Use of fluorescent-labeled lectins for studying progressive stages of fungal decay in Douglas-fir and ponderosa pine Public Deposited

http://ir.library.oregonstate.edu/concern/graduate_thesis_or_dissertations/cv43p026h

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  • The ability of the fluorescent-coupled lectins wheat germ agglutinin (WGA) and concanavalin A (Con A) to react with selected Basidiomycetes, Ascomycetes, and Fungi Imperfecti was evaluated using pure cultures of thirty-five fungi grown on malt extract agar. WGA, which is specific for residues of N-acetylglucosamine present in fungal chitin, reacted with nearly all hyaline fungal structures, but did not react with dematiaceous (dark) structures. Con A, which is specific for a-D-mannosyl and a-D-glucosyl residues, reacted with about one half of the fungi that reacted with WGA. This lectin was less useful for detecting fungal decay. The results indicate that WGA is a highly specific probe for detecting fungal chitin and fungal colonization by non-dematiaceous fungi. The sequence of changes that occurred in wood over progressive stages of fungal decay was also studied using Douglas-fir heartwood [Pseudotsuga menziesii (Mirb.) Franco] and ponderosa pine sapwood (Pinus ponderosa Dougl. ex Laws.). The fungi used were Poria carbonica Overh., a brown-rot fungus, Coriolus versicolor (L. ex Fr.) Quel., a white-rot fungus, and Chaetomium globosum Kunze ex Fr., a soft-rot fungus. Incident light and fluorescence microscopy were used to observe wood sections. Poria carbonica and Coriolus versicolor caused high weight losses at the early stages of exposure, while Chaetomium globosum caused lower weight losses. As expected, weight losses were generally greater in pine blocks. Poria carbonica displayed a greater tendency to attack ponderosa pine than Douglas-fir. The cellulolytic enzymes of the brown-rot fungus in ponderosa pine were capable of penetrating and acting within the cell walls at early stages of decay, while attack of Douglas-fir was concentrated on the more readily accessible ray parenchyma. The attack patterns found with Coriolus versicolor were similar to those found with P. carbonica on Douglas-fir, but bore holes were less common in the advanced stages of white rot attack. The hyphae of C. versicolor were widespread in ponderosa pine in early stages of decay. Passive penetration via the pits was prevalent in the early stages of decay (<4.35%), while active penetration through the cell walls was prevalent in the advanced stages of decay. Chaetomium globosum lacked the ability to cause soft rot damage (Type I or 2) in Douglas-fir blocks and failed to produce soft rot cavities (Type I) in ponderosa pine blocks. The soft rot fungus preferentially invaded the ray cells of both wood species, then grew into the lumens of the adjacent tracheids. FITC-WGA improved hyphal visualization when compared with results obtained using a conventional safranin-O/picro-aniline blue method. Double-staining, combining FITC-WGA and safranin-O stains, improved lectin contrast, increased the visibility of hyaline hyphae, and permitted more detailed examination of fungal colonization.
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