An assay based on the competitive polymerase chain reaction (PCR) was developed to quantify Glomus mosseae, an arbuscular mycorrhizal (AM) fungus, within plant roots. Using previously designed G. mosseae specific primers, a heterologous internal standard was constructed by amplifying Pseudomonas DNA under low stringency annealing conditions. Co-amplification of G. mosseae and internal standard DNA within leek root extracts provided accurate quantification of target DNA. Colonization of leek roots by G. mosseae was monitored in a comparative study by competitive PCR and microscopy, a conventional method of quantification. These two methods gave closely parallel data for G. mosseae colonization from three different inoculum levels over a 6 week period. Results indicate that competitive PCR is a sensitive and accurate method of quantification. The major advantage of competitive PCR over microscopy is that it can quantify specific AM fungi.

The amount of Gaeumannomyces graminis var. tritici inoculum was quantified by slot-blot hybridization assay, using a specific DNA probe, pG158. Estimates of the amount of G. graminis var. tritici DNA in naturally infested and inoculated soils were made and correlated with take-all disease severity as determined by a conventional soil bioassay (expressed as a percentage of seminal roots with G. graminis var. tritici lesions). Disease severity in the glasshouse assay and DNA levels were highly correlated. The level of take-all disease severity in naturally infested soil can be predicted using the equation yi=53·078 log (xi)+88·866, where yi=the predicted value of the disease severity (%) and xi=amount of G. graminis var. tritici DNA (ng) in 100 g soil. The DNA-based assay for quantifying the amount of G. graminis var. tritici has been further developed and improved in terms of the sampling techniques and efficiency. Soils of different types were collected to a depth of 10 cm and organic matter between 0·5–1·4 mm in size was separated from this soil. Using a slot-blot hybridization assay, it was found that G. graminis var. tritici was mostly present in the organic matter fraction greater than a size of 0·5 mm and in soil at a depth of 5 cm. This rapid and reliable DNA-based assay is being used in conjunction with a model of take-all disease development to predict disease levels in the field due to G. graminis var. tritici.

Torrubiella petchii is described from scale insects that infest bamboo leaves in tropical forest. It is compared with four other species of Torrubiella described from Thailand and with other species known to infect scale insects. T. petchii was successfully isolated from whole ascospores. No anamorph was found in the material collected from the field. One isolate produced a Hirsutella anamorph. This is compared with Hirsutella-like anamorphs, collected from the field, of two other scale-insect Torrubiella spp.

3- and 7-hydroxyflavone and their acetic acid esters were tested against Cladosporium herbarum and Penicillium glabrum for antifungal activity. The effect of the hydroxylated flavones was relatively low whereas the ester derivatives were substantially more active.

Three new species of Stenella, S. heterophragmae, S. lantanae and S. buteae occurring on Heterophragma sp., Lantana indica and Butea parviflora, respectively, have been described and illustrated.

The pathogenicity of two isolates of Verticillium lecanii towards non-target invertebrates and six species of aphid was tested in bioassays. There was no evidence of infection in any of the 20 non-target invertebrates. All target aphid species were susceptible to at least one isolate. The level of susceptibility varied between aphid species, with Myzus persicae and Brachycaudus helichrysi showing the greatest susceptibility.

Studies on initial spore deposition, retention and germination on two susceptible aphid species and four resistant non-target insects revealed differences in the behaviour of the pathogen between the two groups. Spore density mm−2 was greater on susceptible insects both immediately post-inoculation and following 24 h incubation. There was no loss of spores from M. persicae over 24 h. All other species showed more than 50% spore loss in the same period. However, even after 90% spore loss, large numbers of spores remained on the bigger insects such as Agonum dorsale. Germination and germ-tube growth were possible on such resistant non-target insects, indicating that resistance to infection occurs after this stage. There was little difference in rate of germination on the two groups. However, differences in germ-tube growth were evident, with the production of longer and narrower germ-tubes on the resistant insects.

A homothallic species of Nectria producing a Cylindrocladium-like anamorph was collected from bark of a fallen tree in the Amazonian forest in Ecuador. The anamorph, which is placed in a new genus, Xenocylindrocladium, is characterized by forming straight, cylindrical, 1-septate conidia borne on penicillate conidiophores with coiled, avesiculate stipe extensions. The teleomorph, which is best accommodated in Nectria, is distinct in forming yellow-orange perithecia with red ostiolar regions, ellipsoidal, smooth, hyaline, 1-septate ascospores, and long-stalked, cylindrical asci with apical discharge mechanisms. Both the teleomorph and anamorph states are newly described as Nectria serpens and Xenocylindrocladium serpens.

Monoclonal antibodies were raised against the mycelium of a litter-decomposing basidiomycete, Mycena galopus, for use in an autecological study of the fungus in forest litter. Two clones (3E12 and 6D8), produced using particulate and whole-cell extracts of the fungus respectively, cross-reacted with several species including other Mycena species, non-Mycena basidiomycetes, zygomycetes and anamorphs or mitosporic fungi. Of these clones, 3E12 appeared to be directed against an unidentified β-glucan and 6D8 against chitin. A third clone (6E1), using as immunogen latex collected in the field from stipes of the basidiomes, was specific to M. galopus mycelium in ELISA and immunofluorescence tests and was probably directed against an unidentified polysaccharide.

Cerato-ulmin (CU) is a low molecular weight hydrophobin protein secreted at high levels in vitro by the highly aggressive Dutch elm disease pathogen, O. novo-ulmi, but at much lower levels by the less aggressive O. ulmi. The recent discovery of an endemic Dutch elm disease fungus in the Himalayas, Ophiostoma himal-ulmi, which is a separate species from O. ulmi and O. novo-ulmi, yet like O. novo-ulmi produces high levels of CU in vitro, suggested that O. novo-ulmi may have acquired its cu gene from O. himal-ulmi. To investigate this possibility we have compared the nucleotide sequences of the cu gene isolates of O. himal-ulmi with those of O. ulmi and of the EAN and NAN races of O. novo-ulmi. Alignments of nucleotide sequences and their derived amino acid sequences are presented, allowing estimations of evolutionary relationships to be made. These comparisons indicate that the cu gene of O. himal-ulmi is less closely related to the cu gene of either O. ulmi or O. novo-ulmi than the cu gene of O. novo-ulmi is to that of O. ulmi; hence the cu gene of O. novo-ulmi is unlikely to have been acquired recently from O. himal-ulmi.

The haustorial morphology of 21 Ravenelia species, two Kernkampella species, and four leguminicolous Uredo and Uraecium species which are probably related to Ravenelia, was investigated by light microscopy. Three different kinds of interaction could be found: (1) species with haustorial complexes comprising dikaryotic haustoria and intracellular hyphae. These species showed strictly intracellular growth of the parasitic mycelium. (2) Species with only dikaryotic haustoria, and (3) species with only intracellular hyphae or monokaryotic haustoria. Complexes of dikaryotic haustoria and intracellular hyphae were found in 17 of the investigated Ravenelia species and three Uredo species. Three Ravenelia species revealed dikaryotic haustoria, and Ravenelia lonchocarpicola had only intracellular hyphae. Complexes of dikaryotic haustoria and intracellular hyphae are probably the prevalent type of parasitic interaction in Ravenelia. The investigated Kernkampella species revealed dikaryotic haustoria.

The process is described of teliospore germination in Tilletia opaca and T. sumatii leading to the production of basidiospores on basidia, and later the development of ballistospores and blastospores on agar. In both species the basidium shows some evidence of being a condensed dichotomously branched structure. The situation in these two smuts is discussed in relation to that in several other species of Tilletia, and some consideration is given to the origin of the basidium and evolutionary trends within the genus.

Restriction fragment length polymorphisms (RFLPs) were identified among total DNA from clonal lines of Uncinula necator when cloned sequences of total U. necator DNA were used as probes. Four probes, pUnP14, pUnP27, pUnE21 and pUnE4, hybridized to multiple-copy sequences and, with the exception of pUnE4, detected genetic variation among clonal lines of U. necator. Clones pUnP14, pUnP27 and pUnE4 produced banding patterns that were stable for DNA extracted from different asexual generations of U. necator clonal lines over at least 15 months. In addition, clones were evaluated for species specificity. Clones pUnP27 and pUnE4 detected only U. necator sequences in total DNA from infected grapevine leaves. Clones pUnP14 and pUnE4 did not hybridize to total DNA from a range of fungi.

Genetic diversity in a sample of the Australian U. necator population was investigated; 15 genotypes were identified among 29 U. necator clonal lines examined. Genetic variation was detected in samples collected within micro-geographical areas, for example, different genotypes representing both mating types of U. necator were detected on a single plant of Vitis amurensis. RFLP analysis of the banding patterns produced using pUnP14, pUnP27 and pUnE21, identified two broad genetic groups, designated A and B. Analysis of DNA fragment patterns obtained using the polymerase chain reaction (PCR) and the plant intron splice junction (ISJ) primer R1 also supported the allocation of U. necator clonal lines into groups A and B.

Seven hundred and eighty two isolates corresponding to 52 fungal taxa have been obtained from 920 segments of healthy and symptomatic twigs of Eucalyptus grandis, incubated on media with water potential −0·138 MPa and −4·19 MPa. Only thirty taxa were isolated at frequencies of 2% and greater. The composition of species isolated from bark of healthy twigs incubated on media with both water potentials was similar, with Aureobasidium pullulans being dominant. Sistrotrema brikmannii was the main species isolated from xylem of healthy twigs at −0·138 MPa whereas Pleospora sp. and Sphaeropsis eucalyptus were the dominant species isolated from this material on medium at −4·19 MPa. Fusicoccum eucalypti and Cytospora chrysosperma were most commonly isolated from bark and xylem of symptomatic twigs. Correspondence analysis showed the fungal species associated with each tissue. Summer drought followed by frequent frost in early autumn may have incited wounds allowing fungal invasion of the bark of twigs.

Two new species, Pseudocercosporella arcuata and P. woodfordiana, on Rubus ellipticus and Woodfordia floribunda, respectively, are described and compared with related taxa.

To imbibe and germinate urediniospores require free water or near saturated atmospheres in which both adsorption of water is maximal and capillary condensation pads form. The rate of spore germination is influenced by the amount of free water available. During imbibition, spores swell and capillary pads form beneath spores in contact with the substratum. These pads greatly increase the area of contact with the substratum. Pad area fluctuates and may depend on the balance between the rate of capillary condensation and the rate of uptake of fluid by spores. When spores, on leaves or cellulose acetate, are misted with water they imbibe rapidly and germinate. Adhesion pads which form under fully imbibed, germinating spores are morphologically similar to capillary pads but they differ in composition. On synthetic surfaces, during imbibition of spores and under microscopic glass beads, pads are composed principally of volatile capillary condensation and are removed when samples are freeze-dried. On host surfaces, when spores are fully imbibed and germinating, pads contain non-volatile soluble materials derived from both pathogen and host. These substances are not removed by extended freeze-drying.

Three fungicolous hyphomycetes are described and illustrated. Anastomyces gen. nov., with the type species A. microsporus sp. nov., is described on ascomata of Apiospora sp. on dead culms of a grass from China. It differs from comparable genera by its fungicolous habitat, sporodochial conidiomata and polylocal conidiogenous cells mostly with 2 loci. Each locus produces one conidium but two conidia formed from the same conidiogenous cells join together to form H-shaped conidia in the earliest stages of their development. Idriella rhododendri sp. nov. grows over other microfungi and is described from leaf litter of Rhododendron ponticum from England. Infundibura adhaerens was found to be commonly associated with ascomata and conidiomata on dead leaves of R. ponticum in England.

Serine proteinase activity was assayed during sporophore maturation and postharvest senescence in Agaricus bisporus. Serine proteinase accumulated largely in stipe tissue early during postharvest storage and in the later stages of maturation of unpicked sporophores. Western blot staining correlated with enzyme activity indicating de novo synthesis. The enzyme could not be detected by Western blotting in freshly harvested sporophores at an early maturation stage. Small increases in activity were detected in the cap tissue during the final stages of senescence, postharvest and in unpicked sporophores. The distribution of the serine proteinase in senescent sporophore tissues (high in stipe, low in cap) was confirmed by histochemical and immunologically stained tissue prints.

Peridermium harknessii, cause of western gall rust of pines, comprises two populations of multilocus electrophoretic types (zymodemes) in the western United States. When stained with a DNA-specific fluorochrome, mature, ungerminated aeciospores from zymodeme I were found to be predominantly binucleate (70%), as were those of the related macrocyclic species, Cronartium quercuum (74%), whereas aeciospores from zymodeme II were predominantly uninucleate (93%). Within each zymodeme, aeciospores with two nuclei had significantly (P=0·01) more DNA than spores with one nucleus, and numbers of nuclei in germlings increased arithmetically over time. These data suggest that aeciospore nuclei in both zymodemes I and II divide mitotically, not meiotically, as is consistent with an asexual life cycle.

Photometric measurements also indicated that the amount of DNA in one nucleus of a uninucleate zymodeme II aeciospore was similar to the total amount of DNA in a binucleate zymodeme I aeciospore. These data, coupled with recent isozyme studies, suggest either that zymodeme II evolved after karyogamy of zymodeme I and an unidentified zymodeme, or that zymodeme I evolved after haploidization of zymodeme II.

Genetic variation of the haploid canker pathogen fungus Gremmeniella abietina was investigated by RAPD marker analysis. A total of 126 single pycnidia isolates of G. abietina were sampled from one Pinus contorta plantation in northern Sweden. Average Nei's gene diversity over 32 putative RAPD loci was 0·29. Multilocus genotype analysis revealed 85 different genotypes among the 126 isolates sampled, of which 66 genotypes were detected only once. Nineteen genotypes were detected more than once among 41 isolates, representing 32·5% of clonal fraction of the sampled population. Gametic disequilibrium tests of the sampled population revealed significant associations between 29·6% (147 of 496) of the RAPD loci. When the clonal fraction was removed from the data set, the number of significant pairwise loci combinations decreased from 147 to 73 (14·7%), indicating a high degree of random mating. These results are consistent with previous field observations suggesting that sexual fruiting bodies of G. abietina were common in northern Scandinavia. The high genetic variability detected in the population also suggests that the population was founded by a relatively large number of unrelated genotypes.

The internal transcribed spacer regions (ITS1 and ITS2) of the ribosomal RNA gene repeat from Phytophthora species were amplified using the polymerase chain reaction and sequenced. Sequences from P. cambivora, P. cinnamomi, P. citricola, P. cryptogea, P. drechsleri, P. fragariae var. fragariae, P. fragariae var. rubi, P. megasperma var. megasperma and P. nicotianae were compared with published sequences and phylogenetic trees were produced. The resultant grouping of species generally agreed with groupings established using classical morphological criteria, primarily sporangial morphology. Amongst species with non-papillate sporangia two clades were evident, one consisting of P. fragariae, P. cambivora and P. cinnamomi and the other of P. megasperma, P. drechsleri and P. cryptogea. The latter three were placed in the tree between the non-papillate groups and the papillate and semi-papillate groups which formed three distinct clades. One group comprised P. citricola, P. citrophthora and P. capsici, another P. megakarya and P. palmivora and a third P. pseudotsugae, P. cactorum, P. idaei, P. nicotianae and P. infestans. More isolates of P. megasperma, P. drechsleri and P. cryptogea will need to be examined to settle more precisely the relationship of these species to the others. PCR amplification of ITS sequences using freeze-thawed mycelial scrapings from pure cultures growing on agar followed by digestion with restriction enzymes is a quick and easy way to compare and identify isolates without the need for laborious DNA extraction procedures. With improved technology, rapid automatic sequencing of PCR-amplified ITS regions is now possible and yields detailed information of relationships within the genus as well as allowing the design of species-specific PCR primers for diagnostic purposes.