This month Mycological Research News features work on chromosome translocations in six closely related Saccharomyces species, and analyses of the largest rDNA data sets of fungi yet published which call into question some proposed phylogenies and natural groupings.

Amongst the 16 papers in this issue are molecular studies exploring the diversity of soil fungi, rDNA and taxonomy of Ganoderma, RAPD analyses of Armillaria mellea populations, relationships of Phytophthora species, variation in P. cactorum, dsRNA viruses in Metarhizium anisopliae isolates, the genetic structure of Cryphonectria parasitica populations, and the array of Trichoderma isolates from Russia, Siberia and Himalaya.

The effects of mycorrhizal and non-mycorrhizal root exudates on the sporulation of Phytophthora fragariae are reported, and the cytology and ultrastructure of Fusarium culmorum and Mycosphaerella graminicola on wheat are documented.

The fruiting inducing factor from Phellinus contiguus has been partially characterized, and the resistance of Acer pseudoplatanus wood to Armillaria mellea degradation is explored.

The distribution of fungi on fallen Pinus sylvestris needles is examined along altitudinal transects, and a remarkable diversity of fungi on Picea abies documented.

Craspedodidymum nigroseptatum sp. nov. is described and a key to the species provided.

Ribosomal DNA sequences provide new molecular approaches to characterise fungal species from the environment. This study presents a culture-independent approach to assessing fungal diversity by direct amplification, cloning, restriction digestion and sequencing of internal transcribed spacers (ITS). We have assessed this novel approach both with culturable soil fungi and with total environmental DNA extracted from the same soil sample. Sixty-seven colonies and 51 cloned amplicons from total DNA were compared on the basis of their ITS restriction patterns. Then, 58 representative ITS sequences were determined and classified by comparing their sequences with ITS sequences of known origin. All culturable fungi were ascomycetes with one basidiomycete exception. In contrast, cloned amplicons from total environmental DNA were identified to ascomycetes, one basidiomycete, one zygomycete and to several fungi belonging to the kingdoms Chromista and Protozoa. The results show that PCR–RFLP of ITS provides an efficient culture-independent molecular tool for ecological studies and to assess fungal diversity.

As part of research to implement methods for the identification of Ganoderma species, this paper describes the use of PCR coupled to restriction digestions, single-strand conformational polymorphism (SSCP) and direct sequencing to assay rDNA polymorphism in South American collections of Ganoderma. The aim of the present study was to determine whether morphologically defined groups could be discriminated by molecular markers and to examine ITS sequence variation. Nineteen isolates of subgenus Ganoderma representing seven morphological taxa, and thirty isolates of subgenus Elfvingia representing three taxa, were studied. ITS I and IT II regions of twenty-one isolates were sequenced and were cladistically analyzed. Seven additional sequences were downloaded from EMBL/Genbank for comparison. In general, agreement between groupings delimited by restriction patterns, SSCP of ITS I and ITS II was evident. Gene trees derived from ITS I, ITS II and from a combined data set, were obtained and compared. Agreement between molecular and morphological data was clear at the subgeneric level; however, at the specific level, this relationship was difficult to visualize.

The genetic variation of Armillaria mellea sensu stricto was studied within and among non-heterothallic populations from Japan and Africa, and heterothallic populations from Europe and North America, using somatic incompatibility tests and RAPD analysis. Non-heterothallic isolates from Japan and Africa were divided into four somatic compatibility (SI) groups (A, B, C and D). SI group A contained half of all Japanese isolates and all African isolates. SI groups B and C each contained four Japanese isolates and D contained one isolate. All European and North American isolates were clearly separated from each other by somatic incompatibility tests. From the RAPD analysis, three distinct groups were separated (non-heterothallic, European and North American groups). The non-heterothallic group was divided into three subgroups (Ia, Ib and Ic). Subgroup Ia and Ib corresponded to SI groups A and B, however, Ic contained SI groups C and D. Subgroup Ia, in which 12 of 14 isolates had the same haplotype, showed a little variation. Non-heterothallic populations were less variable than heterothallic populations. The distribution of the isolates belonging to each SI group overlapped within Japan.

Phylogenetic relationships among Phytophthora species were investigated by sequence analysis of the internal transcribed spacer region I of the ribosomal DNA repeat unit. The extensive collection of isolates included taxa from all six morphological groups recognized by Waterhouse (1963) including molecular groups previously identified using isozymes and mtDNA restriction fragment length polymorphisms. Similar to previous studies, the inferred relationships indicated that molecular groups of P. cryptogea/drechsleri-like and P. megasperma-like taxa are polyphyletic. Morphological groups V and VI, which are differentiated by the presence of amphigynous or paragynous antheridia, are not monophyletic: species of the two groups are interspersed in the tree. Species with papillate and semi-papillate sporangia (groups I–IV) clustered together and this cluster was distinct from those of species with non-papillate sporangia. There was no congruence between the mode of antheridial attachment, sporangial caducity, or homo- or heterothallic habit and the molecular grouping of the species. Our study provides evidence that the antheridial position together with homo- or heterothallic habit does not reflect phylogenetic relationships within Phytophthora. Consequently, confirming studies done previously (Cooke & Duncan 1997), this study provides evidence that the morphological characters used in Phytophthora taxonomy are of limited value for deducing phylogenetic relationships, because they exhibit convergent evolution.

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.

Arbuscular mycorrhizal fungi (AMF) show significant potential for biocontrol of Phytophthora spp., but there is little evidence for the mechanisms involved in the process. This study establishes that microorganism free exudates from roots colonised by AMF result in significantly less sporulation of P. fragariae than those from uncolonised plants. Experiments in vitro showed that after 48 h in the presence of exudates from strawberry roots colonised by Glomus etunicatum and G. monosporum, sporulation of P. fragariae was reduced by ca 67% and 64% relative to sporulation in the presence of uncolonised roots. After 72 h sporulation was reduced by 83% and 89% respectively. These data were then confirmed in an in vivo system in which Phytophthora fragariae was inoculated into the mycorrhizosphere of either uncolonised strawberry plants or those colonised by G. etunicatum. A similar trend was observed, with a 69% reduction in sporulation of P. fragariae after 72 h in the mycorrhizosphere of colonised plants relative to sporulation in the mycorrhizosphere of uncolonised plants.

We have investigated the wheat infection process of Mycosphaerella graminicola (teleomorph of Septoria tritici) through the combined application of differential interference contrast light, confocal laser scanning, and cryo-scanning electron microscopies. Several aspects of fungal development have been documented, including the presence of extracellular cirrus and germling matrices, as well as nutrient-sensitive morphogenesis. The use of confocal laser scanning microscopy was invaluable in documenting the time course of pathogen invasion and symptom development. These data were used to generate summary diagrams of the asexual infection cycle of M. graminicola, to benchmark the efficacy of various crop protection strategies.

The infection of wheat spikes by Fusarium culmorum, one of the agents responsible for wheat head blight, was examined by light and electron microscopy. Macroconidia of the pathogen germinated 6–12 h after inoculation on all host surfaces they contacted. Developing germ tubes did not infect host tissues immediately, but gave rise to hyphae that grew and branched on host surfaces. Hyphal networks were usually formed 2 d after inoculation (dpi) on the inner surfaces of lemma, glume and palea, but not on the outer surfaces of lemma, glume and rachis. Hyphae on the outer surfaces of lemma and glume often grew over their edges to reach their inner surfaces. Penetration of host tissues occurred by infection hyphae on the inner surfaces of lemma, glume and palea, and on the upper part of the ovary. Occasionally, the pathogen invaded the host tissues through stomatal openings on the inner surface. Thereafter, the pathogen spread downwards to rachilla and rachis node by inter- and intracellular growth from above infected tissues. From the rachis node, hyphae extended downward to rachis and upward to peduncle through vascular bundles and cortical parenchyma tissue. When the pathogen arrived at the rachis 4–5 dpi, hyphae grew upwards and downwards inter- and intracellularly in vascular bundles and cortical parenchyma tissue of the rachis. During colonisation of the wheat spike, a series of alterations occurred in host tissues, including degeneration of host cytoplasm and organelles, collapse of parenchyma cells, disintegration or digestion of host cell walls and appearance of electron-dense coating materials on vessel walls.

Several isolates of the entomopathogenic Metarhizium anisopliae that harboured dsRNA viruses of similar electrophoretic band sizes (1.8 and 2.0 kbp) were assessed for homologies of the dsRNA by Northern analysis. The isolates were also characterised genetically by RAPD and VCG. Similarly sized dsRNA, as visualised by electrophoresis, were not always homologous, suggesting that the comparison of dsRNA based solely on electrophoretic banding patterns is an unreliable method of dsRNA characterisation. Several isolates, but not all, harbouring multiple dsRNA patterns, including a 1.8 and 2.0 kbp doublet, also showed homologies to strains harbouring only the dsRNA banding doublet. This suggests that mixed infections of different dsRNA elements are found in M. anisopliae. Genetically similar fungi, based on RAPD banding patterns and vegetative compatibility, were more likely to harbour genetically related dsRNA. The findings suggested that dsRNA elements in M. anisopliae are horizontally transferred to genetically related isolates or are maintained through clonal lineages.

Filtrates from cultures of wild isolates of Phellinus contiguus grown on liquid medium containing malt extract induced precocious fruiting of this resupinate polypore when tested in a bioassay. Active filtrates were analysed using gel filtration, HPLC and GC-MS and were compared with filtrates from single spore progeny, two of which lacked fruiting inducing activity. The active material was polar and heat stable, passed through both anion and cation exchange columns and had a mol. wt less than approx. 1000. Fractionation using HPLC on amino-silica yielded one fraction which was active on its own and a second, later eluting fraction, which had scarcely detectable activity when alone, but when combined with the first fraction had much higher activity than either alone. GC-MS showed that both this first fraction and the most active fraction from P2 gel filtration were rich in mono- and disaccharide sugars. In comparison with fractions from parallel inactive filtrates, all active fractions contained more myo-inositol and a relatively high glucose/disaccharide ratio.

It is suggested that a combination of high concentrations of myo-inositol and apoplastic glucose may be required for initiation of fruiting in P. contiguus. Although additional work is needed, this view is consistent with known features of growth and fruiting in white rot fungi.

The genetic structure of three newly established Cryphonectria parasitica populations (Choëx, Weggis, Murg) was analysed and compared to an older, post-epidemic population (Claro). Vegetative compatibility (vc) type and DNA fingerprint analysis revealed an almost clonal C. parasitica population in Choëx, Weggis and Murg. These stands are all situated in northern Switzerland, outside the main range of European chestnut. Only one vc type and one dominant DNA fingerprint was found in Choëx and Murg. In Weggis, two vc types and two dominating DNA fingerprints were found. The European Cryphonectria hypovirus (CHV1) was not detected in these three populations. In contrast, the population in Claro, situated within the main range of European chestnut in southern Switzerland, had much higher vc type and DNA fingerprint diversity. DNA fingerprints were correlated to vc types in Claro and in Weggis. Mating type determination revealed one strongly dominating mating type in each of the three northern populations, but not in Claro. From these results we conclude that C. parasitica disseminated almost exclusively by means of asexual reproduction in Choëx, Weggis and Murg, whereas in Claro sexual reproduction also played an important role. Additionally, founder effects and restricted gene flow were dominating factors in shaping the genetic structure of the three northern populations.

About 35 Trichoderma species are currently recognised on the basis of morphological and molecular characters. Besides the role of a few of these species in biotechnology, several seem to play prominent roles in soil ecosystems. With a goal of investigating global biodiversity in Trichoderma, we report on the occurrence of Trichoderma spp. in Russia (Moscow and Ural areas), Siberia (Krasnoyarsk area) and the Himalayan mountains – areas from which no Trichoderma isolates are so far available. The ITS 1 and 2 sequence of the rDNA cluster of the 75 isolates obtained was compared with that of ex-type strains and taxonomically established isolates of Trichoderma. Thirty-nine isolates were positively identified as T. atroviride, T. virens, T. hamatum, T. asperellum, T. koningii and T. oblongisporum. A further 26 isolates yielded six closely related ITS1/2 sequence types, which are highly similar yet different from the ex-(neo)type strains of T. harzianum and T. inhamatum. Some of these genotypes (i.e. 1 and 2a) occurred only in Russia/Siberia, whereas others (2b, 3, 4 and 5) were found only in the Himalayas. RAPD analysis was consistent with these genotypes, and revealed genetic homogeneity even between strains from widely separated areas. Parsimony analysis placed these five genotypes, together with T. harzianum, T. inhamatum and the mushroom-aggressive T. harzianum ‘biotype 2’ in a large, unresolved ‘harzianum’ clade. Ten isolates were not safely alignable within known species, and five of them may be undescribed taxa: one isolate from 2700 m elevation in the Himalayas, which clustered in parsimony analysis at a basal position in section Longibrachiatum; and four isolates, displaying two closely related sequence types, forming a separate clade with T. stromaticum. The five remaining isolates also exhibited three unique ITS1 and 2 sequence patterns, but parsimony analysis placed them into the unresolved ‘harzianum’ clade, and their relationship to T. harzianum is thus unclear. The study shows that molecular screening of uninvestigated geographic areas can lead to the identification of isolates with new ITS1 and ITS2 sequence patterns, some of which may be new taxa. It also reveals that T. harzianum is at present the genetically most diverse member of the genus.

The white rotting Armillaria mellea induced a spatially very distinctive pattern of degradation in sycamore wood, under both natural and artificial conditions. Degradation began preferentially within groups of fibres containing intercellular spaces, whereas fibre regions lacking such spaces were undegraded and remained largely intact even when decay had become advanced elsewhere. The two types of fibre region differed not only in the presence of intercellular spaces, and hence in the potential for gas exchange, but also in their degree of lignification. This was higher in the more resistant type, as shown by staining of undecayed wood with toluidine blue-O, by microspectrometry after staining for the Mäule colour reaction, and by uv-microscopy. A spatially similar pattern of cellulose degradation was induced by the brown rotting Laetiporus sulphureus, which is known to cause preferential degradation of less strongly lignified cell walls. By contrast, the white-rotting Ganoderma pfeifferi showed a tendency to degrade the stronger lignified cell walls. Thus, in combination with the application of conventional histological methods, the wood degradation modes observed give additional evidence for stronger and weaker lignified fibre regions within the wood of sycamore.

This work tested the hypothesis that the abundance of fungal species on coniferous needles is correlated to climatic factors at a regional scale. The proportions of Pinus sylvestris needles colonised by Lophodermium pinastri, Cyclaneusma minus and Verticicladium trifidum were examined along two altitudinal transects in 2 successive years. L. pinastri and C. minus colonisation increased with altitude, whereas that of V. trifidum decreased. The data are discussed in relation to the importance of climatic controls on species distributions.

The majority of microbes living on forest trees are still unnamed and our knowledge of their species richness is vague. This paper describes the fungal diversity of the above ground parts of a 61 year old Norway spruce tree lacking visible signs of damage or disease. The problem involved with identification of the fungi to named species was circumvented by classifying them into operational chemotaxonomic units (OCTUs) by using their combined fatty acid and sterol profiles (FAST-profiles). The variation of these units was chosen to correspond to a within-species-variation determined for several morphologically defined taxonomic species occurring on Norway spruce. Ninety-nine OCTUs were identified from 666 fungal isolates obtained. Bacteria were found only occasionally from the inner bark samples (three isolates) and from needles (five isolates). Models describing the accumulation of OCTUs against the number of samples taken were used for extrapolation of the total number of fungal OCTUs in the above ground parts of the tree. Our results suggest that an undamaged, apparently healthy Norway spruce, harbours in its above ground parts nearly two hundred fungal species. The majority were estimated to be needle epiphytes. At the large forest area scale, the species richness may be one order of magnitude higher.

Craspedodidymum nigroseptatum sp. nov. is described and illustrated based on specimens collected on decaying rachides of Oncosperma horridum in Brunei Darussalam. It differs from other species of Craspedodidymum in producing dark brown conidia with three to four thick, darkly-pigmented septa, and a paler basal cell. Conidia are broadly ellipsoidal to obovoid and truncate at the base. Craspedodidymum nigroseptatum is compared with other species in the genus and a key to the genus is provided.