Diallyl Trisulfide, the Antifungal Component of Garlic Essential Oil and the Bioactivity of Its Nanoemulsions Formed by Spontaneous Emulsification.

The aim of this study was to evaluate the chemical compounds of garlic essential oil (EO), and determine the antifungal efficacy of garlic EO and its major components, diallyl trisulfide and its nanoemulsions against wood-rotting fungi, Trametes hirsuta and Laetiporus sulphureus. GC-MS analysis revealed that the major constituents of garlic EO were diallyl trisulfide (39.79%), diallyl disulfide (32.91%), and diallyl sulfide (7.02%). In antifungal activity, the IC50 value of garlic EO against T. hirsuta and L. sulphureus were 137.3 and 44.6 µg/mL, respectively. Results from the antifungal tests demonstrated that the three major constituents were shown to have good antifungal activity, in which, diallyl trisulfide was the most effective against T. hirsuta and L. sulphureus, with the IC50 values of 56.1 and 31.6 µg/mL, respectively. The diallyl trisulfide nanoemulsions showed high antifungal efficacy against the examined wood-rotting fungi, and as the amount of diallyl trisulfide in the lipid phase increases, the antifungal efficacy of the nanoemulsions increases. These results showed that the nanoemulsions and normal emulsion of diallyl trisulfide have potential to develop into a natural wood preservative.

Studies on the characteristics and mechanism of aerobic biodegradation of tetrabromobisphenol A by Irpex lacteus F17.

The study investigated the characteristics of aerobic degradation of tetrabromobisphenol A (TBBPA) by Irpex lacteus F17 (I. lacteus F17) under four different cometabolic substrates (phenol, glucose, sodium pyruvate, and sodium citrate). The biodegradation of TBBPA by I. lacteus F17 could be enhanced via cometabolism, and glucose (8 g/L) was confirmed to be the optimum carbon source. For different initial solution pH ranging from 3.0 to 8.0, the results showed that I. lacteus F17 could be applied to biodegrade TBBPA in a wide pH range of 4.0-8.0, and the degradation rate could reach the maximum 75.31%, while the debromination rate reached the maximum 12.40% under pH 5.0. In addition, it has been confirmed that Mn2+ (50 µmol/L) could promote the secretion of manganese peroxidase and TBBPA biodegradation efficiency. Seven intermediates were identified by gas chromatography-mass spectrometry analysis, and the possible degradation pathways were proposed, which indicated the biodegradation of TBBPA might be subjected to debromination, ß-scission, hydroxylation, deprotonation, and oxidation reactions.

Antifeedant and antiphytopathogenic metabolites from co-culture of endophyte Irpex lacteus, phytopathogen Nigrospora oryzae, and entomopathogen Beauveria bassiana.

Five new tremulane sesquiterpenoids were isolated from co-culture of endophyte Irpex lacteus, phytopathogen Nigrospora oryzae, and entomopathogen Beauveria bassiana. All compounds showed obvious antifeedant activities against silkworm with inhibition percentages of 73-99%, at concentrations of 50 µg/cm2. Compound 11 indicated notable antifeedant activity with inhibition percentage of 93% at concentration of 6.25 µg/cm2 among them. Compounds 2, 3, 4, 8, 9, 15 and 16 indicated anti-fungal activities against I. lacteus with MIC values ≤8 µg/mL, compounds 11, 12, 16-18 showed significant anti-fungal activity against N. oryzae with MICs ≤ 4 µg/mL, and compounds 2, 5, 12 and 18 indicated significant anti-fungal activity against B. bassiana with MICs ≤ 8 µg/mL. In addition, the I. lacteus should unite B. bassiana to inhibit the production of phytotoxins from N. oryzae in the ternary culture.

The Cocultured Nigrospora oryzae and Collectotrichum gloeosporioides, Irpex lacteus, and the Plant Host Dendrobium officinale Bidirectionally Regulate the Production of Phytotoxins by Anti-phytopathogenic Metabolites.

The distinctive nature of the endophyte Irpex lacteus, host plant, and the phytopathogen Collectotrichum gloeosporioides resulted in both negative and positive regulation of the production of phytotoxins from Nigrospora oryzae. The coculture of nonhomologous I. lacteus and N. oryzae resulted in a greater number of anti-phytopathogenic metabolites from the dominant endophyte than the coculture of homologous I. lacteus and N. oryzae. The coculture of the phytopathogen N. oryzae and either the nonhomologous (isolation of I. lacteus and N. oryzae from the different plants) or homologous (isolation of I. lacteus and N. oryzae from the same plant) endophyte I. lacteus from different sources indicated that the nonhomologous I. lacteus grew faster than the homologous I. lacteus, and the production of phytotoxic azaphilone from the phytopathogenic N. oryzae decreased due to the inhibition resulting from being cocultured with nonhomologous I. lacteus. On the other hand, the production of phytotoxic azaphilone was promoted by the coculture of two phytopathogens, N. oryzae and C. gloeosporioides. The extract of the host plant, Dendrobium officinale, also increased anti-phytopathogenic metabolite production. Six new phytotoxic azaphilones from N. oryzae, four new tremulane sesquiterpenes from I. lacteus, and a new polyketone were isolated. The endophyte-phytopathogen, phytopathogen-phytopathogen, and endophyte-phytopathogen-host interactions can induce the chemical diversity of novel anti-phytopathogenic metabolites.

New azaphilones and tremulane sesquiterpene from endophytic Nigrospora oryzae cocultured with Irpex lacteus.

Five new metabolites belonging to two backbones of pulvilloric acid-type azaphilone and tremulane sesquiterpene were obtained and their structures were determined by spectral analysis. Based on the biogenesis analysis, tremulane sesquiterpenes were obtained from Irpex lacteus by the stimulation of mixed-culture. The antifungal selectivities of metabolites produced by fungus against their co-culture fungus and common pathogens, exhibited competitive interaction of this mix-culture. The tremulane sesquiterpene conocenol B produced by I. lacteus through the induction of Nigrospora oryzae showed selectivity of anti-fungal activity against its co-culture fungus, N. oryzae, with MICs at 16 µg/mL and 128 µg/mL against I. lacteus. The fungus can metabolize these new compounds to inhibit the growth of co-culture fungus while not inhibiting its own growth. Compound 5 was active against acetylcholinesterase (AChE) with a ratio of 35% at the concentration of 50 µM.

[Effect of calcium on sporulation of Taiwanofungus camphoratus in submerged fermentation].

Taiwanofungus camphoratus is a valuable and rare medicinal mushroom with various bioactivities, such as liver protection and anti-cancer. T. camphoratus can produce many arthroconidia at the end of submerged fermentation, but molecular mechanism underlying this submerged conidiation remains unknown. In this study, we found that Ca²âº concentration in culture medium significantly affected the arthroconidium production of T. camphoratus. Then, we identified two proteins (CaM and HSP90) involved in Ca²âº/calmodulin signaling pathway and one protein (AbaA) involved in FluG-mediated conidiation pathway by two-dimensional electrophoresis analyses. Furthermore, we proposed a Ca²âº/calmodulin- and FluG-mediated signaling pathway by bioinformatics analysis. By real-time quantitative PCR analyses of 23 key genes in the Ca²âº/calmodulin- and FluG-mediated conidiation pathway, we found that expression levels of 7 genes (crz1, hsp90, flbB, brlA, abaA, wetA and fadA) showed significant responses to Ca²âº concentration in fermentation medium. Our research is beneficial for elucidating the underlying mechanism of submerged fermentation conidiation for T. camphoratus.

Applications of volatile compounds acquired from Muscodor heveae against white root rot disease in rubber trees (Hevea brasiliensis Müll. Arg.) and relevant allelopathy effects.

The bioactive compounds of the volatile metabolite-producing endophytic fungus, Muscodor heveae, were examined by the process of biofumigation for the purposes of controlling white root rot disease in rubber trees (Hevea brasiliensis Müll. Arg.). Volatile organic compounds (VOCs) of M. heveae possess antimicrobial activity against Rigidoporus microporus in vitro with 100 % growth inhibition. The synthetic volatile compounds test confirmed that the major component, 3-methylbutan-1-ol, and the minor compounds, 3-methylbutyl acetate and 2-methylpropanoic acid, inhibited root and shoot growth in the tested plants 3-methylbutan-1-ol showed ED50 value and MIQ value on seed germination of ruzi grass, Arabidopsis thaliana Col-0 and tomato at 10, 5 and 5 µL-1 airspace, respectively. In vivo tests were carried out under greenhouse conditions using M. heveae inoculum fumigated soil that had been inoculated with R. microporus inoculum. After which, all seven treatments were compared. Significant differences were observed with a disease score at 150 d after treatment. Biofumigation by M. heveae showed great suppression of the disease. Biocontrol treatments; RMH40 (40 g kg-1M. heveae inoculum) and RMH80 (80 g kg-1M. heveae inoculum) were not found to be significantly different when compared with fungicide treatment (RT) and the non-infected control, but results were found to be significantly different from R. microporus infested (R) treatment. RMH40 and RMH80 revealed a low disease scores with a high survival rate of rubber tree seedling at 100 %, while R treatment showed the highest disease score of 4.8 ± 0.5 with a survival rate of rubber tree seedling at 25 %. The infected roots, appearing as a white colour. We have concluded that the bioactive VOCs of M. heveae would be an alternative method for the control of white root rot disease in rubber trees.

Optimization of laccase fermentation and evaluation of kinetic and thermodynamic parameters of a partially purified laccase produced by Daedalea flavida.

Studies on laccase production by Daedalea flavida were carried out in static and low-speed shake cultures. The enzyme production was reduced drastically at a high speed of shaking. Optimal production conditions are necessary to assess the quality of laccase suitable for a specific application. Thus, the production of laccase was optimized by the application of response surface methodology. Laccase production was 8-fold and 7.5-fold more in static and low-speed shake conditions, respectively, in an optimal medium composition than in an unoptimized medium. Laccase obtained using the optimal culture medium of D. flavida was tested for its stability at different temperatures and pH conditions. The partially purified enzyme was most stable at 30°C and pH 5. The half-life of laccase is 87 min at 60°C and at pH 6. The kinetic and thermodynamic parameters were evaluated for the inactivation of the partially purified laccase. The entropy change of inactivation of the enzyme is least at pH 4.

Evaluation of the antifungal effects of bio-oil prepared with lignocellulosic biomass using fast pyrolysis technology.

This study was performed to investigate the utility of bio-oil, produced via a fast pyrolysis process, as an antifungal agent against wood-rot fungi. Bio-oil solutions (25-100 wt.%) were prepared by diluting the bio-oil with EtOH. Wood block samples (yellow poplar and pitch pine) were treated with diluted bio-oil solutions and then subjected to a leaching process under hot water (70°C) for 72 h. After the wood block samples were thoroughly dried, they were subjected to a soil block test using Tyromyces palustris and Trametes versicolor. The antifungal effect of the 75% and 100% bio-oil solutions was the highest for both wood blocks. Scanning electron microscopy analysis indicated that some chemical components in the bio-oil solution could agglomerate together to form clusters in the inner part of the wood during the drying process, which could act as a wood preservative against fungal growth. According to GC/MS analysis, the components of the agglomerate were mainly phenolic compounds derived from lignin polymers.

Synthesis and evaluation of N-alkyl-beta-D-glucosylamines on the growth of two wood fungi, Coriolus versicolor and Poria placenta.

Various glucosylamines were synthesized from glucose and different alkyl amine compounds. These amino compounds are beta-D-glucopyranosylamine (GPA), N-ethyl-beta-D-glucopyranosylamine (EtGPA), N-butyl-beta-D-glucopyranosylamine (BuGPA), N-hexyl-beta-D-glucopyranosylamine (HeGPA), N-octyl-beta-D-glucopyranosylamine (OcGPA), N-dodecyl-beta-D-glucopyranosylamine (DoGPA), N-(2-hydroxyethyl)-beta-D-glucopyranosylamine (HEtGPA) and N,N-di(2-hydroxyethyl)-beta-D-glucopyranosylamine (DHEtGPA). They were tested for their antifungal activity against the growth of Coriolus versicolor and Poria placenta. An improvement of the antifungal activity with the increase of alkyl chain length was observed. DoGPA exhibited the best antifungal activity against both strains. It completely inhibited the fungal growth at 0.01x10(-3)molmL(-1) and 0.0075x10(-3)molmL(-1) for C. versicolor and P. placenta, respectively. For other glucosylamines higher concentrations were needed for complete inhibition of fungi.

The role of the ubiquitin-proteasome system in the response of the ligninolytic fungus Trametes versicolor to nitrogen deprivation.

The white rot fungus Trametes versicolor is an efficient lignin degrader with ecological significance and industrial applications. Lignin-modifying enzymes of white rot fungi are mainly produced during secondary metabolism triggered in these microorganisms by nutrient deprivation. Selective ubiquitin/proteasome-mediated proteolysis is known to play a crucial role in the response of cells to various stresses such as nutrient limitation, heat shock, and heavy metal exposure. Previous studies from our laboratory demonstrated that proteasomal degradation of intracellular proteins is involved in the regulation of laccase, a major ligninolytic enzyme of T. versicolor, in response to cadmium. In the present study, it was found that the 6-h nitrogen starvation leads to depletion of intracellular free ubiquitin pool in T. versicolor. The difference in the intracellular level of free monomeric ubiquitin observed between the mycelium extract from the nitrogen-deprived and that from the nitrogen-sufficient culture was accompanied by the different pattern of ubiquitin-dependent degradation. Furthermore, it was found that nitrogen deprivation affected 26S proteasome activities of T. versicolor. Proteasome inhibition by lactacystin beta-lactone, a highly specific agent, increased laccase activity in nitrogen-deprived cultures, but not in nitrogen-sufficient cultures. The present study implicates the ubiquitin/proteasome-mediated proteolytic pathway in the response of T. versicolor to nitrogen deprivation.

Impact assessment of bisphenol A on lignin-modifying enzymes by basidiomycete Trametes versicolor.

The impact of different concentrations of bisphenol A (BPA) was evaluated on growth of the white-rot basidiomycete, Trametes versicolor, and on the expression of genes encoding lignin-modifying enzyme (LME) activities. Effective doses (EDs) were obtained from fungal growth rate to monitor LME activities and the expression levels of their encoding genes. The fungus showed mycelial growth at concentrations of up to 300 microg ml(-1) of BPA with an ED50 value of 185 microg ml(-1). The LME activities were stimulated by BPA concentrations up to 300 microg ml(-1). The lignin peroxidase (LIP) encoding gene may be sensitive to BPA stress.

Biodegradation of Orange G by wood-rot fungi Phanerochaete sordida TXJ-1302A and Tyromyces lauteus TXJ-1302B.

Two strains isolated from the organic layers of forests on Zijin Mountain have indicated a strong capability of decolorization for Orange G on the solid plates. They were identified as Phanerochaete sordida and Tyromyces lauteus according to phenotypic and molecular techniques. Through this study, we try to find the suitable condition and cheapest way for decolorization by two strains. The result shows that malt extract and ammonium sulfate are the best N source for P. sordida and T. lauteus, respectively; 0.95 g per L glucose + 0.05 g per L ethonal are the best C source both for P. sordida and T. lauteus. Oxalate plays an important role as the organic acid chelator which can also enhance the decolorized capability of fungi.

Manganese affects the production of laccase in the basidiomycete Ceriporiopsis subvermispora.

The authors have previously identified and characterized lcs, a gene encoding laccase in the white-rot basidiomycete Ceriporiopsis subvermispora. In this work, the effect of Mn2+ in the production of extracellular laccase in liquid cultures of this fungus has been assessed. It was observed that at low (0-10 microM) concentrations of Mn2+, high titers of lcs-mRNA were obtained, whereas at high (160-194 microM) concentrations of this metal ion, transcripts levels decreased markedly. This phenomenon was observed at different days of growth. On the other hand, Cu2+ or Ag+, but not Zn2+ or Cd2+, led to an accumulation of lcs transcripts only in cultures grown in the absence of Mn2+. A dramatic increase in lcs transcript levels was also obtained with syringic acid, a lignin-related aromatic compound. This effect was more pronounced in cultures lacking Mn2+. In the course of these studies it was observed that Mn2+ stimulates mycelium growth. Thus, although extracellular laccase activity appeared higher in cultures containing 160 or 194 microM Mn2+, i.e. when lcs transcripts were lower, a correlation between lcs-mRNA levels and enzymatic activity was observed when values of the latter were corrected by the amount of mycelium present in the cultures.

Viridenepoxydiol, a new pentasubstituted oxiranyldecene produced by Trichoderma viride.

A new pentasubstituted oxiranyldecene, named viridenepoxydiol, has been isolated (0.9 mg/L) from the culture filtrate of a strain of Trichoderma viride showing in vitro and in vivo antagonistic activity against Sclerotium rolfsii, which is the causal agent of crown and stem rot of artichoke. Viridenepoxydiol was characterized as 3,5,9-trimethyl-2-oxiranyl-dec-8-ene-2,5-diol (3) using spectroscopic methods. It showed inhibitor effect on mycelial growth of S. rolfsii and its minimum inhibitory concentration (over 90% inhibition) was found to be 396 mug/mL. This is the first time that viridenepoxydiol was reported.

Adenosine as an active component of Antrodia cinnamomea that prevents rat PC12 cells from serum deprivation-induced apoptosis through the activation of adenosine A(2A) receptors.

Antrodia cinnamomea (formerly named Antrodia camphorata) is a rare medicinal fungus. We previously reported that it exhibits antioxidative, vasorelaxative, anti-inflammatory, and anti-angiogenic effects. When serum deprivation-induced apoptosis in neuronal-like PC12 cells was used as a stress model, the extract of A. cinnamomea displayed effectiveness in preventing serum-deprived apoptosis. Since our previous data show that the extract of A. cinnamomea contains adenosine (ADO), we attempt to investigate if the active component is ADO and to identify its targeting site in this study. After pre-incubation with ADO deaminase, neither ADO nor the extract of A. cinnamomea exerted any protection, demonstrating that the active component of A. cinnamomea is ADO. Furthermore, an ADO A(2A) receptor (A(2A)-R) antagonist was used and was able to block the protective effects of ADO and the extract of A. cinnamomea, demonstrating that the ADO targeting site in this model is A(2A)-R. Taken together, the protective effect of A. cinnamomea is owed to its active component, ADO, which acts through activation of A(2A)-R to prevent serum deprivation-induced PC12 cell apoptosis.

Antifungal efficacy of chitosan and its thiourea derivatives upon the growth of some sugar-beet pathogens.

Chitosan (CS) was modified by reaction with benzoyl thiocyanate to give a thiourea derivative (TUCS). The antifungal behavior of chitosan and its thiourea derivative was investigated in vitro on the mycelial growth, sporulation and germination of conidia or sclerotia of the following sugar-beet: Beta vulgaris pathogens isolated in Egypt, Rhizoctonia solani Kühn (AG(2-2)) Sclerotium rolfsii Sacc. and Fusarium solani (Mart.) Sacc. All the prepared thiourea derivatives had a significant inhibiting effect on the different stages of development on the germination of conidia or sclerotia of all the investigated fungi in the polymer concentration range of 5-1000 microg ml(-1). In the absence of chitosan and its derivative, R. solani exhibited the fastest growth of the fungi studied. However, growth tolerance of the modified chitosan was highest for F. solani and lowest for R. solani. The most sensitive to the modified chitosan stress with regard to their germination and number produced were the sclerotia of S. rolfsii. It has been found that the TUCS is a much better fungicidal agent (about 60 times more) than the pure CS against most of the fungal strains tested. The molecular weight and the degree of deacetylation were found to have an important effect on the growth activities of the pathogens.

Role of white rot fungus Funalia trogii in detoxification of textile dyes.

Toxic and genotoxic effects of the textile dyes on organisms suggest the need for remediation of dyes before discharging them into the environment. For this reason, the ability of Funalia trogii pellets to detoxify textile dyes was investigated and evaluated. Although, textile dyes are toxic substances for many microorganisms, the pellets were able to decolorize and detoxify the azo dyes used. Astrazon Blue and Red dyes inhibit growth of F. trogii and S. aureus on solid medium in a concentration dependent manner. The toxicity of these dyes on a fungus, F. trogii and a bacterium, S. aureus was significantly decreased after pretreatment with fungal pellets.

Viridepyronone, a new antifungal 6-substituted 2H-pyran-2-one produced by Trichoderma viride.

A new antifungal 6-substituted 2H-pyran-2-one, named viridepyronone, has been isolated from a cultural filtrate of a strain of Trichoderma viride showing antagonistic activity in vitro toward Sclerotium rolfsii, which is the causal agent of crown and stem rot of artichoke. Viridepyronone was characterized as 6-(4-oxopentyl)-2H-pyran-2-one 2 with spectroscopic methods. Bioassays showed that viridepyronone had a good antifungal activity against S. rolfsii, and its minimum inhibitory concentration (over 90% inhibition) was found to be 196 microg/mL. This is the first report of viridepyronone produced by any species of fungi.

Grape seeds: the best lignocellulosic waste to produce laccase by solid state cultures of Trametes hirsuta.

Grape seeds were used by Trametes hirsuta as a substrate for laccase production giving 23 kU l(-1), which was 10-fold the value attained in the cultures with no lignocellulosic waste addition. The dyes, Indigo Carmine and Bromophenol Blue, were easily decolourised (100% in 24 h) by the extracellular liquid obtained in such cultures, whereas Methyl Orange (65% in 24 h) and Phenol Red (36% in 24 h) were more resistant to degradation. This shows the specificity of laccase towards different dye structures.