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Coping with stress: calmodulin and calcineurin in model and pathogenic fungi.

Calcium signaling via calmodulin and calcineurin is critical for the regulation of stress responses in fungi. The functions of calmodulin and calcineurin are largely conserved among pathogenic fungi and model fungi, however, the mechanisms of action have diverged. Saccharomyces cerevisiae is an excellent model for understanding the framework of calcium-mediated signal transduction pathways, and considerable progress has been made in understanding the details of how Ca(2+)-calmodulin and calcineurin control adaptation to environmental stress. Studies using the divergent human pathogenic fungi Candida albicans and Cryptococcus neoformans reveal that calcineurin is critical for virulence, yet it acts via distinct mechanisms in each fungus. These differences in function may reflect the requirements of each pathogen to survive inside the host, and illustrate that studies must be conducted in each organism in order to elucidate the details of the molecular mechanisms of calmodulin and calcineurin-mediated signaling pathways.

Calcineurin↗

Chemical basis of the resistance of barley seeds to pathogenic fungi.

The 5-(n)-alkylresorcinol fraction of the epicuticular waxes of Hordeum vulgare seeds appeared to be responsible for their in-born resistance to pathogenic fungi such as Aspergillus niger and Penicillium crysogenum. The antifungal properties of this fraction were evaluated qualitatively and quantitatively with a novel bioassay where the extreme lipophilicity of these compounds was taken into account. The minimum inhibitory concentration in the fungi tested ranged from 5.6 to 10 micrograms cm-2 for the alkyresorcinols. The behaviour of the different cultivars against these fungi could be predicted by measuring the natural amount of resorcinols of each variety by TLC-scanning densitometry. The ranking of cultivars thus established correlated well with the field behaviour of each cultivar, providing a useful and rapid method for predicting the behaviour against fungi of new varieties being developed.

Chromatography, Thin Layer↗

Bacteria and fungi in aerosols generated by two different types of wastewater treatment plants.

Raw wastewater is a potential carrier of pathogenic microorganisms and may pose a health risk when pathogenic microorganisms become aerosolized during aeration. Two different types of wastewater treatment plants were investigated, and the amounts of cultivable bacteria and fungi were measured in the emitted aerosols. Average concentrations of 17,000 CFU m(-3) of mesophilic, 2,100 CFU m(-3) of TSA-SB bacteria (bacteria associated with certain waterborne virulence factors), 1700 CFU m(-3) of mesophilic and 45 CFU m(-3) of thermotolerant fungi, were found in the aerosol emitted by the aeration tank of the activated sludge plant. In the aerosol of the fixed-film reactor 3000 CFU m(-3) mesophilic and 730CFUm(-3) TSA-SB bacteria, and 180 CFUm(-3) mesophilic and 14 CFU m(-3) thermotolerant fungi were measured. The specific emissions per population equivalent between the two types of treatment plants differed by two orders of magnitude. The microbial flux based on the open water surface area of the two treatment plants was similar. The aerosolization ratios of cultivable bacteria (expressed as CFU m(-3) aerosol/m(-3) wastewater) ranged between 8.4 x 10(-11) and 4.9 x 10(-9). The aerosolization ratio of fungi was one to three orders of magnitude higher and a significant difference between the two types of treatment plants could be observed.

Aerosols↗

Characteristics of indoor and outdoor airborne fungi at suburban and urban homes in two seasons.

Literature has suggested association between damp environments, microbial exposure, and higher prevalence of respiratory symptoms and diseases. The study began by evaluating the airborne fungal concentrations at urban and suburban areas of a typical metropolitan city in southern Taiwan for the estimation of related health risks. A group of representative homes, based on the housing characteristics questionnaires completed earlier, were selected from two parts of the city; urban and suburban. Burkard sampler (BURKARD, Rickmansworth, England) was used to collect airborne fungi onto agar plates with malt-extract. After incubation and identification, concentrations of airborne fungi were calculated as CFU/m3. The geometric mean (GM) concentration for indoors was 8946 (4372-18,306) CFU/m3 in winter and 4381 (1605-11,956) in summer. For outdoors, it was 11,464 (5767-22,788) CFU/m3 in winter and 4689 (1895-11,603) in summer. In summer, the total fungal concentration, both indoors and outdoors of suburban homes, were significantly higher than those of urban homes. The dominant fungi contributing to such a difference were indoor Cladosporium spp. and outdoor Penicillium spp. (P < 0.01). The indoor/outdoor ratio (I/O) was similar in two areas except for Penicillium spp. in winter and Aspergillus spp. in summer; both higher in the suburban area. Significantly higher levels of airborne fungi were observed in this region than those seen in northern Taiwan or other parts of the world. Future investigations are needed to further examine the effects of these exposures on the related health problems.

Air Microbiology↗

Impact of environment and interspecific interactions between spoilage fungi and Aspergillus ochraceus on growth and ochratoxin production in maize grain.

Using layers of irradiated but still fertile maize grain, the effects of water activity (0.995, 0.95 a(w)) and temperature (18, 30 degrees C) on interspecific interactions between Aspergillus ochraceus and five other spoilage fungi were examined. Asp. ochraceus was not competitive against Asp. flavus, Asp. niger, or Alternaria alternata at 18 degrees C when water was freely available (0.995 a(w)), while at 0.95 a(w) it was dominant against Asp. candidus, Asp. flavus and Alt. alternata. At 30 degrees C and 0.995 a(w), Asp. ochraceus was dominated by other fungi, except Alt. alternata, and was mutually antagonistic to Asp. candidus and Eurotium amstelodami. However, at 30 degreees C and 0.95 a(w), it was competitive against Asp. candidus and Alt. alternata, but not against the other species examined. The overall Index of Dominance showed that Asp. ochraceus was not competitive under the conditions examined here. At 18 degrees C ochratoxin production by Asp. ochraceus was inhibited significantly by Asp. candidus (0.995 and 0.95 a(w)) and Asp. niger (0.995 a(w)). When grown on maize grain at 30 degrees C, ochratoxin production by Asp. ochraceus was significantly inhibited by other spoilage fungi when both were grown on maize grain, especially by Asp. niger and E. amstelodami (0.995 a(w)) and Asp. flavus at 0.95 a(w). These results suggest that, to a large extent, A. ochraceus is not as competitive as some other spoilage fungi in primary resource capture on maize grain at a(w) of 0.95 or above, although it may modify resource quality and influence secondary colonisation by other species under the conditions tested.

Alternaria↗

Identification of fungi from dairy products by means of 18S rRNA analysis.

The role of fungi as cause of spoilage of dairy products, such as cheese and yoghurt, has been clearly demonstrated. Despite of this, there is still a lack in rapid methods for the identification of food-associated fungi. In the course of the present work, molecular taxonomical techniques were developed and used to identify yeasts involved in the spoilage of yoghurt and moulds responsible for spoilage of vacuum-packaged hard cheese. Three methods for DNA extraction and purification were evaluated and the fungus-specific primers TR1 and TR2 were used to amplify a 581-bp fragment within the gene, coding for the small ribosomal subunit (18S rRNA) of fungi. The 18S rRNA sequence analysis of fungi isolated from yoghurt and packaged cheese allowed to identify yeast belonging to Zygosaccharomyces microellipsoides and moulds belonging to Penicillium chrysogenum and Cladosporium cladosporoides.

Animals↗

Mycotechnology: the role of fungi in biotechnology.

Fungi have been important in both ancient and modern biotechnological processes. Processes and products that utilize fungi include baking, brewing, and the production of antibiotics, alcohols, enzymes, organic acids, and numerous pharmaceuticals. The advent of recombinant DNA technology and large scale genomics analysis has placed yeasts and filamentous fungi in the forefront of contemporary commercial applications. The term 'mycotechnology' is introduced here to describe the enormous impact of fungi on biotechnology.

Anti-Bacterial Agents↗

Pyrene degradation by yeasts and filamentous fungi.

The saprotrophic soil fungi Fusarium solani (Mart.) Sacc., Cylindrocarpon didymum (Hartig) Wollenw, Penicillium variabile Sopp. and the yeasts Rhodotorula glutinis (Fresenius) Harrison and Rhodotorula minuta (Saito) Harrison were cultured in mineral medium with pyrene. The remaining pyrene concentrations were periodically determined during 20 incubation days, using HPLC. To assess the metabolism of pyrene degradation we added 0.1 microCi of [4,5,9,10] 14C-pyrene to each fungi culture and measured the radioactivity in the volatile organic substances, extractable, aqueous phase, biomass and 14CO2 fractions. The assays demonstrated that F. solani and R. glutinis metabolized pyrene as a sole source of carbon. Differences in their activities at the beginning of the cultures disappeared by the end of the experiment, when 32 and 37% of the original pyrene concentration was detected, for the soil fungi and yeasts, respectively. Among the filamentous fungi, F. solani was highly active and oxidized pyrene; moreover, small but significant degradation rates were observed in C. didymum and P. variahile cultures. An increase in the 14CO2 evolution was observed at the 17th day with cosubstrate. R. glutinis and R. minuta cultures showed similar ability to biotransform pyrene, and that 35% of the initial concentration was consumed at the end of the assay. The same results were obtained in the experiments with or without glucose as cosubstrate.

Biodegradation, Environmental↗

Feasibility of fungi bioaugmentation in composting a flare pit soil.

The feasibility of fungi bioaugmentation in composting of a flare pit soil was studied in lab-scale composters. The preliminary screening tests, using a range of bulking agents and white rot fungi strains, were conducted to determine, best strain and bulking agent for the main experiments. The initial total petroleum hydrocarbon (TPH) of the flare pit soil was found to be 16%. The effect of moisture and bulking agent content and the fungi application on biodegradation of hydrocarbons were then evaluated based on a fractional factorial design over a 3-months period. Analysis of the TPH content of the soil after 98 days (using gravimetric method) showed an average of 29% reduction in most jars. Furthermore, gas chromatograph (GC) analysis of the oil extract from the samples showed 70-99% reduction in the peak area of the selected hydrocarbons. However, statistical analysis of the results did not show any significant effect due to the fungi application or the change in the moisture content (30-50% range). The results showed that the change in the bulking agent content was marginally significant for the hydrocarbon loss.

Biodegradation, Environmental↗

Examination of soils from residential garbage in Betul, India, for fungi by the keratin baiting technique.

A report on an examination of soils from residential garbage of Betul, India, for fungi by the keratin (materials) baiting technique is presented. A total of 69 fungi representing 39 species among 17 genera were isolated. In all the soil samples, only two genera, namely Aspergillus (190.9%), followed by Fusarium (118.18%), were most frequently isolated. Maximum colonisation of fungi was found on hair, followed by horn, feathers and nails. For the first time, the genus Fusoma is reported here as a keratinophilic fungus. The keratin baiting technique was found very effective in detecting a broad spectrum of fungi in biological wastes and compost.

Aspergillus↗

Physiological aspects of fungi isolated from root nodules of faba bean (Vicia faba L.).

The present study was made to isolate and assess some physiological characteristics of root nodule-colonizing fungi. During this study, 17 fungal species were isolated from root nodule samples taken from faba bean plants (Vicia faba L.) collected from different sites at Assiut area (Egypt). The growth of faba bean plants in pots was significantly promoted by soil inoculation with most fungi. Growth was checked in pots with inocula of Cladosporium cladosporioides, Fusarium moniliforme, F: oxysporium, F solani, Macrophominia phaseolina and Rhizoctonia solani which were added separately. All growth-promoting fungi were capable of producing cellulase, pectin lyase, polygalacturonase, protease, urease, amidase, acid phosphatase, alkaline phosphatase and arylsulfatase in growth medium supplemented with the corresponding substrates. Four fungal species, Aspergillus awamori, A. flavus, Penicillium chrysogenum and Trichoderma koningii showed the highest rates of enzyme formation. The effect of the addition of six trace elements to the growth media at 30 micromol/ml on enzyme production revealed some dependency on species, enzyme and metal ion. Cd2+, Hg2+ and Zn2+ generally inhibited enzyme activity. Cu(1+), Fe3+ and Al3+ showed a stimulatory effect. Fungicides (afugan and tilt) and herbicides (brominal and fusilade) at 50 ppm generally promoted enzyme activity, but insecticides (kelthane and fenvalerate) caused some inhibition to enzyme activities. Salinization of the growth media with NaCl strongly inhibited the enzymatic activity of all fungi at concentrations between 0.5 and 1.5%.

Fabaceae↗

Are Ichthyosporea animals or fungi? Bayesian phylogenetic analysis of elongation factor 1alpha of Ichthyophonus irregularis.

Ichthyosporea is a recently recognized group of morphologically simple eukaryotes, many of which cause disease in aquatic organisms. Ribosomal RNA sequence analyses place Ichthyosporea near the divergence of the animal and fungal lineages, but do not allow resolution of its exact phylogenetic position. Some of the best evidence for a specific grouping of animals and fungi (Opisthokonta) has come from elongation factor 1alpha, not only phylogenetic analysis of sequences but also the presence or absence of short insertions and deletions. We sequenced the EF-1alpha gene from the ichthyosporean parasite Ichthyophonus irregularis and determined its phylogenetic position using neighbor-joining, parsimony and Bayesian methods. We also sequenced EF-1alpha genes from four chytrids to provide broader representation within fungi. Sequence analyses and the presence of a characteristic 12 amino acid insertion strongly indicate that I. irregularis is a member of Opisthokonta, but do not resolve whether I. irregularis is a specific relative of animals or of fungi. However, the EF-1alpha of I. irregularis exhibits a two amino acid deletion heretofore reported only among fungi.

Animals↗

Health effects of indoor fungi.

OBJECTIVE: To review the nontoxic harmful effects that poor indoor air quality caused by fungi can have on health. DATA SOURCES: We searched PubMed for publications related to the various topics discussed in this review, and we relied on our knowledge of the field. STUDY SELECTION: Where more than one publication was relevant, we attempted to identify a consensus of the reports and cited the most relevant articles. Priority was given to randomized controlled trials and expert reports when available, although much of the information herein relates to laboratory research. RESULTS: Actively growing fungal colonies can release volatile substances that have an unpleasant smell, leading to psychological responses in the occupants such as fatigue and nausea. Symptoms that are likely caused by indoor fungi include respiratory complaints that involve the nose and lungs, eye symptoms, and mucous membrane irritation. These adverse effects can occur by a variety of mechanisms, including IgE-mediated hypersensitivity, fungal infection, irritant reaction to spores or fungal metabolites, and possibly toxic reaction to mycotoxins. CONCLUSIONS: Reduced fungal exposure can reasonably be expected to improve health. Removal of moisture from the indoors and proper maintenance of air filters can aid in prevention and elimination of fungi from the home environment. Small areas of present contamination can be cleaned with a dilute bleach solution, which kills viable colonies and removes their mycelia. If fungal contamination is not addressed early, substantial damage can occur, requiring professional remediation. Above all, the individual should not panic at the first sight of fungi growing in the home. Regular inspection and cleaning can prevent many fungus-related problems.

Air Pollution, Indoor↗

New and simple plate test for screening relative transfructosylation activity of fungi.

Several microorganisms are reported to have transfructosylation activity due to fructosyltransferase and/or fructofuranosidase activities. However, the search for other fungi with higher transfructosylation activity remains a challenge. So, a presumptive and indirect colorimetric plate assay for the evaluation of transfructosylation activity in fungi was developed which involved the simultaneous determination in the same plate of glucose and fructose released from sucrose. The method entailed the (a) glucose oxidase-peroxidase coupled reaction using phenol and 4-aminoantipyrine for determination of glucose; and (b) fructose dehydrogenase oxidation in the presence of a tetrazolium salt for determination of fructose. The presence of enzymes with transfructosylation activity was identified by the formation of pink (presence of glucose) and blue (presence of fructose) halos around the fungal colony. In conclusion, the results showed that the method is suitable for screening a large number of fungi due to its simplicity, reproducibility and rapidity and also gives a relative quantitative idea of the transfructosylation activity of different fungi species.

Ampyrone↗

Biological control of Ostertagia ostertagi by feeding selected nematode-trapping fungi to calves.

Three nematode-trapping fungi, one Arthrobotrys oligospora and two Duddingtonia flagrans isolates, were fed to Ostertagia ostertagi-infected calves to test their ability to destroy larvae of this parasite in faeces and consequently to reduce the transmission of infective larvae to herbage. The fungi had previously been selected for their capability to pass the alimentary tract of cattle without losing growth and nematode-trapping potentials. Dung was collected from three calves each fed one of the three fungi and placed as 1-kg cow pats on a parasite-free grass plot together with control cow pats from a calf that was not given fungi. The cow pats contained comparable concentrations of parasite eggs. The two D. flagrans isolates were highly effective in that they reduced herbage larval infectivity by 74-85%. In contrast, A. oligospora did not show any effect in the present experiment. Field experiments will demonstrate if D. flagrans represents a potential organism for biological control of bovine gastrointestinal nematodes under practical agricultural management conditions.

Animals↗

Fungi associated with shoots of silver birch (Betula pendula) in Scotland.

The fungi associated with diseased and healthy shoots of young, planted Betula pendula were studied at five sites in Scotland where crown die-back of birch is occurring. At each site, 30 diseased and 30 healthy shoots were sampled twice, with 1 yr-old shoots collected in May 2002, and 4-5-month-old shoots collected in September/October 2002. Overall, 34 different fungal species were recorded, with 92 % and 60 % of all diseased and healthy shoot pieces, respectively, colonised by at least one fungal species. The main colonisers of both diseased and healthy shoots were Melanconium bicolor, Discula betulina and Godronia urceolus, and the fungi most frequently found on diseased shoots only included Marssonina betulae and Fusarium avenaceum. The pathogenicity of these fungi was tested on seedlings of B. pendula and B. pubescens. Melanconium bicolor required a wound for lesion development on both birch species and caused larger lesions on B. pubescens than on B. pendula. Discula betulina caused lesions on stems and shoots of both birch species, including non-wounded current shoots. Marssonina betulae caused lesions and die-back of wounded and non-wounded current shoots of B. pendula seedlings. This fungus was not pathogenic on B. pubescens. Fusarium avenaceum caused restricted lesions on the main stems of both birch species. Godronia urceolus was not pathogenic on either birch species. Further work needs to focus on the role of these fungi in causing crown die-back of young, planted B. pendula in Scotland.

Betula↗

Degradation of starchy endosperm cell walls in nongerminating sterilized barley by fungi.

Strains of fungi from different origins, including isolates of the natural microflora of barley, were screened for their ability to modify barley starchy endosperm cell walls in situ. In an initial step, fungi were selected that degrade the major component of the cell walls, that is, (1-->3),(1-->4)-beta-D-glucan, in vitro on artificial media. Nongerminating, sterilized barley, obtained by gamma-irradiation, was inoculated with such fungi and subjected to solid state fermentation under conditions resembling those of a traditional malting process. For some strains of fungi, a clear correlation between the production of endo-beta-glucanase and the friability of the treated kernels was found. Image analysis of Calcofluor stained longitudinal sections of barley kernels fermented with the endo-beta-glucanase producing strains showed that starchy endosperm cell walls were modified. As malt quality is inversely related to its (1-->3),(1-->4)-beta-D-glucan content, the selected strains have high potential to be used as starter cultures during malt production, contributing to the processing quality of the final product.

Aspergillus↗

Toxigenic saprophytic fungi in marine shellfish farming areas.

Toxigenic saprophytic fungi were isolated from samples of shellfish, sediment and seawater obtained from marine shellfish farming areas. The 456 strains identified included 12 different genera, with a clear predominance (68%) of Penicillium, Aspergillus, Trichoderma and Cladosporium. To assess the risk of poisoning due to the presence of these fungi in shellfish farming areas, the strains were cultured in liquid medium, filtered, and tested on larvae of Artemia salina, a small crustacean highly sensitive to mycotoxins. Thirty-five point five percent of the strains proved active with this test. This study confirms the existence of fungi in shellfish farming areas, as suggested by our earlier work showing that filter-feeding shellfish accumulate toxic metabolites of fungal origin. The presence of fungi in the marine environment represents a real risk of poisoning through the consumption of contaminated shellfish.

Animals↗