Metabolism of chiral ionylideneacetic acids on the abscisic acid biosynthetic pathway in Cercospora.
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A Chiralcel OJ column was used to determine the absolute configuration of naturally occurring alpha-ionylideneacetic acid from Cercospora rosicola and gamma-ionylideneacetic acid from C. cruenta as (R) enantiomers in accordance with their biosynthetic product, (S)-ABA. Both enantiomers of [1, 2-(13)C2]-alpha and gamma-ionylideneacetic acids were prepared and fed to C. rosicola and C. cruenta. Six combinations of feeding experiments comparatively and unequivocally demonstrated stereoselectivity in the biosynthetic conversions, including stepwise hydroxylation at C-1' and 4'. Enzymatic isomerization from the gamma to alpha-intermediate was suggested not to be involved in ABA biosynthesis in C. rosicola. (+info)
Ergosterol biosynthesis in novel melanized fungi from hypersaline environments.
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Halotolerant and halophilic melanized fungi were recently described in hypersaline waters. A close study of the sterol composition of such fungi, namely Hortaea werneckii, Alternaria alternata, Cladosporium sphaerospermum, Cladosporium sp., and Aureobasidium pullulans revealed the dominance of ergosterol and the presence of 29 intermediates of its biosynthesis pathway. The presence or absence of intermediates from distinct synthesis routes gave insight into the operative synthetic pathways from 4,4,14-trimethylcholesta-8,24-dien-3 beta-ol (lanosterol) to ergosterol in melanized fungi and in Saccharomyces cerevisiae, a reference yeast cultured in parallel. In all studied melanized fungi, initial methylation at C-24 took place before C-14 and C-4 demethylation, involving a different reaction sequence from that observed in S. cerevisiae. Further transformation was observed to occur through various routes. In A. alternata, isomerization at C-7 takes place prior to desaturation at C-5 and C-22, and methylene reduction at C-24. In addition to these pathways in Cladosporium spp., H. werneckii, and A. pullulans, ergosterol may also be synthesized through reduction of the C-24 methylene group before desaturation at C-5 and C-22 or vice versa. Moreover, in all studied melanized fungi except A. alternata, ergosterol biosynthesis may also proceed through C-24 methylene reduction prior to C-4 demethylation. -- Mejanelle, L., J. F. Lopez, N. Gunde-Cimerman, and J. O. Grimalt. Ergosterol biosynthesis in novel melanized fungi from hypersaline environments. J. Lipid Res. 2001. 42: 352--358. (+info)
Isolation, and biological properties of a new cell cycle inhibitor, curvularol, isolated from Curvularia sp. RK97-F166.
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A new cell growth inhibitor, curvularol, was isolated from the fermentation broth of Curvularia sp. RK97-F166. Curvularol showed no antibacterial activity, and very weak antifungal activity. However, curvularol inhibited the cell cycle progression of normal rat kidney (NRK) cells in G1 phase at 150 ng/ml. Curvularol induced the morphological reversion of srcts-transformed NRK cells at 100 ng/ml, and inhibited protein synthesis same as cycloheximide. (+info)
New cyclic depsipeptide antibiotics, clavariopsins A and B, produced by an aquatic hyphomycetes, Clavariopsis aquatica. 1. Taxonomy, fermentation, isolation, and biological properties.
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Clavariopsins were isolated from the fermentation broth of Clavariopsis aquatica AJ 117363. Clavariopsins are cyclic depsipeptide antibiotics with the molecular weight of 1,153 and 1,139. Clavariopsins showed in vitro antifungal activity against not only Aspergillus fumigatus but also, although to a lesser extent, A. niger and Candida albicans. (+info)
New cyclic depsipeptide antibiotics, clavariopsins A and B, produced by an aquatic hyphomycetes, Clavariopsis aquatica. 2. Structure analysis.
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The structures of new cyclic decadepsipeptides, clavariopsins A and B, were determined to be cyclo[-(R)-2-hydroxyisovaleryl-L-pipecoyl-L-MeVal-L-Val-L-MeAsp-L-MeIle-L-MeIle-G ly L-MeVal-L-Tyr(OMe)-] and cyclo[-(R)-2-hydroxyisovaleryl-L-pipecolyl-L-Val-L-Val-L-MeAsp-L-Melle-L-MeIle-Gl y-L-MeVal-L-Tyr(OMe)-], respectively, by spectroscopic analyses, especially using 2D NMR techniques. The absolute stereochemistry was elucidated by the advanced Marfey's method and chiral HPLC analysis. (+info)
Inoculum standardization for antifungal susceptibility testing of filamentous fungi pathogenic for humans.
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Two methods of inoculum preparation for filamentous fungi were compared: counting with a hematocytometer and spectrophotometric adjustment. One hundred eighty-two filamentous fungi pathogenic for humans were used. Colony counts were done for all inoculum preparations. The agreement between the hematocytometer counts and the colony counts (CFU per milliliter) was 97.2%. The reproducibility between the hematocytometer counts and the colony counts by means of an intraclass correlation coefficient was 0.70. Pearson's correlation index for hematocytometer counts versus colony counts was 0.56, whereas that for optical density versus colony counts was 0.008. Both methods can be used for inoculum size adjustment. However, the use of the spectrophotometric method requires that each species be standardized separately. (+info)
Effect of culture conditions on ergosterol as an indicator of biomass in the aquatic hyphomycetes.
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Ergosterol is a membrane component specific to fungi that can be used to estimate fungal biomass using appropriate factors of conversion. Our objectives were to determine the limits of use of ergosterol content as a measure of biomass for aquatic hyphomycetes, and to evaluate a previously established ergosterol-to-biomass conversion factor. We varied inoculum quality, growth medium, and degree of shaking of four aquatic hyphomycete species. In cultures inoculated with homogenized mycelium, we found a significant effect of shaking condition and culture age on ergosterol content. In liquid cultures with defined medium, ergosterol content reached 10 to 11 microg/mg of mycelium (dry mass) and varied by factors of 2.2 during exponential growth and 1.3 during stationary phase. The increase in ergosterol content during exponential phase could be attributed, at least in part, to rapid depletion of glucose. Oxygen availability to internal hyphae within the mycelial mass is also responsible for the differences found between culture conditions. Ergosterol concentration ranged from 0.8 to 1.6 microg/mg in static cultures inoculated with agar plugs. Ergosterol content varied by a factor of 4 in two media of different richnesses. For different combinations of these parameters, strong (r(2) = 0.83 to 0.98) and highly significant (P << 0.001) linear relationships between ergosterol and mycelial dry mass (up to 110 mg) were observed. Overall, the ergosterol content varied by a factor of 14 (0.8 to 11 mg/g). These results suggest that care must be taken when the ergosterol content is used to compare data generated in different field environments. (+info)
Utility of inoculum counting (Walshe and English criteria) in clinical diagnosis of onychomycosis caused by nondermatophytic filamentous fungi.
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Opportunistic onychomycosis caused by nondermatophytic molds may differ in treatment from tinea unguium. Confirmed diagnosis of opportunistic onychomycosis classically requires more than one laboratory analysis to show consistency of fungal outgrowth. Walshe and English in 1966 proposed to extract sufficient diagnostic information from a single patient consultation by counting the number of nail fragments positive for inoculum of the suspected fungus. Twenty fragments were plated per patient, and each case in which five or more fragments grew the same mold was considered an infection by that mold, provided that compatible filaments were also seen invading the nail tissue by direct microscopy. This widely used and often recommended method has never been validated. Therefore, the validity of substituting any technique based on inoculum counting for conventional follow-up study in the diagnosis of opportunistic onychomycosis was investigated. Sampling of 473 patients was performed repeatedly. Nail specimens were examined by direct microscopy, and 15 pieces were plated on standard growth media. After 3 weeks, outgrowing dermatophytes were recorded, and pieces growing any nondermatophyte mold were counted. Patients returned on two to eight additional occasions over a 1- to 3-year period for similar examinations. Onychomycosis was etiologically classified based on long-term study. Opportunistic onychomycosis was definitively established for 86 patients. Counts of nondermatophyte molds in initial examinations were analyzed to determine if they successfully predicted both true cases of opportunistic onychomycosis and cases of insignificant mold contamination. There was a strong positive statistical association between mold colony counts and true opportunistic onychomycosis. Logistic regression analysis, however, determined that even the highest counts predicted true cases of opportunistic onychomycosis only 89.7% of the time. The counting criterion suggested by Walshe and English was correct only 23.2% of the time. Acremonium infections were especially likely to be correctly predicted by inoculum counting. Inoculum counting could be used to indicate a need for repeat studies in cases of false-negative results from laboratory direct microscopy. Inoculum counting cannot serve as a valid substitute for follow-up study in the diagnosis of opportunistic onychomycosis. It may, nonetheless, provide useful information both to the physician and to the laboratory, and it may be especially valuable when the patient does not present for follow-up sampling. (+info)