This study was conducted to reduce contamination ratio of oyster mushroom bottle cultivation. The optimalconditions of substrate sterilization for reducing of contamination ratio were at 121oC for 90min. In addition, UV-C irradiation isgood for lower contamination ratio to continue over 6 hours at cooling and inoculation room after sterilization. Thecontamination ratio and density of microorganisms of substrate were showed 0% after sterilization at 121oC for 90min.Trichoderma sp., main pathogen of mushrooms, was detected from substrate after sterilized during 2 or 4 hours at 101oC and 105oC,respectively. The amount of electricity used was the lowest at 121oC for 90min than that of other sterilization conditions. The UV-Cirradiation treatment was used UV-C lamp(40 watts) in the inoculation room(56m3). The density of bacteria did not detected after UV-C irradiation for 6 hours. And the death ratio of bacteria and Trichoderma sp. was 99.9% after UV-C irradiation for 6 hours. However,in the same UV-C irradiation time, the death ration of Cladosporium sp. was 90.9%. Therefore, the death ratio of fungi was lower thanthat of bacteria at the same UV-C irradiation treatment.