Observations by electron microscopy of the micromorphological alterations caused in wood by fungal strains in which the cellulase system had been repressed or inhibited showed a substantial destruction of the cellulose network. The question thus arises which agents, other than enzymes, could be responsible for the modification of polysaccharides during fungal attack. Among the small diffusible agents possibly involved, activated oxygen species generated from hydrogen peroxide have been suggested. In particular the powerful oxidizing hydroxyl radical (· OH), which can be generated by reductive cleavage of H2O2 or by reduction of oxygen, has been reported to take part in degradation of lignin by white‐ and brown‐rot fungi. Manganic ions which are generated by manganese‐dependent peroxidase (MnP) represent another diffusible oxidizing agent. To investigate whether these agents, produced by lignin‐degrading enzymes, could also participate in wood polysaccharide degradation, the action of hydroxyl radicals and of some Mn(III)‐organic acid complexes on carbohydrate and polysaccharide model compounds and also on wood and pulps, was studied. The action of ·OH, generated by the iron‐catalyzed Fenton's reagent, on the glucuronoxylan from birch wood resulted in an extensive degradation of the hemicellulose (about 50% loss of carbohydrate after 180 min). A study of the degraded products suggests that internal cleavage occurs in the xylan chain accompanied by sugar ring degradation, the latter becoming predominant on the soluble oligomeric products. In a parallel study, the susceptibility of carbohydrates to oxidation by manganic ions showed that monosaccharide degradation occurred rapidly in strongly acidic medium. The mechanism which was studied with a Mn(III)‐pyrophosphate complex, involves, as an initial step, the oxidative cleavage of C1 with the formation of a lower carbohydrate, and then proceeds by a recurrent series of oxidations. However, the reaction seems to be controlled by the formation of a Mn(III)‐pyrophosphate complex with the sugar, which prevents further oxidation. The two oxidizing agents were applied to birch wood wafers. The hydroxyl radicals had only a relatively weak degradative action on the cell walls. However, when the radicals were generated in situ, the extent of degradation as visualized by electron microscopy, gave images comparable to those obtained on fungal‐degraded wood. Samples treated with manganic pyrophosphate and manganic organic acid complexes also showed typical degradation indicating that lignin was primarily degraded and also that hemi‐celluloses had also been partly damaged.

Polyclonal antibodies were raised against proteinase K and were used to immunolocalize the major extracellular proteinase of the sap-staining fungus Ophiostoma piceae (Munch) H . and P. Sydow. Immunodot blotting showed that the IgG antibodies recognized both enzymes but reacted more strongly with proteinase K than with the 0. piceae proteinase. Immunogold labelling and transmission electron microscopy revealed that the 0. piceae proteinase was localized in the cell walls of 0. piceae grown either in liquid media or wood.

SUMMARYThe growth and proteolytic activity of the sap-staining fungus Ophiostoma piceae were characterized in the wood of four tree species: lodgepole pine {Pinus contorta var. latifolia Dougl.), aspen (Populus treniutoides Michx.), Douglas fir {Pseudotsuga menziesii (Mirb.) Franco) and Western hemlock [Tsuga heterophvlla Sarg.). Proteolytic activity and growth were highest in aspen. Proteolytic activity was minimal in both Douglas fir and Western hemlock. Polyclonal antibodies raised against the major proteinase produced by O.picaea were used to immunolocalize the enzyme after fungal growth in lodgepole pine and aspen. Immunogold labelling revealed that the proteinase was localized mainly in the cell wall of O. piceae. Gold particles concentrated in a sheath which was well defined after 10 d. Labelling appeared greater in the inoculated aspen than in lodgepole pine.