Two main antifungal metabolites resistomycin and tetracenomycin D were isolated and purified from a termite-associated Streptomyces canus BYB02 by column chromatography. The structures of isolated compounds were determined on the basis of extensive spectroscopic analysis. Resistomycin possessed strong activities against mycelial growth of Valsa mali (IC(50) = 1.1 μg/mL) and Magnaporthe grisea (IC(50) = 3.8 μg/mL), which were comparable to those of referenced cycloheximide, with IC(50) values of 2.3 and 0.3 μg/mL, respectively. A further spore germination test showed that resistomycin exhibited potent reduction in spore germination for M. grisea , with an IC(50) value of 5.55 μg/mL. Finally, the in vivo antifungal activity experiment showed that resistomycin possessed significant preventive efficacy against rice blast, which was more potent than that of referenced carbendazim, with control efficacies of 66.8 and 58.7%, respectively. The present results suggest that resistomycin has potential to be used as a fungicide.