The present study used different applications as Fe3O4 NPs, yeast, Bacillus megaterium (Bio-Arc® 6% WP), Azoxystrobin, (Amistar® 25% SC), and Penaconazole (Topas® 100 10%EC) to control the natural infection of powdery mildew on lettuce growing under field conditions during winter growing seasons (2020 and 2021). The obtained data showed that disease severity was significantly decreased in all treated lettuce plants compared to the untreated plants. In addition, all applications enhanced plant height, number of leaves/plant, and weight of each plant. The different applications significantly increased chlorophyll, carotene, phenolic, and protein contents compared to the control. In addition, the above applications increased the activities of catalase (CAT) and polyphenol oxidase (PPO) in treated plants. Finally, Azoxystrobin fungicide had the best effect compared to the different applications and the control while Bio-Arc was less effective.