Chestnut blight disease caused by Cryphonectria parasitica is widely distributed throughout chestnut tree plantations in Korea. We surveyed 65 sites located at 9 provinces in South Korea, and isolated 248 virulent and 3 hypovirulent strains of chestnut blight fungus. Hypovirulent strains had dsRNA virus in the cytoplasm, which is one of the typical characteristics of hypovirulent strains. In addition, they showed more characteristics of hypovirulent strains, i.e., suppressed conidiation, reduced pigmentation in colony color, and reduced phenol oxidase activity as well as reduced pathogenicity. Hypovirulent strains, KCPH-22, KCPH-135 and KCPH-136, had a genomic dsRNA band with the molecular weight of 12.7 kb, which is the L-dsRNA of CHV1. They also had a 2.7 kb defective dsRNA band. Single conidia isolated from hypovirulent strains were cultured and various phenotypes and absence of dsRNA bands were obtained from single conidial cultures, which means that hypovirulence transmission is unstable in asexual reproduction and variations in viral heredity by asexual reproduction. Biocontrol trial using hypovirulent strains was also carried out in the chestnut tree plantations, and canker expansion in the treated trees was stopped and healed by callus formation at the margin of the canker. These results show the potentials in successful biocontrol of chestnut blight if the vegetatively compatible hypovirulent strains could be directly used around the canker formed by compatible virulent strains.