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[Ecology of opportunistic fungi].

As most of the pathogenic fungi, opportunistic fungi have a saprophytic life in the soil. The knowledge of their life in natural conditions may allow to avoid their proliferation in the vicinity of man. Examples of ecology were taken among Aspergilli and mycetoma-inducing fungi. The ecology of these fungi was studied in Senegal-Mauritania where mycetoma is fairly frequent and well studied. Cultures of L. senegalensis and L. tompkinsii were very often isolated from dry thorns of acacias or other thorn trees. M. mycetomi, the most frequent mycetoma-inducing fungus, was very difficult to culture from wood or soil specimens. A special selective sulphide medium had to be used. Some strains of M. mycetomi were isolated from specimens taken off the surface of termitaria.

Aspergillus↗

A review of the volatile metabolites of fungi found on wood substrates.

The holdings of eight collections of fungi have been examined for organisms isolated from wood and/or trees. Further selection of these fungi has been made according to their reported ability to produce volatile, biologically active metabolites. It is emphasized that the isolates in the collections do not necessarily produce such metabolites. The list of fungi fulfilling these conditions is slightly augmented by reports we have found in the literature, where the fungi concerned have not yet been deposited. The biochemistry of these compounds is considered with particular emphasis on their biosynthesis including that by Homo sapiens. The physiological and toxicological activity of these metabolites is reviewed especially with reference to their potential role in the complex symbioses existent in, for example, a tree. The review concludes with a discussion of areas of botany deserving increased attention in the hope that this will stimulate further work. The statements in the review are based on 173 references.

Acetylene↗

Production of oxalic acid by some fungi infected tubers.

Oxalic acid (as oxalate) was detected in four tubers commonly used for food in Nigeria-Dioscorea rotundata (White yam), Solanum tuberosum (Irish potato), Ipomoea batatas (Sweet potato), and Manihot esculenta (cassava). Whereas healthy I. batata had the highest oxalic acid content, healthy M. esculenta contained the lowest. When all tubers were artifically inoculated with four fungi-Penicillium oxalicum CURIE and THOM, Aspergillus niger VAN TIEGH, A. flavus and A. tamarii KITA, there was an increase in oxalate content/g of tuber tissue. The greatest amount of oxalate was produced by P. oxalicum in D. rotundata tuber. Consistently higher amounts of oxalate were produced by the four fungi in infected sweet potato tuber than in any other tuber and consistently lower amounts of oxalate were produced by the four fungi in Irish potato tuber. Differences in the carbohydrate type present in the tubers and in the biosynthesis pathway are thought to be responsible for variation in the production of oxalate in the different tubers by the four fungi used.

Fungi↗

Entomopathogenous fungi degrade epicuticular hydrocarbons of Triatoma infestans.

Studies were undertaken to analyze the ability of entomopathogenous fungi to degrade insect hydrocarbons. Strains of Beauveria bassiana and Metarhizium anisopliae pathogenic to the blood-sucking bug Triatoma infestans were grown on hydrocarbon and non-hydrocarbon insect lipid extracts and on synthetic hydrocarbon-enriched media as the sole carbon source. Entomopathogenous fungi were shown to utilize hydrocarbons as the only carbon source for their growth. Insect-derived hydrocarbons served more efficiently as metabolic fuel rather than synthetic compounds of similar structure. [3H]n-Pentacosane, [11,12-3H]3,11-dimethylnonacosane, and [14C]n-hexadecane were catabolized into different amounts of polar lipids, free fatty acids, and acylglycerols. In experiments using the branched alkane, labeled hydrocarbons of different chain length than the precursor were also synthesized. Evidence of complete catabolism was obtained by a significant release of 14CO2 from [1-14C]n-hexadecane. 14CO2 production might be used as a simple method to compare hydrocarbon utilization by fungal strains. These data demonstrate that entomopathogenous fungi are able to transform a variety of hydrocarbon structures into different lipid products, part of which may be subsequently utilized for energy production and for the biosynthesis of cellular components. These data are the first evidence of hydrocarbon catabolism and synthesis in entomopathogenous fungi.

Alkanes↗

A comparison of sampling media for environmental viable fungi collected in a hospital environment.

Quantitative evaluation of fungal exposure is often conducted by analysis of the composition of microbes in air samples and calculation of the concentrations afterward. The collecting medium that favors the growth for most saprophytic fungi is considered to be the ideal choice in most circumstances. Currently, the culture medium most frequently adopted in environmental sampling for airborne fungi is MEA (malt extract agar) recommended by the ACGIH for its suitability for most fungal growth. DG18 (dichloran glycerol-18), developed in 1980, is suggested for growth at lower water activity (a(w)=0.95) specifically and is not as commonly used in general studies. This investigation collected airborne viable fungi using a single stage/N6 Andersen impactor with MEA and DG18 agar plates attached simultaneously to the same set of samplers. The sampling locations were at 17 sites within a central air-conditioned hospital. After incubation and morphological identification, concentrations of airborne fungi and bacteria were expressed as CFU/m(3) (colony forming units/m(3)). There are 405 DG18 plates and 378 plates available for statistical analysis. Results show that the airborne fungal concentrations, shown by geometric mean (GM), are higher from the DG18 plates than from the MEA plates. The total fungal concentrations is 68.6 vs 12.94 CFU/m(3), and for Aspergillus spp., the concentration is 1.58 vs 0.72 CFU/m(3); for Penicillium spp., 3.37 vs 0.71; and for yeast, 5.09 vs 0.49 CFU/m(3). In addition, the number of different genera present is greater on the DG18 plates than on the MEA plates, on average, 2.85 types vs 1.72. This study suggests that in a hospital environment with 24-h, central air conditioning, DG18 plates appear to be more effective in collecting more fungal colonies in terms of both quantity and types of genera. Such a finding is presumed to be attributed to the characteristic of DG18 in slowing colony growth so that the dominating genus will not over occupy the culture plate surface before the less competitive genus can fully develop. Future studies on related biological mechanisms are essential to conclude whether the above results sustain when sampling is conducted in other environments.

Air Pollutants↗

Entomopathogenic fungi associated with Ixodes ricinus ticks.

The objective of this study was to demonstrate the occurrence of entomopathogenic fungi on Ixodes ricinus ticks in relation to the tick stage, engorgement and season. Ticks were collected from the vegetation, from small rodents and from deer. All entomopathogenic fungi found belonged to the Hyphomycetes. Paecilomyces farinosus and Verticillium lecanii were the predominant species. Other species, found only on engorged females were: Beauveria bassiana, B. brongniartii, P. fumosoroseus and V. aranearum. Eight out of 1833 ticks collected from the vegetation and three out of 269 engorged nymphs were infected with fungi. Thirty-three out of 149 engorged females were infected, whereas males and engorged larvae were not infected. Throughout the season, a significantly higher proportion of ticks collected in autumn were infected. Entomopathogenic fungi may have a significant impact on the size of the I. ricinus population, since females were the most frequently infected stage.

Animals↗

Differentiation of species and strains of entomopathogenic fungi by random amplification of polymorphic DNA (RAPD).

Polymerase chain reaction (PCR)-based technology, involving random amplification of polymorphic DNA (RAPD), was used to assess the genomic variability between 24 isolates of deuteromycetous fungi (Metarhizium anisopliae, Metarhizium flavoviride, unidentified strains of Metarhizium and Beauveria bassiana) which were found to infect grasshoppers or locusts. M. flavoviride showed little intraspecific variability in PCR-amplified fragments when compared to M. anisopliae. The high level of variability in PCR-amplified fragments contained within M. anisopliae was similar to the total variability between B. bassiana, M. anisopliae and M. flavoviride, and suggests that M. anisopliae may include a number of cryptic species. Four polymorphic RAPD fragments were used to probe the genomic DNA of the various species and strains. On the basis of these probes the fungi can be grouped into M. flavoviride, M. anisopliae, or B. bassiana. According to PCR-amplified fragments, previously-unidentified Metarhizium strains were characterized as M. flavoviride. There was little evidence that these fungi, all isolated from, or virulent towards, grasshoppers or locusts, showed host-selection in PCR-amplified fragments. Nor was geographical origin a criterion for commonality based on PCR-amplified fragments. PCR-fragment-pattern polymorphisms and the construction of probes from one or more of these fragments may provide a useful and rapid tool for identifying species and strains of entomopathogenic fungi.

Animals↗

Isolation of fungi from the pelage of cats and dogs using the hairbrush technique.

A total of 178 cats and 59 dogs in Palmerston North, New Zealand were sampled for the presence of keratinophilic fungi on their pelage; 57.8% had fungi. The fungi were classified in 20 genera with the predominant species being members of the genera; Chrysosporium, Microsporum and Trichophyton. Cats were the major carriers of keratinolytic fungi. 18.5% of the cats and 5.1% of the dogs were either carriers or infected with M. canis. Microsporum canis was a frequent isolate and its distribution had a seasonal variation, with a peak appearing in May-June.

Animals↗

Studies on pathogenic dematiaceous fungi. 1. Isolation from natural sources.

Two hundred and twenty-six samples of woody materials, vegetable matter and soil were processed by the direct plating and mouse inoculation technique for the isolation of pathogenic dematiaceous fungi. The species of fungi isolated were Fonsecaea pedrosoi - 13, Cladosporium carrionii - 7 and Phialophora verrucosa -4 isolates. The mouse inoculation technique was found to be much better than direct plating for the recovery of these fungi. Woody plant materials proved to be a good sample source for pathogenic dematiaceous fungi contributing about 90% of the isolates. All the isolates were pathogenic for mice as evidenced by the presence of dark nodular lesions containing fungal elements in the organs of experimentally infected animals.

Animals↗

Association of anurans with pathogenic fungi.

In a study of 450 Amazonian anurans, we isolated yeasts and yeast-like fungi from 54 animals (Bufo granulosus, B. marinus, Dendrophrynyscus sp., Hyla geographica, H. lanciformes, Ololygon rubra, Adenomera hylaedactyla, Eleutherodactylus fenestratus, Leptodactylus fuscus, L. ocellatus, L. pentadactylus). The internal organs of these animals did not show any macroscopic anomaly nor histopathology. We recovered 105 fungal isolates from the anuran liver, lung, kidney, spleen, heart and gonad. The isolates were made up of 30 fungal species, 9 of which (48 isolates, 46%) were fungi with known pathogenic potentials, namely: Candida guilliermondii, C. parapsilosis, C. tropicalis, C. glabrata, Geotrichum candidum, Aureobasidium pullulans, Wangiella dermatitidis, Trichosporon cutaneum and Exophiala werneckii. Eleven animals harbored identical fungi in more than one of their internal organs; seven animals had more than one fungal species colonizing a single organ. Our findings indicated probable natural subclinical infections of candidiasis, geotrichosis or phaeohyphomycosis, and also symbiotic presence of non-pathogenic fungi among neotropical anurans.

Animals↗

The effect of freezing and the influence of isolation medium on the recovery of pathogenic fungi from sputum.

The primary objective of this study was to determine whether freezing sputa in dry ice had any effect on the recovery of pathogenic fungi. Sputa seeded with each of five fungi (Histoplasma capsulatum, Blastomyces dermatitidis, Cryptococus neoformans, Coccidioides immitis, and Aspergillus fumigatus) were frozen and stored for 24, 48, and 72 hours on dry ice. H. capsulatum was killed, and only a few colonies of B. dermatitidis and C. neoformans were isolated from these sputa. However, A. fumigatus and C. immitis withstood the effects of freezing. A second objective was to compare the recovery of all five fungi from seeded sputa stored at room temperature for 24, 48, and 72 hours, on yeast extract-phosphate agar with NH4OH and on Sabhi agar. The yeast extract-phosphate agar with NH4OH was superior to Sabhi agar, for the isolation of all fungi studied, except A. fumigatus.

Agar↗

Classical and molecular approaches as a powerful tool for the characterization of rumen polycentric fungi.

Ribosomal ITS1 and ITS2 fragments from 8 isolates of polycentric rumen anaerobic fungi were PCR-amplified and sequenced; the sequences obtained were aligned with published data and phylogenetic analyses were performed. Analysis of the ITS1 fragment clearly differentiated between the two polycentric genera Orpinomyces and Anaeromyces and this classification is supported by morphological observation. A multi-order phylogram based on ITS2 sequences proved that anaerobic rumen fungi are separated from aerobic chytrids, which form a well-supported monophylum with the highest possible bootstrap proportion values of 100%. Sequence analysis of ITS regions is a powerful tool for classification of anaerobic fungi but morphological description of strains is still necessary because some genera of rumen fungi display a high genetic heterogeneity.

Animals↗

Formation of glucoside conjugate of acetaminophen by fungi separated from soil.

The phase II metabolite of acetaminophen in filamentous fungi and actinomycetes separated from soil was investigated. Fifty-four filamentous fungi and twenty-seven actinomycetes were screened to transform acetaminophen. The metabolites of acetaminophen were assayed using liquid chromatography-tandem mass spectrometry. The only metabolite was subject to enzymatic hydrolysis to confirm its structure. Acetaminophen was converted into glucoside conjugate, by filamentous fungi JX1-60, LN17-2, LN20-1 and the yield of the conjugate was 60.01%, 44.27%, 100%, respectively, and no phase I metabolites were detected. Glucoside conjugation of acetaminophen in filamentous fungi differs from the phase II metabolism of glucuronidation in humans. The fungus LN20-1 could be a suitable model to synthesize glucoside conjugate of acetaminophen.

Acetaminophen↗

Contribution of hydrolytic enzymes produced by saprophytic fungi to the decrease in plant toxicity caused by water-soluble substances in olive mill dry residue.

We studied the influence of saprophytic fungi on the toxic effect that the water-soluble substances in dry residues from olive (ADOR) have on the growth of plants. All saprophytic fungi were able to decrease the phytotoxicity of ADOR, although the toxicity of this residue did not decrease in the same way. Penicillium chrysogenum was able to reduce the toxicity of ADOR when this residue was applied at the highest dose of 15%. Fusarium lateritum, F. graminearum and Mucor racemosus were able to reduce the toxicity of ADOR when this residue was applied at the intermediate doses. However, F. oxysporum decreased the phytotoxicity of ADOR only when the residue was applied at the lowest dose of 2.5%. All saprophytic fungi tested produce endoglucanase, endopolymetylgalacturonase and endoxiloglucanase when grown in the presence of ADOR. A close relationship was found between the decrease in the phytotoxicity of ADOR and the amount of hydrolytic enzymes produced by the saprophytic fungi. These results shows that hydrolytic enzymes can be important in the degradation of phytotoxic substances present in olive mill dry residue.

Biodegradation, Environmental↗

Degradation of fresh ryegrass by methanogenic co-cultures of ruminal fungi grown in the presence or absence of Fibrobacter succinogenes.

The ability of five ruminal fungi in syntrophic co-culture with the methanogen Methanobrevibacter smithii to degrade perennial ryegrass ( Lolium perenne) stem fragments and leaf blades was studied to determine the susceptibilities of non-autoclaved fresh tissues to fungal degradation. Autoclaving did not significantly increase fungal degradation of stem fragments but strongly increased degradation of leaf blades by a species of Caecomyces. In methanogenic co-cultures, non-autoclaved stem fragments were degraded more extensively by Neocallimastix frontalis and Piromyces isolates than by Caecomyces isolates. The N. frontalis and Piromyces isolates showed the greatest rates of stem degradation. When interactions between Fibrobacter succinogenes and methanogenic co-cultures of fungi growing on ryegrass stem were investigated, N. frontalis inhibited F. succinogenes. This has not been observed previously. In contrast, a Caecomyces species interacted positively with F. succinogenes to increase stem degradation, suggesting that F. succinogenes and Caecomyces spp. may have complementary fibrolytic activities. All five fungi tested failed to grow on fresh non-autoclaved leaf blades. In a repeat experiment with leaves from a separate harvest, leaf blades were degraded by N. frontalis but not by a Caecomyces species. We suggest that ryegrass leaf blades may contain natural anti-fungal compounds. Our results confirm the superiority of fungi in the degradation of intact stem and indicate that in vitro studies with non-autoclaved forage tissues may yield new information on forage factors affecting rumen microbes.

Anaerobiosis↗

Differential RNA accumulation of two beta-tubulin genes in arbuscular mycorrhizal fungi.

RNA was isolated from spores of different arbuscular mycorrhizal (AM) fungi and used for RT-PCR with degenerate primers for beta-tubulin genes. PCR products were cloned and the sequence of several clones was analysed for each fragment. Comparison of sequences identified two loci for beta-tubulin genes with different GC content and codon usage. Btub1 sequences were most similar to beta-tubulin genes from the Oomycota, while Btub2 sequences showed highest similarity to sequences from the Zygomycota. RT-PCR experiments were carried out to monitor RNA accumulation patterns of Btub1 and Btub2 in asymbiotic germinating spores and in symbiotic extraradical hyphae of three different AM fungi. This indicated that Btub1 is constitutively expressed in Gigaspora rosea, but down-regulated during symbiosis in Glomus mosseae and Glomus intraradices. In contrast, Btub2 showed constitutive expression in the two Glomus species, but down-regulation in G. rosea. Further analysis of different fungi indicated that Btub2 primers could be used to specifically monitor RNA accumulation of AM fungi in environmental samples.

Founder Effect↗

Variation in aluminum resistance among arbuscular mycorrhizal fungi.

Arbuscular mycorrhizal (AM) fungi mediate interactions between plants and soils, and are important where nutrient or metal concentrations limit plant growth. Variation in fungal response to edaphic conditions may influence the effectiveness of the plant-mycorrhizal association in some soil environments. Andropogon virginicus (broomsedge) colonizes disturbed sites in the eastern United States, including acidic mine soils where aluminum (Al) is phytotoxic, and Al resistance in broomsedge has been associated with colonization by the AM fungus Glomus clarum. In the present study, inter- and intra-specific variation to confer Al resistance to broomsedge was assessed among selected species of AM fungi. Broomsedge seeds were grown in sand culture inoculated with one of five isolates of three species of fungi (G. clarum, Acaulospora morrowiae, and Scutellospora heterogama). Plants were exposed to 0 or 400 microM Al in nutrient solution and harvested after 4 or 9 weeks of growth. Mean infection percentage, plant biomass, and plant tissue Al and phosphorus (P) concentrations were measured. G. clarum conferred the greatest Al resistance to broomsedge, with the lowest variability among isolates for colonization and growth inhibition by Al [tolerance indices (TI) between 22.4 and 92.7%]. Broomsedge plants colonized by A. morrowiae were consistently the most sensitive to Al, with little variation among isolates (TI between 1.6 and 12.1%). Al resistance by S. heterogama isolates was intermediate and wide-ranging (TI between 3.9 and 40.0%). Across all AM fungal isolates, resistance was associated with high rates of colonization and low tissue Al concentrations of broomsedge plants. The functional diversity in Al resistance displayed by these AM fungi reflect variation in acclimation mechanisms operating in the mycorrhizal symbiosis under environmental stress.

Aluminum↗

Evaluation of the viability of pathogenic filamentous fungi after prolonged storage in sterile water and review of recent published studies on storage methods.

We have evaluated the survival and potential morphological alterations of 45 species of pathogenic filamentous fungi that had been stored in sterile water following Castellani's method in the National Collection of Pathogenic Fungi (NCPF). Storage duration varied from 2 months to over 21 years. Ninety percent of stored organisms were shown to be viable. Viability was largely independent of the duration of storage, but did apparently vary to some degree in an organism-specific manner. In addition, certain fungi were shown to have undergone morphological alterations during storage, and exhibited significant degrees of pleomorphism upon revival. This was especially marked for several isolates of dermatophytes, where storage resulted in loss of recognisable colonial features, and overproduction of sterile mycelium with aberrant or no conidia. These findings suggest that while Castellani's method remains an easy and inexpensive method for long-term preservation of most fungi, water storage should be supplemented by a second storage method to increase the chances of retaining both viability and morphological stability over long periods.

Mitosporic Fungi↗