packed hyphal elements. Finally, when the stem was 2–3 mm long, pre-apothecia emerged in the apical end, with ridges and pits having distinguished types of paraphyses. Extracelluar mucilage covered the ridge layer and helped give the tissue its shape and rigidity.