A Target-Site-Specific Screening System for Antifungal Compounds on Appressorium Formation in Magnaporthe grisea

ABSTRACT Chemical fungicides are a major method of control for plant diseases in spite of potential negative effects on the environment and the appearance of resistant strains. Development of new chemical fungicides has been largely dependent upon in vivo efficacy tests in the greenhouse or in fields, which is in contrast to target-oriented in vitro screening systems widely used in the pharmaceutical field. To establish a target-site-specific screening system for antifungal compounds, specific inhibition on appressorium formation of the rice blast fungus Magnaporthe grisea was employed. For many plant-pathogenic fungi, including M. grisea, appressorium formation is an essential step to infect host plants. Among 1,000 culture filtrates of members of the class Actinomycetes and fungi, five (A5005, A5008, A5314, A5387, and A5397) from the class Actinomycetes showed differential inhibitory effects on appressorium formation of M. grisea in a dosage-dependent manner. Three (A5005, A5314, and A5387) of these were further fractionated into ethyl acetate and water fractions. The ethyl acetate fraction of A5005 and both the ethyl acetate and water fractions from A5314 and A5387 inhibited appressorium formation, while conidial germination remained little affected. Inhibition of appressorium formation by the ethyl acetate or water fraction was reversed by the exogenous addition of cyclic AMP. Significantly reduced numbers of conidia with appressoria were observed on rice leaves in the presence of culture filtrates. Furthermore, these culture filtrates also exhibited significant disease control of rice blast in the greenhouse. This rapid and target-oriented screening system could be adopted to screen candidate compounds for rice blast control and could be applicable for other appressorium-forming, plant-pathogenic fungi.