Changes in mycelial structure of Botrytis cinerea induced by removal of the glucan matrix

Aims : β-1,3-glucanase is one of the main pathogenesis related proteins of plants, involved in plant-pathogen interactions. Its effect on fungal pathogens is not entirely known. The hyphae of Botrytis cinerea are covered by an extra cellular matrix, mainly composed of a β-1,3-D-glucan. This matrix also contains a variety of enzymes, lipids and melanin which may play a role in fungal virulence. Methods and results : Cultures of Botrytis cinerea are made in presence of β-1,3-glucanase. The structure of the mycelium of Botrytis cinerea after exposure to β-1,3-glucanase during growth was examined by staining with Schiff's reagent and using the electron microscope. Without glucanase, hyphae have a normal diameter and were surrounded by a glucan matrix. Cytoplasm is dense and contains little vacuoles. The glucanase treatment removed most of the glucan sheath, but did not kill the fungus. The structure of the hyphae was changed by the treatment and their diameter increased. Membrane structure showed marked changes, the cytoplasm of the cells was less dense, but more inclusions were observed, including an increase in what appeared to be lipids. Conclusion : The appearance of the mycelium, whose glucan sheath has been removed, was that of cells under stress. The possible implications of the function of the glucan sheath during the interaction of Botrytis cinerea with its host during pathogenesis are discussed. Significance and impact of study : These changes following glucanase treatment would lead to a fungal mycelium which will be more sensitive to antifungal agents and might suggest ways of combating Botrytis infections by preventing the formation of the extra-cellular matrix.