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Distribution of the three symbiotic protozoa in Formosan subterranean termite, Coptotermes formosanus Shiraki (Isoptera: Rhinotermitidae)
1993 - IRG/WP 93-10010
Six colonies (three each from laboratory and field) of Formosan subterranean termite, Coptotermes formosanus Shiraki, were served for investigating the abundance and distribution of three symbiotic protozoa in the hindgut of workers. The total protozoan number amounted to 6,000-10,000 per a worker, and the order of the abundance of the three protozoa and the proportional distribution of each species in the hindgut were common among the colonies. Pseudotrichonympha grassii Koidzumi was the smallest in number (800-2,200 per a worker) and was preferentially distributed in the anterior part of the hindgut. Holomastigotoides hartmanni Koidzumi was medial in number (1,200-3,000), and the distribution was relatively uniform all through the hindgut. Spirotrichonympha leidyi Koidzumi was the most abundant in number (2,800-5,000) and was found mainly in the posterior part. These results appeared to support that the prominent localization of each protozoan species in the worker´s hindgut could be related to the nutritional metabolism in Coptotermes formosanus.
T Yoshimura, K Tsunoda, M Takahashi


Cytochemical localization of hydrogen peroxide in brown rot fungus Tyromyces palustris by cerium chloride technique
1999 - IRG/WP 99-10299
Cerium chloride (CeCl3) was used to localize H2O2 cytochemically for studying relationship between ultrastructural and functional characteristics of cellulose degradation by brown rot fungi. This technique proved very useful in localizing discrete electron-densereactionproducts at high resolution with minimal nonspecific deposition. The cytochemical localization of extracellular H2O2 by CeCl3 using TEM demonstrated the presence of H2O2 within the fungal hyphae. Furthermore, our results give an indication of the diffusion of extarcellular H2O2 from brown-rot decay fungi into the intact wood cell walls in the early stages of decay.
Yoon Soo Kim, Seung-Gon Wi


Analysing the characteristic role of moisture content for drying and fluid flow in Sitka spruce. - Part 1: The drying process of sapwood and heartwood of two different thickness of Sitka spruce using a kiln. - Part 2: Effects of moisture content on longitudinal permeability of Sitka spruce in vertical variation of the tree
2000 - IRG/WP 00-40173
The characteristic role of the moisture content in Sitka spruce (Picea sitchensis (Bong.) Carr.) that grown in the United Kingdom was examined by this study on the basis of (1) the reduction of moisture content in two different thickness of sapwood and heartwood by kiln drying process, and (2) the effects of moisture content to the longitudinal void volume filled of tanalith-C by the full-cell process from base (1 m) to apex (3 m) of the tree in sapwood zone. Accordingly, conclusions on indication of the drying process of sapwood and heartwood, and vertical variation of longitudinal flow with effects of moisture were listed separately: (1) Comparison of Drying Characteristic of Sapwood and Heartwood: The two different thickness (300x30x30 mm3 and 300x20x20 mm3) of sapwood and heartwood of Sitka spruce was dried using the suggested drying schedule in kiln. The reduction of moisture was schematically diagrammed according to sapwood and heartwood stakes. The reduction of moisture followed the same downward trend that sapwood (S) loses more moisture than heartwood (H) although the small stakes of S and H lost moisture rapidly compared with the large ones. (2) Vertical Variation of Moisture Content and Longitudinal Permeability: The 90 kiln dried defect free sapwood stakes (150x25x25 mm3) of Sitka spruce was taken from base to apex of the trees at 1, 2 and 3 m above ground level. After having the determination of moisture content in each experimental stake, the treatment was carried out by the full-cell process with CCA preservative (Tanalith-C) using a model pressure treatment plant. Significant differences observed among the tree heights from 1 to 3 m showing that slightly increases of moisture content from base to apex and conversely decreases of longitudinal void volume filled by preservative fluid.
I Usta


Localization of oxalate decarboxylase in the brown-rot fungus Postia placenta
1996 - IRG/WP 96-10161
Oxalate decarboxylase, the enzyme that breaks oxalic acid down into formic acid and carbon dioxide, was recently detected in mycelial extracts of the brown-rot fungus Postia placenta. Differential centrifugation was used to demonstrate that the enzyme is loosely associated with the hyphal surface. Enzyme activity can be removed by washing the hyphae with a low pH buffer. Only low levels of activity were detected in soluble and membrane-bound intracellular fractions. The presence of the enzyme on the hyphal surface and possibly in the hyphal sheath supports the hypothesis that this brown-rot fungus actively regulates the pH and oxalic acid concentration of its environment.
J A Micales


Oxalic acid quantification, oxaloacetase assay and ESI localization of P, C, and Fe from the brown rot fungus Postia placenta
1994 - IRG/WP 94-10063
The mechanism by which brown-rot fungi initiate depolymerization of holocellulose in wood remains unknown. Recently, oxalic acid (OA) has received considerable attention in cellulose breakdown by brown-rot fungi. The OA could serve as a proton donor for hydrolytic or an electron donor for oxidative (Fenton's reaction-H2O2/Fe2+) cleavages of cellulose. The acid may originate via oxaloacetase's action upon oxaloacetate. We report electron microscopic imaging (ESI) to localize Fe and HPLC/oxalic kit colorimetry to purify/quantify OA from hyphae upon agar, southern pine wood blocks (WBs) or in liquid culture. Comparative ESI at 25, 59, 110, 222, and 710 ev of hyphae grown upon agar or WBs demonstrated hyphal Fe (710 ev). Although Fe was not visualized in cell walls of uninoculated WBs, it was in certain wood cell walls of inoculated WBs. The Fe distribution differed from C and P. Oxaloacetase activity was not observed in either Amicon YM10 filter-retained intra-or extracellular fractions of liquid cultured hyphae or in homogenates from decayed WBs. In contrast, HPLC detected OA in both Postia placenta liquid cultures and decayed WBs. The less sensitive oxalic kit (mg vs. ug) did not detect OA in liquid cultures. These results constitute additional evidence for an OA Fe2+/H2O2-Fenton's mechanism for brown rot-induced cellulose degradation. However, OA's origin was not established.
C R Jordan, W V Dashek, T L Highley


New perspectives on the biology of the tropical powderpost beetle, Minthea rugicollis (Walk.)
1994 - IRG/WP 94-10085
Minthea rugicollis (Walk.) is one of the most important pests of seasoned hardwoods in the tropics. The species owes its ubiquity largely to its insidious development within a nutrient-filled environment and also to a strong coevolutionary specialization with its natural habitat, wood. Such an environment provides a buffer to extrinsic fluctuations and accounts for a wider range of tolerance by immature stages to variations in climatic conditions than would otherwise be possible. Aspects of culture methods, characteristic habits and external tolerances are emphasised so as to generate new perspectives in understanding the fundamental biology of this organism and to improve current wood preservation strategies.
F Abood, R J Murphy, R W Berry


Immuno-electron microscopic localization of extracellular metabolites in spruce wood decayed by brown-rot fungus Postia placenta
1990 - IRG/WP 1441
Degradation by Postia placenta in spruce and birch wood was shown to occur not only in the wood cell wall but also in the middle lamellae region. Middle lamellae was often found to be degraded along the centerline so that cells could separate along this line. Extracellular membrane structures were found surrounding the hyphae and this matrix labelled positively with antisera produced to Postia placenta extracellular metabolites. This matrix was also visible in the secondary wall of degraded birch wood. Antisera labelling was also noted in the secondary cell walls of the wood cells, but not in the middle lamellae region.
Y S Kim, B Goodell, J Jellison


Immunolocalization of extracellular metabolites from Poria placenta
1988 - IRG/WP 1361
Polyclonal antisera produced to Poria placenta extracellular metabolites was used in immuno-fluoresence microscopy and immuno-gold TEM studies. In the fluorescence work, labelling of Poria placenta hyphae in wet fixed wood material was observed but not in infected wood which was oven dried prior to sectioning and immunolabelling. TEM studies provided better resolution, with gold labelling detected in the extracellular slime layer surrounding hyphae. Labelling occurred within the wood cell wall, but little non-specific labelling was noted within the cytoplasm of the fungal hyphae. A vesicle-like body within the hyphae did exhibit specific labelling.
B Goodell, G F Daniel, J Jellison, T Nilsson


Immuno-scanning electron microscopic localization of extracellular polysaccharidases within the fibrillar sheath of the brown rot fungus Postia placenta
1991 - IRG/WP 1497
Extracellular polysaccharidases of the brown-rot fungus Postia placenta were localized using colloidal gold labeled monoclonal antibodies to the B-1,4-xylanase (32-36kDa) fraction of Postia placenta. Postia placenta was grown from agar onto glass coverslips, immunolabeled with or without prior fixation, and examined by SEM. Enzymes were localized on the hyphal surface and on the clumped fibrillar elements (mycofibrils) of the hyphal sheath following fixation with enzymes. If fixation was omitted, labeling was diffuse and not localized on individual or clumped mycofibrils. We conclude that extracellular decay enzymes are weakly bound (non-covalently), but not identical to, the linear mycofibrillar elements of the hyphal sheath. Enzymes appear to dissociate into the water soluble glucan matrix of the sheath during incubation in physiological buffers when fixation is omitted.
F Green III, C A Clausen, M J Larsen, T L Highley


Cytoplasmic and extracellular localization of manganese II dependent peroxidase(s) in white rot fungi during degradation of woody materials
1989 - IRG/WP 1416
The manner by which lignin is degraded in-situ in natural substrates by white rot fungi still remains a controversial issue particularly the distribution and role(s) played by lignin degrading enzymes (i.e. manganese II peroxidase and lignin peroxidase). In the present study, use was made of anti-manganese II peroxidase and immunolabelling techniques in conjunction with transmission electron microscopy (TEM) to study the spatial distribution of manganese II peroxidase during degradation of wood and woody fragments by Phanerochaete chrysosporium and Lentinus edodes. Intracellularly, manganese II peroxidase was found localized in the peripheral regions of the fungal cell cytoplasm in association with both the outer cell membrane and membranes within characteristic vesicular bodies. In addition the enzyme was frequently found localized at the interfacial regions of the cell membrane and inner fungal cell wall. Using double immunolabelling procedures and in addition anti-lignin peroxidase, the cytoplasmic distribution of the two lignin degrading enzymes was compared. Both enzymes showed a fairly similar peripheral cytoplasmic localization although manganese II peroxidase tended to be more concentrated compared to lignin peroxidase in peripheral vesicular bodies. Extracellularly, and in solid wood samples manganese II peroxidase was found localized in all wood cell wall regions of either Betula verrucosa, Populus sp. or Fagus sylvatica decayed by either Phanerochaete chrysosporium or Lentinus edodes at both early and late stages of degradation. In particular, manganese II peroxidase was localized in characteristic zones of degradation produced within the secondary wood cell wall regions. These regions displayed a more open structure compared to unattacked wood cell walls and were easily penetrated by lignin degrading enzymes as judged by infiltration and double immunolabelling studies with highly purified and partially purified manganese II and lignin peroxidases. With Lentinus edodes a very characteristic pattern of lignin degradation was noted in which the middle lamella regions between wood cells was selectively degraded. In these regions manganese II peroxidase was found concentrated and associated with its degrading matrix. An extracellular distribution of manganese II peroxidase associated with wood fragments was also observed in liquid cultures of Phanerochaete chrysosporium grown under conditions optimal for peroxidase production. Despite the immersed conditions, similarities between the patterns of attack and extracellular distribution of the enzyme as for solid wood were noted. With both solid wood and wood fragments, manganese II peroxidase penetration was restricted to regions showing structurally modification, and penetration into undecayed cell walls was not observed. The present work suggests a close substrate-enzyme association during wood cell wall and lignin degradation under natural conditions, and in addition, a close correlation between changes in the micromorphology of decay and manganese II peroxidase distribution. Possible reasons for the failure of previous and similar immunolabelling studies to show such a correlation with lignin degrading enzymes are briefly discussed.
G F Daniel, B Pettersson, T Nilsson, J Volc