Histological changes in the cell wall structure during wood decay by Trametes hirsuta and Trametes versicolor in neem (Azadirachta Indica A. Juss)

ABSTRACT

Structural alterations in the wood cell walls of neem inoculated with by Trametes hirsuta and T. versicolor were studied by microscopic methods. In vitro decayed wood showed extensive weight loss of test blocks (26.7 and 41.38% by T. versicolor and T. hirsuta, respectively) at the end of 3 months. Selective delignification in the initial phase followed by simultaneous removal of lignin was evident in test blocks inoculated with both the species. The separation of middle lamellae and patches of cellulosic polysaccharides stained blue with Astra blue in the delignified region of the fiber wall during early stages indicates selective mode of decay. In contrast, the occurrence of erosion troughs with characteristic U-notch in tangential sections is a characteristic feature of simultaneous rot that was apparent after 3 months of incubation. The decay pattern occurred concomitantly in all the xylem elements irrespective of general resistance pattern shown by vessel and axial parenchyma cells. At an advance stage, both species of Trametes showed formation of erosion channels along the microfibrils angle of cellulose which is considered as characteristics of soft rot decay type. The sharing of white rot and soft rot decay pattern by both the fungi suggest a phylogenetic link between both groups of fungi.     

The Delignification Pattern of Ailanthus excelsa Wood by Inonotus hispidus (Bull.: Fr.) P. Karst.

In vitro laboratory decay tests on Ailanthus excelsa Roxb. wood revealed that I. hispidus exhibits a combination of both white-rot and soft-rot patterns of wood decay. Early stages of wood decay showed dissolution of the middle lamella as well as defibration and localized delignification of fiber walls; vessels, axial, and ray parenchyma remained unaltered. Delignification commenced from the middle lamellae at the cell corners without any marked effect on the primary and secondary wall layers. In later stages of growth, the species produces typical soft-rot decay pattern by forming erosion channels through the S₂ layers of fiber walls, transverse bore holes in the cell walls, and erosion channels alongside/following the orientation of cellulose microfibrils. The rays showed signs of cell wall alterations only after the extensive damage to the fiber walls. After 120 days of incubation, the vessels also showed localized delignification, the erosion of pits, and separation from associated xylem elements. The extensive weight losses under natural and in vitro decayed wood as well as the very soft nature of severely degraded wood indicate that I. hispidus alters wood strength and stiffness.     

A histological investigation of Oriental beech wood decayed by Pleurotus ostreatus and Trametes versicolor

Chemical, light and electron microscopic studies were carried out on wood of Oriental beech (Fagus orientalis Lipsky) decayed by the white‐rot fungi Pleurotus ostreatus and Trametes versicolor for 30, 60 and 120 days according to the modified European standard EN 113. Mass loss as well as lignin, cellulose and carbohydrate content were determined before and after fungal attack. There were no significant differences of wood mass loss and chemical composition between both fungi at the end of incubation. After each incubation period, small specimens were stained for microscopic studies. The micromorphology of fungal cell wall degradation was rather similar for both fungi. Both decreased the cell wall thickness to the same extent. The accumulation of hyphae as well as the rupture of cell walls was also similar. The occurrence of hyphae, cavities in the pits and vessel walls followed nearly the same patterns. The parenchyma cells were completely destroyed. Altogether, both fungi produced a simultaneous white rot in Oriental beech wood.     

Effect of heat treatment on extracellular enzymatic activities involved in beech wood degradation by Trametes versicolor

Effect of heat treatment on extracellular enzymes involved in wood degradation by Trametes versicolor was investigated. Heat-treated and untreated beech blocks were exposed to T. versicolor on malt-agar medium and extracellular enzymatic activities investigated. A strong ABTS oxidizing activity has been detected during the first stage of colonization in both cases, while cellulase activities are mainly detected in the case of untreated beech wood. Further investigations carried out on holocellulose, isolated using sodium chlorite delignification procedure and subjected to heat treatment or not, indicate that commercially available cellulases and xylanases are able to hydrolyse untreated holocellulose, while heat-treated holocellulose was not affected. All these data suggest that chemical modifications of wood components during heat treatment disturb enzymatic system involved in wood degradation.     

Degradation of lipophilic wood extractive constituents in Pinus sylvestris by the white-rot fungi Bjerkandera sp. and Trametes versicolor

The white-rot fungi Trametes versicolor and Bjerkandera spp. are among the most frequent decomposers of angiosperm wood in forest ecosystems and in wood products in service. Wood extractives have a major impact on wood properties and wood utilization. This work evaluated the ability of two white-rot fungal strains (Bjerkandera sp. strain BOS55 and T. versicolor strain LaVec94-6) to degrade the main lipophilic extractive constituents in Scots pine (Pinus sylvestris L.). The time course of wood decay and wood extractive degradation was monitored in stationary batch assays incubated for eight weeks. The strains tested eliminated high levels of total resin, 34 to 51% in two weeks. Wood triglycerides were the most readily degraded extractive components (over 93% elimination in only two weeks). Free fatty acids and resin acids, which are potential fungal inhibitors, were also rapidly decomposed by the fungal strains. Sterols were used more slowly, nonetheless, the fungal degradation of this extractive fraction ranged from 50 to 88% after four weeks. Received 19 March 1999     

Micromorphological characteristics of decayed wood and laccase produced by the brown-rot fungus <Emphasis Type="Italic">Coniophora puteana</Emphasis>

Microscopic examination showed the cell wall decay pattern produced by the brown-rot fungus Coniophora puteana to be different from the degradation pattern known to be typical for brown-rot fungi. Erosion and thinning of cell walls in patterns considered to be characteristic of white-rot decay were observed. In particular, the fungal strain COP 20242 degraded secondary cell wall layers extensively, and also degraded lignin-rich middle lamellae. Some strains of C. puteana produced soft-rot type cavities in the S2 layer. All strains of C. puteana employed in the present work showed a positive reaction to tannic acid in the Bavendamm test, indicating the production of laccase. Microscopic and enzymatic studies provided evidence to suggest that the wood decay by C. puteana is unique both in terms of micromorphological and enzymatic patterns of cell wall degradation. This is because brown-rot fungi are not generally known to form cavities in the cell walls or to produce lignin-degrading enzymes. These observations suggest that lignin degradation capacity of brown-rot fungi may be greater than previously considered.     

Non-destructive monitoring of early stages of white rot by Trametes versicolor in Fraxinus excelsior

? Non-destructive detection of fungal decay in living trees is relevant for forest management of valuable species, hazard tree assessment, and research in forest pathology. A variety of tomographic methods, based on stress wave timing, radioactive radiation, or electrical resistivity have been used to detect decay in standing trees non-destructively. But apart from mobile gamma ray computed tomography (Habermehl and Ridder, 1993) which is virtually unavailable, the detection of incipient stages of decay is still not possible.

? Wood moisture and electrolyte content influence the electric resistivity of wood. Both are changed by fungal decay. Therefore electric resistivity tomography (ERT) should detect decay in its early stages. Then it could be used to monitor the spatial and temporal progress of degradation.

? We infected four Fraxinus excelsior trees with Trametes versicolor using wooden dowels and measured two-dimensional electric resistivity tomograms 3, 10, 13 and 21 months after infection. Immediately after the last electric resistivity measurement trees were felled for further analyses of stem cross-sections. Wood moisture content and raw density had significantly increased in infected areas, but dry density had not significantly changed after 21 months. Areas of very low electric resistivity around the infected wounds correlated very well with infected wood in the stem cross-sections. Increasing areas of low electric resistivity around the infected wounds during consecutive measurements indicate increasing areas of infected wood.

? We conclude that the growth of white rot by Trametes versicolor can be monitored with electric resistivity tomography (ERT) beginning from incipient stages, even before wood density decreases. ERT could therefore be a powerful research tool for decay dynamics as well as a method for diagnosing wood decay in forestry and arboriculture.

     

Identification of wood‐decay fungi and assessment of damage in log depots of Western Black Sea Region (Turkey)

The aim of this study was to determine and quantify the wood‐decay fungi found on logs of forest tree species (beech, oak, hornbeam, Scots pine and fir) stored in log depots located in six different provinces in the Western Black Sea Region of Turkey. Additionally, it was aimed to determine the natural durability of some important wood species against the most commonly detected wood‐decay fungi. Eighteen families, 31 genera and 45 species belonging to the division Basidiomycota were detected; Antrodia crassa was identified for the first time in Turkey. The abundance of Panus neostrigosus, Polyporus meridionalis, Trametes hirsuta, T. versicolor and Stereum hirsutumincreased significantly with the holding time of the logs (r = 0.99, 0.87, 0.53, 0.57 and 0.78, respectively, p < 0.05). The majority of the fungal species were detected on logs stored in depots for 4–6 years (66%). The percentage of fungal species found on the logs with a holding time of three years or less was 29%, whereas the percentage for those detected on logs stored for seven or more years was 31%. Among the wood species, the greatest number of fungal species (29) and highest amount of fungi (2,539) occurred on beech wood. Natural durability tests showed that T. versicolor caused the greatest loss of wood mass, with an average of 23%. Field studies and natural durability tests performed in the laboratory showed that beech wood lost the most mass among the timber species studied.     

Detection of incipient decay in tree stems with sonic tomography after wounding and fungal inoculation

Picus^® acoustic tomography was used to map incipient stages of fungal decay in the sapwood of standing Douglas fir, beech, oak, and sycamore trees 2, 16, and 27 months after wounding and artificial inoculation with brown-, soft-, and white-rot decay fungi. Some wood properties were additionally measured before (velocity of sound) and after (moisture content, weight loss, and density of sound, discoloured and/or decayed wood) tree felling (28 months). With the exception of Trametes versicolor in sycamore, wood decay was not evident from the tomograms in any host-fungus combination. In comparison to measurements after two months, the device recorded a reduction in sound velocity in some host-fungus combinations after 16 and 27 months. In beech, there was a significant reduction in sound velocity after inoculation with Ganoderma applanatum, Kretzschmaria deusta, and Trametes versicolor. Similarly, a reduction in sound velocity was recorded in sycamore inoculated with Kretzschmaria deusta and Trametes versicolor. In all these combinations, losses in wood weight and wood density were also found. Results showed that the detection of incipient fungal decay at the periphery of tree stems needs to be improved such that tomograms of the Picus^® acoustic tomograph are capable of identifying decay progressing from the sapwood inwards.     

Decline of Paulownia tomentosa caused by Trametes hirsuta in Serbia

During the monitoring of the health status of nurseries and plantations in Serbia, a decline in a 5‐year‐old Paulownia tomentosa plantation was recorded. Trees displayed symptoms of dieback, massive breaking at different positions, the appearance of decay and fungal fruitbodies at the stems. Using standard isolation methods, white colonies with cottony surfaces and regular growth were obtained from the decaying wood samples, and after the oxidation degree analysis, it was determined that these isolates belonged to Davidson's group 6, indicating white rot basidiomycetes. To identify the isolated fungus, the ITS region of one selected isolate was sequenced. Based on the morphological analysis of the obtained colonies, collected fruitbodies and ITS sequence analysis, this fungus was identified as Trametes hirsuta. This is the first report of T. hirsuta on Paulownia tomentosa in Serbia. Irregularly performed pruning as the possible infection route and the implications of these findings are discussed.     

Degradation of endosulfan and endosulfan sulfate by white-rot fungus <Emphasis Type="Italic">Trametes hirsuta</Emphasis>

Endosulfan, an organochlorine insecticide, and its metabolite endosulfan sulfate are persistent in environments and are considered toxic. We investigate the possible nontoxic bioremediation of endosulfan. An endosulfandegrading fungus that does not produce endosulfan sulfate was selected from eight species of white-rot fungi. High degradation of endosulfan and low accumulation of endosulfan sulfate were found in cultures of Trametes hirsuta. A degradation experiment using endosulfan sulfate as the substrate revealed that T. hirsuta is able to further degrade endosulfan sulfate following the oxidative conversion of endosulfan to endosulfan sulfate. Endosulfan and endosulfan sulfate were converted to several metabolites via hydrolytic pathways. In addition, endosulfan dimethylene, previously reported as a metabolite of the soil bacterium Arthrobacter sp., was detected in T. hirsuta culture containing endosulfan sulfate. Our results suggest that T. hirsuta has multiple pathways for the degradation of endosulfan and endosulfan sulfate and thus has great potential for use as a biocatalyst in endosulfan bioremediation.     

In vitro decay of Aextoxicon punctatum and Fagus sylvatica woods by white and brown-rot fungi

Summary The in vitro decay of Aextoxicon punctatum and Fagus sylvatica wood by the fungi Trametes versicolor, Ganoderma australe, Phlebia chrysocrea and Lentinus cyathiformis was studied by the agar-block method, and then the decayed woods were analyzed by chemical and spectroscopic techniques. The results demonstrated the strong resistance of the A. punctatum wood to the brown-rot fungus L. cyathiformis; the resistance might be related to the low S/G lignin ratio in this Austral hardwood. Wood decay by the Austral white-rot fungi G. australe and P. chrysocrea was rather limited, and preferential degradation of lignin was not produced although all the fungi studied increased wood digestibility. The most characteristic white and brown-rot decay patterns were observed during the in vitro decay with T. versicolor and L. cyathiformis, respectively. Trametes versicolor caused high weight losses and reduced the lignin content of the wood, whereas L. cyathiformis produced a preferential removal of xylan. No important changes in the solid-state ¹³C NMR spectra were observed after wood degradation by T. versicolor, but this technique evidenced an increase in aromatic carbon by L. cyathiformis. This increase was higher than that found in the Klason lignin content, suggesting the presence of altered lignin fractions in the brown-rotted wood.The authors are indebted to Prof. H. D. Lüdemann for the facilities at the Institut für Biophysik und physikalische Biochemie (Regensburg), to A. Navarrete (INIA, Madrid) for her collaboration, and to C. F. Warren (ICE, Alcalá de Henares) for her linguistic assistance. The computer program for spectra treatment was developed by G. Almendros (Centro de Ciencias Medioambientales, CSIC, Madrid). This investigation has been funded by the Spanish Biotechnology Program (Grant BIO88-0185)     

Cellular UV-microspectrophotometric investigations on pine wood ( Pinus taeda and Pinus elliottii) delignification during biopulping with Ceriporiopsis subvermispora (Pilát) Gilbn. & Ryv. and alkaline sulfite/anthraquinone tr

Scanning UV-microspectrophotometry was used to investigate the topochemistry of lignin removal from pine wood (Pinus taeda and P. elliottii) chips during biopulping involving wood treatment with Ceriporiopsis subvermispora (Pilát) Gilbn. & Ryv. followed by alkaline sulfite/anthraquinone delignification. A delignification front starting from the lumen towards the compound middle lamella was clearly observed in micrographs recorded from individual cell wall layers of wood samples biotreated for 30 days. Lignin was removed without cell wall erosion. UV-micrographs of wood samples cooked for a short time (90 min pulping) showed that the S2 of biotreated samples are more homogeneously delignified compared to the S2 of the undecayed controls. Similarly, the compound middle lamella and cell corners are also more delignified in biotreated samples. On the other hand, UV-micrographs of samples cooked for a long time (150 min pulping at 170°C) showed that there are no significant differences in the contents of residual lignin retained in the S2 of undecayed and biotreated wood samples.     

Micromorphological characteristics of bamboo (<Emphasis Type="Italic">Phyllostachys pubescens</Emphasis>) fibers degraded by a brown rot fungus (<Emphasis Type="Italic">Gloeophyllum trabeum</Emphasis>)

The decay pattern in bamboo fibers caused by a brown rot fungus, Gloeophyllum trabeum, was examined by microscopy. The inner part of the polylaminate secondary wall was degraded, while the outer part of the secondary wall remained essentially intact. Degradation in bamboo fiber walls without direct contact with the fungal hyphae was similar to wood decay caused by brown rot fungi. Degradation in polylaminate walls was almost confined to the broad layers whereas the narrow layers appeared resistant. The p-hydroxylphenyl unit lignin in middle lamella, particularly in the cell corner regions, was also degraded. The degradation of lignin in bamboo fibers was evidenced by Fourier transform infrared spectra. The present work suggests that the decay of bamboo fiber walls by G. trabeum was influenced by lignin distribution in the fiber walls as well as the polylaminate structures.     

Anatomical characterization of decayed wood in standing light red meranti and identification of the fungi isolated from the decayed area

To further our understanding of wood decay in living light red meranti (Shorea smithiana) trees, microscopic characteristics of the cell and cell wall degradations of S. smithiana wood in the presence of the decay fungi, the identity of the causal fungi, and the decay potential and pattern by an isolated fungus were investigated. Cell wall degradations, including cell wall thinning, bore holes formation, rounded pit erosion, and eroded channel opening were clearly observed under light and scanning electron microscopy. In transverse view, many large voids resulting from a coalition of degraded wood tissue appeared in the decayed canker zone. All these observations suggest the well-known simultaneous decay pattern caused by white-rot fungi. By phylogenetic analysis based on the sequences of internal transcribed spacer region of ribosomal DNA, a basidiomycete fungus isolated from the decayed wood was identified as Schizophyllum commune. The degradation caused by this fungus on sound S. smithiana wood in an in situ laboratory decay test was classified as the early stage of simultaneous decay, and showed a similar pattern to that observed in the wood samples naturally decayed.     

Comparative study on the durability of heat-treated White Birch (Betula papyrifera) subjected to the attack of brown and white rot fungi

Abstract

The effect of heat treatment on decay resistance of white birch was evaluated for different incubation periods ranging from 2 to 12 weeks using three species of brown rot and one species of white rot fungus. The results of weight loss tests showed that the white rot fungus, Trametes versicolor, effectively degraded the untreated wood (73.5%). While the degradation of untreated wood by brown rot fungi species, Gloephyllum trabeum (11.6%) and Conifora puteana (6.2%), was considerably less compared to T. versicolor, the third brown rot fungi studied, Poria placenta, caused an appreciable degradation of the same species (52.4%). The results clearly showed that the heat treatment reduced the effect of fungi attack on white birch. Increasing the heat treatment temperature from 195 to 215°C resulted in reduction of weight loss, consequently, reduction in fungal attack. As an example, the weight loss reductions due to T. versicolor, P. placenta, G. trabeum and C. puteana attack was 62.2%, 71.3%, 89.6% and 100%, respectively, compared to the weight loss of untreated wood when it is heat treated at 215°C. Thus, these results confirmed that the heat treatment increased the biological resistance of white birch.     

Evaluation of Schizophyllum commune Fr. potential for biodegradation of lignin: A light microscopic analysis

Lignin biodegradation potential of Schizophyllum commune Fr. is studied by using sound wood blocks of Ailanthus excelsa, Azadirachta indica, Tectona grandis, Eucalyptus sp. and Leucaena leucocephala. Initially, in vitro wood decay test showed minor weight loss, but it became rapid after one month. After 120 days of incubation, weight loss was minimum in T. grandis (24.05%) whereas it was maximum in A. excelsa (34.44%). Treated test blocks were characterised by enlargement of pits on ray cell wall, formation of additional boreholes in rays, separation of fibres and cell wall thinning and formation of ‘U’-shape notches. Fungal hyphae moved through the xylem cell lumen, and intercellular spaces formed in response to separation of fibres. Hyphae traverse in adjacent cell through the cell wall pits or by making additional boreholes. In all the species studied, xylem fibres and parenchyma (axial and ray) cells were more susceptible while vessels were resistant to fungal attack. In advanced stage of decay, fibres and axial parenchyma lost their rigidity while vessel walls showed uneven thinning. In the tension wood, G-fibres remained unaffected initially but loosening and separation of gelatinous layer facilitated fungal action and showed similar pattern of cell wall deterioration. Among the wood of different species studied, Tectona was more resistant whereas Ailanthus was more susceptible to fungal attack.     

Changes in structural and chemical components of wood delignified by fungi

Summary Cerrena unicolor, Ganoderma applanatum, Ischnoderma resinosum and Poria medulla-panis were associated with birch wood that had been selectively delignified in the forest. Preferential lignin degradation was not uniformly distributed throughout the decayed wood. A typical white rot causing a simultaneous removal of all cell wall components was also present. In the delignified wood, 95 to 98% of the lignin was removed as well as substantial amounts of hemicelluloses. Scanning and transmission electron microscopy were used to identify the micromorphological and ultrastructural changes that occurred in the cells during degradation. In delignified areas the compound middle lamella was extensively degraded causing a defibration of cells. The secondary wall, especially the S₂ layer, remained relatively unaltered. In simultaneously white-rotted wood all cell wall layers were progressively removed from the lumen toward the middle lamella causing erosion troughs or holes to form. Large voids filled with fungal mycelia resulted from a coalition of degraded areas. Birch wood decayed in laboratory soil-block tests was also intermittently delignified. Selective delignification, sparsely distributed throughout the wood, and a simultaneous rot resulting in the removal of all cell wall components were evident. Scanning electron microscopy appears to be an efficient technique for examining decayed wood for fungi with the capacity to selectively delignify wood.The authors would like to thank Kathy Zuzek for technical assistance and Dr. M. Larsen, Forest Prod. Lab., Madison, for identifying the sporophores of Poria medulla-panis. This research was founded in part by a grant from the USDA Forest Service, Forest Products Laboratory and from the Graduate School, University of Minnesota     

Studies on the delignification of spruce wood by organosolv pulping using SEM-EDXA and TEM

Summary Samples of spruce wood derived from various stages of organosolv pulping were studied by SEM-EDXA and TEM. During the first stage (methanol-water) the lignin content of the secondary walls decreased slowly, whereas in the compound middle lamellae only the reactivity of lignin increased. During the subsequent stage (methanol-NaOH) the delignification proceeded fast in both layers but the residual lignin content in the compound middle lamellae remained higher than in the secondary walls.     

Effect of thermal modification temperature on the mechanical properties,dimensional stability,and biological durability of black spruce (Picea mariana)

This study was aimed at evaluating the effect of thermal modification temperature on the mechanical properties, dimensional stability, and biological durability of Picea mariana. The boards were thermally modified at different temperatures 190, 200 and 210 °C. The results indicated that the thermal modification of wood caused a significant decrease in the modulus of rupture (MOR) after 190 °C, while the modulus of elasticity (MOE) seemed less affected with a slight increase up to 200 °C and slight decrease with further increase in temperature. The hardness of the thermally modified wood increased in the axial direction. This increase was also observed in tangential and axial directions but at a lesser extent. The final value was slightly higher in axial direction and lower in radial and tangential directions compared to those of the untreated wood. Dimensional stability improved with thermal modification in the three directions compared to the dimensional stability of unmodified wood. The fungal degradation results showed that the decay resistance of thermally modified wood against the wood-rotting fungi Trametes versicolor and Gloephyllum trabeum improved compared to that of the untreated wood. By contrast, the thermal modification of P. mariana had a limited effect on the degradation caused by the fungus Poria placenta.