This work aimed to evaluate in vitro antagonistic activities and mechanisms of endophytic yeasts against phytopathogenic fungi. A total of 407 yeast strains isolated from tissue of rice, corn, and sugarcane leaves were evaluated for their antagonistic activities against ten phytopathogenic fungi. Only strains of Wickerhamomyces anomalus and Kodamaea ohmeri were found to inhibit the growth of phytopathogenic fungi. Wickerhamomyces anomalus (seven strains) showed antagonistic activity against Curvularia lunata (cause of dirty panicle disease of rice), three Fusarium moniliforme strains (cause of bakanae disease of rice, stalk rot disease of corn, and red rot disease of sugarcane), and Rhizoctonia solani (cause of sheath blight disease of rice). Whereas four Kodamae ohmeri strains inhibited growth of F. moniliforme (cause of bakanae disease of rice). Antagonistic mechanisms of W. anomalus were based on the production of volatile organic compounds (VOCs) (mainly 3-methyl-1-butyl acetate and 3-methyl-1-butanol), fungal cell wall-degrading enzymes (β-1,3-glucanase and chitinase), and siderophores as well as phosphate and zinc oxide solubilization. As for K. ohmeri, the production of VOCs (mainly 3-methyl-1-butanol), β-1,3-glucanase and chitinase were hypothesized as possible mechanisms. The antagonistic activity of W. anomalus against these phytopathogenic fungi and of K. ohmeri against F. moniliforme, and the analysis of the VOCs produced by K. ohmeri are reported for the first time. Two W. anomalus strains, DMKU-RE13 and DMKU-CE52, were evaluated for controlling rice sheath blight disease caused by R. solani in rice plants in the greenhouse and found to suppress the disease 55.2-65.1%, whereas 3% validamycin suppressed this disease by 88.5%.