DNA Aptamers Targeting BcSOD1: A Novel Strategy for Controlling Botrytis cinerea in Sustainable Agriculture.

Botrytis cinerea, the necrotrophic fungus responsible for grey mould disease, is a major threat to global crop production. Control strategies mainly rely on chemical fungicides, but resistance development limits their long-term effectiveness. This study introduces, for the first time in crop protection, the use of DNA aptamers as a novel and sustainable strategy. Aptamers are short, single-stranded DNA molecules that bind specific targets with high affinity, acting like 'chemical antibodies'. Using SELEX technology, two aptamers, SOD9.14F and SOD9.26F, were designed to target BcSOD1, a superoxide dismutase enzyme essential for fungal virulence and ROS detoxification. Molecular modelling predicted that both aptamers bind within BcSOD1's catalytic pocket. Both aptamers inhibited BcSOD1 enzymatic activity (97.5%) and reduced germination (67%), fungal biomass (58%) and lesion formation (42%) in B. cinerea-infected tomato leaves (Solanum lycopersicum) and apple fruits (Malus domestica). Fluorescence microscopy confirmed aptamer binding to conidia surfaces. No antifungal effect was observed in the ΔBcsod1 mutant or with the non-structured control aptamer Ap.AGA, supporting target specificity. RNA-Seq analysis revealed that SOD9.26F interfered with fungal oxidative stress responses and metabolism. Additionally, aptamer application primed tomato plants, activating defence-related gene expression. Interestingly, Ap.AGA aptamer triggered partial priming, suggesting a broader DNA-induced effect. These findings validate BcSOD1 as an antifungal target and highlight aptamers as dual-action agents: impairing fungal development and enhancing plant immunity. This study positions DNA aptamers as specific, effective and sustainable tools for integrated management of grey mould in agriculture.