Pre-Emptive Upregulation of Antimicrobial Peptides by Dietary Propolis Improves Ethanol Tolerance in Drosophila melanogaster

Ethanol is a pervasive chemical stressor in fermentative environments and represents a major ecological challenge for Drosophila melanogaster, a species that naturally inhabits decaying fruits. Although ethanol tolerance has traditionally been attributed to detoxification and antioxidant pathways, accumulating evidence indicates that immune-related genes, particularly those encoding immune deficiency (IMD) pathway-associated antimicrobial peptides (IMD-AMPs), contribute importantly to chemical stress adaptation. Previous studies have demonstrated that IMD-AMP induction is required for ethanol tolerance; however, whether elevated IMD-AMP expression alone is sufficient to enhance tolerance has remained unresolved. In this study, we investigated the functional significance of IMD-AMP upregulation in ethanol tolerance using dietary propolis as an experimental immune-modulating agent. D. melanogaster were reared throughout their life cycle on propolis-supplemented diets and subsequently exposed to ethanol. Propolis-fed flies exhibited significantly enhanced survival under ethanol stress compared with control flies. Notably, this increased tolerance was not accompanied by upregulation of classical ethanol metabolism genes or broad induction of antioxidant-related genes. Instead, propolis feeding increased baseline and early-stage expression of IMD-AMP genes, including Diptericin A (DptA), Diptericin B (DptB), Attacin (AttC), and Metchnikowin (Mtk) before and during ethanol exposure. These findings suggest IMD-AMP upregulation is positively associated with enhanced ethanol tolerance in D. melanogaster. Our results establish a proactive role for immune-related pathways in chemical stress resistance and extend the functional scope of AMPs beyond pathogen defense. This study identifies IMD-AMPs as key effectors linking immune activation to physiological adaptation under ethanol-induced chemical stress.