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PubMed2000

Differential C isotope discrimination by fungi during decomposition of C(3)- and C(4)-derived sucrose.

Stable isotope analysis is a major tool used in ecosystem studies to establish pathways and rates of C exchange between various ecosystem components. Little is known about isotopic effects of many such components, especially microbes. Here we report on the discovery of an unexpected pattern of C isotopic discrimination by basidiomycete fungi with far-reaching consequences for our understanding of isotopic processing in ecosystems where these microbes mediate material transfers across trophic levels. We measured fractionation effects on three ecologically relevant basidiomycete species under controlled laboratory conditions. Sucrose derived from C(3) and C(4) plants is fractionated differentially by these microbes in a taxon-specific manner. The differentiation between mycorrhizal and saprotrophic fungi observed in the field by others is not explained by intrinsic discrimination patterns. Fractionation occurs during sugar uptake and is sensitive to the nonrandom distribution of stable isotopes in the sucrose molecule. The balance between respiratory physiology and fermentative physiology modulates the degree of fractionation. These discoveries disprove the assumption that fungal C processing does not significantly alter the distribution of stable C isotopes and provide the basis for a reevaluation of ecosystem models based on isotopic evidence that involve C transfer across microbial interfaces. We provide a mechanism to account for the observed differential discrimination effects.

Carbon Isotopes↗
PubMed2000

Effects of metal phytoextraction practices on the indigenous community of arbuscular mycorrhizal fungi at a metal-contaminated landfill.

Phytoextraction involves use of plants to remove toxic metals from soil. We examined the effects of phytoextraction practices with three plant species (Silene vulgaris, Thlaspi caerulescens, and Zea mays) and a factorial variation of soil amendments (either an ammonium or nitrate source of nitrogen and the presence or absence of an elemental sulfur supplement) on arbuscular mycorrhizal (AM) fungi (Glomales, Zygomycetes) at a moderately metal-contaminated landfill located in St. Paul, Minn. Specifically, we tested whether the applied treatments affected the density of glomalean spores and AM root colonization in maize. Glomalean fungi from the landfill were grouped into two morphotypes characterized by either light-colored spores (LCS) or dark-colored spores (DCS). Dominant species of the LCS morphotype were Glomus mosseae and an unidentified Glomus sp., whereas the DCS morphotype was dominated by Glomus constrictum. The density of spores of the LCS morphotype from the phytoremediated area was lower than the density of these spores in the untreated landfill soil. Within the experimental area, spore density of the LCS morphotype in the rhizosphere of mycorrhizal maize was significantly higher than in rhizospheres of nonmycorrhizal S. vulgaris or T. caerulescens. Sulfur supplement increased vesicular root colonization in maize and exerted a negative effect on spore density in maize rhizosphere. We conclude that phytoextraction practices, e.g., the choice of plant species and soil amendments, may have a great impact on the quantity and species composition of glomalean propagules as well as on mycorrhiza functioning during long-term metal-remediation treatments.

Biodegradation, Environmental↗
PubMed2000

Antibiosis between ruminal bacteria and ruminal fungi.

Cellulose digestion, bacterial numbers, and fungal numbers were monitored over time in vitro by using a purified cellulose medium with and without antibiotics (penicillin and streptomycin). All fermentations were inoculated with a 1:10 dilution of whole rumen contents (WRC). Without antibiotics, cellulose digestion was higher (P < 0.01) at 24, 30, 48, and 72 h; fungi had almost disappeared by 24 h, while bacterial concentrations increased over 100-fold in 24 h and then decreased gradually up to 72 h. In those fermentations with added antibiotics, fungal concentrations increased 4-fold by 30 h and up to 42-fold at 72 h; bacterial concentrations were markedly reduced by 24 h and remained low through 72 h. Similar results were obtained with ground alfalfa as a substrate. In further studies, the in vitro fermentation of purified cellulose without antibiotics was stopped after 18 to 20 h, and the microbial population was killed by autoclaving. Antibiotics were added to half of the tubes, and all tubes were reinoculated with WRC. After 72 h, extensive cellulose digestion had occurred in those tubes without antibiotics, as compared to very low cellulose digestion with added antibiotics. The extent of this inhibition was found to increase in proportion to the length of the initial fermentation period, suggesting the production of a heat-stable inhibitory factor or factors. The inhibitory activity was present in rumen fluid, could be extracted from lyophilized rumen fluid (LRF) with water, and was stable in response to proteolytic enzymes. In addition, the water-extracted residue of LRF was found to contain growth factor activity for rumen fungi in vitro.

Animals↗
PubMed2001

Impact of biocontrol Pseudomonas fluorescens CHA0 and a genetically modified derivative on the diversity of culturable fungi in the cucumber rhizosphere.

Little is known about the effects of Pseudomonas biocontrol inoculants on nontarget rhizosphere fungi. This issue was addressed using the biocontrol agent Pseudomonas fluorescens CHA0-Rif, which produces the antimicrobial polyketides 2,4-diacetylphloroglucinol (Phl) and pyoluteorin (Plt) and protects cucumber from several fungal pathogens, including Pythium spp., as well as the genetically modified derivative CHA0-Rif(pME3424). Strain CHA0-Rif(pME3424) overproduces Phl and Plt and displays improved biocontrol efficacy compared with CHA0-Rif. Cucumber was grown repeatedly in the same soil, which was left uninoculated, was inoculated with CHA0-Rif or CHA0-Rif(pME3424), or was treated with the fungicide metalaxyl (Ridomil). Treatments were applied to soil at the start of each 32-day-long cucumber growth cycle, and their effects on the diversity of the rhizosphere populations of culturable fungi were assessed at the end of the first and fifth cycles. Over 11,000 colonies were studied and assigned to 105 fungal species (plus several sterile morphotypes). The most frequently isolated fungal species (mainly belonging to the genera Paecilomyces, Phialocephala, Fusarium, Gliocladium, Penicillium, Mortierella, Verticillium, Trichoderma, Staphylotrichum, Coniothyrium, Cylindrocarpon, Myrothecium, and Monocillium) were common in the four treatments, and no fungal species was totally suppressed or found exclusively following one particular treatment. However, in each of the two growth cycles studied, significant differences were found between treatments (e.g., between the control and the other treatments and/or between the two inoculation treatments) using discriminant analysis. Despite these differences in the composition and/or relative abundance of species in the fungal community, treatments had no effect on species diversity indices, and species abundance distributions fit the truncated lognormal function in most cases. In addition, the impact of treatments at the 32-day mark of either growth cycle was smaller than the effect of growing cucumber repeatedly in the same soil.

Alanine↗
PubMed2003

Fungal lipid accumulation and development of mycelial structures by two arbuscular mycorrhizal fungi.

We monitored the development of intraradical and extraradical mycelia of the arbuscular mycorrhizal (AM) fungi Scutellospora calospora and Glomus intraradices when colonizing Plantago lanceolata. The occurrence of arbuscules (branched hyphal structures) and vesicles (lipid storage organs) was compared with the amounts of signature fatty acids. The fatty acid 16:1omega5 was used as a signature for both AM fungal phospholipids (membrane constituents) and neutral lipids (energy storage) in roots (intraradical mycelium) and in soil (extraradical mycelium). The formation of arbuscules and the accumulation of AM fungal phospholipids in intraradical mycelium followed each other closely in both fungal species. In contrast, the neutral lipids of G. intraradices increased continuously in the intraradical mycelium, while vesicle occurrence decreased after initial rapid root colonization by the fungus. S. calospora does not form vesicles and accumulated more neutral lipids in extraradical than in intraradical mycelium, while the opposite pattern was found for G. intraradices. G. intraradices allocated more of its lipids to storage than did S. calospora. Thus, within a species, the fatty acid 16:1omega5 is a good indicator for AM fungal development. The phospholipid fatty acid 16:1omega5 is especially suitable for indicating the frequency of arbuscules in the symbiosis. We propose that the ratio of neutral lipids to phospholipids is more important than is the presence of vesicles in determining the storage status of AM fungi.

Fatty Acids↗
PubMed2003

18S rRNA gene variation among common airborne fungi, and development of specific oligonucleotide probes for the detection of fungal isolates.

In this study, we sequenced 18S rRNA genes (rDNA) from 49 fungal strains representing 31 species from 15 genera. Most of these species are common airborne fungi and pathogens that may cause various public health concerns. Sequence analysis revealed distinct divergence between Zygomycota and Ascomycota. Within Ascomycota, several strongly supported clades were identified that facilitate the taxonomic placement of several little-studied fungi. Wallemia appeared as the group most diverged from all the other Ascomycota species. Based on the 18S rDNA sequence variation, 108 oligonucleotide probes were designed for each genus and species included in this study. After homology searches and DNA hybridization evaluations, 33 probes were verified as genus or species specific. The optimal hybridization temperatures to achieve the best specificity for these 33 probes were determined. These new probes can contribute to the molecular diagnostic research for environmental monitoring.

Air Microbiology↗
PubMed2004

Heavy-metal stress and developmental patterns of arbuscular mycorrhizal fungi.

The rate of global deposition of Cd, Pb, and Zn has decreased over the past few decades, but heavy metals already in the soil may be mobilized by local and global changes in soil conditions and exert toxic effects on soil microorganisms. We examined in vitro effects of Cd, Pb, and Zn on critical life stages in metal-sensitive ecotypes of arbuscular mycorrhizal (AM) fungi, including spore germination, presymbiotic hyphal extension, presymbiotic sporulation, symbiotic extraradical mycelium expansion, and symbiotic sporulation. Despite long-term culturing under the same low-metal conditions, two species, Glomus etunicatum and Glomus intraradices, had different levels of sensitivity to metal stress. G. etunicatum was more sensitive to all three metals than was G. intraradices. A unique response of increased presymbiotic hyphal extension occurred in G. intraradices exposed to Cd and Pb. Presymbiotic hyphae of G. intraradices formed presymbiotic spores, whose initiation was more affected by heavy metals than was presymbiotic hyphal extension. In G. intraradices grown in compartmentalized habitats with only a portion of the extraradical mycelium exposed to metal stress, inhibitory effects of elevated metal concentrations on symbiotic mycelial expansion and symbiotic sporulation were limited to the metal-enriched compartment. Symbiotic sporulation was more sensitive to metal exposure than symbiotic mycelium expansion. Patterns exhibited by G. intraradices spore germination, presymbiotic hyphal extension, symbiotic extraradical mycelium expansion, and sporulation under elevated metal concentrations suggest that AM fungi may be able to survive in heavy metal-contaminated environments by using a metal avoidance strategy.

Cadmium↗
PubMed2004

Wood-destroying soft rot fungi in the historic expedition huts of Antarctica.

Three expedition huts in the Ross Sea region of Antarctica, built between 1901 and 1911 by Robert F. Scott and Ernest Shackleton, sheltered and stored the supplies for up to 48 men for 3 years during their explorations and scientific investigation in the South Pole region. The huts, built with wood taken to Antarctica by the early explorers, have deteriorated over the past decades. Although Antarctica has one of the coldest and driest environments on earth, microbes have colonized the wood and limited decay has occurred. Some wood in contact with the ground contained distinct microscopic cavities within secondary cell walls caused by soft rot fungi. Cadophora spp. could be cultured from decayed wood and other woods sampled from the huts and artifacts and were commonly associated with the soft rot attack. By using internal transcribed spacer sequences of ribosomal DNA and morphological characteristics, several species of Cadophora were identified, including C. malorum, C. luteo-olivacea, and C. fastigiata. Several previously undescribed Cadophora spp. also were found. At the Cape Evans and Cape Royds huts, Cadophora spp. commonly were isolated from wood in contact with the ground but were not always associated with soft rot decay. Pure cultures of Cadophora used in laboratory decay studies caused dark staining of all woods tested and extensive soft rot in Betula and Populus wood. The presence of Cadophora species, but only limited decay, suggests there is no immediate threat to the structural integrity of the huts. These fungi, however, are widely found in wood from the historic huts and have the capacity to cause extensive soft rot if conditions that are more conducive to decay become common.

Antarctic Regions↗
PubMed2004

Development and use of flow cytometry for detection of airborne fungi.

Traditional methods for the enumeration of airborne fungi are slow, tedious, and rather imprecise. In this study, the possibility of using flow cytometry (FCM) for the assessment of exposure to the fungus aerosol was evaluated. Epifluorescence microscopy direct counting was adopted as the standard for comparison. Setting up of the method was achieved with pure suspensions of Aspergillus fumigatus and Penicillium brevicompactum conidia at different concentrations, and then analyses were extended to field samples collected by an impinger device. Detection and quantification of airborne fungi by FCM was obtained combining light scatter and propidium iodide red fluorescence parameters. Since inorganic debris are unstainable with propidium iodide, the biotic component could be recognized, whereas the preanalysis of pure conidia suspensions of some species allowed us to select the area corresponding to the expected fungal population. A close agreement between FCM and epifluorescence microscopy counts was found. Moreover, data processing showed that FCM can be considered more precise and reliable at any of the tested concentrations.

Air Microbiology↗
PubMed2005

Hydroxylation of the herbicide isoproturon by fungi isolated from agricultural soil.

Several asco-, basidio-, and zygomycetes isolated from an agricultural field were shown to be able to hydroxylate the phenylurea herbicide isoproturon [N-(4-isopropylphenyl)-N',N'-dimethylurea] to N-(4-(2-hydroxy-1-methylethyl)phenyl)-N',N'-dimethylurea and N-(4-(1-hydroxy-1-methylethyl)phenyl)-N',N'-dimethylurea. Bacterial metabolism of isoproturon has previously been shown to proceed by an initial demethylation to N-(4-isopropylphenyl)-N'-methylurea. In soils, however, hydroxylated metabolites have also been detected. In this study we identified fungi as organisms that potentially play a major role in the formation of these hydroxylated metabolites in soils treated with isoproturon. Isolates of Mortierella sp. strain Gr4, Phoma cf. eupyrena Gr61, and Alternaria sp. strain Gr174 hydroxylated isoproturon at the first position of the isopropyl side chain, yielding N-(4-(2-hydroxy-1-methylethyl)phenyl)-N',N'-dimethylurea, while Mucor sp. strain Gr22 hydroxylated the molecule at the second position, yielding N-(4-(1-hydroxy-1-methylethyl)phenyl)-N',N'-dimethylurea. Hydroxylation was the dominant mode of isoproturon transformation in these fungi, although some cultures also produced traces of the N-demethylated metabolite N-(4-isopropylphenyl)-N'-methylurea. A basidiomycete isolate produced a mixture of the two hydroxylated and N-demethylated metabolites at low concentrations. Clonostachys sp. strain Gr141 and putative Tetracladium sp. strain Gr57 did not hydroxylate isoproturon but N demethylated the compound to a minor extent. Mortierella sp. strain Gr4 also produced N-(4-(2-hydroxy-1-methylethyl)phenyl)-N'-methylurea, which is the product resulting from combined N demethylation and hydroxylation.

Agriculture↗
PubMed1968

Assimilation of alkanes and alkenes by fungi.

A group of filamentous fungi were assayed for their ability to utilize a series of n-alkanes and 1-alkenes as the sole source of carbon. Although strains of Cunninghamella exhibited profuse growth on most of the hydrocarbons tested, the majority of fungi tested were able to produce definite growth on one or more of the compounds. The hydrocarbons with a 14-carbon chain length appeared to be more consistently utilized than any other. Strains of Aspergillus appeared to differ in their capacity to utilize individual members of the hydrocarbon series. Thin-layer chromatographic analyses of ether extracts from C. blakesleeana grown on n-tetradecane and 1-tetradecene were similar and revealed the presence of a monocarboxylic acid, a primary alcohol, and a secondary alcohol.

Alkanes↗
PubMed1969

Mouse toxicty of fungi of tobacco.

A bioassay for fungal toxins based on the intraperitoneal injection of test materials into mice was used to screen 976 cultures isolated from tobacco and grown in a high-protein baby cereal and also to determine whether samples of tobacco damaged by fungi are more toxic than samples of apparently sound tobacco. Of 236 fungal isolates from noncured tobacco, 79% were lethal when homogenized cultures of these isolates were tested. Forty-nine per cent of 740 fungi isolated from cured tobacco were lethal. Of the genera from which 30 or more isolates were tested, Epicoccum, Alternaria, and Penicillium had the highest percentage of toxic isolates from non-cured tobacco, whereas Epicoccum, Aspergillus, and Alternaria had the highest percentage from cured tobacco. Samples of tobacco naturally infected with brown spot, caused by Alternaria tenuis, did not have a significantly different LD(50) value after 48 hr than comparable disease-free samples. However, animals which died from doses near the LD(50) dose of tobacco infected with Alternaria generally died in 24 to 48 hr with signs associated with a depressant rather than a stimulant, such as nicotine, which caused death in 15 to 30 min. These signs were duplicated by injecting homogenized pure cultures of Alternaria. These studies, although inconclusive with regard to the effects of fungal contaminants on the quality or usability of tobacco, have developed evidence that suggests the advisability of a study on smoke or smoke condensates from moldy and nonmoldy tobacco.

Animals↗
PubMed1974

Storage of stock cultures of filamentous fungi, yeasts, and some aerobic actinomycetes in sterile distilled water.

Castellani's procedure for maintaining cultures of filamentous fungi and yeasts in sterile distilled water was evaluated. Four hundred and seventeen isolates of 147 species belonging to 66 genera of filamentous fungi, yeasts, and aerobic actinomycetes were maintained in sterile distilled water at room temperature over periods ranging from 12 to 60 months in four independent experiments. Of the 417 cultures, 389 (93%) survived storage in sterile distilled water. The selection of good sporulating cultures and sufficient inoculum consisting of spores and hyphae suspended in sterile distilled water were the most important factors influencing survival in water over a longer period of time. The technique was found to be simple, inexpensive, and reliable.

Actinomycetales↗
PubMed1974

Fermentation of feedlot waste filtrate by fungi and streptomycetes.

The soluble and dispersed nitrogen and carbon components in the filtrate fraction of cattle feedlot waste are a potential nutrient source from which single-cell protein could be produced for animal feeds. The ability of more than 200 fungi and streptomycetes to grow in this liquid was determined; these included isolates from the waste and associated sources, as well as organisms maintained in the Culture Collection of the Agricultural Research Service in Peoria, Ill. Utilization of waste nutrients was measured by changes in nitrogen content and chemical oxygen demand. Only 20% of the organisms were able to grow appreciably in the filtrate. Of these, dry-weight yields varied from 0.6 to 2.7 g of mycelium per liter; from 21 to 50% of the nitrogen in the filtrates was used during growth, whereas chemical oxygen demand levels diminished from 4 to 60%. In general, streptomycetes isolated from the feedlot used nutrients from the filtrates better than fungi did. Addition of readily available carbon sources such as glucose or whey significantly increased (as much as sixfold) cell yields of selected organisms and promoted better utilization of nitrogen (from two- to threefold); the effect on chemical oxygen demand varied (0 to 33% increase).

Animals↗
PubMed2004

Surface glycans of Candida albicans and other pathogenic fungi: physiological roles, clinical uses, and experimental challenges.

Although fungi have always been with us as commensals and pathogens, fungal infections have been increasing in frequency over the past few decades. There is a growing body of literature describing the involvement of carbohydrate groups in various aspects of fungal disease. Carbohydrates comprising the cell wall or capsule, or as a component of glycoproteins, are the fungal cell surface entities most likely to be exposed to the surrounding environment. Thus, the fungus-host interaction is likely to involve carbohydrates before DNA, RNA, or even protein. The interaction between fungal and host cells is also complex, and early studies using whole cells or crude cell fractions often produced seemingly conflicting results. What was needed, and what has been developing, is the ability to identify specific glycan structures and determine how they interact with immune system components. Carbohydrate analysis is complicated by the complexity of glycan structures and by the challenges of separating and detecting carbohydrates experimentally. Advances in carbohydrate chemistry have enabled us to move from the foundation of composition analysis to more rapid characterization of specific structures. This, in turn, will lead to a greater understanding of how fungi coexist with their hosts as commensals or exist in conflict as pathogens.

Animals↗
PubMed1993

Compounds active against cell walls of medically important fungi.

A number of substances that directly or indirectly affect the cell walls of fungi have been identified. Those that actively interfere with the synthesis or degradation of polysaccharide components share the property of being produced by soil microbes as secondary metabolites. Compounds specifically interfering with chitin or beta-glucan synthesis have proven effective in studies of preclinical models of mycoses, though they appear to have a restricted spectrum of coverage. Semisynthetic derivatives of some of the natural products have offered improvements in activity, toxicology, or pharmacokinetic behavior. Compounds which act on the cell wall indirectly or by a secondary mechanism of action, such as the azoles, act against diverse fungi but are usually fungistatic in nature. Overall, these compounds are attractive candidates for further development.

Aminoglycosides↗
PubMed1996

Activation of the complement system by pathogenic fungi.

Fungi have been studied as prototype activators of the complement cascade since the early 1900s. More recently, attention has focused on the role of the complement system in the pathogenesis of fungal infections. The interactions of Cryptococcus neoformans and Candida albicans with the complement system are the most widely characterized; however, all pathogenic fungi examined to date have the ability to initiate the complement cascade. The molecular mechanisms for initiation and regulation of the complement cascade differ from one fungus to another, most likely reflecting differences in the structure of the outer layers of the cell wall. The molecular bases for such differences remain to be identified. Studies of mycoses in experimental animals with induced or congenital deficiencies in the complement system demonstrate that complement is an important innate system for control of fungal infection. Contributions to host resistance include opsonization and generation of inflammatory mediators. Inflammation induced by chemotactic products of the complement system may contribute to the pathogenesis of some fungal infections.

Aspergillosis↗
PubMed2004

Novel p-type ATPases mediate high-affinity potassium or sodium uptake in fungi.

Fungi have an absolute requirement for K+, but K+ may be partially replaced by Na+. Na+ uptake in Ustilago maydis and Pichia sorbitophila was found to exhibit a fast rate, low Km, and apparent independence of the membrane potential. Searches of sequences with similarity to P-type ATPases in databases allowed us to identify three genes in these species, Umacu1, Umacu2, and PsACU1, that could encode P-type ATPases of a novel type. Deletion of the acu1 and acu2 genes proved that they encoded the transporters that mediated the high-affinity Na+ uptake of U. maydis. Heterologous expressions of the Umacu2 gene in K+ transport mutants of Saccharomyces cerevisiae and transport studies in the single and double Deltaacu1 and Deltaacu2 mutants of U. maydis revealed that the acu1 and acu2 genes encode transporters that mediated high-affinity K+ uptake in addition to Na+ uptake. Other fungi also have genes or pseudogenes whose translated sequences show high similarity to the ACU proteins of U. maydis and P. sorbitophila. In the phylogenetic tree of P-type ATPases all the identified ACU ATPases define a new cluster, which shows the lowest divergence with type IIC, animal Na+,K(+)-ATPases. The fungal high-affinity Na+ uptake mediated by ACU ATPases is functionally identical to the uptake that is mediated by some plant HKT transporters.

Adenosine Triphosphatases↗