Endo-xylanase GH11 activation by the fungal metabolite eugenitin

Eugenitin, a chromone derivative and a metabolite of the endophyte Mycoleptodiscus indicus, at 5 mM activated a recombinant GH11 endo-xylanase by 40 %. The in silico prediction of ligand-binding sites on the three-dimensional structure of the endo-xylanase revealed that eugenitin interacts mainly by a hydrogen bond with a serine residue and a stacking interaction of the heterocyclic aromatic ring system with a tryptophan residue. Eugenitin improved the GH11 endo-xylanase activity on different substrates, modified the optimal pH and temperature activities and slightly affected the kinetic parameters of the enzyme.

The COP9 signalosome counteracts the accumulation of cullin SCF ubiquitin E3 RING ligases during fungal development

Defects in the COP9 signalosome (CSN) impair multicellular development, including embryonic plant or animal death or a block in sexual development of the fungus Aspergillus nidulans. CSN deneddylates cullin-RING ligases (CRLs), which are activated by covalent linkage to ubiquitin-like NEDD8. Deneddylation allows CRL disassembly for subsequent reassembly. An attractive hypothesis is a consecutive order of CRLs for development, which demands repeated cycles of neddylation and deneddylation for reassembling CRLs. Interruption of these cycles could explain developmental blocks caused by csn mutations. This predicts an accumulation of neddylated CRLs exhibiting developmental functions when CSN is dysfunctional. We tested this hypothesis in A. nidulans, which tolerates reduced levels of neddylation for growth. We show that only genes for CRL subunits or neddylation are essential, whereas CSN is primarily required for development. We used functional tagged NEDD8, recruiting all three fungal cullins. Cullins are associated with the CSN1/CsnA subunit when deneddylation is defective. Two CRLs were identified which are specifically involved in differentiation and accumulate during the developmental block. This suggests that an active CSN complex is required to counteract the accumulation of specific CRLs during development.

Production of xylanase and beta-xylosidase from autohydrolysis liquor of corncob using two fungal strains

Agroindustrial residues are materials often rich in cellulose and hemicellulose. The use of these substrates for the microbial production of enzymes of industrial interest is mainly due to their high availability associated with their low cost. In this work, corncob (CCs) particles decomposed to soluble compounds (liquor) were incorporated in the microbial growth medium through autohydrolysis, as a strategy to increase and undervalue xylanase and beta-xylosidase production by Aspergillus terricola and Aspergillus ochraceus. The CCs autohydrolysis liquor produced at 200 A degrees C for 5, 15, 30 or 50 min was used as the sole carbon source or associated with untreated CC. The best condition for enzyme synthesis was observed with CCs submitted to 30 min of autohydrolysis. The enzymatic production with untreated CCs plus CC liquor was higher than with birchwood xylan for both microorganisms. A. terricola produced 750 total U of xylanase (144 h cultivation) and 30 total U of beta-xylosidase (96-168 h) with 0.75% untreated CCs and 6% CCs liquor, against 650 total U of xylanase and 2 total U of beta-xylosidase in xylan; A. ochraceus produced 605 total U of xylanase and 56 total U of beta-xylosidase (168 h cultivation) with 1% untreated CCs and 10% CCs liquor against 400 total U of xylanase and 38 total U of beta-xylosidase in xylan. These results indicate that the treatment of agroindustrial wastes through autohydrolysis can be a viable strategy in the production of high levels of xylanolytic enzymes.

Fresh Pasta Production Enriched with Spirulina platensis Biomass

The aim of this work was to study the enrichment of Spirulina platensis in wheat flour to prepare fresh pasta to evaluate the green color and nutritional enrichment in addition to functional properties due to the presence of the bioactive compounds in the cyanobacterium. The pastas were evaluated for the centesimal composition, microbiological contamination, sensorial acceptance and technological characteristics such as cooking time, water absorption, volume displacement and loss of solids. The superior protein contents and the satisfactory technological and sensorial attributes compared with the control with no cyanobacterium showed the usefulness of incorporating S. platensis biomass in the fresh pastas. The microbiological quality was in compliance with the legislation in force. The sensorial quality was considered satisfactory ("liked very much") and purchase intention high ("probably would buy").

Biotechnological Utilization of Biodiesel-Derived Glycerol for the Production of Ribonucleotides and Microbial Biomass

Ten yeast strains were evaluated concerning their capabilities to assimilate biodiesel-derived glycerol in batch cultivation. The influence of glycerol concentration, temperature, pH and yeast extract concentration on biomass production was studied for the yeast selected. Further, the effect of agitation on glycerol utilization by the yeast Hansenula anomala was also studied. The yeast H. anomala CCT 2648 showed the highest biomass yield (0.30 g g(-1)) and productivity (0.19 g L-1 h(-1)). Citric acid, succinic acid, acetic acid and ethanol were found as the main metabolites produced. The increase of yeast extract concentration from 1 to 3 g L-1 resulted in high biomass production. The highest biomass concentration (21 g L-1), yield (0.45 g g(-1)) and productivity (0.31 g L-1 h(-1)), as well as ribonucleotide production (13.13 mg g(-1)), were observed at 700 rpm and 0.5 vvm. These results demonstrated that glycerol from biodiesel production process showed to be a feasible substrate for producing biomass and ribonucleotides by yeast species.

Functional Characterization of an Aspergillus fumigatus Calcium Transporter (PmcA) that Is Essential for Fungal Infection

Aspergillus fumigatus is a primary and opportunistic pathogen, as well as a major allergen, of mammals. The Ca+2-calcineurin pathway affects virulence, morphogenesis and antifungal drug action in A. fumigatus. Here, we investigated three components of the A. fumigatus Ca+2-calcineurin pathway, pmcA,-B, and -C, which encode calcium transporters. We demonstrated that CrzA can directly control the mRNA accumulation of the pmcA-C genes by binding to their promoter regions. CrzA-binding experiments suggested that the 5'-CACAGCCAC-3' and 5'-CCCTGCCCC-3' sequences upstream of pmcA and pmcC genes, respectively, are possible calcineurin-dependent response elements (CDREs)-like consensus motifs. Null mutants were constructed for pmcA and -beta and a conditional mutant for pmcC demonstrating pmcC is an essential gene. The Delta pmcA and Delta pmcB mutants were more sensitive to calcium and resistant to manganese and cyclosporin was able to modulate the sensitivity or resistance of these mutants to these salts, supporting the interaction between calcineurin and the function of these transporters. The pmcA-C genes have decreased mRNA abundance into the alveoli in the Delta calA and Delta crzA mutant strains. However, only the A. fumigatus Delta pmcA was avirulent in the murine model of invasive pulmonary aspergillosis.

Determination of the intrinsic kinetic parameters of sulfide-oxidizing autotrophic denitrification in differential reactors containing immobilized biomass

Nitrogen removal coupled with sulfide oxidation has potential for the treatment of effluents from anaerobic reactors because they contain sulfide, which can be used as an endogenous electron donor for denitrification. This work evaluated the intrinsic kinetics of sulfide-oxidizing autotrophic denitrification via nitrate and nitrite in systems containing attached cells. Differential reactors were fed with nitrified synthetic domestic sewage and different sulfide concentrations. The intrinsic kinetic parameters of nitrogen removal were determined when the mass transfer resistance was negligible. This bioprocess could be described by a half-order kinetic model for biofilms. The half-order kinetic coefficients ranged from 0.425 to 0.658 mg N-1/2 L-1/2 h(-1) for denitrification via nitrite and from 0.190 to 0.609 mg N-1/2 L-1/2 h(-1) for denitrification via nitrate. In this latter, the lower value was due to the use of electrons donated from intermediary sulfur compounds whose formation and subsequent consumption were detected. (C) 2011 Elsevier Ltd. All rights reserved.

Metal biosorption onto dry biomass of Arthrospira (Spirulina) platensis and Chlorella vulgaris: Multi-metal systems

Binary and ternary systems of Ni2+, Zn2+, and Pb2+ were investigated at initial metal concentrations of 0.5, 1.0 and 2.0 mM as competitive adsorbates using Arthrospira platensis and Chlorella vulgaris as biosorbents. The experimental results were evaluated in terms of equilibrium sorption capacity and metal removal efficiency and fitted to the multi-component Langmuir and Freundlich isotherms. The pseudo second order model of Ho and McKay described well the adsorption kinetics, and the FT-IR spectroscopy confirmed metal binding to both biomasses. Ni2+ and Zn2+ interference on Pb2+ sorption was lower than the contrary, likely due to biosorbent preference to Pb. In general, the higher the total initial metal concentration, the lower the adsorption capacity. The results of this study demonstrated that dry biomass of C. vulgaris behaved as better biosorbent than A. platensis and suggest its use as an effective alternative sorbent for metal removal from wastewater. (C) 2012 Elsevier B.V. All rights reserved.

Chiral HPLC analysis of donepezil, 5-O-desmethyl donepezil and 6-O-desmethyl donepezil in culture medium: application to fungal biotransformation studies

An high performance liquid chromatography (HPLC) method for the enantioselective determination of donepezil (DPZ), 5-O-desmethyl donepezil (5-ODD), and 6-O-desmethyl donepezil (6-ODD) in Czapek culture medium to be applied to biotransformation studies with fungi is described for the first time. The HPLC analysis was carried out using a Chiralpak AD-H column with hexane/ethanol/methanol (75:20:5, v/v/v) plus 0.3 % triethylamine as mobile phase and UV detection at 270 nm. Sample preparation was carried out by liquid-liquid extraction using ethyl acetate as extractor solvent. The method was linear over the concentration range of 100-10,000 ng mL(-1) for each enantiomer of DPZ (r a parts per thousand yenaEuro parts per thousand 0.9985) and of 100-5,000 ng mL(-1) for each enantiomer of 5-ODD (r a parts per thousand yenaEuro parts per thousand 0.9977) and 6-ODD (r a parts per thousand yenaEuro parts per thousand 0.9951). Within-day and between-day precision and accuracy evaluated by relative standard deviations and relative errors, respectively, were lower than 15 % for all analytes. The validated method was used to assess DPZ biotransformation by the fungi Beauveria bassiana American Type Culture Collection (ATCC) 7159 and Cunninghamella elegans ATCC 10028B. Using the fungus B. bassiana ATCC 7159, a predominant formation of (R)-5-ODD was observed while for the fungus C. elegans ATCC 10028B, DPZ was biotransformed to (R)-6-ODD with an enantiomeric excess of 100 %.

Evaluating methods for the isolation of marine-derived fungal strains and production of bioactive secondary metabolites

In the present investigation we evaluate methods for the isolation and growth of marine-derived fungal strains in artificial media for the production of secondary metabolites. Inoculation of marine macroorganisms fragments in Petri dishes proved to be the most convenient procedure for the isolation of the largest number of strains. Among the growth media used, 3% malt extract showed the best result for strains isolation and growth, and yielded the largest number of strains from marine macroorganisms. The percentage of strains isolated using each of the growth media which yielded cytotoxic and/or antibiotic extracts was in the range of 23-35%, regardless of the growth media used. Further investigation of extracts obtained from different marine-derived fungal strains yielded several bioactive secondary metabolites, among which (E)-4-methoxy-5-(3-methoxybut-1-enyl)-6-methyl-2H-pyran-2-one is a new metabolite isolated from the Penicillium paxilli strain Ma(G)K.

Characterization of immobilized biomass by amplified rDNA restriction analysis (ARDRA) in an anaerobic sequencing-batch biofilm reactor (ASBBR) for the treatment of industrial wastewater

The performance of an anaerobic sequencing-batch biofilm reactor (ASBBR- laboratory scale- 14L )containing biomass immobilized on coal was evaluated for the removal of elevated concentrations of sulfate (between 200 and 3,000 mg SO4-2·L-1) from industrial wastewater effluents. The ASBBR was shown to be efficient for removal of organic material (between 90% and 45%) and sulfate (between 95% and 85%). The microbiota adhering to the support medium was analyzed by amplified ribosomal DNA restriction analysis (ARDRA). The ARDRA profiles for the Bacteria and Archaea domains proved to be sensitive for the determination of microbial diversity and were consistent with the physical-chemical monitoring analysis of the reactor. At 3,000 mg SO4-2·L-1, there was a reduction in the microbial diversity of both domains and also in the removal efficiencies of organic material and sulfate.

Fresh pasta production enriched with Spirulina platensis biomass

The aim of this work was to study the enrichment of Spirulina platensis in wheat flour to prepare fresh pasta to evaluate the green color and nutritional enrichment in addition to functional properties due to the presence of the bioactive compounds in the cyanobacterium. The pastas were evaluated for the centesimal composition, microbiological contamination, sensorial acceptance and technological characteristics such as cooking time, water absorption, volume displacement and loss of solids. The superior protein contents and the satisfactory technological and sensorial attributes compared with the control with no cyanobacterium showed the usefulness of incorporating S. platensis biomass in the fresh pastas. The microbiological quality was in compliance with the legislation in force. The sensorial quality was considered satisfactory ( liked very much ) and purchase intention high ( probably would buy ).

Intracellular biosynthesis and removal of copper nanoparticles by dead biomass of yeast isolated from the wastewater of a mine in the brazilian Amazonia

In this study was developed a natural process using a biological system for the biosynthesis of nanoparticles (NPs) and possible removal of copper from wastewater by dead biomass of the yeast Rhodotorula mucilaginosa. Dead and live biomass of Rhodotorula mucilaginosa was used to analyze the equilibrium and kinetics of copper biosorption by the yeast in function of the initial metal concentration, contact time, pH, temperature, agitation and inoculum volume. Dead biomass exhibited the highest biosorption capacity of copper, 26.2 mg g(-1), which was achieved within 60 min of contact, at pH 5.0, temperature of 30°C, and agitation speed of 150 rpm. The equilibrium data were best described by the Langmuir isotherm and Kinetic analysis indicated a pseudo-second-order model. The average size, morphology and location of NPs biosynthesized by the yeast were determined by scanning electron microscopy (SEM), energy dispersive X-ray spectroscopy (EDS) and transmission electron microscopy (TEM). The shape of the intracellularly synthesized NPs was mainly spherical, with an average size of 10.5 nm. The X-ray photoelectron spectroscopy (XPS) analysis of the copper NPs confirmed the formation of metallic copper. The dead biomass of Rhodotorula mucilaginosa may be considered an efficiently bioprocess, being fast and low-cost to production of copper nanoparticles and also a probably nano-adsorbent of this metal ion in wastewater in bioremediation process

Biosynthesis and uptake of copper nanoparticles by dead biomass of Hypocrea lixii isolated from the metal mine in the Brazilian Amazon Region

A biological system for the biosynthesis of nanoparticles (NPs) and uptake of copper from wastewater, using dead biomass of Hypocrea lixii was analyzed and described for the first time. The equilibrium and kinetics investigation of the biosorption of copper onto dead, dried and live biomass of fungus were performed as a function of initial metal concentration, pH, temperature, agitation and inoculum volume. The high biosorption capacity was observed for dead biomass, completed within 60 min of contact, at pH 5.0, temperature of 40 °C and agitation speed of 150 rpm with a maximum copper biosorption of 19.0 mg g(-1). The equilibrium data were better described using the Langmuir isotherm and kinetic analysis indicated that copper biosorption follows a pseudo-second-order model. The average size, morphology and location of NPs biosynthesized by the fungus were determined by scanning electron microscopy (SEM), energy dispersive X-ray spectroscopy (EDS) and transmission electron microscopy (TEM). NPs were mainly spherical, with an average size of 24.5 nm, and were synthesized extracellularly. The X-ray diffraction (XRD) analysis confirms the presence of metallic copper particles. Infrared spectroscopy (FTIR) study revealed that the amide groups interact with the particles, which was accountable for the stability of NPs. This method further confirmed the presence of proteins as stabilizing and capping agents surrounding the copper NPs. These studies demonstrate that dead biomass of Hypocrea lixii provides an economic and technically feasible option for bioremediation of wastewater and is a potential candidate for industrial-scale production of copper NPs.

Aerosols in Amazonia: Natural Biogenic Particles and Large Scale Biomass Burning Impacts.

The Large Scale Biosphere Atmosphere Experiment in Amazonia (LBA) is a long term (20 years) research effort aimed at the understanding of the functioning of the Amazonian ecosystem. In particular, the strong biosphere-atmosphere interaction is a key component looking at the exchange processes between vegetation and the atmosphere, focusing on aerosol particles. Two aerosol components are the most visible: The natural biogenic emissions of aerosols and VOCs, and the biomass burning emissions. A large effort was done to characterize natural biogenic aerosols that showed detailed organic characterization and optical properties. The biomass burning component in Amazonia is important in term of aerosol and trace gases emissions, with deforestation rates decreasing, from 27,000 Km2 in 2004 to about 5,000 Km2 in 2011. Biomass burning emissions in Amazonia increases concentrations of aerosol particles, CO, ozone and other species, and also change the surface radiation balance in a significant way. Long term monitoring of aerosols and trace gases were performed in two sites: a background site in Central Amazonia, 55 Km North of Manaus (called ZF2 ecological reservation) and a monitoring station in Porto Velho, Rondonia state, a site heavily impacted by biomass burning smoke. Several instruments were operated to measured aerosol size distribution, optical properties (absorption and scattering at several wavelengths), composition of organic (OC/EC) and inorganic components among other measurements. AERONET and MODIS measurements from 5 long term sites show a large year-to year variability due to climatic and socio-economic issues. Aerosol optical depths of more than 4 at 550nm was observed frequently over biomass burning areas. In the pristine Amazonian atmosphere, aerosol scattering coefficients ranged between 1 and 200 Mm-1 at 450 nm, while absorption ranged between 1 and 20 Mm-1 at 637 nm. A strong seasonal behavior was observed, with greater aerosol loadings during the dry season (Jul-Nov) as compared to the wet season (Dec-Jun). During the wet season in Manaus, aerosol scattering (450 nm) and absorption (637 nm) coefficients averaged, respectively, 14 and 0.9 Mm-1. Angstrom exponents for scattering were lower during the wet season (1.6) in comparison to the dry season (1.9), which is consistent with the shift from biomass burning aerosols, predominant in the fine mode, to biogenic aerosols, predominant in the coarse mode. Single scattering albedo, calculated at 637 nm, did not show a significant seasonal variation, averaging 0.86. In Porto Velho, even in the wet season it was possible to observe an impact from anthropogenic aerosol. Black Carbon was measured at a high 20 ug/m³ in the dry season, showing strong aerosol absorption. This work presents a general description of the aerosol optical properties in Amazonia, both during the Amazonian wet and dry seasons.