The nematophagous fungus Pochonia chlamydosporia is a potential biocontrol agent against root knot and cyst nematodes. Genetic transformation of the fungus to introduce visual marker genes, novel traits, or changes in expression levels of endogenous genes, would greatly enhance understanding of its behaviour on nematode-infested roots and of its interactions with other soil and rhizosphere microorganisms. A transformation system for the introduction of novel genes into P. chlamydosporia has been developed. Methods to generate protoplasts, introduce DNA and regenerate transformed viable fungal mycelium have been optimised, using plasmids carrying the green fluorescent protein marker gene gfp and the hygromycin resistance gene hph. Cultures of P. chlamydosporia were resistant to high levels of a range of fungal inhibitors, including hygromycin, that are commonly used with dominant selectable marker genes in the transformation of other fungi. However, regenerating protoplasts transformed with hph could be selected by their ability to grow through an agar overlay containing 1 mg ml−1 hygromycin. Green fluorescence was observed in protoplasts and regenerating mycelium after transformation with gfp, but the GFP phenotype was lost on subculture. Maintenance of introduced genes was not stable, and during subculture, PCR assays indicated that the transformants lost both hph and gfp. When these genes were introduced on the same plasmid, segregation of hph and gfp was observed prior to their loss. It was unclear whether the introduced plasmids were able to replicate autonomously in P. chlamydosporia, or if they integrated transiently into the fungal genome. Possible reasons for the instability of the transformants are discussed.

A key to 17 species in the genus Hygrocybe, subgenus Pseudohygrocybe, sections Coccineae and Neohygrocybe sensu Boertmann is provided for the Greater Antilles. Five new species and five taxa that are new reports for the region are described. The new species in section Coccineae are H. pseudoadonis, H. viridiphylla, and H. zonata. The new species in section Neohygrocybe are H. albomarginata and H. ovinoides. The new reports are H. caespitosa, H. coccinea, H. cf. miniata, H. papillata, and H. subovina. Three new combinations are proposed: Hygrocybe mycenoides, H. papillata and H. subovina.

Creolophus cirrhatus, Hericium erinaceus and H. coralloides were paired against over 20 other wood decay fungi from beech (Fagus sylvatica) covering a range of ecological strategies, on 2% malt agar (MA), 0.5% MA, 0.5% MA adjusted to −1.25 MPa by addition of KCl, 0.5% MA adjusted to pH 4 with KOH/H3PO4 and 0.5% MA under 5% O2/30% CO2 all at 20 °C. Creolophus cirrhatus and H. coralloides were also paired against 17 other species in wood, incubated at 20 ° for 36–38 wk. They were average to good combatants, deadlocking with or replacing over half of the antagonists, H. erinaceus being slightly more combative than the other two species. Outcomes in wood were representative of those in agar. Abiotic conditions altered the outcomes slightly, reduced water potential favouring C. cirrhatus and H. erinaceus. Overall extension rate of C. cirrhatus and H. erinaceus often increased in the presence of antagonists. There was prolific production of fruit bodies in controls and interactions, though notably not under altered gaseous regime. Results are discused in an ecological context.

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.

During a survey of 375 strains of the Xylariales, one isolate (F127076) was observed to have an ITS1 size of 833 bp. This size exceeds the average ITS1 size in the Xylariales (mean=209±57 bp). Comparison of the DNA sequence with GenBank and with a proprietary DNA database revealed low homology with Xylaria hypoxylon ATCC 42768, and with one undescribed Xylaria species. When the ITS2 sequence was compared, these isolates were 96–98% homologous. Sequences of other variable genes confirmed the relatedness among these strains. A closer observation of the ITS sequence of this isolate revealed the presence of three repeated domains of 250 bp plus one truncated domain, showing 52–75% homology. Sequence similarity suggests that the repeated domain is derived from the fusion of the ITS1 with a DNA fragment derived from the γ-glutamyltranspeptidase gene. This example suggests that the rate of evolution of ITS1 can be independent of the rate of evolution of other genes, even when this variability is not a result of slipped strand misspairing events like in other Xylariales. This observation also indicates that recombination with other nuclear genes could participate in the evolution of the internal transcribed spacer.

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.

Dark septate root endophytes (DSE) are an artificial assemblage of fungi that have darkly pigmented, septate hyphae and that are frequent or distinctive intracellular associates of roots of apparently healthy plants. Based on isolates obtained from the roots of Salix spp., the distribution of a common DSE fungus, Phialocephala fortinii, was examined along a latitudinal transect in Canada running from the high arctic to the 49° N parallel. Non-sporulating isolates were provisionally identified as P. fortinii through analysis of DNA sequence data of the ITS2 region of rDNA. P. fortinii was isolated frequently from boreal and arctic habitats, but rarely from grassland habitats. Patterns of genetic variation were examined through analysis of amplified fragment length polymorphisms (AFLP). All AFLP profiles were unique with the majority of genetic variation occurring among individuals within the collecting sites at each latitude. Neighbour-joining analysis of genetic distances yielded eight well-supported clusters, three of which included individuals from more than one latitude. Some linkage disequilibrium, possibly due to partial clonality, was detected.

Competitive interactions among bark beetle associated fungi are potentially influenced by abiotic factors. Water potential, in particular, undergoes marked changes over the course of beetle colonization of tree hosts. To investigate the impact of water potential on competition among three southern pine beetle associated fungi, Ophiostoma minus, Entomocorticium sp. A and Ceratocystiopsis ranaculosus, we utilized artificial media with water potentials of 0, −5, −10, and −20 MPa. Growth of all three fungi, when grown alone, decreased on media with lower water potentials. Growth rates of all three fungi were likewise reduced in competition experiments. At −5 to −10 MPa, C. ranaculosus (a fungus with beneficial effects toward southern pine beetle) was nearly equal in competitive ability to O. minus (a fungus with antagonistic effects towards southern pine beetle). This was not true on control media, nor at other water potentials tested. The range of water potentials used in our assays was similar to the range of water potentials we measured in loblolly pines within a southern pine beetle infestation. This study indicates that water potential may alter the outcome of competitive interactions among bark beetle-associated fungi in ways that favour bark beetle success.

Phoma medicaginis was isolated as the dominant endophyte from surface-sterilized shoots of Medicago sativa and M. lupulina growing outdoors. Plants were either symptomless or showed signs of infection in the shape of limited lesions which sometimes contained melanized pycnidial initials. Rapid colonization of host tissue and sporulation were observed within 9 d on dead plant material upon incubation in a moist chamber. Such colonized material, but not freshly harvested living tissue, contained brefeldin A (1.7 μg g−1D.W.). This toxin was also produced in pure culture (20 mg l−1) and in artificially inoculated autoclaved M. sativa stems (3 mg g−1D.W.=920 μg ml−1). The latter concentration of brefeldin A should be similar to that produced within a fruiting lesion of P. medicaginis and suppressed spore germination and growth of nine of 11 common phylloplane fungi tested. This metabolite may thus have a function in substrate defence after the switch from the endophytic to the saprotrophic period in the life-cycle of P. medicaginis following the death of infected host tissue.

The new species, Termitomyces bulborhizus, is described from south-west China. The stipe of the basidiome is swollen into a prominent bulbous base at ground level, and this, together with the floccules on the stipe surface, distinguishes this species within the genus. T. bulborhizus is compared with closely related species, including three originally described from China, and a key to T. bulborhizus and related species is provided.

Molecular techniques have become increasingly important for the identification of arbuscular mycorrhizal fungi (AMF). In this work Acaulospora longula and Glomus mosseae have been detected in plant roots from pastures using specific nucleotide primers for the two species. Part of the 5′ end of the large subunit of the ribosomal RNA gene was amplified by nested PCR and sequenced. The distribution of the fungi within three different plant species, Plantago lanceolata, Trifolium repens, and Holcus lanatus, and two different types of grassland, have been studied. Neither the fungi nor the plants showed specific preference for their symbiotic partnership.

Terrestrial invertebrates in central Amazonian floodplains must cope with annual long-term inundation. Parasites should be affected mainly indirectly through the specific life-cycles of their hosts. We studied the temporal structure of a beetle–fungus system at a central Amazonian blackwater floodplain (Rio Negro, Brazil). The host species Phaeoxantha aequinoctialis (Coleoptera, Carabidae, Cicindelinae) showed a seasonal, univoltine life-cycle triggered by the annual flood pulse. Infestation frequency of its fungal parasite, Laboulbenia phaeoxanthae (Ascomycota, Laboulbeniales), varied seasonally. However, the seasonality was opposed in host and parasite: the lowest infestation frequencies were observed during periods of highest beetle abundance and vice versa. Periods of lowest beetle abundance coincided with the end of the old generation, those with highest abundance with the appearance of new adults. The resulting annual patterns of a slow spread in the host population resembled the few records of temporal patterns from temperate regions. It is explicitly demonstrated that older adult female beetles are more frequently infested than younger ones. Future studies may reveal whether this is simply the result of specific host life-cycles (driven by a flood pulse, winter, or other factors), or might also be related to potentially easier infestation in older individuals.

Morphologically different thalli of Laboulbenia richardiana sp. nov. (Laboulbeniales, Ascomycota) were found on various parts of the body of the South American fly Richardia teevani. These thalli lack a separate cell IV and have similar appendages and perithecial apices. They are interpreted here as ‘growth forms’ of a single biological species. Spore characteristics and insect behavior support this view, rather than the hypothesis of several different species displaying position specificity.

Silver scurf is an important postharvest disease affecting potato tubers worldwide, caused by Helminthosporium solani. In the present study, key steps of infection of potato tubers (cv. ‘Dark Red Norland’) by H. solani were described using transmission (TEM) and scanning electron microscopy (SEM). The fungus entered potato tubers mainly via hyphae, although germ tubes were also able to directly penetrate the tubers. An extracellular sheath was observed around hyphae growing over the surface of tubers and the host cell wall appeared lyzed at the point of penetration. Observations suggested that both mechanical and enzymatic processes are involved in periderm penetration. Hyphae of H. solani, 9 h after tuber inoculation, were present intracellularly mostly in the periderm and in some cortical cells. Two days after inoculation, host cells were invaded and both infected and neighbouring host cells showed signs of necrosis (disrupted cytoplasm, absence of typical organelles or endomembrane systems, collapsed peridermal cells) that were not observed in healthy control tubers. Four days after inoculation, completing the infection cycle, conidiophores emerged from peridermal cells directly by erupting through the host cell walls.

Growth and interactions of Piptoporus quercinus were studied in artificial culture to give insights into the ecological characteristics of a species which is both rare and widespread. P. quercinus grew slowly under all conditions, the maximum ranging between 1.9 and 3.15 mm d−1, depending on isolate, at 25 °C pH 3.75. Upper limits for growth were 25–30 ° or above 30 °, depending on isolate, and pH 6.6. Lower limits for growth were 5–10 °, between −1.75 and −2.5 MPa; there was still growth at pH 1.81. Preincubating plates at 5 ° for 77 d and then incubating at 20 ° resulted in a significantly greater extension rate (2.89 mm d−1) than when incubated continuously at 20 ° (1.70 mm d−1) for one isolate though not for another isolate. P. quercinus survived exposure to low temperature (−18 ° for 7 d), showing renewed growth, albeit limited, after a lag of 37 d. It was a poor combatant, being unable to replace any of the eight fungi (with a range of ecological roles) against which it was paired, and being replaced or partially replaced by most, on 2% MA at pH 5.49, at pH 3.75 and at −1.25 MPa. It did, however, inhibit extension rate of some other heart rot fungi. Results are discussed in an ecological context.