Black spot disease is a significant worldwide disease on the rose plant. Due to this infection, the leaves become yellow and eventually fall off. The occurrence of this disease has become a major problem, especially in landscape purpose. Therefore, this research was conducted to isolate fungal species from black spot disease in rose and identify using morphological characteristics. Then, all the isolates were tested for pathogenicity to confirm Koch’s postulates. In this study, four fungal isolates have been successfully isolated from black spot disease in rose namely Rhizoctonia sp. (one isolate), Colletotrichum sp. (two isolates) and Penicillium sp. (one isolate). Based on pathogenicity test result using potato dextrose agar (PDA) plug technique, fungus UMTT27R (Penicillium sp.) showed highly pathogenic on rose’s leaves with disease severity (DS) = 88.89% followed by UMTT13R (Colletotrichum sp.) with DS=72.22%, UMTT21R (Colletotrichum sp.) with DS=66.67% and UMTT4R (Rhizoctonia sp.) with DS=61.11%. Correct identification of fungal pathogens is very important to strategize a proper method to control the black spot disease in rose cultivation.