Production Of Pycnidia Research Articles

Production of Macrophomina phaseolina Conidia by Multiple Soybean Isolates in Culture.

Macrophomina phaseolina is the cause of charcoal rot of soybean (Glycine max). Resistance to M. phaseolina in commercial soybean cultivars is not common but is needed in locations where the disease is chronic and severe. The objective of this study was to develop a reliable method to produce sufficient M. phaseolina conidia that can be used to inoculate soybean plants in a high-throughput resistance-screening program. Production of pycnidia is not common on most culture media, such as potato dextrose agar, but can be produced on media containing plant parts or oilseed extracts. Seven semi-defined media were tested to induce pycnidia production. Results indicated that the number of pycnidia that were produced by eight M. phaseolina isolates was dependent on induction medium; however, peanut butter extract-saturated filter paper placed over soynut butter extract agar (PESEA) allowed for greater pycnidia and conidia production than the other media tested. Production of pycnidia on PESEA ranged from 269 to 1,082 per plate. There were no differences among isolates in germination of conidia produced on PESEA, which averaged 83 ± 2% germination. A conidial suspension from one M. phaseolina isolate produced on PESEA and inoculated onto soybean radicles significantly distinguished (P < 0.01) 'DT97-4290', a soybean genotype with partial resistance to charcoal rot, from a susceptible genotype, 'LS98-0358'. Results of this study indicated that multiple isolates of M. phaseolina from soybean produced sufficient amounts of conidia on PESEA to use as inoculum. This conidia inoculum production method will facilitate soybean charcoal rot resistance screening evaluation with different soybean isolates.

Microsphaeropsis olivacea as an etiological agent of human skin infection

Microsphaeropsis olivacea is reported as the agent of a case of human skin infection in an otherwise healthy woman. This fungus has not been reported previously as causing disease in humans. It was identified on the basis of its production of pycnidia and conidial structures typical of the Coelomycetes, and by its light brown, ellipsoid to cylindrical and thick walled conidia. The in vitro inhibitory activity of amphotericin B, fluconazole, flucytosine, itraconazole, ketoconazole and miconazole was determined.

Breaching by a new strain of Leptosphaeria maculans of anatomical barriers in cotyledons of Brassica napus cultivar Surpass 400 with resistance based on a single dominant gene

Infection processes were examined to investigate the breach by a strain of Leptosphaeria maculans of anatomical barriers in cv. Surpass 400, a cultivar containing single dominant gene-based resistance (SDGBR). Two strains, UWA 192 and UWA P11, were used to inoculate cvs. Surpass 400 and Westar. The pre-penetration and penetration behaviour of both strains was similar in both cultivars. However, they differed significantly after penetration. When UWA P11 infected cv. Surpass 400 through stomata, guard cells rapidly died within a few hours and the surrounding mesophyll cells became necrotic, constituting a hypersensitive reaction (HR). Hyphal growth continued, albeit slowly, through the intercellular palisade mesophyll and spongy mesophyll spaces, but hyphae rarely spread beyond the HR region, and did not sporulate. Polyphenolic compounds accumulated in the area bordering the HR. However, when UWA 192 infected through stomata, symptoms were not evident until 10–12 days postinoculation (dpi) and were typically characterized by pale tan to grey circular lesions in which abundant pycnidia were produced by 14 dpi. Subsequently, hyphae extensively spread beyond the lesion border, reaching the veins and progressing down the petiole towards the stem. Where the SDGBR remained effective (i.e. against strain UWA P11) death of cells was restricted to a few palisade cells within the HR, even though hyphae were present in the lower tissue layers of the cotyledon. In contrast, where the SDGBR was not present (cv. Westar) or was overcome (cv. Surpass 400 with UWA 192), extensive death of epidermal and upper and lower palisade cells occurred throughout infected areas of the cotyledon, with subsequent abundant production of pycnidia. Polyphenolic compounds, which are also associated with resistance, did not accumulate in this instance. It was evident that the ability of the host to instigate the HR mechanism displayed by cv. Surpass 400 was lost with UWA 192 resulting in a “normal” susceptible response. This is the first study of the specific processes involved in the breaching of the HR in cv. Surpass 400.

Production of Diplodia scrobiculata and Diplodia pinea pycnidia on ground Austrian pine needle agar medium

The in vitro production of fruiting structures represents an important tool for the morphological identification of fungal genera or species. It is also important for the controlled production of spores to be used in experiments. However, some fungal species do not readily sporulate in pure culture. In the present study we induced the production of pycnidia of Diplodia scrobiculata and D. pinea, two species recalcitrant to sporulation in pure culture, by growing them on two media containing ground Austrian pine needle. The two fungal species grew equally rapidly on both media and pycnidial primordia were produced on the medium surface after only 4 days at room temperature. Conidia matured in less than two weeks and their germination rate at 25°C was about 96%, indicating high viability.

Saprophytic Colonization of Citrus Twigs by Diaporthe citri and Factors Affecting Pycnidial Production and Conidial Survival.

Melanose, caused by Diaporthe citri, produces reddish brown lesions on the fruit, leaves, and twigs of citrus trees, and greatly reduces the marketability of fresh fruit. Most of the inoculum is produced in pycnidia on dead twigs in the tree canopy, which exude large numbers of conidia in slimy masses. In this study, detached twigs inoculated with conidia were readily colonized and produced large numbers of pycnidia within 30 to 40 days when they were soaked 3 to 4 h on alternate days. Conidial production was measured by wetting twigs in a rain tower periodically and collecting the conidia in the runoff water. Production began after 80 days and continued for nearly 300 days. In other experiments, production of mature pycnidia on detached twigs was greatest at 94 to 100% relative humidity (RH) and at 28°C. Low RH and temperature, however, favored survival of conidia in exuded masses on twigs. In the field, colonization of detached twigs by D. citri was high in rainy season, moderate in spring and early fall, and minimal in late fall and winter. Twig colonization was positively related to the number of rain days and average temperature, but not to total rainfall. In another experiment, inoculated twigs placed in the tree canopy developed pycnidia and then produced conidial masses for about 200 days. D. citri is a serious pathogen, but a weak parasite, that survives primarily by colonization and reproduction on dead twigs.

Caracterização fisiológica, cultural e patogênica de diferentes isolados de Lasiodiplodia theobromae

Lasiodiplodia theobromae é responsável por doenças em inúmeras plantas cultivadas, causando perdas significativas de produção, notadamente no Nordeste brasileiro. Estudos básicos sobre esse fungo são necessários para se compreender melhor as interações patógeno-hospedeiro e traçar estratégias de controle. Este trabalho teve como objetivo caracterizar oito isolados de L. theobromae, obtidos de diferentes hospedeiros, quanto ao crescimento micelial, produção e fertilidade de picnídios, aspectos morfológicos das colônias e patogenicidade. Os isolados variaram em todas as características estudadas. O meio aveia-ágar proporcionou a maior produção e fertilidade de picnídios, sobressaindo-se os isolados obtidos de manga e maracujá. O meio cenoura-ágar não promoveu a produção dessas estruturas. Quanto ao crescimento micelial, o meio V-8 promoveu maior velocidade de crescimento dos isolados. Em relação à coloração da colônia houve grande variação, predominando a coloração branco-acinzentada. Quanto à formação dos picnídios, verificou-se que a maioria dos isolados formaram tais estruturas. Em ensaios de inoculação cruzada, todos os isolados mostraram-se patogênicos aos hospedeiros testados.

Correlative analysis of Mycosphaerella graminicola pathogenicity and cell wall‐degrading enzymes produced in vitro: the importance of xylanase and polygalacturonase

Eight Mycosphaerella graminicola isolates were investigated for correlations between pathogenicity and the in vitro production of cell wall‐degrading enzymes. Isolate pathogenicity was evaluated in terms of lesion and production of pycnidia in wheat leaves. Additionally, the isolates were compared over time for their ability to produce in vitro significant levels of xylanase (EC 3·2·1·8), β‐xylosidase (EC 3·2·1·37), β‐1,3‐glucanase (EC 3·2·1·6), cellulose (EC 3·2·1·4) and polygalacturonase (EC 3·2·1·15) activities when grown in a liquid medium. Correlation tests and principal component analysis revealed a significant correlation between the in vitro production of xylanase and pectinase and pathogenicity components. Xylanase was correlated to necrosis frequency (r = 0·795), β‐xylosidase was correlated to the mean of the lesion length (r = −0·787), whereas polygalacturonase was correlated to the time when 50% of the leaves contained a lesion (r = 0·776), the lesion frequency (r = 0·646) and the time when 50% of the leaves showed pycnidia (r = −0·711). The results suggest that these two groups of cell wall‐degrading enzymes are therefore likely to be key determinants of pathogenicity in M. graminicola.

Influence of culture media and environmental factors on mycelial growth and pycnidial production of Sphaeropsis pyriputrescens

Sphaeropsis pyriputrescens, the causal agent of Sphaeropsis rot of pears and apples, is a recently described species. In this study the effects of culture media, temperature, water potential, pH and light on mycelial growth and pycnidial production of S. pyriputrescens were evaluated. Apple juice agar and pear juice agar were most suitable for mycelial growth of all six isolates tested. Cornmeal agar was not suitable for either mycelial growth or pycnidial production. The fungus grew from −3 to 25 C, with optimum growth at 20 C and no growth at 30 C. The fungus grew at water potential as low as −5.6 MPa on potassium chloride-amended potato-dextrose agar (PDA). Hyphal extension was not observed at −7.3 MPa after 10 d incubation, but growth resumed when the inoculum plugs were placed on PDA. The fungus grew at pH 3.3–6.3 and optimum growth was at pH 3.3–4.2. No mycelial growth was observed at pH above 7.2 after 10 d incubation, but growth resumed when the inoculum plugs were transferred onto PDA. Regardless of medium tested, few pycnidia formed at 20 C in the dark. Pycnidial production was enhanced significantly by fluorescent light, but continuous light appeared to reduce pycnidial production, depending on the medium. Oatmeal agar (OMA) was most suitable for production of pycnidia and conidia. Pycnidia that formed on 3 wk old OMA cultures at 20 C under 12 h light/12 h dark produced abundant conidia, and the technique is recommended for inoculum production.

Phacidiopycnis washingtonensis--a new species associated with pome fruits from Washington State

A new species of Phacidiopycnis associated with pome fruits is described. The fungus causes fruit rot on apples during storage and is associated with a twig dieback and canker disease of crabapple trees and dead twigs of pear trees. To characterize the biology of the fungus and compare it with Ph. piri, the type species of the genus, effects of nine media and light on mycelial growth and pycnidial production, mycelial growth in response to temperature and mode of conidial germination in response to nutrient were determined. Apple-juice agar, pear-juice agar, prune-juice agar, potato-dextrose agar (PDA) and malt-extract agar, Czapek-Dox agar and oatmeal agar (OMA) favored mycelial growth. Cornmeal agar (CMA) did not favor mycelial growth. Light effect on pycnidial formation was medium dependent. Abundant pycnidia with mature conidia formed in 14 d old PDA and OMA cultures at 20 C, regardless of light, whereas none or very few pycnidia formed on other media in the dark. Fluorescent light stimulated formation of pycnidia except on CMA. The fungus grew at -3-25 C, with optimum growth at 15-20 C. Conidia germinated either by forming germ tubes or less often by budding. Budding of conidia occurred in 1 and 10% pear-juice solutions but not in 100% pear-juice solution. Six isolates of Ph. washingtonensis from different species of pome fruits had identical ITS sequences. The sizes of the ITS region were the same for both Ph. washingtonensis and Ph. piri, and four polymorphic nucleotide sites were found in the ITS region between Ph. washingtonensis and Ph. piri. The similarity in ITS sequences between these two taxa is confirmatory evidence for the erection of the new species of Phacidiopycnis associated with pome fruits we describe here.

Phacidiopycnis washingtonensis—a new species associated with pome fruits from Washington State

A new species of Phacidiopycnis associated with pome fruits is described. The fungus causes fruit rot on apples during storage and is associated with a twig dieback and canker disease of crabapple trees and dead twigs of pear trees. To characterize the biology of the fungus and compare it with Ph. piri, the type species of the genus, effects of nine media and light on mycelial growth and pycnidial production, mycelial growth in response to temperature and mode of conidial germination in response to nutrient were determined. Apple-juice agar, pear-juice agar, prune-juice agar, potato-dextrose agar (PDA) and malt-extract agar, Czapek-Dox agar and oatmeal agar (OMA) favored mycelial growth. Cornmeal agar (CMA) did not favor mycelial growth. Light effect on pycnidial formation was medium dependent. Abundant pycnidia with mature conidia formed in 14 d old PDA and OMA cultures at 20 C, regardless of light, whereas none or very few pycnidia formed on other media in the dark. Fluorescent light stimulated formation of pycnidia except on CMA. The fungus grew at −3–25 C, with optimum growth at 15–20 C. Conidia germinated either by forming germ tubes or less often by budding. Budding of conidia occurred in 1 and 10% pear-juice solutions but not in 100% pear-juice solution. Six isolates of Ph. washingtonensis from different species of pome fruits had identical ITS sequences. The sizes of the ITS region were the same for both Ph. washingtonensis and Ph. piri, and four polymorphic nucleotide sites were found in the ITS region between Ph. washingtonensis and Ph. piri. The similarity in ITS sequences between these two taxa is confirmatory evidence for the erection of the new species of Phacidiopycnis associated with pome fruits we describe here.

Influence of culture media and environmental factors on mycelial growth and pycnidial production of Sphaeropsis pyriputrescens

Sphaeropsis pyriputrescens, the causal agent of Sphaeropsis rot of pears and apples, is a recently described species. In this study the effects of culture media, temperature, water potential, pH and light on mycelial growth and pycnidial production of S. pyriputrescens were evaluated. Apple juice agar and pear juice agar were most suitable for mycelial growth of all six isolates tested. Cornmeal agar was not suitable for either mycelial growth or pycnidial production. The fungus grew from -3 to 25 C, with optimum growth at 20 C and no growth at 30 C. The fungus grew at water potential as low as -5.6 MPa on potassium chloride-amended potato-dextrose agar (PDA). Hyphal extension was not observed at -7.3 MPa after 10 d incubation, but growth resumed when the inoculum plugs were placed on PDA. The fungus grew at pH 3.3-6.3 and optimum growth was at pH 3.3-4.2. No mycelial growth was observed at pH above 7.2 after 10 d incubation, but growth resumed when the inoculum plugs were transferred onto PDA. Regardless of medium tested, few pycnidia formed at 20 C in the dark. Pycnidial production was enhanced significantly by fluorescent light, but continuous light appeared to reduce pycnidial production, depending on the medium. Oatmeal agar (OMA) was most suitable for production of pycnidia and conidia. Pycnidia that formed on 3 wk old OMA cultures at 20 C under 12 h light/12 h dark produced abundant conidia, and the technique is recommended for inoculum production.

Caracterização de Diaporthe citri em diferentes meios de cultura, condições de temperatura e luminosidade

Estudou-se o crescimento micelial de dez isolados de Diaporthe citri, utilizando-se seis meios de cultura (aveia-ágar, maltose-peptona-ágar, batata-dextrose-ágar, folha de laranja-dextrose-ágar, folha de limão-dextrose-ágar, milho-ágar) à temperatura de 22 ± 2 °C e fotoperíodo de 12 h claro/12 h escuro. O cultivo em meio de batata-dextrose-ágar (BDA) foi conduzido em cinco temperaturas diferentes (10, 15, 20, 25 e 30 °C). Três diferentes regimes de luminosidade (12 h claro/12 h escuro, claro contínuo, escuro contínuo) foram utilizados para verificar o crescimento do fungo. Foram observadas variações na produção de picnídios e de massa micelial nos diferentes meios de cultura, temperaturas e regimes de luminosidade testados, sendo que, para a maioria dos isolados, o meio de cultura de aveia-ágar, a faixa de 20 a 25 °C e o regime de claro contínuo induziram maior crescimento micelial. A produção de picnídios foi maior para o regime de luz contínua. O teste de patogenicidade foi feito por inoculação de discos de micélio de 5 mm de diâmetro em ferimentos em ramos e caule de limão 'Feminelo' (Citrus limon) enxertado em citrumelo 'Swingle'(Poncirus trifoliolata x Citrus paradisi) e plantas de limão 'Cravo' (C. limonia) enxertados com laranja 'Valência' (C. sinensis). Após sete dias, houve o aparecimento de exsudação de goma nas plantas inoculadas com os isolados, mas não na testemunha. Todos os isolados mostraram-se patogênicos, sendo os isolados PC2 e PC5, os que causaram comprimento de lesão maior nas plantas.

Effects of culture media and environmental factors on mycelial growth and pycnidial production of Potebniamyces pyri

Potebniamyces pyri (anamorph Phacidiopycnis piri) is the causal agent of Phacidiopycnis rot of apples and pears. The disease has recently been recognized in pears in the USA. Little information on the basic biology of the fungus is available. In this paper, we report the effects of culture media, temperature, water potential and pH on mycelial growth, and the effects of media and light on pycnidia production. Prune juice agar was the best for rapid mycelial growth. Pear juice agar, apple juice agar, potato dextrose agar, and oatmeal agar (OMA) also favoured mycelial growth. Czapek-Dox agar was not suitable for mycelial growth. The fungus was able to grow at temperatures from -3 to 25 degrees C. Optimal mycelial growth occurred between 15 and 20 degrees. The average radial growth rate on OMA was 3.9 mm d(-1) at 15 degrees and 4.4 mm d(-1) at 20 degrees. Mycelial growth was not observed after 10 d at 30 degrees, but growth resumed at 20 degrees. The fungus failed to resume growth at 20 degrees after being incubated for 10 d at 35 degrees. The fungus was able to grow at water potentials as low as -4 MPa but no growth took place at -7.3 MPa. Active mycelial growth was observed on OMA at pH between 3.2 and 6.1. Optimal growth was observed at pH around 4 and no growth was observed at pH 7.1. No pycnidia or very few formed on the nine media at 20 degrees in the dark. Fluorescent light significantly stimulated formation of pycnidia. OMA was the best medium for production of pycnidia and macroconidia. Pycnidia that formed on 8-wk-old OMA cultures incubated at 20 degrees under 12 h dark/12 h light produced abundant macroconidia and the technique is recommended for inoculum production of conidia for research.

Factors Affecting Pycnidium Production of Diaporthe citri on Detached Citrus Twigs.

Melanose, caused by Diaporthe citri, produces black-to-reddish brown lesions on twigs, leaves, and fruit of citrus and reduces the external quality of fruit destined for the fresh market. Inoculum for infection is produced primarily in pycnidia formed on dead twigs, and conidia are dispersed by rainwater. In laboratory studies, the effect of moisture, temperature, twig size, and melanose severity on pycnidium production on detached twigs was investigated. Pycnidium production was greatest when twigs were soaked for 3 to 4 h on alternate days three times per week and the temperature was 28°C. Production was greatest on twigs 3 to 5 mm in diameter and less on thinner or thicker twigs. Pycnidium production was related linearly to melanose severity on the twigs, and almost no pycnidia were produced on asymptomatic twigs. In the field, pycnidium production was greatest on detached, melanose-affected twigs placed in the canopy monthly during January to April than it was on twigs placed in the canopy during other months. The largest number of pycnidia was produced from May to August when fruit is most susceptible. The number of pycnidia produced was related significantly to degree-days above 20°C and weakly related to cumulative rainfall. Knowledge of inoculum production peaks may assist in timing of pruning and fungicide sprays.

Factors Influencing the Efficacy of Myclobutanil and Azoxystrobin for Control of Grape Black Rot.

We studied several factors influencing the efficacy of the demethylation inhibitor (DMI) fungicide myclobutanil and the strobilurin fungicide azoxystrobin for control of grape black rot, caused by the pathogen Guignardia bidwellii (anamorph Phyllosticta ampelicida). The distribution of sensitivities to myclobutanil among G. bidwellii isolates from an "organic" vineyard (no previous exposure to synthetic fungicides, n = 50) and from a commercial vineyard with a history of DMI applications (n = 60) was determined in vitro. There was little difference between the two populations, and the range of sensitivities was narrow; for the composite population of 110 isolates, the value of the mean effective dose for 50% inhibition (ED50) was 0.04 mg/liter, and the most- and least-sensitive isolates were separated by a factor of 16. When applied from 2 to 6 days after inoculating grape seedlings with a suspension containing either 2 × 104 or 1 × 106 conidia per ml, myclobutanil (60 mg/liter) provided complete control of lesion development. When applied beyond 6 days after inoculation but prior to lesion appearance (9 to 11 days after inoculation, depending on temperature), it provided complete control of pycnidium production in those lesions that developed subsequently. In contrast, when applied 2 to 10 days after inoculation with 2 × 104 conidia per ml, azoxystrobin (128 mg/liter) provided only 78 to 63% control of lesion formation and erratic control of pycnidium formation, although conidium production was reduced by 85 to 68% across this range of treatments. Relatively little control was provided by azoxystrobin treatments following inoculation with 1 × 106 conidia per ml. On leaf disks treated with azoxystrobin at 20 mg/liter prior to inoculation, 8 to 43% of conidia from five G. bidwellii isolates germinated, and 4 to 19% formed appressoria. However, these processes were completely to near-completely inhibited when salicylhydroxamic acid (SHAM), which inhibits an alternative respiration pathway utilized to circumvent the activity of strobilurin fungicides, was added to the inoculum at 100 mg/liter. Thus, alternative respiration apparently allowed the conidia to germinate and form appressoria on azoxystrobin-treated leaves. When grape seedlings were sprayed with commercially formulated azoxystrobin at 200 mg/liter and inoculated the next day with G. bidwellii conidia, little or no disease was evident 4 weeks later. However, G. bidwellii pycnidia formed on up to 50% of the leaves from such plants when they were killed with paraquat 1 to 7 days after inoculation. These results suggest that latent infections became established on azoxystrobin-treated leaves and became active after the plants were killed with paraquat.

Effect of environmental factors on growth, pycnidial production and spore germination of Microsphaeropsis isolates with biocontrol potential against apple scab

The potential as biocontrol agents of the apple scab pathogen, Venturia inaequalis and the effect of environmental factors on growth, pycnidial production and spore germination of four isolates of Microsphaeropsis were examined on culture media and leaf discs. V. inaequalis ascospore production was reduced by 52, 77, 89 and 95% on leaf discs treated with the isolates IMI 294735, P176A, DAOM 198536 and P130A, respectively. For all isolates, optimum temperature for mycelial growth was 25 °C. More pycnidia were produced on culture media than on leaf discs. On culture media, optimum temperature was from 15 to 25 °, while on leaf discs it varied among isolates. Pycnidial production was inhibited by darkness and a minimum of 8 h light d −1 was required. Spores of isolates P130A and DAOM 198536 germinated at temperatures above 15 ° and at pH 4–5, as compared to 10 ° and pH 4–8 for isolates P176A and IMI 294735. The best candidates were P130A and DAOM 198536 and optimum conditions for growth, pycnidial production and spore germination were temperatures between 20 and 25 °, pH approx. 4 and light.

Some Nutritional Factors Affecting Production of Biomass and Antifungal Metabolites of Coniothyrium minitans

The ability of the mycoparasite Coniothyrium minitans to utilize a range of C and N sources and vitamins for growth, pycnidial formation and antifungal metabolite production was examined using a defined liquid medium. Coniothyrium minitans was able to use all the C sources tested, with the exception of D -xylose, and all the N sources tested, although growth was generally better on organic N sources rather than NO 3 -N. Increasing C:N ratios from 9:1-202:1 with N constant (2.0 g L -l L -alanine) resulted in steadily increasing yields, whereas increasing C:N ratios with C constant (40.0 g L -l D -glucose) gradually decreased yield. Addition of thiamine to the glucose-alanine basal medium resulted in the greatest increase in growth but biomass was still less than that achieved using an undefined molasses-yeast medium. Pycnidial production was generally low or failed to occur in the basal medium + C + N sources in the absence of vitamins, but addition of thiamine consistently led to abundant pycnidial formation. Molasses-yeast static culture provided greater biomass and conidial yields than molasses-yeast shaken culture. Incorporation of C. minitans culture medium into potato dextrose broth (10% v/v) resulted in consistent reduction in growth of Sclerotinia sclerotiorum irrespective of C, N or vitamin content of the basal medium or whether molasses-yeast medium was used. This is the first report of consistent production of antifungal metabolites by C. minitans .

Comparison of the laccases, molecular marker proteins, and induction of pycnidia by three species of botryosphaeriaceous fungi

Three species of botryosphaeriaceous fungi,Botryosphaeria sp. isolate MAMB-5,Botryosphaeria ribis andLasiodiplodia theobromae, were compared for the production of pycnidia and laccases. Laccases were produced both intra- and extra-cellularly when the fungi were cultivated on basal medium in the presence and absence of veratryl alcohol, withBotryosphaeria sp. MAMB-5 showing the highest enzyme titres. Electrophoretic examination of intracellular marker proteins (esterases and phosphatases) and laccases indicated that the three species were genetically distinctly different, although the laccase zymograms for the three fungi showed similarity. The production of pycnidia occurred under continuous lighting at 28°C, but conditions differed among the three fungal species. Production could be induced on artificial media (potato-dextrose and oat agar) under stress-induced conditions where the mycelium was stimulated by physical abrasion, and in the case ofBotryosphaeria sp. isolate MAMB-5 on eucalypt woodchips. Evidence is presented that veratryl alcohol facillitated the secretion of intracellular-localised laccases into the extracellular medium.

Microsphaeropsis olivaceaas an etiological agent of human skin infection

Microsphaeropsis olivacea is reported as the agent of a case of human skin infection in an otherwise healthy woman. This fungus has not been reported previously as causing disease in humans. It was identified on the basis of its production of pycnidia and conidial structures typical of the Coelomycetes, and by its light brown, ellipsoid to cylindrical and thick walled conidia. The in vitro inhibitory activity of amphotericin B, fluconazole, flucytosine, itraconazole, ketoconazole and miconazole was determined.

Variability of Ascochyta fabae in South Australia

Fifty-two isolates of Ascochyta fabae were established from 23 collections made in 3 States of Australia and were purified through 2 cycles of single-spore isolation. The isolates were evaluated for spore size, spore production, colony diameter, aerial mycelium, and pycnidia production. Variation in all of these traits among related single-spore cultures was comparable to that among unrelated ones and only colony diameter varied significantly among isolates. Spore size was 3–6 by 10–26 µm. Eight of these 52 isolates were chosen for further investigations of pathogenicity characteristics using 8 populations of faba bean. Plants were scored daily for rate of appearance of symptoms and then 15 and 21 days after inoculation for lesion size and number, production of pycnidia on the lesions and overall disease score. Leaves and stems reacted differently to the disease, with one isolate producing many leaf lesions but few stem lesions on one bean accession but many stem lesions on another. Lesion size was not strongly correlated with the other measures of disease. Resistant accessions had longer incubation periods, fewer total lesions and fewer pycnidia-producing lesions than susceptible accessions. The 8 isolates on the 8 bean accessions showed 7 distinct patterns of resistance. The results showed that in southern Australia, A. fabae exhibited great variability which was incompatible with classification into biologically meaningful pathotypes.