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PubMed1998

Agrobacterium tumefaciens-mediated transformation of filamentous fungi.

Agrobacterium tumefaciens transfers part of its Ti plasmid, the T-DNA, to plant cells during tumorigenesis. It is routinely used for the genetic modification of a wide range of plant species. We report that A. tumefaciens can also transfer its T-DNA efficiently to the filamentous fungus Aspergillus awamori, demonstrating DNA transfer between a prokaryote and a filamentous fungus. We transformed both protoplasts and conidia with frequencies that were improved up to 600-fold as compared with conventional techniques for transformation of A. awamori protoplasts. The majority of the A. awamori transformants contained a single T-DNA copy randomly integrated at a chromosomal locus. The T-DNA integrated into the A. awamori genome in a manner similar to that described for plants. We also transformed a variety of other filamentous fungi, including Aspergillus niger, Fusarium venenatum, Trichoderma reesei, Colletotrichum gloeosporioides, Neurospora crassa, and the mushroom Agaricus bisporus, demonstrating that transformation using A. tumefaciens is generally applicable to filamentous fungi.

Agrobacterium tumefaciens↗
PubMed2025

Limited contributions of bacteria and fungi to coral nutrition revealed by amino acid δ13C analysis.

Corals often form reef ecosystems that support diverse marine life, but they are sensitive to environmental fluctuations that can affect their nutrient acquisition. While coral-associated microbes (e.g., Symbiodiniaceae, bacteria and fungi) may supplement nutrients to coral hosts via metabolite translocation and nutrient recycling, the extent to which these microbial partners contribute to coral autotrophy or heterotrophy remains unclear. Here, we seasonally measure the carbon isotopes of amino acids (δ13CAA) in reef-building coral Pocillopora damicornis and its nutrient sources (e.g., Symbiodiniaceae and particulate organic matter). Regional Bayesian mixing models show that P. damicornis increased autotrophy (from 67.1 to 80.5%), but decreased particulate feeding (from 32.9 to 19.5%) from the cool season to the warm season. Stable essential δ13CAA values (valine, leucine and isoleucine) suggest limited seasonal changes in microbial contributions. Linear discriminant analysis, which combines current and published data from basal organisms (e.g., bacteria and fungi) to coral consumers, also reveals limited bacterial and fungal contributions to coral nutrition. Thus, we advocate that coral nutrition is primarily determined by Symbiodiniaceae translocation and particulate feeding. As these nutritional pathways are highly subject to environmental fluctuations, corals lacking trophic flexibility may suffer more from malnutrition and even population decline under global environmental change.

Anthozoa↗
PubMed2002

Mate recognition in fungi.

The ascomycete and basidiomycete fungi have contributed much to our understanding of eukaryotic cell biology. The study of mate recognition, in particular, has provided detailed understanding of cell signalling pathways and cell-specific gene transcription. Sexual dimorphism has little relevance to mating in these organisms, indeed specialised cells for mating are found only in filamentous ascomycetes and even here, a single individual produces both male and female structures. None the less, most species have genetic barriers to prevent selfing. The genes that determine self-incompatibility divide populations into different mating types, and only individuals with different mating types can engage in sexual reproduction. Ascomycetes have just two mating types, but basidiomycetes may have several thousands. Despite apparent differences in the biology and numbers of mating types in these fungi, it is becoming increasingly apparent that many components of their mating pathways are highly conserved.

Fungi↗
PubMed2003

Reproducibility of allergen, endotoxin and fungi measurements in the indoor environment.

Measurements of biocontaminants in settled house dust once a year are commonly used to assess long-term exposure. To examine stability over time and seasonal variation, we measured concentrations of mite and cat allergens, endotoxin and mold spores in living room floor dust in 745 German homes collected twice a year in two different seasons. The study population consisted of adults and children living in five different areas in Germany. All dust samples were collected in a standardized manner from the living room floor and taken during the years 1995 to 1998. The median interval between the two dust samplings was approximately 7 months. Mite and cat allergens were measured in settled house dust by monoclonal antibodies, endotoxin by the limulus amebocyte lysate method, and total spore counts by cultural methods. Crude Pearson's correlation coefficients between log-transformed concentrations in the first and second dust samples ranged between 0.65 and 0.75 for allergens, 0.59 for endotoxin and only 0.06 for total spore counts. The strongest and most consistent seasonal effects were observed for fungi with highest levels in July-September. Cat allergen concentrations were found consistently to be increased in January-March. Mite allergens did not show a strong and consistent seasonal pattern. We conclude that repeated measurements of mite and cat allergens and endotoxin in settled house dust improve the estimate for annual mean concentrations. However, even a single observation of these biocontaminants may be a good proxy for a 1-year exposure since repeated measures were highly correlated. However, repeated measurements of fungi levels were only weakly correlated and thus repeated observations for assessment of annual means of total spore counts are needed.

Air Pollution, Indoor↗
PubMed1999

Determination of selected microbial volatile organic compounds by diffusive sampling and dual-column capillary GC-FID--a new feasible approach for the detection of an exposure to indoor mould fungi?

A new, analytically valid procedure is described to assess the exposure of human beings to the so-called microbial volatile organic compounds (MVOCs) in air. The method can be used routinely for large sample numbers and is especially valuable as a basis for further research on the correlation between single MVOCs and indoor mould growth. The procedure is based on the fact that fungi produce a variety of volatile organic compounds, such as 3-methylbutan-1-ol, 3-methylbutan-2-ol, fenchone, heptan-2-one, hexan-2-one, octan-3-one, octan-3-ol, pentan-2-ol, alpha-terpineol, and thujopsene, which they emit into the indoor environment. Using diffusive samplers, these MVOCs are adsorbed onto charcoal during a sampling interval of four weeks. The described method is thus superior to existing methods which use short-term active sampling. After desorption with carbon disulfide, the MVOCs were determined by dual-column gas chromatography with flame ionization detection using the large-volume injection technique for sample introduction. The detection limits ranged between 0.15 and 0.53 microgram m-3, within-series precision was found to range between 6.5 and 19.0%, and recovery was between 77 and 118%. The procedure has been successfully applied in the context of a large field study to measure the indoor MVOC exposure in children's rooms of 132 dwellings. The objective of the study was to examine the relation between indoor mould growth, the indoor MVOC exposure and the prevalence of adverse health effects. Information about mould formation has been obtained by a questionnaire and by the determination of colony forming units of mould fungi in mattress dust. With the exception of 3-methylbutan-2-ol, fenchone, nonan-2-one, octan-2-one, and thujopsene, indoor air concentrations of all MVOCs under investigation were significantly higher inside damp and mouldy dwellings. From the primary MVOCs under investigation, 3-methylbutan-1-ol, hexan-2-one, heptan-2-one, and octan-3-ol were found to be most reliable indicators for mould formation. A correlation was also found between selected MVOCs and the occurrence of mould species in mattress dust. Aspergillus sp. correlated with heptan-2-one, hexan-2-one, octan-3-ol, octan-3-one, and alpha-terpineol, while the occurrence of Eurotium sp. was correlated with higher indoor air concentrations of 3-methylbutan-1-ol, 3-methylbutan-2-ol, heptan-2-one, hexan-2-one, octan-3-ol, and thujopsene. Children living in dwellings with elevated MVOC levels had a higher prevalence of asthma, hay fever, wheezing, and irritations of the eyes. These positive associations persisted after controlling for confounding factors such as age, sex, body-mass index, number of siblings, social status, passive smoking, type of heating, and ventilation habits. However, they were not statistically significant. This lack of significance may be a result of the small number of investigated samples.

Air Pollution, Indoor↗
PubMed2002

Evaluation of PCR primers and PCR conditions for specific detection of common airborne fungi.

We examined the selectivity of 53 sets of primers for environmental monitoring of indoor air quality. Thirty-six fungal strains, representing 26 species from 14 genera of commonly occurring fungi, and 16 different bacterial strains, representing both gram-negative and gram-positive species, were included in the experiment. We verified the specificity of 28 of the 53 sets of primers, which were classified as universal fungal, universal bacterial, group or species specific. The PCR conditions required for optimal specificity were also determined. These results can serve as a guide for the step-wise PCR-based detection and identification of airborne fungi commonly found in indoor environments.

Air Pollution, Indoor↗
PubMed2006

Acetylcholinesterase inhibitors from plants and fungi.

This review describes 183 compounds obtained from plants and fungi which have been shown to inhibit acetylcholinesterase. The mechanism of action of cholinesterase, together with the binding sites, and, where this is known, the mode of action of inhibitors is described. The relative activities of the different compounds are recorded. The strongest inhibitors are generally alkaloids although some meroterpenoids from fungi have also been found to be active and display better selectivity.

Biological Products↗
PubMed2004

Structural elucidation of novel phosphocholine-containing glycosylinositol-phosphoceramides in filamentous fungi and their induction of cell death of cultured rice cells.

Novel ZGLs (zwitterionic glycosphingolipids) have been found in and extracted from the mycelia of filamentous fungi ( Acremonium sp.) isolated from soil. Five ZGLs (ZGL1-ZGL5) were structurally elucidated by sugar compositional analysis, methylation analysis, periodate oxidation, matrix-assisted laser-desorption ionization-time-of-flight MS, (1)H-NMR spectroscopy and fast-atom bombardment MS. Their chemical structures were as follows: GlcN(alpha1-2)Ins1-P-1Cer (ZGL1), Man(alpha1-6)GlcN(alpha1-2)Ins1-P-1Cer (ZGL2), Man(alpha1-6)Man(alpha1-6)GlcN(alpha1-2)Ins1-P-1Cer (ZGL3), PC-->6Man(alpha1-6)GlcN(alpha1-2)Ins1- P -1Cer (ZGL4), and PC-->6Man(alpha1-6)Man(alpha1-6)GlcN(alpha1-2)Ins1-P-1Cer (ZGL5) (where Cer is ceramide and PC is phosphocholine). In addition, one acidic glycosphingolipid, which was the precursor of ZGLs, was also characterized as inositol-phosphoceramide. The core structure of the ZGLs, GlcN(alpha1-2)Ins1- P, is rather different from those found in other fungi, such as Man(alpha1-2)Ins1- P and Man(alpha1-6)Ins1- P. Interestingly, the terminal mannose residue of ZGL4 and ZGL5 was modified further with a PC group. The presence of PC-containing glycosylinositol-phosphoceramides has not been reported previously in any organism. The ceramide constituents of both ZGLs and acidic glycosphingolipid were essentially the same, and consisted of a 4-hydroxyoctadecasphinganine (phytosphingosine) as the sole sphingoid base and 2-hydroxytetracosanoic acid (>90%) as the major fatty acid. ZGLs were found to cause cell death in suspensions of cultured rice cells. The cell death-inducing activity of ZGLs is probably due to the characteristic glycan moiety of Man(alpha1-6)GlcN, and PC-containing ZGLs had high activity. This study is the first to demonstrate that fungal glycosylinositol-phosphoceramides induce cell death in cultured rice cells.

Acidic Glycosphingolipids↗
PubMed2002

Community analysis of arbuscular mycorrhizal fungi associated with Ammophila arenaria in Dutch coastal sand dunes.

A polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) approach for the detection and characterization of arbuscular mycorrhizal fungi (AMF) 18S ribosomal DNA (rDNA) was developed and applied to the study of AMF communities associated with the main sand-stabilizing plant species of the Dutch sand dunes, marram grass (Ammophila arenaria, L.). DNA was extracted directly from plant roots, soil or isolated AMF spores, and prominent bands resulting from AMF-specific DGGE profiles were excised for sequence analysis. This strategy provided a robust means of detecting and identifying AMF-like species without the use of trap plant cultivation methods. A number of Glomus-like and Scutellospora-like sequences was detected, including a putatively novel Glomus species, and differences were observed in the dominant AMF-like populations detected in healthy vs. degenerating stands of A. arenaria and in bulk sand dune soil. It has previously been suggested that plant pathogens, such as fungi and nematodes, may contribute to the decline of A. arenaria. Although no causal relationship can be drawn between the observed differences in the dominantly detected AMF-like populations and the vitality of plant growth, these results indicate that mutualistic interactions between this plant and AMF should not be overlooked when examining the role of soil-borne microorganisms in vegetation dynamics. In addition, there were discrepancies observed between the AMF-like groups detected in spore populations vs. direct 18S rDNA analysis of root material, corroborating previous suggestions that spore inspection alone may poorly represent actual AMF population structure.

DNA, Fungal↗
PubMed1999

Prevalence and residential determinants of fungi within homes in Melbourne, Australia.

BACKGROUND: Recent epidemiological studies suggest that the adverse respiratory health effects caused by the inhalation of fungal propagules are substantial. Knowledge of the prevalence and environmental determinants of indoor fungal levels is essential in designing effective avoidance measures. AIM: To investigate the prevalence of fungi and the influence of residential characteristics on levels of fungi within homes in Melbourne, Australia. METHODS: Floor dust and air samples were collected from bedrooms in 485 houses over 1 year. The dust was analysed for ergosterol, a marker of cumulative fungal biomass exposure. Total and genera-specific fungal propagules were identified in air samples. Details of the relevant residential characteristics were documented using a questionnaire. Independent predictors (P < 0.05) of ergosterol and total fungal propagules were identified by multiple linear regression. RESULTS: Fifty-five percent of the houses had viable fungal propagules exceeding 500 CFU/m3. Cladosporium and Penicillium were identified as the most prevalent and abundant fungal genera in indoor air. The median ergosterol level in bedroom floor was 3.8 microg/g of dust. Multivariate analysis showed that total fungal propagules in indoor air were lower in bedrooms with a ceiling fan, without visible mould, and those that were more frequently vacuumed, had a solid fuel fire, had windows closed at the time of the sampling or lacked pets. The presence of more than one cat had the greatest effect on total fungal propagules. Ergosterol levels were significantly lower in homes without old fitted carpets, visible mould or pets and those with frequent airing and regular use of an extractor fan in the kitchen. Old wall-to-wall carpets had the greatest effect on ergosterol. CONCLUSIONS: High indoor fungal exposures were associated with infrequent ventilation or vacuuming, presence of pets, visible mould and old carpets.

Adult↗
PubMed2003

Inhibition of human pathogenic fungi by ethnobotanically selected plant extracts.

In this study, 36 extracts derived from 29 plant species selected using an ethnobotanical approach were tested for antifungal activity against a taxonomically diverse group of 13 human pathogenic fungi. We compared the inhibitory characteristics of these plant extracts with those of the commonly used antifungals, amphotericin B and ketoconazole, and the plant-derived antifungal, berberine. Several plant extracts, notably those from Zingiber officinale (ginger) and Juglans cinerea (butternut), had pronounced antifungal activity against a wide variety of fungi, including strains that were highly resistant to amphotericin B and ketoconazole. Further exploration of Z. officinale as an antifungal is warranted as this species is generally regarded as safe for human consumption.

Amphotericin B↗
PubMed2003

The maintenance and preservation of keratinophilic fungi and related dermatophytes.

A total of 239 fungi belonging to nine genera and 23 species were maintained on agar slopes in a refrigerator at +8 +/- 1 degrees C and in sterile distilled water, under liquid paraffin, dried on silica gel, in sterile soil at room temperature. A limited number has been stored by freeze-drying between 1985 and 1998. It is recommended that each isolate should be maintained by at least two of these methods. The choice of the method will depend to a large extent on the equipment and facilities available in the laboratory. It is reported that simple methods can be used for preservation of keratinophilic fungi and related dermatophytes.

Arthrodermataceae↗
PubMed2004

Identification of medically important fungi by the Pyrosequencing technology.

The Pyrosequencing technology was used for identification of different clinically relevant fungi. The tests were performed on amplicons derived from the 18S rRNA gene using polymerase chain reaction (PCR) universal primers for amplification. Sequencing was performed up to 40 bases in a variable region with a designed general sequencing primer and the Pyrosequence data were analyzed by BLAST sequence search in the GenBank database. DNA from a total of 21 fungal specimens consisting of nine strains of clinically relevant fungi and 12 clinical specimens from patients suffering from proven invasive fungal infections were PCR-amplified and analyzed by gel electrophoresis, PCR-enzyme-linked immunosorbent assay (ELISA) and the Pyrosequencing technology. All data obtained by the Pyrosequencing technology were in agreement with the results obtained by PCR-ELISA using species/genus-specific oligonucleotides and were as well in accordance with the culture results. The results demonstrate that the Pyrosequencing method is a reproducible and reliable technique for identification of fungal pathogens.

Aspergillus↗
PubMed2004

Infections of the central nervous system by melanized fungi: a review of cases presented between 1999 and 2004.

Several types of infections of the central nervous system by melanized fungi can be distinguished: (a) single-organ infection of the cerebrum, (b) extension into the cerebrum from adjacent cavities, (c) fungal presence in the cerebrospinal fluid, or (d) meningitis. The fungal order Chaetothyriales (containing Exophiala-like black yeasts and relatives) is particularly rich in fungi causing cerebral infections. Cases by the main agents, Cladophialophora bantiana, Exophiala dermatitidis, and Ramichloridium mackenziei, published during the last 5 years are reviewed. Most of these infections prove to be fatal. Resection of the lesions in combination with antimycotic therapy may reduce mortality.

Adolescent↗
PubMed2001

Mycotoxin production from fungi isolated from grapes.

AIMS: In order to assess the potential for producing mycotoxins, fungi were isolated from wine producing grapes. METHODS AND RESULTS: The isolates were identified and Penicillium expansum, the most well recognized mycotoxin producer, was analysed for mycotoxin production by TLC. Many of the strains produced patulin and/or citrinin, often depending on whether they were grown on a grape or yeast extract sucrose media. CONCLUSION: Citrinin was produced by all strains grown in the yeast extract sucrose medium, but only one strain (from 51) was able to produce this compound in grape juice medium. Patulin was produced in the yeast extract medium by 20 strains and in grape juice medium by 33 strains. SIGNIFICANCE AND IMPACT OF THE STUDY: The presence of mycotoxins in wine producing grapes is discussed. Grapes contamination with patulin seems not to contribute to wine contamination, and no ochratoxin producing fungi was identified.

Beverages↗
PubMed2002

Conidium production by insect pathogenic fungi on commercially available agars.

AIMS: Conidium production by three species of insect pathogenic fungi, Metarhizium anisopliae, Beauveria bassiana and Verticillium lecanii, was assessed on various depths and types of commercially available agars. METHODS: Conidium production was assessed after 14 d of growth on commercially available media as well as at three different agar depths. RESULTS: Metarhizium anisopliae and B. bassiana isolates showed greatest conidium production on potato dextrose agar (PDA) at a depth of 2 mm, whereas V. lecanii showed greatest conidium production on yeast extract-peptone-dextrose agar (YPDA) regardless of agar depth. Optimum conidium production for M. anisopliae and B. bassiana was not only dependent upon the isolate used but also on the medium type and agar depth. SIGNIFICANCE AND IMPACT OF THE STUDY: Conidia are the infective structures for insect pathogenic fungi and this study suggests a rationale basis for consistent conidium production for laboratory and commercial practices.

Agar↗
PubMed1992

Distribution of radioactivity of 14C-amino acids added to the medium in cells and metabolites in cultures of rumen fungi.

A mixture of L-(U-14C) amino acids was added to cultures of 11 strains of rumen anaerobic fungi belonging to Neocallimastix frontalis, Neocallimastix joyonii, Sphaeromonas communis and Piromonas communis. Fungi were grown in a complex medium with glucose for 4 days. The radioactivity was found in cellular protein (27.7-65.3% of the total radioactivity recovered), lactate (16.9-41.8%), volatile fatty acids (7.4-25.7%) and ethanol (4.6-10.5%). A small amount of radioactivity was recovered in lipids (0.2-1.8%) and CO2 (0.3-1.0%). The results suggest that the assimilation of amino acids by growing fungal cells was quantitatively comparable with their dissimilation to metabolites.

Amino Acids↗
PubMed1995

Effect of anaerobic fungi on glycoside hydrolase and polysaccharide depolymerase activities, in sacco straw degradation and volatile fatty acid concentrations in the rumen of gnotobiotically reared lambs.

Four naturally born lambs were placed in sterile isolators 24 h after birth, before the natural establishment of the cellulolytic microorganisms. At the age of 4 weeks, a cellulolytic bacterial population of approximately 10(8) cells g-1 of rumen contents was established by inoculation with a 10(-6) dilution of ruminal contents taken from an adult sheep. A pure culture of Neocallimastix frontalis MCH3 and Piromyces communis FL was inoculated into the rumen 5 months after birth and a stable population of 10(3)-10(4) zoospores g-1 developed; the cellulolytic bacteria and fungi established populations in the 4 lambs that were similar to those observed in conventional animals. The presence of fungi led to an increase in the activity of most of the glycoside and polysaccharide hydrolases of the particle-associated microbial populations. However, this effect was not accompanied by an increase in the in sacco degradation of wheat straw or an increase in the volatile fatty acid concentration in the rumen contents.

Anaerobiosis↗