Callose synthesis as a tool to screen chitosan efficacy in inducing plant resistance to pathogens

Abstract -The effectiveness of chitosans with different molecular weights (MW 6-753 kD) to elicit callose synthesis in Phaseolus vulgaris was evaluated, and correlated with their capability in inducing resistance to tobacco necrosis virus (TNV). To rapidly screen the pattern and amount of callose apposition, leaf fragments were floated in a cell culture multi-wells dish, each well filled with a different chitosan dissolved at variable concentration (0.1-0.2%), Aniline blue staining, performed 12 h after treatment, showed that chitosans with MW of 76,120 and 139 kD were the most effective in inducing callose synthesis in comparison with those having lower or higher MW. Callose appositions were randomly scattered in the mesophyll tissues, forming a homogeneous network of bright fluorescent spots. TNV inoculation of chitosan-treated bean plants showed that the efficacy of chitosans as resistance elicitors positively correlated with their ability in inducing callose apposition, with the 76-kD one being the most effective, with a 95% reduction of viral lesions, Microscopic and ultrastructural alterations in leaf fragments floated on chitosan and inoculated with TNV indicated that the mechanism of induced resistance involves, besides callose, a network of hypersensitive-like reactions, elicited by the compound, that impair virus spreading.