Abstract:The identification of the pathogens isolated from the canker of eucalyptus (Eucalyptus grandis) were carried out. Seventeen diseased eucalyptus samples were observed in Shilin and Honghe in Yunnan Province (south-western China). Seventeen isolates of possible pathogenic fungi were isolated from trunks and branches of infected eucalyptus plants. Bases on the pathogenicity experiment, the results showed that YNE1-YNE17 were the pathogens of eucalyptus (E.grandis) canker disease. The morphological identification showed that YNE1-YNE17 was identified as Botryosphaeria dothidea.By analysis of the rDNA internal transcribed spacer region (ITS1-5.8S-ITS2). BLAST searches at GenBank showed a high identity with reference sequences (99%-100%). Representative sequences of both regions were deposited to GenBank (Accession No. FJ517657, FJ517658). Morphological and molecular identification confirmed this species as B. dothidea. B. dothidea was first reported as the pathogen of eucalyptus canker and shoot blight disease in China.