Soft Rot Fungi Research Articles

Trials on the Efficacy of Micronized Copper in Australia

Alkaline copper quat (ACQ) is an established wood preservative that is formulated with solubilized copper in amine solvent. This article describes three separate trials in Australia that investigated whether substituting soluble copper with micronized copper affects performance. ACQ and micronized copper quat (MCQ) performed similarly in Pinus radiata against four brown-rot fungi in a soil-block bioassay, while MCQ performed slightly better against two white-rot fungi in Eucalyptus delegatensis. A 2.3-year in-ground stake trial in the wet tropics at Innisfail also found that ACQ and MCQ performed comparably in P. radiata and Corymbia maculata. This was a severe test site with attack caused by soft-rot fungi, white-rot fungi, and termites. An H3 (outside, aboveground) field test against termites in Darwin showed that ACQ- or MCQ-treated P. radiata and C. maculata performed similarly against Coptotermes acinaciformis and Mastotermes darwiniensis. These trials demonstrated that MCQ performs comparably to ACQ under the test conditions used.

Microbial attack of acetylated wood in field soil trials

Microscopic evaluation of microbial attack in the acetylated wood after a long term exposure to the field soil was the main concern of the current research. Stakes of wood were cut from beech, poplar and Scots pine wood and acetylated by using acetic anhydride to achieve different weight percent gains (WPGs). Afterwards, the stakes were exposed to the field soil for 350 weeks according to EN 252. Small specimens were cut from the stakes, sectioned and stained for microscopic studies. Small blocks were also prepared for scanning electron microscopy (SEM). Microscopic observations indicated that the soft rot was dominant rot fungi in the field soil and decayed the non-acetylated wood severely and attack also the moderately acetylated wood slowly. Bacteria were also in close association with the soft rot fungi to degrade the acetylated and the non-acetylated wood. In the moderately acetylated wood, white rot decay was at developing stage, while it was at initial stage at the highest WPGs. Microscopy revealed that the initial stage of the decay was begun at the highest WPGs and it was at developing stage of the decay in the moderately acetylated wood. The less development of the decay in the acetylated wood comparing the non-acetylated wood proved very good protection of the wood against the soil microorganisms due to the acetylation after 7 years period. Soft rot and white rot decays were observed in the non-acetylated and the acetylated wood at the same time. No brown rot decay was observed in those woods. Bacteria were seen in all wood types. The bacteria cooperated generally with the soft rot fungi to degrade the non-acetylated wood or limit themselves to pits at the higher degrees of the acetylation.

Thermogravimetric analysis of fungus-degraded lime wood

Lime wood samples decayed under the attack of the soft rot fungus Chaetomium globosum, for different durations, have been obtained. The degree of decay was determined by mass loss, which was of 50.4% after 133 days. The samples were analyzed by thermogravimetry. The degradative modification of the wood due to presence of the fungus was evidenced by structural changes, also affecting thermal properties both with respect to wood hydrophilicity (evidenced mainly in the desorption process) and decomposition behavior. The increased characteristic temperatures for water desorption and decreased temperatures for decomposition processes, as well as the lower thermal stability, could be explained by the formation of reactive species due to oxidation reactions induced by enzymes and the demethoxylation of lignin structure.

Enzyme Production by Wood-Rot and Soft-Rot Fungi Cultivated on Corn Fiber Followed by Simultaneous Saccharification and Fermentation

This research aims at developing a biorefinery platform to convert lignocellulosic corn fiber into fermentable sugars at a moderate temperature (37 °C) with minimal use of chemicals. White-rot (Phanerochaete chrysosporium), brown-rot (Gloeophyllum trabeum), and soft-rot (Trichoderma reesei) fungi were used for in situ enzyme production to hydrolyze cellulosic and hemicellulosic components of corn fiber into fermentable sugars. Solid-substrate fermentation of corn fiber by either white- or brown-rot fungi followed by simultaneous saccharification and fermentation (SSF) with coculture of Saccharomyces cerevisiae has shown a possibility of enhancing wood rot saccharification of corn fiber for ethanol fermentation. The laboratory-scale fungal saccharification and fermentation process incorporated in situ cellulolytic enzyme induction, which enhanced overall enzymatic hydrolysis of hemi/cellulose components of corn fiber into simple sugars (mono-, di-, and trisaccharides). The yeast fermentation of the hydrolyzate yielded 7.8, 8.6, and 4.9 g ethanol per 100 g corn fiber when saccharified with the white-, brown-, and soft-rot fungi, respectively. The highest ethanol yield (8.6 g ethanol per 100 g initial corn fiber) is equivalent to 35% of the theoretical ethanol yield from starch and cellulose in corn fiber. This research has significant commercial potential to increase net ethanol production per bushel of corn through the utilization of corn fiber. There is also a great research opportunity to evaluate the remaining biomass residue (enriched with fungal protein) as animal feed.

The effectiveness of a biological treatment with Rhizopus oryzae and of a photo-Fenton oxidation in the mitigation of toxicity of a bleached kraft pulp mill effluent

Huge efforts have been made both in adopting more environmental-friendly bleaching processes, and in developing advanced oxidation processes and more effective biological treatments for the reduction of deleterious impacts of paper mill effluents. Even so, the success of such treatments is frequently reported in terms of chemical parameters without a proper evaluation of the effluent's toxicity mitigation. This is the first study reporting an exhaustive evaluation of the toxicity of a secondary bleached kraft pulp mill effluent, after either tertiary treatment with the soft-rot fungi Rhizopus oryzae or with a photo-Fenton oxidation, using a battery of freshwater species. As it has been reported the photo-Fenton/UV treatment has proved to be the most effective in reducing the colour and the COD (chemical oxygen demand) of the effluent. Nevertheless, extremely low EC 50 values were reported for almost all species, after this tertiary treatment. The treatment with R. oryzae was less effective in terms of colour removal and COD reduction, but proved to be the most promising in reducing toxicity.

Fungal colonization on excavated prehistoric wood: Implications for in-situ display

Excavations at the Neolithic settlement of Dispilio, Greece, have revealed significant amounts of waterlogged wood, some of which is being considered for in-situ display. This study investigated the presence of fungi and their biodeterioration patterns on the excavated material. Data will be used to assess the current threat from degradation and to aid in the design of appropriate control measures for any future display strategy. Fungi were isolated using different culture media and species were identified from slide cultures using light and fluorescence microscopy. Wood micro-morphology was examined using scanning electron and light microscopy. Seventeen fungal species were identified, all of which are typical terrestrial species, suggesting that they have developed in the exposed, post-excavation environment, rather than under waterlogged burial conditions. According to the literature many of the isolated species are potential wood deteriogens. In addition to fungi, abundant bacterial growth was observed in most samples. Microscopic examination of wood cells showed patterns of decay associated with soft-rot fungi, and tunnelling and erosion bacteria, suggesting past attack related to the waterlogged burial environment. The results obtained indicated fungal colonization of exposed timbers, presenting a potential threat to their long-term survival. Any in-situ display strategy, such as continuous or periodic water-spraying of the excavated wood, should include measures to monitor and control fungal growth.

Biological treatment of the effluent from a bleached kraft pulp mill using basidiomycete and zygomycete fungi

Three white-rot fungi ( Pleurotus sajor caju, Trametes versicolor and Phanerochaete chrysosporium) and one soft-rot fungi ( Rhizopus oryzae) species confirmed their potential for future applications in the biological treatment of effluents derived from the secondary treatment of a bleached kraft pulp mill processing Eucalyptus globulus. Among the four species P. sajor caju and R. oryzae were the most effective in the biodegradation of organic compounds present in the effluent, being responsible for the reduction of relative absorbance (25–46% at 250 nm and 72–74% at 465 nm) and of chemical oxygen demand levels (74 to 81%) after 10 days of incubation. Laccase (Lac), lignin (Lip) and manganese peroxidases (MnP) expression varied among fungal species, where Lac and LiP activities were correlated with the degradation of organic compounds in the effluent treated with P. sajor caju. The first two axes of a principal component analysis explained 88.9% of the total variation among sub-samples treated with the four fungus species, after different incubation periods. All the variables measured contributed positively to the first component except for the MnP enzyme activity which was the only variable contributing negatively to the first component. Absorbances at 465 nm, LiP and Lac enzyme activities were the variables with more weight on the second component. P. sajor caju revealed to be the only species able to perform the biological treatment without promoting an increment in the toxicity of the effluent to the Vibrio fischeri, as it was assessed by the Microtox® assay. The opposite was recorded for the treatments with the other three species of fungus. EC 50–5 min values ranging between 28 and 57% (effluent concentrations) were recorded even after 10 to 13 days of treatment with P. chrysosporium, R. oryzae or with T. versicolor.

Fungal diversity and deterioration in mummified woods from the ad Astra Ice Cap region in the Canadian High Arctic

Non-permineralized or mummified ancient wood found within proglacial soil near the ad Astra Ice Cap (81°N, 76°W), Ellesmere Island, Canada was investigated to ascertain the identification of the trees, current morphological and chemical characteristics of the woods and the fungi within them. These woods, identified as Betula, Larix, Picea and Pinus, were found with varying states of physical and chemical degradation. Modern microbial decomposition caused by soft rot fungi was evident and rDNA sequencing of fungi obtained from the samples revealed several species including Cadophora sp., Exophiala sp., Phialocephala sp., as well as others. Analytical 13C-labeled tetramethylammonium hydroxide thermochemolysis showed the lignin from the ancient wood was in a high degree of preservation with minor side chain alteration and little to no demethylation or ring hydroxylation. The exposure of these ancient woods to the young soils, where woody debris is not usually prevalent, provides carbon and nutrients into the polar environment that are captured and utilized by unique decay fungi at this Arctic site.

Host responses in the xylem of trees after inoculation with six wood-decay fungi differing in invasivenessThis article is one of a collection of papers based on a presentation from the Stem and Shoot Fungal Pathogens and Parasitic Plants: the Values of Biological Diversity session of the XXII International Union of Forestry Research Organization World Congress meeting held in Brisbane, Queensland,

Host responses, i.e., formation of reaction and barrier zones, were studied in the xylem of Douglas-fir, beech, oak, and sycamore trees, after wounding and artificial inoculation with brown-, soft-, and white-rot fungi. The objective of this study was to determine whether strongly invasive wood-decay fungi trigger a higher magnitude of host response than weakly invasive fungi. Differences in active host response, observed microscopically, depended on wood anatomy. Restriction of discoloration and decay by reaction zones was primarily influenced by the content and distribution of parenchyma cells within the sapwood of each host. By contrast, barrier-zone anatomy showed similarities to the basic xylem structure of each host, except for some cell types that were either reduced in number or absent. Regardless of the decay fungus inoculated, individual trees of each host responded differently. With the exception of beech trees inoculated with the soft-rot fungus Kretzschmaria deusta (Hoffm.: Fr.) P. Martin and the white-rot fungus Trametes versicolor (L.: Fries) Pilát, host response appeared to be nonspecific, as the degree of fungal invasiveness did not influence the magnitude of host response within the xylem of investigated trees.

Action of fungal β-glucosidase on the mono-O-methylated p-nitrophenyl β-d-glucopyranoside 2nd ICC 2007, Tokyo, Japan, October 25–29, 2007

Abstract The action of β-glucosidase on the methyl derivatives of p-nitrophenyl β-d-glucopyranoside (pNP-β-Glc), which were regio-specifically substituted at O-2, O-3, O-4, and O-6 positions, was studied. Specifically, several β-glucosidases isolated from brown-rot, white-rot, soft-rot fungi, and almond were investigated. These β-glucosidases did not act on the 2, 3, and 4-O-methyl derivatives, while the 6-O-methyl one was hydrolyzed by all the enzymes to some extent. The results indicate that the methyl group at O-2, O-3, and O-4 of the glucopyranoside strongly inhibits the recognition by the β-glucopyranoside, while the enzymes do not discriminate the structure difference between pNP-β-Glc and its methyl derivative at O-6.

Lignin degradation in wood-feeding insects.

The aromatic polymer lignin protects plants from most forms of microbial attack. Despite the fact that a significant fraction of all lignocellulose degraded passes through arthropod guts, the fate of lignin in these systems is not known. Using tetramethylammonium hydroxide thermochemolysis, we show lignin degradation by two insect species, the Asian longhorned beetle (Anoplophora glabripennis) and the Pacific dampwood termite (Zootermopsis angusticollis). In both the beetle and termite, significant levels of propyl side-chain oxidation (depolymerization) and demethylation of ring methoxyl groups is detected; for the termite, ring hydroxylation is also observed. In addition, culture-independent fungal gut community analysis of A. glabripennis identified a single species of fungus in the Fusarium solani/Nectria haematococca species complex. This is a soft-rot fungus that may be contributing to wood degradation. These results transform our understanding of lignin degradation by wood-feeding insects.

Resistance of Trichoderma harzianum to the biocide tebuconazol – Proposed biodegradation pathways

Abstract Wood preservatives are moving from metal-based to totally organic chemical systems. A major problem with organic biocides is biodegradation by wood-inhabiting but non-decaying microorganisms during the long service expected from treated wood. While data for triazoles suggest good efficacy against a wide range of basidiomycete fungi, this class of preservatives are less effective against the ascomycetes (soft-rot fungi, molds, and stains). The objective of this study was to examine the biotransformation of tebuconazol as one of the triazoles by the mold Trichoderma harzianum. After inoculation for 21 days, 68% of the tebuconazol was degraded in vitro. MS and IR analysis of isolated metabolites indicated that the major pathway is cleavage of the triazole ring on tebuconazol. Additionally, oxidation reactions occurred on the tert-butyl moiety. The oxidation products are subsequently metabolized by acetylation. Biotransformation resulting in non-toxic degradates is one of the mechanisms employed by this fungus in tolerating tebuconazol.

Suitability of Eucalyptus hybrid, Melia azedarach and Mangifera indica treated with CCA, ACA and copper lignate in cooling tower

Eucalyptus hybrid, Mangifera indica and Melia azedarach were treated with different concentrations of CCA, ACA and Copper lignate. Pinus roxburghii (Pine) was also treated with the same preservatives and used as reference species. Two experiments for different periods i.e. 4 and 5 years were conducted in the same cooling tower. Observations in terms of weight loss were taken. It was found that Melia azedarach (Melia), Pinus roxburghii and Eucalyptus hybrid (Eucalyptus) performed excellently when treated with CCA at 16 kg/m3 retention throughout the period of investigation. While ACA (16 kg/m3) and CCA (i) (consisting As2O3.2H2O) treated samples were protected considerably but were not comparable with CCA treated samples. Copper lignate (1.0, 1.5 and 2.0 kg/m3) treated samples (10 x 2.5 x 0.6 cm3) of all species, could not perform satisfactorily. Visual observations of Mangifera indica (Mango) samples indicated heavy attack of soft rot fungus on the sample in the 3rd year of experiment itself, thus indicating none of the preservative treatment being able to protect Mango wood samples. Thus, Eucalyptus hybrid (heartwood) and Melia azedarach treated with CCA can be recommended for smaller size samples used in cooling towers whereas for bigger size of wood used in cooling towers, where strength is important criteria, bigger samples i.e. stake test (45 x 5 x 4 cm3) is required.

A family 51 α- l-arabinofuranosidase from Penicillium purpurogenum: purification, properties and amino acid sequence

The soft rot fungus Penicillium purpurogenum secretes a wide variety of xylanolytic enzymes to the medium, among them three α- l-arabinofuranosidases. This work refers to arabinofuranosidase 2 (ABF 2). This enzyme was purified to homogeneity and characterized; it is a glycosylated monomer with a molecular weight of 70 000 and an isoelectric point of 5.3. When assayed with p-nitrophenyl α- l-arabinofuranoside ( pNPAra) the enzyme followed Michaelis–Menten kinetics with a K M of 0.098 m m. The optimum pH is 5 and the optimal temperature 60 °C. ABF 2 showed weak activity on natural polymeric substrates, such as sugar beet arabinan, debranched arabinan, and arabinoxylan. These results, together with its low K M ( pNPAra) and its activity towards short arabinooligosaccharides, suggest that the enzyme belongs to the exo α- l-arabinosyl hydrolases not active on polymers. The abf2 gene and its cDNA were sequenced, and the gene was found to possess seven introns. The mature protein is 618 amino acids long with a calculated molecular weight of 67 212. Amino acid sequence alignments show that the enzyme belongs to family 51 of the glycosyl hydrolases, although it differs in some properties from other enzymes of this family.

Decay development in living sapwood of coniferous and deciduous trees inoculated with six wood decay fungi

Development of decay and/or discoloration was assessed in the functional sapwood of one coniferous and three deciduous trees after wounding and artificial inoculation with six wood decay fungi. Living stems of mature Douglas fir, beech, oak, and sycamore trees were wounded in spring 2002 and immediately inoculated with brown, white, and soft rot fungi. Extent of discoloration and decay, wood weight loss, and total phenols in the reaction zone (zone of active response at a dynamic interface between living sapwood and wood colonized by decay fungi) were assessed 16 and 28 months after inoculation. In all hosts, extent of discoloration and decay was greater in the axial than in the radial and tangential directions. After both sampling times, less sapwood had become dysfunctional, and there were greater total phenols in the reaction zone of Douglas fir and oak than in beech and sycamore. Moreover, there was an increase rather than a reduction in dry weight in Douglas fir and oak wood samples. The white rot fungi Ganoderma adspersum, Ganoderma applanatum, and Ganoderma resinaceum were weakly invasive, i.e. behaved as saprotrophs in Douglas fir and oak. By contrast, in beech and sycamore the white rot fungus Trametes versicolor and the soft rot fungus Kretzschmaria deusta (ex Ustulina deusta) were strongly invasive 28 months after inoculation. After this time, both fungal species had induced the highest axial, radial, and tangential extent of dysfunctional wood in beech and sycamore, were capable of defeating the barrier zone (structurally homogeneous barrier comprising cells laid down by the cambium after wounding as response to damage), colonized the sapwood formed after wounding, and can therefore be classified as facultative parasites.

Identification of white-rot and soft-rot fungi increasing ethanol production from spent sulfite liquor in co-culture withSaccharomyces cerevisiae

To identify fungi that are capable of increasing ethanol production from lignocellulose in spent sulfite liquor. In a batch fermentation study, the fungal mix could produce 24.61 g l(-1) ethanol using spent sulfite liquor as substrate. The fungal mix grew well on glucose, xylose, hemicellulose and cellulose. In addition, we were able to identify the fungal mix by use of PCR-amplification of DNA and sequencing, and they were identified as Chalara parvispora and Trametes hirsuta/T. versicolor. In a reconstitution study, the identified fungi were shown to produce equal amount of ethanol as the fungal mix. We were also able to show that C. parvispora could produce ethanol from xylose. The present study has shown that ethanol production from biomass can be increased by use of C. parvispora and T. versicolor when compared with fermentation using only S. cerevisiae. The study shows that refining biomass by ethanol production from spent sulfite liquor, a lignocellulose material, can be increased by adding C. parvispora and T. versicolor, and it is thus of great potential economical impact.

Degradation of thermo-hygro-mechanically (THM)-densified wood by soft-rot fungi

Abstract Thermo-hygro-mechanical (THM)-densified wood is more resistant to colonisation and degradation by brown-rot fungi than untreated wood. Colonisation and degradation by soft-rot fungi was investigated in treated Norway spruce (Picea abies) and treated beech (Fagus sylvatica) to assess their suitability for utility class 4. Three different treatments were applied: thermal-hygro (TH) treatment, mechanical densification and THM-treatment including densification and post-treatment under saturated steam conditions at different temperatures. For comparison, additional wood specimens were treated with two concentrations of a chromium-copper (CC) wood preservative. After 32 weeks incubation, weight losses induced by soft-rot fungi were lowest in wood treated with CC. Highest weight losses were recorded from TH-treated wood, in which soft-rot erosion attack (type 2) was exclusively observed in spruce. In comparison to controls, significantly lower weight losses by soft-rot fungi were recorded in THM-treated spruce wood, but no such differences were found in beech wood. Microscopical examination showed that in THM-treated wood of spruce, soft-rot type 1 commenced from the outer wood surfaces and cavity formation was not found in deeper regions of the wood samples. THM-treated beech wood was more susceptible to degradation than that of spruce which can be partly explained by the higher syringyl lignin content in beech wood, which is more susceptible to all kinds of degradation. Hyphal colonisation and soft-rot was facilitated within deeper regions of beech wood mainly in the non-occluded lumina of parenchyma cells in multiseriate xylem rays. It can be concluded that TH-treated spruce wood and THM-treated beech wood is susceptible to soft-rot and therefore inappropriate for utility class 4.

Fundamental Differences Between two Fiber Types in Acer

Our earlier studies on Acer have shown morphological and chemical differences between two fiber types. Our new research applies the soft-rot cavity method using the soft-rot fungus Chaetomium globosum to measure the microfibril angle (MFA) in these two fiber types in fifteen species of Acer (maple). Microfibril angles in fiber type 1 were significantly larger than those in fiber type 2. The greatest difference (11.6°) was noted in radial sections of Acer floridanum, in which the average MFA (n =100) obtained from soft-rot cavities for fiber type 1 was 23.3 degrees (95 % C.I. ±1.4°) and for fiber type 2 11.7 degrees (95 % C.I. ±1.2°). The second greatest difference (10.0°) was in Acer nigrum, where the average MFA for type 1 was 16.4 degrees (95 % C.I. ±1.07°) and for type 2 6.4 degrees (95 % C.I. ± 0.64°). When transverse sections were examined with polarized light, areas of fiber type 1 were darker (less birefringent) than those of type 2, which indicates that the MFA of the two cell types differs. After four weeks of exposure to C. globosum the fibers of type 1 were more intensely attacked by the fungus than were the fibers of type 2, suggesting a difference in cell wall chemistry. Fluorescence spectra of the two types of fibers support that observation. Differences in MFA supplement the differences in morphology and chemistry and demonstrate that for Acer species the two fiber types are fundamentally different. These two types of fiber differ in the distribution of their pits. In type 1 the pits are concentrated mainly toward the center of the fiber while in type 2 the pits are distributed along the fiber length.

Early soft rot colonization of Scots sapwood pine in above-ground exposure

The early colonization of Scots pine ( Pinus sylvestris L.) sapwood exposed above ground (staple bed) was studied. Two different types of exposures were used, one in an open field and the other in a shaded field. Decay type and degree of degradation due to soft rot, and mass and strength loss of wood were correlated. Fungal species in Scots pine sapwood were identified by sequencing, using the fungal nuclear ribosomal DNA (nrDNA) after 24 months. The most abundant decay type found was soft rot, which also agreed with the mass loss (7–8%). Pine sapwood did not differ significantly between the two sites regarding the average mass loss during the time of exposure. The early colonization of wood by soft rot fungi together with mass loss indicates that this fungal type might be more common in above-ground conditions than recognized earlier.

An eleven amino acid residue deletion expands the substrate specificity of acetyl xylan esterase II (AXE II) from Penicillium purpurogenum

The soft-rot fungus Penicillium purpurogenum secretes to the culture medium a variety of enzymes related to xylan biodegradation, among them three acetyl xylan esterases (AXE I, II and III). AXE II has 207 amino acids; it belongs to family 5 of the carbohydrate esterases and its structure has been determined by X-ray crystallography at 0.9 A resolution (PDB 1G66). The enzyme possesses the alpha/beta hydrolase fold and the catalytic triad typical of serine esterases (Ser90, His187 and Asp175). AXE II can hydrolyze esters of a large variety of alcohols, but it is restricted to short chain fatty acids. An analysis of its three-dimensional structure shows that a loop that covers the active site may be responsible for this strict specificity. Cutinase, an enzyme that hydrolyzes esters of long chain fatty acids and shows a structure similar to AXE II, lacks this loop. In order to generate an AXE II with this broader specificity, the preparation of a mutant lacking residues involving this loop (Gly104 to Ala114) was proposed. A set of molecular simulation experiments based on a comparative model of the mutant enzyme predicted a stable structure. Using site-directed mutagenesis, the loop's residues have been eliminated from the AXE II cDNA. The mutant protein has been expressed in Aspergillus nidulans A722 and Pichia pastoris, and it is active towards a range of fatty acid esters of up to at least 14 carbons. The availability of an esterase with broader specificity may have biotechnological applications for the synthesis of sugar esters.