The concept of species and species complexes of lichenized fungi is reviewed in the light of recent molecular approaches. Species concepts based on chemistry, morphology, reproductive mode, photobiont choice and habitat preference provide working hypotheses from which the delimitation of species or relationships within species complexes can be tested with molecular data. In studies in which a single locus such as ITS is used, a phylogenetic species concept can be applied only when the sequence data delimits genetically discrete clades that correlate with phenotypic characters or biogeographical distribution. This will be the case when sufficient time has passed for genetic isolation to result in the coalescence of different character states among the sibling species of a complex. In the case of recently diverged species, a single locus may not accurately separate species, and is not sufficient evidence to reject putative species based on a phenotypic species concept. In such cases, several genetic loci must be used to delimit species by a phylogenetic species concept. These phylogenetic species may corroborate phenotypic characters or biogeography, or they may be cryptic for all observed characters. Molecular analyses at the species level will lead to the re-evaluation of phenotypic characters for species delimitation, and will improve the understanding of speciation in lichenized fungi. The results from new molecular approaches have implications for taxonomic revisions. Formal changes in nomenclature based solely on molecular evidence should be made conservatively, and only after studies based on a thorough sampling with sufficient data of all kinds is performed, and after the taxonomic history of the group has been reviewed. In the meantime, informal names to describe new phylogenetic hypotheses are sufficient. Formally named cryptic species cause problems in distributional studies, where old material and literature references are included, but are useful in high resolution studies such as ecophysiology, especially where phenotypic differences may exist among cryptic species.

Cellulose-acetate electrophoresis (CAE) was used to investigate isozyme polymorphisms of 34 Group 1 and Group 2 isolates of Fusarium graminearum. Of the 26 enzyme systems screened, alkaline phosphatase, NADP-dependent glutamate dehydrogenase, peptidase D and phosphoglucomutase were found to be suitable for distinguishing members of Group 1 from members of Group 2. A great deal of similarity in the isozyme patterns among the isolates of a same group, independently of geographic origin, indicated that isolates within a given group are descendant from a same ancestral population. Significant differences in the isozyme patterns between the isolates of Group 1 and Group 2 indicated, however, that the two groups were of different ancestry. Group 1 isolates have since been named F. pseudograminearum. Due to the short run time, minimal need of equipment, small sample volume and staining requirements, CAE generally provides a rapid and accurate method for isozyme analysis of Fusarium species.

Genetic variation within a collection of 25 isolates of Monascus from red rice and sofu was assessed with RAPD markers using genetic distances (1-Jaccard's coefficient) calculated for all combinations of isolates. Cluster analysis based on genetic distance was performed using the Ward's minimum variance method. Five distinct clusters were revealed based on genetic distances between and among clusters. The robustness (reproducibility) of the cluster assignments was tested by resampling (bootstrap) analysis. Cluster distribution was visualized as a three-dimensional graph based on multiple correspondence analysis. A dendrogram, based on the clusters, was constructed to examine the relationships. Three clusters accounted for 21 isolates; the fourth cluster consisted of three isolates which were quite distinct from each other and all other isolates. The fifth cluster, a single isolate from Japan, was very different from all others in RAPD patterns and was used as an outgroup. Resampling analysis indicated that the 25 isolates represented four genetic lineages of red rice fungi, suggesting that a relatively narrow genetic source of Monascus isolates is used in food products in Asia.

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.

We have designed primers for identification of the economically important plant pathogenic Rhizoctonia solani, for AG 2 and for each subgroup and an ecological type of AG 2. These specific primers have been designed based on specific sequences of the ITS regions in the R. solani species complex. The PCR procedure involves amplification of the 5.8S ribosomal DNA and part of the ITS regions, using the designed primers in combination with the general fungal primers ITS1F and ITS4B. Two of the primers amplify under optimal PCR conditions R. solani AG 1, AG 2, AG 3, AG 4, AG 5 and binucleate Rhizoctonia (BNR), and six more primers amplify specifically R. solani AG 2, the subgroups, AG 2-1, AG 2-2 and AG 2-3, and the ecological type AG 2-t. In this study DNAs from R. solani AG 2 and AG 4 growing on infected radish were amplified similarly to DNAs from axenic cultures. PCR detection has time saving advantages over traditional isolation methods for detection of Rhizoctonia on infected plant tissue and provides a powerful tool of identification.

DNA polymorphisms among Magnaporthe grisea isolates from rice and from finger millet were examined using restriction analysis of genomic DNA. The rice-infecting isolates produced 50–70 resolvable EcoRI fragments with the rice blast-derived repetitive sequence probe MGR586, while isolates infecting finger millet showed only one prominent band. Probes derived from a repetitive sequence (grasshopper) from a finger millet also distinguished the two pathogenic forms; grh pES 3–3 (35–70 EcoRI RFLPs) and grh pKE 2–2 (2–12 EcoRI RFLPs) hybridized exclusively with isolates infecting finger millet. Probes of two avirulence genes derived from rice blast genomes hybridised exclusively with rice blasts. These distinctions in Indian blast populations matched those observed elsewhere in Asia and in the Americas. No genomes were observed in the field that shared RFLPs from rice-infecting and finger millet-infecting isolates, even among isolates from a Himalayan area where the two forms have coexisted for centuries. Pathogenicity tests showed that the blast fungus from the two hosts did not cross-infect, nor did the two forms cross in the laboratory. The results confirm that the rice and millet-infecting M. grisea populations in India are distinct.

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.

Employing a single DNA primer targeted to clustered tRNA genes, a PCR system has been developed which can be used to uniquely identify different strains of the zygomycete Mortierella alpina. The procedure enjoys advantages over the RAPD-PCR technique in that the single primer operates robustly within a range of PCR conditions, and can be used successfully for members of many fungal genera. As a result of the tendency of some repetitive DNA elements to integrate near to tRNA gene clusters, the possibility that this technique may be extended to identify new examples of retrotransposons and tRNA-derived short interspersed repetitive elements (SINES) is discussed.

The genetic relationships of 56 isolates from the taxa traditionally grouped in Fusarium sections Fusarium (syn. Discolor) and Roseum were studied using the Restriction Fragment Length Polymorphism (RFLP) technique, by Southern hybridisation with random genomic and mitochondrial DNA probes originating from Fusarium species. Pairwise distances between taxa were calculated from the 957 RFLP bands scored on autoradiograms, using Dice's coefficient in the RAPDistance computer program. A strong genetic relationship was observed between F. graminearum, F. culmorum and F. crookwellense. However, the morphologically similar taxon, F. pseudograminearum shared only approximately 40% of genomic DNA RFLP bands with these taxa, demonstrating affinity, but less direct genetic similarity. Of all taxa examined F. pseudograminearum and F. graminearum shared the greatest similarity in mitochondrial DNA RFLP patterns. F. avenaceum subsp. avenaceum, subsp. aywerte and subsp. nurragi displayed very little genetic resemblance with each other, or with F. heterosporum and the cereal infecting pathogens. F. torulosum was closest genetically to subsp. avenaceum, with approximately 22% of genomic DNA RFLP bands in common. On the basis of these results F. avenaceum subsp. aywerte and subsp. nurragi are elevated to species rank, as F. aywerte comb.. nov. and F. nurragi comb. nov., and F. avenaceum subsp. avenaceum is returned to its former species status.

This study shows for the first time that Tolypocladium species produce efrapeptins, a group of toxic peptides, in vivo but the quantities are too small to account for insect death, suggesting that these insecticidal compounds work in concert with other pathogenicity determinants. There is inter- and intraspecific variation in efrapeptin production in vitro by Tolypocladium species. T. parasiticum produced only efrapeptin E, in small quantities. Efrapeptins were detectable 48 h after inoculation and increased with biomass. The relative amounts of individual efrapeptins (C, D, E, F, G) produced by T. niveum in vitro were D > E > F > C > G but in vivo they were D > F > C > E > G. Efrapeptins were toxic to a wide range of insects when injected into the haemocoel. Mortality was dose-related. Efrapeptins also exhibited limited antifungal and antibacterial activity. Micrococcus luteus was considered an excellent indicator of efrapeptin presence in culture filtrate extracts because of its extreme sensitivity to these compounds.

Fungal biomass was detected in peat soils from throughout the Everglades based on the presence of ergosterol. Ergosterol concentrations in soils were not detectably affected by the dominant plant, Cladium jamaicense or Typha domingensis, or phosphorus content of soils. In situ decomposition of decaying leaves, measured by respiration, was high (maximum 484 μl O2 h−1 g−1 dry biomass). Approximately 30% of respiration was by bacteria, and the rest was by fungi and other eukaryotes. Respiration rates were essentially the same for decomposing leaves of both plant species, with higher rates early in the decomposition process. Respiration rates were relatively unaffected by the nutrient status of the site, except for eukaryotic respiration on Cladium, which was usually higher at a high nutrient site. Ergosterol concentration increased in decaying leaves through time and was unrelated to the nutrient level except for Cladium, where it was higher at the high nutrient site. Eukaryotic respiration was not correlated with ergosterol concentration in decomposing leaves of Typha, but was positively correlated with ergosterol for Cladium.

Several species of Typhula cause diseases of cereals and grasses at low temperatures. Taxonomic separation of T. phacorrhiza, T. incarnata and T. ishikariensis was investigated using RAPD, ITS-RFLP and nucleotide sequencing of the ITS region. Using 10 RAPD primers on genomic DNA or eight restriction enzymes on PCR-amplified ITS regions, banding patterns were generated for each of 19 isolates from five species or varieties. Polymorphisms were found between species and varieties, with fewer among isolates within species or varieties. RAPD analysis suggested that T. ishikariensis var. idahoensis may be distinct from the other varieties (var. ishikariensis and var. canadensis), although this was not supported by ITS-RFLP. To confirm relationships, ITS nucleotides were obtained from single isolates of each of the five Typhula taxa, and sequence alignments were subjected to distance and parsimony analysis. Although there were differences in branching between dendrograms derived from sequencing vs. RAPD or ITS-RFLP, sequence data supported the distinction of the three main Typhula species, with the T. ishikariensis varieties grouping together and showing differences at an infraspecific level. ITS sequence data also indicated the closer taxonomic affinity of Typhula species to the agaricoid Tricholoma sejunctum compared to the clavarioid Thelephora terrestris.

The ITS regions of 14 isolates of endophytic fungi obtained from the root systems of four Woollsia pungens plants were sequenced and compared to sequences of Hymenoscyphus ericae, Oidiodendron maius and to sequences from other fungal taxa currently available in the GenBank and EMBL nucleotide databases. Sequence data suggested that six distinct taxa were present in the root systems of the four plants and that up to four of these were present in the root system of a single plant. One of the endophytes was identified as an Oidiodendron species, while most other isolates probably belong to the Leotiales.

Black mould lesions were caused by Alternaria alternata in 76% of 228 tomato fruit with characteristic sunken black lesions collected from fields of processing tomatoes in California. Analysis of 29 RAPD primers revealed a high level of genetic diversity among the 69 isolates tested. Two major phenetic groups (Group 1 with 55 isolates and Group 2 with 14) were identified independently by PCA and by UPGMA of Jaccard similarity coefficients. Only 34 of 137 RAPD markers were monomorphic for all isolates and the genetic similarity between the two groups was 50%. Co-infection of black mould lesions by genetically distinct strains of A. alternata occurred in two of 10 isolates tested. There was no evidence for geographic clustering of isolates with high levels of genetic similarity, suggesting that isolates are widely dispersed across California. Only one isolate was identified which also caused stem canker disease on a susceptible tomato cv., suggesting that these strains play a minor role in causing black mould on processing tomatoes in California. This isolate and two other known stem canker isolates were clustered together with 11 other isolates in Group 2. Group 2-specific bands were also identified in a survey of seven additional isolates known to produce host-specific toxins.

Extensive fungal growth has been detected on the walls and other building constructions in the inner parts of the Shelter of the damaged fourth Unit of the Chernobyl Nuclear Power Plant in 1997–98. The mycobiota comprised 37 species of 19 genera. Zygomycetes and ascomycetes were represented by one species each: Mucor plumbeus and Chaetomium globosum, respectively. Two mitosporic fungi commonly found were Cladosporium sphaerospermum and Penicillium hirsutum. Alternaria alternata, Aureobasidium pullulans, Aspergillus versicolor, Acremonium strictum, and Cladosporium herbarum were also encountered. Penicillium ingelheimense, Phialophora melinii, Doratomyces stemonitis and Sydowia polyspora were isolated from the Shelter and are recorded from the Ukraine for the first time. Comparison of the species growing under both severe and relatively weak radioactive contamination revealed a dominance of melanin-containing species in heavily contaminated sites; biodiversity and prevalence coefficients supported this.

Comparison of all presently available data on gasteromycetoid structure and development allows reassessment and leads to rectification of some questionable ideas about the relation of gasteromycetes to Agaricales. The gasteroid features which are dealt with are those which are particularly relevant to this subject. Some important, but specifically taxonomic, features are left out of the discussion. The loss of violent spore discharge is the only criterion to characterise basidiomycetes which have been united under the term gasteromycetes. Gasteromycetoid adaptations have probably arisen in the distant past at successive levels of basidiomycete evolution. Sometimes the shape of gasteromycetes is reminiscent of one of the groups from which they have descended and in other cases they have been subjected to an unique evolution in response to challenge by particular ecological requirements regarding spore dispersal.

Recombination between two strains of Metarhizium anisopliae, subcultured on artificial medium and passed through an insect host was investigated. When grown on agar culture, or passed through a host, strains did not show detectable genetic changes. When passed through an insect in the presence of the other strain, genotypic changes in the isolates, as detected by esterase electrophoresis, restriction digestion of total genomic DNA and RAPD-PCR, were apparent. The instability of molecular markers suggests recombination between strains.

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 phylogeny of the nuclear large ribosomal subunit (LSU) rDNA from Peronosporomycetes (Oomycota) was studied. Five orders Rhipidiales, Leptomitales, Saprolegniales, Pythiales and Peronosporales were included in the study in order to reveal phylogenetic relationships within the group. Based on maximum parsimony, neighbour-joining and maximum-likelihood the Peronosporomycetes seem to have evolved as two major lineages. One lineage includes members of the Rhipidiales, Leptomitales and Saprolegniales and the other includes the members of Pythiales and Peronosporales. However, the inclusion of Rhipidiales in Saprolegniomycetidae was not well supported. Leptomitales were placed basal to the Saprolegniales in the LSU rDNA based tree. In the Saprolegniaceae sensu lato the primary zoospore seems to have been lost several times in the evolution of this family. Species of Aphanomyces were placed on the most basal branch in this family. The obligate parasite, Albugo, was placed on the most basal branch within the Peronosporomycetidae. Phytophthora showed a closer relationship to Peronospora and Peronophythora than to Pythium suggesting that this genus should be removed from Pythiales to Peronosporales.