Strawberry powdery mildew, caused by the obligate biotrophic fungus Podosphaera aphanis, is one of the most devastating diseases in strawberry, resulting in severe reduction in fruit yield and quality. Exploring and utilizing powdery mildew-resistant strawberries is an important disease management strategy. The present study evalu-ated the resistance of 19 strawberry varieties to Podosphaera aphanis NWAU1 (Pa. NWAU1) based on disease index. We have comprehensively investigated the different cytological and physiological responses of different resistant strawberries. Compared with highly susceptible (HS) strawberries 'Benihoppe' and 'Tianxianzui', the moderately resistant (MR) strawberries 'Heilongjiang' and 'Yellow Wonder' exhibited higher levels of callose and H2O2 accumulation after inoculation with Pa. NWAU1. The activities of SOD, POD, GPX, and the GSH level in HS strawberries were increased after inoculation with Pa. NWAU1. The SA and JA levels in resistant straw-berries were almost similar. However, the ACC level varied in HS and MR strawberries. Our results indicate that increased levels of callose, H2O2 and ACC resulted in resistance to Pa. NWAU1 in MR strawberries. The higher antioxidant enzyme activities may contribute to the sensitivity to Pa. NWAU1 in HS strawberries. Our study provides a reliable and comprehensive method for evaluating and understanding potential defense responses to powdery mildew resistance in different strawberries.