Botryosphaeria canker and fruit rot caused by the fungus Botryosphaeria dothidea is one of the most destructive diseases of apple worldwide. Autophagy is an evolutionarily conserved self-degradation process that is important for maintaining homeostasis to ensure cellular functionality. To date, the role of autophagy in B. dothidea is not well elucidated. In this study, we identified and characterized the autophagy-related protein Atg1 in B. dothidea. The BdAtg1 deletion mutant ΔBdAtg1 showed autophagy blockade and phenotypic defects in mycelial growth, conidiation, ascosporulation and virulence. In addition, ΔBdAtg1 exhibited an increased number of nuclei in the mycelial compartment. Comparative transcriptome analysis revealed that inactivation of BdAtg1 significantly influenced multiple metabolic pathways. Taken together, our results indicate that BdAtg1 plays an important role in vegetative differentiation and the pathogenicity of B. dothidea. The results of this study will provide a reference for the development of new target-based fungicides.