Structural basis for activation of fungal sterol receptor Upc2 and azole resistance.

Fungal transcription factor Upc2 senses ergosterol levels and regulates sterol biosynthesis and uptake. Constitutive activation of Upc2 causes azole resistance in Candida species. We determined the structure of ergosterol-bound Upc2, revealing the ligand specificity and transcriptional regulation. Ergosterol binding involves conformational changes of the ligand-binding domain, creating a shape-complementary hydrophobic pocket. The conserved helix α12 and glycine-rich loop are critical for sterol recognition by forming the pocket wall. The mutations of the glycine-rich loop inhibit ligand binding by steric clashes and constitutively activate Upc2. The translocation of Upc2 is regulated by Hsp90 chaperone in a sterol-dependent manner. Ergosterol-bound Upc2 associates with Hsp90 using the C-terminal tail, which retains the inactive Upc2 in the cytosol. Ergosterol dissociation induces a conformational change of the C-terminal tail, releasing Upc2 from Hsp90 for nuclear transport by importin α. The understanding of the regulatory mechanism provides an antifungal target for the treatment of azole-resistant Candida infections.

Investigation of the Defective Growth Pattern and Multidrug Resistance in a Clinical Isolate of Candida glabrata Using Whole-Genome Sequencing and Computational Biology Applications.

Candida glabrata is increasingly isolated from blood cultures, and multidrug-resistant isolates have important implications for therapy. This study describes a cholesterol-dependent clinical C. glabrata isolate (ML72254) that did not grow without blood (containing cholesterol) on routine mycological media and that showed azole and amphotericin B (AmB) resistance. Matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) and whole-genome sequencing (WGS) were used for species identification. A modified Etest method (Mueller-Hinton agar supplemented with 5% sheep blood) was used for antifungal susceptibility testing. WGS data were processed via the Galaxy platform, and the genomic variations of ML72254 were retrieved. A computational biology workflow utilizing web-based applications (PROVEAN, AlphaFold Colab, and Missense3D) was constructed to predict possible deleterious effects of these missense variations on protein functions. The predictive ability of this workflow was tested with previously reported missense variations in ergosterol synthesis genes of C. glabrata. ML72254 was identified as C. glabrata sensu stricto with MALDI-TOF, and WGS confirmed this identification. The MICs of fluconazole, voriconazole, and amphotericin B were >256, >32, and >32 µg/mL, respectively. A novel frameshift mutation in the ERG1 gene (Pro314fs) and many missense variations were detected in the ergosterol synthesis genes. None of the missense variations in the ML72254 ergosterol synthesis genes were deleterious, and the Pro314fs mutation was identified as the causative molecular change for a cholesterol-dependent and multidrug-resistant phenotype. This study verified that web-based computational biology solutions can be powerful tools for examining the possible impacts of missense mutations in C. glabrata. IMPORTANCE In this study, a cholesterol-dependent C. glabrata clinical isolate that confers azole and AmB resistance was investigated using artificial intelligence (AI) technologies and cloud computing applications. This is the first of the known cholesterol-dependent C. glabrata isolate to be found in Turkey. Cholesterol-dependent C. glabrata isolates are rarely isolated in clinical samples; they can easily be overlooked during routine laboratory procedures. Microbiologists therefore need to be alert when discrepancies occur between microscopic examination and growth on routine media. In addition, because these isolates confer antifungal resistance, patient management requires extra care.

Improved osmotic stress tolerance in brewer's yeast induced by wheat gluten peptides.

The influences of three wheat gluten peptides (WGP-LL, WGP-LML, and WGP-LLL) on the osmotic stress tolerance and membrane lipid component in brewer's yeast were investigated. The results demonstrated that the growth and survival of yeast under osmotic stress were enhanced by WGP supplementation. The addition of WGP upregulated the expressions of OLE1 (encoded the delta-9 fatty acid desaturase) and ERG1 (encoded squalene epoxidase) genes under osmotic stress. At the same time, WGP addition enhanced palmitoleic acid (C16:1) content, unsaturated fatty acids/saturated fatty acids ratio, and the amount of ergosterol in yeast cells under osmotic stress. Furthermore, yeast cells in WGP-LL and WGP-LLL groups were more resistant to osmotic stress. WGP-LL and WGP-LLL addition caused 25.08% and 27.02% increase in membrane fluidity, 22.36% and 29.54% reduction in membrane permeability, 18.38% and 14.26% rise in membrane integrity in yeast cells, respectively. In addition, scanning electron microscopy analysis revealed that the addition of WGP was capable of maintaining yeast cell morphology and reducing cell membrane damage under osmotic stress. Thus, alteration of membrane lipid component by WGP was an effective approach for increasing the growth and survival of yeast cells under osmotic stress. KEY POINTS: •WGP addition enhanced cell growth and survival of yeast under osmotic stress. •WGP addition increased unsaturated fatty acids and ergosterol contents in yeast. •WGP supplementation improved membrane homeostasis in yeast at osmotic stress.

Physiological and transcriptional profiling of surfactin exerted antifungal effect against Candida albicans.

Given the risk of Candida albicans overgrowth in the gut, novel complementary therapies should be developed to reduce fungal dominancy. This study highlights the antifungal characteristics of a Bacillus subtilis-derived secondary metabolite, surfactin with high potential against C. albicans. Surfactin inhibited the growth of C. albicans following a 1-hour exposure, in addition to reduced adhesion and morphogenesis. Specifically, surfactin did not affect the level of reactive oxygen species but increased the level of reduced glutathione. Surprisingly, ethanol production was increased following 2 h of surfactin exposure. Surfactin treatment caused a significant reduction in intracellular iron, manganese and zinc content compared to control cells, whereas the level of copper was not affected. Alongside these physiological properties, surfactin also enhanced fluconazole efficacy. To gain detailed insights into the surfactin-related effects on C. albicans, genome-wide gene transcription analysis was performed. Surfactin treatment resulted in 1390 differentially expressed genes according to total transcriptome sequencing (RNA-Seq). Of these, 773 and 617 genes with at least a 1.5-fold increase or decrease in transcription, respectively, were selected for detailed investigation. Several genes involved in morphogenesis or related to metabolism (e.g., glycolysis, ethanol and fatty acid biosynthesis) were down-regulated. Moreover, surfactin decreased the expression of ERG1, ERG3, ERG9, ERG10 and ERG11 involved in ergosterol synthesis, whereas genes associated with ribosome biogenesis and iron metabolism and drug transport-related genes were up-regulated. Our data demonstrate that surfactin significantly influences the physiology and gene transcription of C. albicans, and could contribute to the development of a novel innovative complementary therapy.

Improving the inhibition of ß-amyloid aggregation by withanolide and withanoside derivatives.

Here, we have studied the ameliorative effects of Withania somnifera derivatives (Withanolide A, Withanolide B, Withanoside IV, and Withanoside V) on the fibril formation of amyloid-ß 42 for Alzheimer's disease. We analyzed reduction in the aggregation of ß amyloid protein with these Ashwagandha derivatives by Thioflavin T assay in the oligomeric and fibrillar state. We have tested the cytotoxic activity of these compounds against human SK-N-SH cell line for 48 h, and the IC 50 value found to be 28.61 ± 2.91, 14.84 ± 1.45, 18.76 ± 0.76 and 30.14 ± 2.59 µM, respectively. After the treatment of the cells with half the concentration of IC 50 value, there was a remarkable decrease in the number of apoptotic cells stained by TUNEL assay indicating the DNA damage and also observed significant decrease of reactive oxygen species. Also, the binding and molecular stability of these derivatives with amyloid ß was also studied using bioinformatics tools where these molecules were interacted at LVFFA region which is inhibition site of amyloid-ß1 42. These studies revealed that the Withanolides and Withanosides interact with the hydrophobic core of amyloid-ß 1-42 in the oligomeric stage, preventing further interaction with the monomers and diminishing aggregation.

Do the Main Components of the Sclerotia of Umbrella Polypore Mushroom, Polyporus umbellatus (Agaricomycetes), Correlate with Armillaria Associates?

Polyporus umbellatus is a traditional Chinese medicinal mushroom. The growth of P. umbellatus sclerotia requires the rhizomorphs of Armillaria spp. to supply nutrition. Whether the main components (MC) of sclerotia of P. umbellatus are related to the phylogeny of Armillaria associates or other environmental factors is largely unknown. In this study, we collected 17 sclerotia and soil samples from northeast to southwest China. In total, 17 Armillaria associates were isolated, and sclerotial MC contents and soil characteristics (total N, P, K, and organic matter) were determined. The analysis revealed that the MC content of P. umbellatus did not resemble a Brownian motion process in phylogeny of Armillaria associates, but were significantly influenced by the total N content of the soil. These results provide clear evidence that sclerotia of P. umbellatus associating with phylogenetic related Armillaria associates possess differing MC content. The mechanisms of nutrient exchange in P. umbellatus-Armillaria associations now require further elucidation.

Biosorption of sterols from tobacco waste extract using living and dead of newly isolated fungus Aspergillus fumigatus strain LSD-1.

Sterols are verified to be able to produce polycyclic aromatic hydrocarbons during its pyrolysis. In this study, a kind of Aspergillus fumigatus (LSD-1) was isolated from cigar leaves, and the biosorption effects on the stigmasterol, ß-sitosterol, campesterol, cholesterol, and ergosterol by using living and dead biomass of LSD-1 were investigated. The results showed that both living and dead biomass could efficiently remove these sterols in aqueous solution and tobacco waste extract (TWE). Interestingly, compared with the living biomass of LSD-1, the dead biomass of LSD-1 not only kept a high adsorption efficiency but also did not produce ergosterol. Overall, dead biomass of LSD-1 was a more suitable biosorbent to sterols in TWE. Furthermore, Brunner-Emmet-Teller (BET), Fourier transformed infrared spectrometer (FTIR) and scanning electron microscope (SEM) analysis were used to explore the biosorption process of living and dead biomass and their differences, suggesting that the biosorption of sterols was a physical process.

Assessment of chemically characterised Myristica fragrans essential oil against fungi contaminating stored scented rice and its mode of action as novel aflatoxin inhibitor.

The study reports chemically characterised Myristica fragrans essential oil (MFEO) as plant based food preservative against fungal and aflatoxin B1 (AFB1) contamination of scented rice varieties. The chemical profile of MFEO revealed elemicin (27.08%), myristicine (21.29%) and thujanol (18.55%) as major components. The minimum inhibitory and minimum aflatoxin inhibitory concentrations of MFEO were 2.75 and 1.5 mg/ml, respectively. The MFEO was efficacious against a broad spectrum of food deteriorating fungi. MFEO caused decrease in ergosterol content of fungal plasma membrane and enhanced leakage of cellular ions, depicting plasma membrane as the site of action. The MFEO caused reduction in cellular methylglyoxal content, the aflatoxin inducer. This is the first report on MFEO as aflatoxin suppressor. The essential oil may be recommended as plant based food preservative after large scale trials and reduction in methylglyoxal suggests its application for development of aflatoxin resistant varieties through green transgenics.

Effects of UV-C treatment and ultrafine-grinding on the biotransformation of ergosterol to vitamin D2, physiochemical properties, and antioxidant properties of shiitake and Jew's ear.

In the present study, the impact of ultraviolet (UV)-C treatment and ultrafine grinding on the conversion of ergosterol to vitamin D2, physiochemical properties, and antioxidant properties of shiitake and Jew's ear was assessed. After exposure to UV-C, vitamin D2 contents of both the mushroom samples has increased significantly (p < 0.05). Whereas, ultrafine grinding along with UV-C treatment has a synergistic effect on bioconversion of ergosterol to vitamin D2 and this effect is more prominent in low dose UV-C irradiation groups (2 kJ/m2). Ultrafine grinding significantly (p < 0.05) improved the water holding capacity (WHC), water solubility index (WSI) and polysaccharide dissolution rate (PDR). However, UV-C treatment led to insignificant changes in the physiochemical properties of mushroom samples. A significant improvement was also observed in the antioxidant profiles especially tannin contents of mushrooms followed by the ultrafine grinding and UV-C treatment.

The effect of isoflavaspidic acid PB extracted from Dryopteris fragrans (L.) Schott on planktonic and biofilm growth of dermatophytes and the possible mechanism of antibiofilm.

ETHNOPHARMACOLOGICAL RELEVANCE: Dryopteris fragrans (L.) Schott (D. fragrans), a deciduous perennial herb, has been traditionally used for treatment of various skin diseases in Heilongjiang province of China for many years. Phloroglucinol derivatives extracted from D. fragrans were the most effective fraction against dermatophytes. Isoflavaspidic acid PB is a typically phloroglucinol derivative which extracted from D. fragrans and has been reported to exert anti-fungal activities against several dermatophytes. AIM OF THE STUDY: This study aimed to evaluate anti-fungal and anti-biofilm activity of isoflavaspidic acid PB on planktonic and biofilm growth of dermatophytes and explore possible mechanisms of anti-biofilm. MATERIALS AND METHODS: Minimal inhibitory concentrations (MIC) and minimal fungicidal concentrations (MFC) of isoflavaspidic acid PB against 25 isolates of dermatophytes were determined by the Clinical and Laboratory Standards Institute (CLSI) M38-A2 method. The effects of isoflavaspidic acid PB on dermatophytes biofilm formation and pre-formed biofilm were assessed by 2.3-bis (2-methoxy-4-nitro-5-sulfophenyl)-5-[carbonyl (phenylamino)]-2H-tetrazolium hydroxide (XTT) assay. Morphology of mature biofilm were observed by Scanning Electron Microscope (SEM). Biomass, exopolysaccharide and ergosterol content of mature biofilm were analyzed by gravimetric analysis, anthranone sulfuric acid method and Ultra Performance Liquid Chromatography (UPLC) assay respectively. RESULT: The MIC and MFC ranges of isoflavaspidic acid PB against 25 isolates of dermatophytes were 20-80 µg/mL and 40-80 µg/mL respectively. Isoflavaspidic acid PB (2 MIC) inhibited not only Trichophyton biofilm formation (54.8% ∼ 81.2%) but also the metabolic activity of mature biofilm (20.7% ∼ 44.2%). The result of SEM showed that isoflavaspidic acid PB (8 MIC) could destroy the morphology of hyphae seriously. Comparing with control group, biomass, exopolysaccharide and ergosterol content of the mature biofilm under isoflavaspidic acid PB (8 MIC) were significantly decreased (P < 0.01). CONCLUSION: Isoflavaspidic acid PB had anti-fungal and fungicidal activities against dermatophytes. Isoflavaspidic acid PB could inhibit the biofilm of Trichophyton. The mechanism might be related to the decline of the biofilm biomass, exopolysaccharide and ergosterol content. These results showed that isoflavaspidic acid PB could be explored for promising anti-biofilm drugs.

The antifungal activity and mechanisms of action of quantified extracts from berries, leaves and roots of Phytolacca tetramera.

BACKGROUND: Phytolacca tetramera is an endemic plant from Argentina that is currently at serious risk because its environment is subjected to a high anthropic impact. A previous study has shown that berry extracts obtained from this plant display antifungal activity against multiple human-pathogenic fungi when tested with a non-standardized method. Further evidences of the antifungal properties of other parts of the plant and studies of mechanism of antifungal action of the antifungal chemically characterized extracts are required. PURPOSE: This study aimed to gain further evidence of the antifungal activity of P. tetramera berry, leaf and root extracts in order to find the most active extract to be developed as an Herbal Medicinal Antifungal Product. The medicinal usefulness of P. tetramera extracts as antifungal agents will serve as an important support to create concience and carry out actions tending to the preservation of this threatened species and its environment. MATERIALS AND METHODS: Chemical analysis of all P. tetramera extracts, including quantitation of selected markers, was performed through UHPLC-ESI-MS/MS and UPLC-ESI-MS techniques according to the European Medicines Agency (EMA). The antifungal activity of the quantified extracts was tested with the standardized CLSI microbroth dilution method against Candida spp. Antifungal mechanisms of the most active extract were studied by examination of morphological changes by phase-contrast and fluorescence microscopies and both, cellular and enzymatic assays targeting either the fungal membrane or the cell wall. RESULTS: The antifungal activity of twelve P. tetramera extracts was tested against Candida albicans and Candida glabrata. The dichloromethane extract from berries (PtDEb) showed the best activity. Phytolaccagenin (PhytG) and phytolaccoside B (PhytB) were selected as the main active markers for the antifungal P. tetramera extracts. The quantitation of these active markers in all extracts showed that PtDEb possessed the highest amount of PhytG and PhytB. Finally, studies on the mechanism of antifungal action showed that the most active PtDEb extract produces morphological changes compatible with a damage of the cell wall and/or the plasma membrane. Cellular and enzymatic assays showed that PtDEb would not damage the fungal cell wall by itself, but would alter the plasma membrane. In agreement, PtDEb was found to bind to ergosterol, the main sterol of the fungal plasma membrane. CONCLUSION: Studies of the anti-Candida activity of P. tetramera extracts led to the selection of PtDEb as the most suitable extract, confirming the antifungal properties of the threatened species P. tetramera. The new data give a valuable reason for the definitive protection of this sp. and its natural environment thus allowing further studies for the future development of an Herbal Medicinal Antifungal Product.

Two natural molecules preferentially inhibit azole-resistant Candida albicans with MDR1 hyperactivation.

Antifungal drug resistance is a significant clinical problem, and antifungal agents that can evade resistance are urgently needed. In infective niches, resistant organisms often co-existed with sensitive ones, or a subpopulation of antibiotic-susceptible organisms may evolve into resistant ones during antibiotic treatment and eventually dominate the whole population. In this study, we established a co-culture assay in which an azole-resistant Candida albicans strain was mixed with a susceptible strain labeled with green fluorescent protein to mimic in vivo conditions and screen for antifungal drugs. Fluconazole was used as a positive control to verify the validity of this co-culture assay. Five natural molecules exhibited antifungal activity against both susceptible and resistant C. albicans. Two of these compounds, retigeric acid B (RAB) and riccardin D (RD), preferentially inhibited C. albicans strains in which the efflux pump MDR1 was activated. This selectivity was attributed to greater intracellular accumulation of the drugs in the resistant strains. Changes in sterol and lipid compositions were observed in the resistant strains compared to the susceptible strain, and might increase cell permeability to RAB and RD. In addition, RAB and RD interfered with the sterol pathway, further aggregating the decrease in ergosterol in the sterol synthesis pathway in the MDR1-activated strains. Our findings here provide an alternative for combating resistant pathogenic fungi.

Synergistic effects of tea polyphenol epigallocatechin 3-O-gallate and azole drugs against oral Candida isolates.

BACKGROUND: The antifungal drug resistance has become an emerging problem in the management of candida infections worldwide. The objective of this study was to examine the efficacy of epigallocatechin 3-O-gallate (EGCG) alone and in combination with fluconazole/ketoconazole drugs against oral Candida isolates. METHODS: Minimum inhibitory concentration (MIC) and minimum fungicidal concentrations (MFC) of EGCG against 60 oral Candida isolates and 4 ATCC strains were determined. Synergism of EGCG with azole drugs was evaluated by checkerboard micro-dilution method and calculated fractional inhibitory concentration index (FICI). Candida cells' ultrastructure was studied by electron microscopy. RESULTS: MIC and MFC values of EGCG were in the range of 3.91-15.63 and 15.63-31.25µg/mL, respectively. Minimum biofilm inhibitory concentration (MBIC) range of EGCG (62.5-125µg/mL), was less than the ketoconazole (64-256µg/mL) and fluconazole (128-512µg/mL). The combination of EGCG with fluconazole/ketoconazole exhibited synergistic effects (ΣFICI≤0.50). EGCG with azole drugs showed high sensitivity against the tested isolates in growth curve assays. Against the biofilm, the susceptibility of fluconazole/ketoconazole significantly increased (3 to 5 fold), after combination with EGCG (MBIC/4) (P≤0.001). Electron microscopy of EGCG treated cells showed deformation of cell structure, ruptured cell wall and release of intracellular content. In molecular docking experiments, a strong interaction was observed between EGCG and fungal cell membrane molecule ergosterol. CONCLUSION: We conclude that EGCG synergistically enhanced the antifungal potential of azole drugs. The synergistic potential of EGCG might be helpful in preventing the development of drug resistance, in lowering the drug dosage, and thus minimizing adverse effects.

Lipidomic adaptations of the Metarhizium robertsii strain in response to the presence of butyltin compounds.

Metarhizium robertsii, a butyltin-resistant filamentous fungus, can rapid and complete biodegradation of di- (DBT) and tributyltin (TBT) under conditions of intensive aeration and ascorbic acid supplementation. In this paper, lipidomic investigations were performed to find the membrane adaptations necessary for effective butyltins degradation. HPLC-MS/MS analysis showed that the phospholipid profile was greatly modified during M. robertsii batch cultivation (pO2 ≥ 20%), contributing to increased membrane fluidity and facilitated mass transfer, which could enhance butyltins biodegradation. Intensified biosynthesis of phospholipids, sphingolipids and ergosterol by the mycelia exposed to butyltins was noted. DIOC6(3) fluorescence intensity for TBT-treated mycelium increased 9-fold pointing to membrane hyperpolarization. Fluorescent studies showed improved membrane rigidity and integrity in response to butyltins presence. Vitamin C supplementation restored membrane composition and dynamic properties, followed by supposed acceleration of transport of monobutyltin and its biodegradation thus protecting the M. robertsii cells against oxidative and nitrosative stress.

Elucidating the active interaction mechanism of phytochemicals withanolide and withanoside derivatives with human serum albumin.

Withania somnifera (Ashwagandha) is an efficient medicinal plant known in Ayurveda and Chinese medicine since ancient times, whose extracts are consumed orally as food supplement or as a health tonic owing to its several restorative properties for various CNS disorders, inflammation, tumour, stress, rheumatism etc. In this study, we have analyzed the binding interaction of four derivatives of Withania somnifera (Withanolide A, Withanolide B, Withanoside IV and Withanoside V) with HSA because of their important pharmacological properties. To unravel the binding between derivatives of Withania somnifera and HSA, fluorescence spectroscopy was used. Binding studies were further studied by molecular docking and dynamics and results confirmed greater stability upon binding of derivatives with HSA. Circular dichroism data illustrated change in the secondary structure of protein upon interaction with these derivatives, particularly the helical structure was increased and ß-sheets and random coils were decreased. Furthermore, morphological and topological changes were observed using AFM and TEM upon binding of ligands with HSA indicating that HSA-withnoside/withanolide complexes were formed. All the results cumulatively demonstrate strong binding of withanosides and withanolides derivatives with serum albumin, which should further be explored to study the pharmacokinetics and pharmacodynamics of these derivatives.

Microbial Sterolomics as a Chemical Biology Tool.

Metabolomics has become a powerful tool in chemical biology. Profiling the human sterolome has resulted in the discovery of noncanonical sterols, including oxysterols and meiosis-activating sterols. They are important to immune responses and development, and have been reviewed extensively. The triterpenoid metabolite fusidic acid has developed clinical relevance, and many steroidal metabolites from microbial sources possess varying bioactivities. Beyond the prospect of pharmacognostical agents, the profiling of minor metabolites can provide insight into an organism's biosynthesis and phylogeny, as well as inform drug discovery about infectious diseases. This review aims to highlight recent discoveries from detailed sterolomic profiling in microorganisms and their phylogenic and pharmacological implications.

Traditionally practiced medicinal plant extracts inhibit the ergosterol biosynthesis of clinically isolated dermatophytic pathogens.

OBJECTIVE: Dermatophytes are resistant to some available antibiotics. Development of new plant drugs to control drug resistant microbes is urgently needed. This study evaluates the antidermatophytic potential of 18 selected medicinal plants used by traditional healers in Theni and Virudhunagar Districts of Tamil Nadu, India. MATERIALS AND METHODS: Selected plant parts were collected, shade dried and powdered. Plant powders were extracted with ethanol and their antifungal potency was investigated against and clinical dermatophytes. The antioxidant effect of the extracts was screened using DPPH assay. Minimal Inhibitory Concentration (MIC) and Minimal Fungicidal Concentration (MFC) were estimated for the extracts. Ten plant extracts showed maximum MFC and they were selected to study their efficacy in interfering with ergosterol biosynthesis. Fluconazole-35µg/mL known fungicide was used as control. The most active extracts were taken for biocompatibility studies using 3T3-L1 fibroblast cell lines. RESULTS: The ethanol extract of Phyllanthus reticulates leaves showed good antifungal activity compared to other plant extracts. The MIC and MFC for Phyllanthus reticulatus were 62.5 and 250µg/mL against M. pachydermatitis and T. rubrum respectively. The ethanol extracts of Phyllanthus reticulatus, Coldenia procumbens, Thespesia populnea and Senna alata significantly lowered the release of ergosterol by 16.37, 19.53, 24.79, and 21.44%, respectively. The ethanol extract of Phyllanthus reticulatus leaves was more biocompatible to host cells than other active extracts. CONCLUSION: Our study indicated that the ethanol extract of Phyllanthus reticulates leaves showed promising activity against dermatophytes. It could be a potential material for future development of antidermatophytic agents.

In Vitro Expression of Toll-Like Receptors and Proinflammatory Molecules Induced by Ergosta- 7,22-Dien-3-One Isolated from a Wild Mexican Strain of Ganoderma oerstedii (Agaricomycetes).

Compounds showing pharmacological activity on the immune system are of interest because of their therapeutic potential in the treatment of many diseases. However, data from primary human immune cells and in vivo studies are limited. The aim of this study was to analyze the ability to induce the expression of Toll-like receptors (TLRs) and proinflammatory molecules on cells involved in the immune system using the compound ergosta-7,22-dien-3- one, isolated from a wild Mexican strain of Ganoderma oerstedii. According to our study, ergosta-7,22-dien-3-one did not present any cytotoxic effect on HeLa or J774A.1 cells, and it was able to stimulate nitric oxide production, induce the expression of genes, and induce the production of TLRs, cytokines, chemokines, and cellular adhesion molecules in J774A.1 cells, based on reverse-transcriptase polymerase chain reaction and enzyme-linked immunosorbent assay. Here we report a new pro-inflammatory property of ergosta-7,22-dien-3-one, which should be considered as a possible adjuvant property in view of its biological activity.

Bioactive compounds and antioxidant activity exhibit high intraspecific variability in Pleurotus ostreatus mushrooms and correlate well with cultivation performance parameters.

Experimental data related with oyster mushroom production and nutritional properties usually derive from the examination of only one strain, and hence their representativeness/usefulness is questionable. This work aims at assessing intraspecific variability in Pleurotus ostreatus by studying 16 strains, under the same conditions, in respect to essential cultivation and mushroom quality aspects, and by defining the impact of intrinsic/genetic factors on such parameters. Hence, mushroom yield, earliness, crop length, biological efficiency, productivity, and their content in selected macro and microconstituents (e.g. fatty acids, sterols, individual phenolic compounds, terpenic acids, glucans) as well as their antioxidant properties (i.e., antiradical activity, ferric reducing potential, inhibition of serum oxidation) were assayed. The effect of intrinsic/genetic factors was evident, especially as regards earliness, yield of each production flush and mushroom weight, whereas biological efficiency was not particularly influenced by the cultivated strain. Moreover, phenolics, ergosterol and antiradical activity demonstrated significant variability among strains in contrast to what was observed for fatty acids, ß-glucans and ferric reducing potential. The observed heterogeneity reveals the limitations of using a low number of strains for evaluating mushroom production and/or their content in bioactive compounds, and as evidenced, it is valuable for breeding and commercial purposes.

Improvement of a continuous ethanol fermentation from sweet sorghum stem juice using a cell recycling system.

The process variables (aeration rate and recycle ratio) of a continuous ethanol fermentation with a cell recycling system (CRS) by Saccharomyces cerevisiae NP 01 from sweet sorghum stem juice were optimized using response surface methodology (RSM). The relationship between intracellular composition and fermentation efficiency was also investigated. RSM results revealed that the optimum aeration rate and recycle ratio were 0.25vvm and 0.625, respectively. The validation experiment under the optimum conditions indicated high precision and reliability of the experiment, achieving an actual ethanol concentration (PE) of 99.28g/l, which was very close to the predicted value (98.01g/l), and a very high ethanol productivity (QP) of 7.94g/lh. The intracellular composition of the yeast cells (i.e., unsaturated fatty acids (UFAs), total fatty acids (TFAs), ergosterol and trehalose) was positively related to the fermentation efficiency and yeast adaptive response under ethanol stress. A higher ratio of UFAs/TFAs and ergosterol strongly promoted yeast viability and ethanol fermentation. Additionally, high trehalose content was observed when the yeast was subjected to stress conditions.