Effects of gadolinium and tin to the production of oxidative enzymes and the growth of five basidiomycetous fungi

Effects of gadolinium (Gd) and tin (Sn) on the growth and production of oxidative enzymes with five basidiomycetous fungi were tested. For this study we have selected well-known white-rot fungi Obba rivulosa and Kuehneromyces mutabilis, in addition to this we have tested three new isolates, the white-rot fungus Phlebia subochracea, the litter-degrading fungus Gymnopus dryophilus and the brown-rot fungus Heliocybe sulcata. This approach allowed us to find possible new sources for oxidative enzymes, such as laccases and versatile peroxidases (VPs). All five tested fungi grew in the presence of Gd (0-200 mg/l) or Sn (0-200 mg/l) on ABTS (2,2’-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid) containing plates. The growth rate of H. sulcata was tolerant to Gd and Sn (0- 200 mg/l). The growth rates of P. subochracea and G. dryophilus were sensitive to Gd (5-200 mg/l) and Sn (5-200 mg/l). O. rivulosa, K. mutabilis, P. subochracea and G. dryophilus formed colour zones on the ABTS plates indicating that these fungi produced oxidative enzymes, most probably laccases. The brown-rot fungus H. sulcata did not form colour zone on the ABTS plate indicating that this fungus did not produce laccase. The production of laccase with G. dryophilus and K. mutabilis was tolerant to Gd (0-200 mg/l) and Sn (0-200 mg/l). The production of laccase with P. subochracea was sensitive to Gd (5-200 mg/l) and Sn (5-200 mg/l). P. subochracea decolorized the dye Reactive Black 5 without or with Gd and Sn (0- 200 mg/l) indicating the production of VP. O. rivulosa, K. mutabilis, G. dryophilus and H. sulcata did not produce VP. The production of VP by P. subochracea was sensitive to 200 mg/l Gd and Sn.Peer reviewe

Ecosystem services of soil microbial communities - developing sustainable cultivation methods for agriculture

The aim of the project is to develop innovative and feasible agro-environmental technology for improvement of environmental sustainability of cultivation methods. This will be achieved by increasing soil crop cover taking advantage of ecosystem services of soil microbes to suppress crop pathogens as an alternative system for chemical control, and to optimal degradation of crop residues

Conversion of Milled Pine Wood by Manganese Peroxidase from Phlebia radiata

Purified manganese peroxidase (MnP) from the white-rot basidiomycete Phlebia radiata was found to convert in vitro milled pine wood (MPW) suspended in an aqueous reaction solution containing Tw-een 20, Mn2+, bon-chelating organic acid (malonate), and a hydrogen peroxide-generating system (glucose-glucose oxidase). The enzymatic attack resulted in the polymerization of lower-molecular-mass, soluble wood components and in the partial depolymerization of the insoluble bulk of pine wood, as demonstrated by high-performance size exclusion chromatography (HPSEC). The surfactant Tween 80 containing unsaturated fatty acid redsidues promoted the disintegration of bulk MPW. HPSEC showed that the depolymerization yielded preferentially lignocellulose fragments with a predominant molecular mass of ca. 0.5 kDa. MnP from P. radiata (MnP3) turned out to be a stable enzyme remaining active for 2 days even at 37 degreesC with vigorous stirring, and 65 and 35% of the activity applied was retained in Tween 20 and Tween 80 reaction mixtures, respectively. In the course of reactions, major part of the Mn-chelator malonate was decomposed (85 to 87%), resulting in an increase of pH from 4.4 to > 6.5. An aromatic nonphenolic lignin structure (beta -O-4 dimer), which is normally not attacked by MnP, was oxidizible in the presence of pine wood meal. This finding indicates that certain wood components may promote the degradative activities of MnP in a way similar to that promoted by Tween 80, unsaturated fatty acids, or thiols

Degradation of Humic Acids by the Litter-Decomposing Basidiomycete Collybia dryophila

The basidiomycete Collybia diyophila K209, which colonizes forest soil, was found to decompose a natural humic acid isolated from pine-forest litter (LHA) and a synthetic C-14-Iabeled humic acid (14 C-HA) prepared from [U-C-14] catechol in liquid culture. Degradation resulted in the formation of polar, lower-molecular-mass fulvic acid (FA) and carbon dioxide. RA decomposition was considerably enhanced in the presence of Mn 21 (200 muM), leading to 75% conversion of LHA and 50% mineralization of C-14-HA (compared to 60% and 20%, respectively, in the absence of Mn 21). There was a strong indication that manganese peroxidase (MnP), the production of which was noticeably increased in Mn2+-supplemented cultures, was responsible for this effect. The enzyme was produced as a single protein with a pI of 4.7 and a molecular mass of 44 kDa. During solid-state cultivation, C diyophila released substantial amounts of water-soluble FA (predominantly of 0.9 kDa molecular mass) from insoluble litter material. The results indicate that basidiomycetes such as C diyophila which colonize forest litter and soil are involved in humus turnover by their recycling of highmolecular-mass humic substances. Extracellular MnP seems to be a key enzyme in the conversion process

Degradation of Benzo[a]pyrene by the Litter-Decomposing Basidiomycete Stropharia coronilla : Role of Manganese Peroxidase

The litter-decomposing basidiomycete Stropharia coronilla, which preferably colonizes grasslands, was found to be capable of metabolizing and mineralizing benzo[a]pyrene (BaP) in liquid culture. Manganese(II) ions (Mn2+) supplied at a concentration of 200 {micro}M stimulated considerably both the conversion and the mineralization of BaP; the fungus metabolized and mineralized about four and twelve times, respectively, more of the BaP in the presence of supplemental Mn2+ than in the basal medium. This stimulating effect could be attributed to the ligninolytic enzyme manganese peroxidase (MnP), whose activity increased after the addition of Mn2+. Crude and purified MnP from S. coronilla oxidized BaP efficiently in a cell-free reaction mixture (in vitro), a process which was enhanced by the surfactant Tween 80. Thus, 100 mg of BaP liter-1 was converted in an in vitro reaction solution containing 1 U of MnP ml-1 within 24 h. A clear indication was found that BaP-1,6-quinone was formed as a transient metabolite, which disappeared over the further course of the reaction. The treatment of a mixture of 16 different polycyclic aromatic hydrocarbons (PAHs) selected by the U.S. Environmental Protection Agency as model standards for PAH analysis (total concentration, 320 mg liter-1) with MnP resulted in concentration decreases of 10 to 100% for the individual compounds, and again the stimulating effect of Tween 80 was observed. Probably due to their lower ionization potentials, poorly bioavailable, high-molecular-mass PAHs such as BaP, benzo(g,h,i)perylene, and indeno(1,2,3-c,d)pyrene were converted to larger extents than low-molecular-mass ones (e.g., phenanthrene and fluoranthene)

Impact of copper and zinc on the growth of saprotrophic fungi and the production of extracellular enzymes

Peer reviewe

Decolorization of six synthetic dyes by fungi

To find out ability of fourteen basidiomycetes and four ascomycetes strains to grow in the presence of synthetic colour dyes and to degrade them, fungi were cultivated on the malt agar plates containing 0.5 g kg-1 dye, either Remazol Brilliant Blue R, Remazol Brilliant Yellow GL, Remazol Brilliant Orange 3 R, Reactive Blue 4, Remazol Brilliant Red F3B or Reactive Black 5. Fungi representing basidiomycetes were Phlebia radiata (FBCC 43), Tremella encephala (FBCC 1145), Dichomitus squalens (FBCC 312), Physisporinus rivulosus (syn. Obba rivulosa, FBCC 939), Cerrena unicolor (FBCC 387), Pleurotus abieticola (FBCC 517), Phanerochaete velutina (FBCC 941), Agrocybe praecox (FBCC 476), Trametes pubescens (FBCC 735), Pleurotus ostreatus (FBCC 498), Fomitopsis pinicola (FBCC 18), Postia placenta (= syn. Rhodonia placenta, FBCC 112), Gloeophyllum trabeum (FBCC 328) and Piptoporus betulinus (FBCC 1191). Ascomycetes belonged to genera Alternaria (HAMBI 3289), Epicoccum (HAMBI 3291), Fusarium (HAMBI 3292) or Chaetomium (HAMBI 3291). The growth rate of P. rivulosus belonged to three highest among the 14 tested basidiomycetes with five dyes, but not in the case of Remazol Brilliant Red F3B containing plates. The growth rate of A. praecox belonged to three lowest among the 14 tested basidiomycetes on Remazol Brilliant Blue R, Remazol Brilliant Yellow GL, Remazol Brilliant Red F3B, Reactive Black 5 and malt agar. The growth rate of Chaetomium sp. was the highest among the four ascomycetes on all tested plates. Decolorization was seen with 7 basidiomycetous strains on Remazol Brilliant Blue R, with 7 basidiomycetes on Remazol Brilliant Orange 3 R, with 8 basidiomycetes on Reactive Blue 4 and 11 basidiomycetes on Reactive Black 5 containing plates. T. encephala did not decolorize any of the tested six dyes. Epicoccum sp. and Chaetomium sp. decolorized Reactive Black 5 dye containing plates. None of the fourteen basidiomycetous or four ascomycetes were able to degrade all the tested six dyes.Peer reviewe

Production of Manganese Peroxidase and Organic Acids and Mineralization of 14C-Labelled Lignin (14C-DHP) during Solid-State Fermentation of Wheat Straw with the White Rot Fungus Nematoloma frowardii

The basidiomycetous fungus Nematoloma frowardii produced manganese peroxidase (MnP) as the predominant ligninolytic enzyme during solid-state fermentation (SSF) of wheat straw. The purified enzyme had a molecular mass of 50 kDa and an isoelectric point of 3.2. In addition to MnP, low levels of laccase and lignin peroxidase were detected. Synthetic 14C-ring-labelled lignin (14C-DHP) was efficiently degraded during SSF. Approximately 75% of the initial radioactivity was released as 14CO2, while only 6% was associated with the residual straw material, including the well-developed fungal biomass. On the basis of this finding we concluded that at least partial extracellular mineralization of lignin may have occurred. This conclusion was supported by the fact that we detected high levels of organic acids in the fermented straw (the maximum concentrations in the water phases of the straw cultures were 45 mM malate, 3.5 mM fumarate, and 10 mM oxalate), which rendered MnP effective and therefore made partial direct mineralization of lignin possible. Experiments performed in a cell-free system, which simulated the conditions in the straw cultures, revealed that MnP in fact converted part of the 14C-DHP to 14CO2 (which accounted for up to 8% of the initial radioactivity added) and 14C-labelled water-soluble products (which accounted for 43% of the initial radioactivity) in the presence of natural levels of organic acids (30 mM malate, 5 mM fumarate)