The circumscription of the family Agaricaceae, its genera and their characters are investigated. Analyses of the ITS and LSU sequence data, separately and in combination result in exclusions and reorganization. Notholepiota areolata, a secotioid species from New Zealand, Cystoagaricus with brown, irregular-knobby spores, and Leucopholiota with gelatinized hymenophoral trama and amyloid spores, fall outside the family Agaricaceae. In the reorganization, long accepted tribe and genus definitions are amended, viz.: (1) Macrolepiota and Chlorophyllum separately are monophyletic, but not together; (2) Leucoagaricus and Leucocoprinus together form one large monophyletic clade, that also includes polyphyletic Sericeomyces; (3) Lepiota, Cystolepiota and Melanophyllum together form a monophyletic clade; (4) Secotiation has happened within several clades (e.g. Agaricus and Chlorophyllum), though not in the Leucoagaricus/Leucocoprinus clade, nor in the lineage of Macrolepiota proper; gasteroid and most secotioid species in the course of adaptation acquire coloured spores; and (5) the monovelangiocarpic Chamaemyces fracidus appears to be basal to the family and provides clues to the morphology of the ancestral Agaricaceae. Trama and developmental characters characterize the different clades, whereas spore characters play a minor role.

This work aimed to evaluate the genetic diversity of Botrytis cinerea in Chile and to determine whether the two genetically different groups transposa and vacuma, described in France, are present in the country. Isolates collected in Chile from grapes, tomato, kiwifruit and blueberry were analysed using molecular markers. We developed a PCR test to identify the two groups of B. cinerea, transposa (with the transposable elements Boty and Flipper) and vacuma (with neither). As described in France, both kind of isolates were found to be present and sympatric in Chile. Isolates containing the transposable element Boty alone (boty isolates) were also detected. The frequencies of transposa, boty and vacuma isolates were significantly different on kiwifruit compared to grapes, tomato and blueberry. RAPD analysis revealed a high degree of genetic diversity, with no widespread clonal lineages. Dendrograms, analysis of molecular variance and Fst values revealed the existence of genetic differentiation in our sample between isolates from the different host plants. PCR-RFLP markers also showed that isolates sampled from grapes and tomato were genetically differentiated. Additional sampling is required to confirm these findings.

21 cactus species occurring in various localities within Arizona were screened for the presence of fungal endophytes. 900 endophyte isolates belonging to 22 fungal species were isolated. Cylindropuntia fulgida had the maximum endophyte species diversity, while C. ramosissima harboured the maximum number of endophyte isolates. Alternaria sp., Aureobasidium pullulans, and Phoma spp. were isolated from several cactus species. The diversity of the endophyte assemblages was low and no host specificity among endophytes was observed. However, the frequencies of colonization of the few endophyte species recovered were high and comparable to those reported for tropical plant hosts. Species of Colletotrichum, Phomopsis, and Phyllosticta, which are commonly isolated as endophytes from plants of more mesic habitats, were absent from these cacti.

The effect of the flavonoids chrysin, isorhamnetin, kaempferol, luteolin, morin and rutin on pre-symbiotic growth, such as spore germination, hyphal length, hyphal branching and the formation of auxiliary cells and secondary spores, of the arbuscular mycorrhizal fungi Gigaspora rosea, G. margarita, Glomus mosseae and G. intraradices was studied. According to the effect on each fungal growth parameter, the tested compounds could be classified to be genus and/or species specific or specific, for a certain developmental stage of pre-symbiotic growth. A clear arbuscular mycorrhizal genus specific, and even species specific, effect of some flavonoids was observed. However, this specificity of a flavonoid could not be generalized but differs depending on the pre-symbiotic stage of the AM fungus. Moreover, our show that for a better understanding of the role of compounds in the AM symbiosis, studies should not be conducted only with one AM fungus looking at one fungal growth parameter such as spore germination or hyphal length, but should be wider, including several growth parameters and several AM fungi.

During the last 15 years, about 2000 strains of 200 species of 98 genera of fungi have been isolated from around the Chernobyl Atomic Energy Station. Many of these microfungi are capable of growing into and decomposing ‘hot particles’; carbon based radioactive graphite from the reactor and there are suggestions that some fungi actively direct their growth toward sources of radioactivity, possibly attracted to the carbon skeleton of these structures. In our experiments, we eliminated the confounding effects of carbon as a fungal resource, by developing experimental protocols that expose fungal spores and their germinating hyphae to directional sources of ionizing radiation allowing us to measure fungal response to ionizing radiation per se. We show that both beta and gamma radiation promote directional growth of hyphae towards the source of ionizing radiation.

Three fungal species responsible for anthracnose canker, perennial canker, and bull's-eye rot of apple have been considered members of the genus Pezicula for a number of years. Recent studies, however, have provided evidence to (re-)classify these species as Neofabraea. There has been a long historical debate regarding the taxonomy of two of these fungi. In Europe, both Neofabraea malicorticis and N. perennans have generally been considered N. malicorticis, while in North America a species distinction has been maintained. Phylogenetic analyses of Neofabraea isolates were based on DNA sequences of the internal transcribed spacer region of nuclear rDNA (38 isolates), the mitochondrial rDNA small subunit (partial; 48 isolates), the β-tubulin gene (partial; 25 isolates), and a combined data set (21 isolates). Our work provides evidence for the existence of four distinct Neofabraea apple pathogens including N. malicorticis, N. perennans, N. alba, and a putative new Neofabraea species that was isolated in both Europe and eastern North America. Our results indicate that the primary Neofabraea species causing tree cankers and bull's-eye rot in North America are N. malicorticis and N. perennans in the west, and N. alba in eastern Canada. N. perennans, N. alba, and the undescribed Neofabraea species were found in Europe but the presence of N. malicorticis was not confirmed by our limited sampling. Inclusion of Rosa spp. in the host range of N. malicorticis is merited.

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.

The following topics are addressed: the interfaces of the Xylariaceae with related groups and the ‘essence of xylariaceousness’; speciation of Xylariaceae in the tropics; the distribution of Xylariaceae in the tropics; xylariaceous fungi categorized into ecological groups based on substrate type and timing of colonization and on modes of water conservation; xylariaceous fungi as latent pathogens and endophytes; novel morphological features of xylariaceous fungi and their roles under tropical conditions.

The association of ascomycetes with Fucus serratus was investigated by comparing the broad-based molecular and cultural diversities of healthy and dead fronds. Four PCR primer pairs were used to amplify the 18S (primers NS1-FR1; NS1-EF3) or 28S rRNA (primers NL209-NL912; NL359-NL912) genes directly from the DNA of algal thalli. Two novel primer pairs, NL209-NL912 and NL359-NL912 giving product sizes of 700 and 559 bp respectively, were designed to amplify the 28S rDNA from ascomycetes specifically. All primer combinations amplified DNA from 33 reference taxa isolated from Fucus serratus, and the products generated by primers NS1-FR1GC and NL359-NL912GC were separated in 18–38% and 38–60% denaturant gradients respectively after DGGE. The 18S rDNA DGGE system resolved eight bands from algal DNA, but many of the sequences separated were not fungal, whereas the 28S rDNA system resolved seven bands that were all identified as ascomycetes. Phylogenetic analysis and BLAST search results of environmental sequences revealed the presence of four main ascomycete groups: (1) the Halosphaeriales, (2) the Hypocreales, (3) an unidentified Lulworthiales complex, and (4) the Pleosporales. Few fungal isolates were detected molecularly suggesting that fungal colonisation of fronds was limited, mainly to species in dead casts.

Evidence is presented that the biogenesis of sclerotia in Sclerotium rolfsii is associated with lipid peroxidation. Sclerotial initials show 100-fold increase in lipid peroxides of their total lipids as compared with young mycelia grown under reducing conditions (in 2-mercaptoethanol) in dark and without Fe(II). There was a direct relationship between the number of sclerotia formed and lipid peroxidation levels in the mycelial colonies. Lipid peroxides of possible membranous and cytoplasmic origin were found in sclerotial exudate. In this paper a new approach is advanced for the understanding of the mechanism of sclerotial formation in Sclerotium rolfsii and in other fungi. The data in this work on lipid peroxidation – the most frequently evoked consequence of oxygen free radicals – as well as the data of past experiments, strongly suggest that this phenomenon may be associated with oxidative stress caused by growth conditions.

Ophiostoma ulmi and O. novo-ulmi are partly reproductively isolated, morphologically, behaviourally and molecularly distinct species responsible for the first and current pandemics of Dutch elm disease, respectively. Among >11000 isolates sampled from Dutch elm disease sites across Eurasia and North America since 1973, nine could not be assigned to O. ulmi or O. novo-ulmi. Of these isolates one (P129) was from Poland and eight (d10, d11, e12, e27, e28, e37, f30 and g3) were from a single bark sample in Portugal. These nine isolates were termed ‘fast-waxy’ isolates because of their unusual cultural characteristics. The possibility that they were interspecific hybrids was investigated. When compared with O. ulmi and O. novo-ulmi for colony pattern, growth-rate, optimum temperature for growth, vascular wilt ability, elm bark colonizing ability, cerato-ulmin toxin production, ability to fertilize (as [male ]) O. novo-ulmi, and ability to be fertilized (as [female]) by O. ulmi, they exhibited a combination of O. ulmi-like, O. novo-ulmi-like, intermediate or novel characters (female sterility) consistent with their being hybrids. P129 and representative Portuguese isolates d10 and e27 each exhibited a different combination of characters, indicating each was a different hybrid genotype. When d10 and e27 were independently crossed to the same O. novo-ulmi isolate, differences in their F1 progeny sets for growth-rate and pathogenicity distributions were consistent with their being different recombinant genotypes. A molecular analysis of P129, d10 and e27 using RAPDs of genomic DNA, rDNA RFLPs and cerato-ulmin gene sequences confirmed that each was a unique interspecific hybrid. The mechanism of origin of these hybrids and their evolutionary significance are discussed. Combined experimental and circumstantial evidence indicates that they are relatively unfit, rare and probably transient, and that they arise when O. novo-ulmi invades territory occupied by O. ulmi and replaces it. Nonetheless, the possibility that the hybrids act as a genetic bridge, facilitating transfer of novel vegetative compatibility loci and other loci form O. ulmi to O. novo-ulmi at recent epidemic fronts, requires investigation.

Bayesian and maximum parsimony phylogenetic analyses of 92 collections of the genera Basidiophora, Bremia, Paraperonospora, Phytophthora and Plasmopara were performed using nuclear large subunit ribosomal DNA sequences containing the D1 and D2 regions. In the Bayesian tree, two main clades were apparent: one clade containing Plasmopara pygmaea s. lat., Pl. sphaerosperma, Basidiophora, Bremia and Paraperonospora, and a clade containing all other Plasmopara species. Plasmopara is shown to be polyphyletic, and Pl. sphaerosperma is transferred to a new genus, Protobremia, for which also the oospore characteristics are described. Within the core Plasmopara clade, all collections originating from the same host family except from Asteraceae and Geraniaceae formed monophyletic clades; however, higher-level phylogenetic relationships lack significant branch support. A sister group relationship of Pl. sphaerosperma with Bremia lactucae is highly supported. Within Bremia lactucaes. l., three distinct clades are evident, which only partly conform to the published host specificity groups. All species of the genera Basidiophora, Bremia, Paraperonospora and Plasmopara included in the present study were investigated for haustorial morphology, and all had ellipsoid to pyriform haustoria, which are regarded as a diagnostic synapomorphy of the whole clade. Aspects of coevolution and cospeciation within the downy mildew pathogens with ellipsoid to pyriform haustoria are briefly discussed.

Alternaria alternata, A. radicina and A. dauci are important seed-borne fungi on carrot, with the first two species having a high toxigenic potential, for which a specific and sensitive detection method is required. Because both the traditional deep-freeze-blotter method and plating on selective medium are time consuming and laborious, a PCR-based assay was developed. Sequences of the internal transcribed spacer regions of the ribosomal gene repeat (rDNA) from 45 different Alternaria isolates were determined, a restriction fragment length polymorphism analysis was performed and a phylogenetic tree was constructed. Based on the sequences, specific primers for detection and identification of the three Alternaria species on carrot seeds and roots were designed. The primers were highly sensitive and were shown to be able to differentiate between the three Alternaria species. A. alternata and A. radicina could be detected in DNA isolated from carrot material applying the specific primers, even at low infection levels. The PCR-assay was compared to the deep-freeze-blotter method (DFBM) and plating on Alternaria radicina Selective Agar (ARSA, for A. radicina) by testing naturally infected seed samples and root material. Results of the PCR-assay were similar to those of the blotter method and plating on ARSA for the detection of A. alternata and A. radicina. A positive correlation was found between the percentage of seed infection established by the blotter method and the intensity of the amplified, specific product. The PCR-assay based on the specific primers developed seems to be a good alternative for the deep-freeze-blotter method and plating on ARSA, especially when time is an important issue.

The phylogeny of Lecanoromycetes (Ascomycota, Fungi) is investigated utilizing parsimony and Bayesian Markov Chain Monte Carlo analyses, of combined nLSU rDNA and mtSSU rDNA sequence datasets. The results suggest that Acarosporaceae, Candelariaceae, Phlyctis and Pycnora are not members of the monophyletic Lecanorales, and that Timdalia and Pleopsidium are members of a monophyletic Acarosporaceae. Pycnora, Candelariaceae and Acarosporaceae form a monophyletic group. Umbilicariaceae, Hypocenomyce scalaris, H. friesii, Ophioparmaceae, Boreoplaca, Elixia and Fuscidea form either a basal paraphyletic assemblage in Lecanoromycetes, or a monophyletic group which is the sister-group to Lecanorales and the rest of Lecanoromycetes (excluding Acarosporaceae). The Acarosporaceae forms a group with Pycnora and Candelariaceae, which may be outside the Lecanoromycetes. Chaetothyriales, Verrucariales, Eurotiales, Lichinales and Mycocaliciales form a monophyletic group, but with low support. We briefly discuss incongruence between datasets from different genetic markers, comparing the differences between the separate parsimony analyses, where the ILD test indicated a very significant incongruence. The phylogenetic significance of ascus-types that have influenced most recent Ascomycota classifications heavily is also discussed, and we finally point out risks with formalizing classifications too early.

Species in the large mushroom genus Russula are important ecologically as ectomycorrhizal fungi and economically as comestibles. Most infrageneric classification schemes of this genus have originated in Europe, but because of nomenclatural history and an evolving suite of characters these systems remain largely incongruent. Using ribosomal DNA sequences for 87 species representing all infrageneric taxa described from Europe, the phylogenetic position and relationships among these species were examined. Cladistic analysis of the ITS1, 5·8S, and ITS2 regions showed a cluster of five to six small to large clades basal in the topology and one large apical clade arising from the deeper nodes, none of which has been previously recognized in toto at the subgeneric level. Two of these groups, the Compactae and Pallidosporinae, which have been previously recognized as subsections of section Compacta, did not appear to be closely related. Bootstrap support and Bremer decay values indicated that collapse of the tree into monophyly at the deeper nodes would result in two large groups which are consistent with the classical subgeneric concept of the Eurussulae and a narrowed Compactae. The topology confirmed some previously described infrageneric taxa at the section level including the Tenellae and Heterophyllae and at the subsection level including the Cupreinae, Laricinae, Lilaceinae, Integroidinae, Violaceinae, Sphagnophilinae, Viridantinae, Emeticinae, Subvelatae, Pallidosporinae, and portions of the Polychromae and Sardoninae. The molecular analysis also indicated many interesting new combinations or relationships not previously conceived. Mapping of characters such as spore print colour, taste, and presence of acid resistant incrustations, which have been used to define infrageneric taxa in Russula, onto the phylogeny identified interesting patterns consistent with hypotheses regarding plesiomorphic and apomorphic characters. However, because of potential loss or reversal of character states, this analysis did not support their unequivocal use in infrageneric classification.

The natural dihydrate form of calcium sulphate, CaSO4.2H2O (gypsum), particularly found in gypsiferous soils and in certain building constructions, was found to be effectively solubilized by both Aspergillus niger and Serpula himantioides. When the fungi were grown on malt extract agar (MEA) medium amended with up to 1% (w/v) of gypsum, solubilization activity was evident by the appearance of a clear zone (haol) around and/or under the growing colonies. The haloes were found subsequently to enclose concentric rings of newly formed crystalline precipitate. The pH profile of the medium underneath the growing mycelium of A. niger was not affected by the presence of gypsum, while the S. himantioides pH profile displayed a slight reduction in pH values when grown on gypsum-containing media compared with the control. Electron microscopy and X-ray microanalysis of the crystals revealed the formation of calcium-containing crystals with the characteristic morphology typical of different forms of calcium oxalate. The complexing activity of fungal-produced organic acids with calcium was suggested to be the prevalent mechanism of solubilization with the production of oxalic acid resulting in precipitation of oxalate. Such activity resulted in the release of available sulphate which increased over the incubation period. The concentrations of sulphate released from 0·5% (w/v) gypsum after growth of A. niger and S. himantioides for 8 d were 14·7±0·7 and 7·4±0·9 μmol cm−3 respectively. These results are discussed in the light of their relevance to land reclamation and plant nutrition, as well as the weathering of building materials containing gypsum.

Universally primed PCR (UP-PCR) fingerprinting combined with UP-PCR product cross hybridization, and ITS1 ribotyping were used to study the genetic relatedness of strains of Trichoderma and Gliocladium for two purposes: (1) to evaluate the ability of the methods to discriminate closely related strains and as tools to group strains which is necessary to facilitate; (2) identification of markers for development of specific detection assays for selected strains. Included among the strains were one T. harzianum, two T. virens, and one G. roseum that had been selected previously for their antagonistic ability against soil-borne phytopathogens. Similarity among strains, found by cross dot blot hybridization using UP-PCR amplification products, was used to group them into 15 genetic entities. ITS1 ribotyping of the strains was performed by digestion of the PCR amplified rDNA spacer region and electrophoresis of the products. The differences obtained from ribotyping as well as the differences in mobility of the intact spacer region were used for grouping of the strains. The UP-PCR hybridization groups and the ITS1 based groups proved to be consistent, but the resolution of the UP-PCR based approach was superior. The results demonstrate that the combination of UP-PCR and ribotyping can aid in clarifying species distinction in Trichoderma and Gliocladium and has the potential to become a valuable tool for studies of diversity and genetic structure of populations of these fungi. Furthermore, identification of single strains by the specific UP-PCR fingerprint seems feasible.

Thirty-eight strains of the entomopathogenic Beauveria bassiana, isolated from diverse species of Lepidoptera (Pyralidae) or Coleoptera (Curculionidae, Chrysomelidae, and Scolytidae) from various geographical sites, were examined by RFLP and RAPD analyses. Similar groupings were recovered from both approaches and these showed clear relationships between the population structure of B. bassiana and some defined host species. Strains isolated from members of the Pyralidae were recovered as two main groups, one group consisted of all strains isolated from Ostrinia irrespective of their origin. The second group consisted primarily of strains isolated from Diatraea in Cuba. All strains isolated from the curculionid genus Sitona clustered as a single distinct group, separated from strains from other curculionids. In contrast, strains isolated from the pyralid genus Maliarpha, and the coleopteran Chrysomelidae, gave heterogenous patterns and were not recovered as distinct groups. Groups from cluster analysis and non-hierarchic ordination methods were compared and the relative merits of the different grouping strategies are discussed.

This paper continues a revision of generic concepts in the parmelioid lichens using molecular data in order to reach a consensus among lichenologists over which segregates proposed over the last two decades should be accepted. Here we employ data from three gene portions to provide a basis for a revised generic concept of the brown parmelioid lichens hitherto classified in Melanelia. The phylogeny was studied using a Bayesian analysis of a combined data set of nuclear ITS, LSU rDNA and mitochondrial SSU rDNA sequences. 173 new sequences were obtained from 38 specimens of 15 Melanelia species, 37 related parmelioid species, and eight non-parmelioid species. The results indicate that Melanelia is not monophyletic but falls into four different clades. The genus Melanelia is restricted here to a small group of saxicolous lichens related to the type species M. stygia, and with bifusiform conidia, while the remaining species, most of which are primarily corticolous and have mainly cylindrical to filiform conidia, belong to two other clades recognised as two new genera: Melanelixia and Melanohalea, to accommodate the M. exasperata and M. glabra groups, respectively. 27 new combinations are made. The epicortex of Melanelixia species have pores or special structures termed here ‘fenestrations’, while most Melanohalea species are pseudocyphellate. Pleurosticta links to the Melanohalea clade but without strong support, and the phylogenetic position of M. disjuncta and its related species remains uncertain, linking with the Xanthoparmelia (syn. Neofuscelia) clade but also without strong support.

Podospora anserina and Coprinopsis cinerea (syn. Coprinus cinereus) are endowed with a defence system able to differentiate self vs. non-self and involving the generation of peroxide. Indeed, they produce peroxide when confronted with a filamentous fungus, only in non-self confrontations. Both species are not able to recognize yeasts and show a differential response to bacteria. The accumulation of peroxides in the ascomycete Podospora anserina requires an NADPH oxidase and a MAP kinase cascade, previously shown to be involved in fruit body formation, cell differentiation and cell degeneration. Confrontation is accompanied by the death of the contestant hyphae only in specific combinations of species. As in animals and plants, data suggest that peroxide is likely involved in signalling rather than playing a direct toxic role. Fungi display more complex behaviours than generally acknowledged, i.e. they are able to recognize potential contestants and built up defence reactions involving evolutionary conserved enzymes.