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  • 1
    ISSN: 1432-0983
    Keywords: Phialophora ; Chitin synthase ; Multiple genes ; Chromoblastomycosis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Based on conserved amino-acid regions predicted for the chitin synthases (Chs) of Saccharomyces cerevisiae, two different primer sets were synthesized and used in polymerase chain reactions (PCRs) to amplify 614-bp and 366-bp sequences from genomic DNA of the zoopathogenic fungus Phialophora verrucosa. DNA-sequencing and Southern-blotting analyses of the 614-bp DNA amplification products suggested that portions of two distinct P. verrucosa chitin synthase genes (PvCHS1, PvCHS2), coding for two different zymogenic-type PvChs isozymes, had been identified. The deduced amino-acid sequence of each fell into different Chs classes, namely class I and class II. In addition, the 366-bp DNA segment was shown to code for a conserved region having homology with the CSD2/CAL1 gene of S. cerevisiae, which encodes a nonzymogenic-type enzyme, Chs3, in that fungus. The amino-acid sequence derived from PvCHS3 exhibits 88.2% similarity and 78.4% identity to the same amino-acid region of the S. cerevisiae enzyme. These results provide a critical first step toward investigating the molecular and pathogenic importance of CHS gene regulation in this fungus and for exploring steps leading to Chs function as potential targets for the design of new therapeutic agents.
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  • 2
    ISSN: 1432-0983
    Keywords: Key wordsWangiella ; Exophiala ; Transformation ; Gene expression ; Polyketide synthase
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract To explore potential virulence factors in the dematiaceous (melanized) fungus Wangiella dermatitidis, we established a gene expression system with properties of homologous transformation and color identification. Using a polyketide synthase gene (WdPKS1) fragment for targeting, we found that 52% of transformants became albinos easily distinguishable from nonspecific transformants. Southern analysis confirmed that the integrations were at the WdPKS1 locus, which however did not affect transformant growth. With a heterologous promoter, P-glaA, enhanced expression of lacZ was found at 37 °C. Our results indicated that this system allows the efficient production of isogenic strains for gene function analysis in W. dermatitidis.
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  • 3
    ISSN: 1432-0789
    Keywords: Iron ; Phosphorus ; Siderophore ; Hydroxamate ; Chelates
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Geosciences , Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Summary Hydroxamate siderophores (HS) are microbially produced, ferric-specific chelates, known to occur in soil, and to be capable of providing iron to higher plants. This study examined the potential for HS to influence the diffusion of both iron and phosphorus to plant roots in soil. The HS desferrioxamine-B (DFOB) and desferriferrichrome (ferrichrome) were compared with the synthetic chelates ethylenediamine [di(o-hydroxyphenylacetic)acid] (EDDHA) and ethylenediamine-tetraacetic acid (EDTA), and citrate, oxalate, and distilled water in their ability to increase diffusion of iron using a simulated root technique. Chelate solutions were pumped through porous fiber bundles imbedded in soil previously labeled with55Fe. In a sandy loam of pH 7.5,55Fe diffusion caused by 10−4 M DFOB was twice that of water, but similar to that caused by 10−4 M EDDHA. However, 10−3 M EDDHA resulted in greater diffusion than 10-3 M DFOB. The diffusions resulting from equimolar quantities of citrate, oxalate, and EDTA were similar to that with distilled water. In a clay soil of pH 5.2 previously labeled with55Fe and32 P, the response in55Fe diffusion to chelate treatments was: 10−4 M EDDHA 〉 10−4 M ferrichrome 〉 10−3 M DFOB 〉 10−4 M DFOB 〉 water. Both ferrichrome and EDDHA caused2 P diffusion to increase substantially over that of distilled water. These results suggest that hydroxamate siderophores present in the rhizosphere could effectively increase the level of soluble iron for root uptake and possibly increase phosphorus uptake by solubilization of phosphorus from iron phosphates at acid pH.
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  • 4
    ISSN: 1432-0614
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Genetic transformation of Wangiella dermatitidis was studied using three plasmid vectors (pAN7-1, pWU44, and pKK5) and both electroporation and polyethyleneglycol-mediated methods. pAN7-1 contains the E. coli hygromycin B (HmB) phosphotransferase (hph) gene. Expression of the hph gene confers resistance to antibiotic HmB. Selection for resistance, indicative of transformation, resulted in 10–203 HmB-resistant colonies/μg pAN7-1 on medium containing 100 μg HmB/ml. Strains of W. dermatitidis used in this study have innate sensitivity to HmB at a critical inhibitory concentration of 20–40 μg/ml. Vectors pWU44 and pKK5 contain a URA5 gene from Podospora anserina. A ura5 auxotroph of W. dermatitidis was transformed to prototrophy with pWU44 or pKK5 by complementation. Transformation frequencies for these two plasmids were between 17–50 transformants/μg vector DNA. Southern blotting analysis and polymerase chain reaction detection of DNA from putative transformants confirmed transformation.
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  • 5
    ISSN: 1432-0614
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract  Genetic transformation of Wangiella dermatitidis was studied using three plasmid vectors (pAN7-1, pWU44, and pKK5) and both electroporation and polyethyleneglycol-mediated methods. pAN7-1 contains the E. coli hygromycin B (HmB) phosphotransferase (hph) gene. Expression of the hph gene confers resistance to antibiotic HmB. Selection for resistance, indicative of transformation, resulted in 10–203 HmBresistant colonies/μg pAN7-1 on medium containing 100 μg HmB/ml. Strains of W. dermatitidis used in this study have innate sensitivity to HmB at a critical inhibitory concentration of 20–40 μg/ml. Vectors pWU44 and pKK5 contain a URA5 gene from Podospora anserina. A ura5 auxotroph of W. dermatitidis was transformed to prototrophy with pWU44 or pKK5 by complementation. Transformation frequencies for these two plasmids were between 17–50 transformants/μg vector DNA. Southern blotting analysis and polymerase chain reaction detection of DNA from putative transformants confirmed transformation.
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  • 6
    Electronic Resource
    Electronic Resource
    [s.l.] : Nature Publishing Group
    Nature 287 (1980), S. 833-834 
    ISSN: 1476-4687
    Source: Nature Archives 1869 - 2009
    Topics: Biology , Chemistry and Pharmacology , Medicine , Natural Sciences in General , Physics
    Notes: [Auszug] Soil samples (0-10 cm deep) were collected from 57 sites representing numerous biomes and soil types. After extraction (1:1 H2O: soil, w/w) at 4 C for 1 h, samples were centrifuged, the supernatants dried (90 C), and the residues rehydrated, filter sterilized, and bioassayed using the HS ...
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  • 7
    ISSN: 1365-3040
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: To eliminate the confounding effects of microorganisms and to examine the direct utilization of microbial siderophores as iron sources by higher plants, a hydroponic cultural system and methodology was developed to grow plants with axenic roots. This report presents a description of this system, and also its use to determine the efficacy of the microbial siderophore ferrioxamine B (FOB), compared to the synthetic iron chelate FeEDTA, and the phytosiderophores (PS) of barley as an iron source for alleviating iron stress in the model dicot cucumber. It was observed that FOB gave superior plant biomass and was preferentially utilized to restore chlorophyll synthesis in long-term experiments when chelates were supplied at 5mmol m−3 concentrations and nutrient solution was buffered against pH change at 7.4 with solid phase CaCO3. In addition, autoradiograms indicated that 59Fe from FOB was rapidly translocated to shoots through vascular tissues and was specifically distributed to regions of rapid growth and to iron-stressed, but still expanding young leaves. The siderophore itself could be detected within 2h in xylem exudates, regardless of whether or not plants were exposed to metabolic inhibitors. It was concluded that the FOB and iron were taken up by the axenic roots of cucumber in a highly efficient manner, most likely as the iron-siderophore complex, and at rates that could be significant to dicot nutrition. The results also suggested that cucumber may transport FOB through the transpiration stream to upper parts of plants, where the iron would be reductively released from the siderophore for shoot nutrition.
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  • 8
    ISSN: 1573-5036
    Keywords: chelate ; ecology ; membrane transport ; plant iron nutrition ; plant-microbial interactions ; rhizosphere
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract Most bacteria, fungi, and some plants respond to Fe stress by the induction of high-affinity Fe transport systems that utilize biosyrthetic chelates called siderophores. To competitively acquire Fe, some microbes have transport systems that enable them to use other siderophore types in addition to their own. Bacteria such as Escherichia coli achieve this ability by using a combination of separate siderophore receptors and transporters, whereas other microbial species, such as Streptomyces pilosus, use a low specificity, high-affinity transport system that recognizes more than one siderophore type. By either strategy, such versatility may provide an advantage under Fe-limiting conditions; allowing use of siderophores produced at another organism's expense, or Fe acquisition from siderophores that could otherwise sequester Fe in an unavailable form. Plants that use microbial siderophores may also be more Fe efficient by virtue of their ability to use a variety of Fe sources under different soil conditions. Results of our research examining Fe transport by oat indicate parity in plant and microbial requirements for Fe and suggest that siderophores produced by root-colonizing microbes may provide Fe to plants that can use the predominant siderophore types. In conjunction with transport mechanisms, ecological and soil chemical factors can influence the efficacy of siderophores and phytosiderophores. A model presented here attempts to incorporate these factors to predict conditions that may govern competition for Fe in the plant rhizosphere. Possibly such competition has been a factor in the evolution of broad transport capabilities for different siderophores by microorganisms and plants.
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  • 9
    Publication Date: 2014-04-17
    Description: Black or dark brown (phaeoid) fungi cause cutaneous, subcutaneous, and systemic infections in humans. Black fungi thrive in stressful conditions such as intense light, high radiation, and very low pH. Wangiella ( Exophiala ) dermatitidis is arguably the most studied phaeoid fungal pathogen of humans. Here, we report our comparative analysis of the genome of W. dermatitidis and the transcriptional response to low pH stress. This revealed that W. dermatitidis has lost the ability to synthesize alpha-glucan, a cell wall compound many pathogenic fungi use to evade the host immune system. In contrast, W. dermatitidis contains a similar profile of chitin synthase genes as related fungi and strongly induces genes involved in cell wall synthesis in response to pH stress. The large portfolio of transporters may provide W. dermatitidis with an enhanced ability to remove harmful products as well as to survive on diverse nutrient sources. The genome encodes three independent pathways for producing melanin, an ability linked to pathogenesis; these are active during pH stress, potentially to produce a barrier to accumulated oxidative damage that might occur under stress conditions. In addition, a full set of fungal light-sensing genes is present, including as part of a carotenoid biosynthesis gene cluster. Finally, we identify a two-gene cluster involved in nucleotide sugar metabolism conserved with a subset of fungi and characterize a horizontal transfer event of this cluster between fungi and algal viruses. This work reveals how W. dermatitidis has adapted to stress and survives in diverse environments, including during human infections.
    Electronic ISSN: 2160-1836
    Topics: Biology
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