S19 - Session O3 - HIGS and SIGS approaches targeting Cyp51 and SdhD of B. cinerea to control grey mould disease in strawberry and grapevine

Authors: Silvia Sabbadini *, Francesca Negrini, Luca Capriotti, Alice Ciofini, Angela Ricci, Daniel E. Gebremichael, Elena Baraldi, Bruno Mezzetti

Strawberry ( Fragaria x ananassa ) and grapevine ( Vitis spp.) are among the most important berry crops cultivated worldwide. However, their fruits are affected by the well-known fungal pathogen Botrytis cinerea , which causes huge losses during their cultivation, but also in the storage, transport, and commercialization phases. Currently, the control of B. cinerea infection is mainly based on the use of commercial fungicides, which are dangerous for the environment and human health; while there is a lack of real effective technologies of the fresh produce to limit post-harvest losses. Innovative in-planta and topically applied interfering RNA molecules can provide realistic alternatives to current pest and disease control, as they are characterized by a high efficacy, good safety profiles and minimal environmental impact. In this study, a hairpin-based gene construct has been designed and prepared targeting two genes of B. cinerea , i.e. the cytochrome p450 51 ( cyp51 ) and the subunit D of the succinate dehydrogenase ( sdhD ), under the control of 35S promoter. Well-established in vitro regeneration and transformation protocols have been applied to obtain HIGS lines of both strawberry ( F. x ananassa ) cv Sveva, and grapevine ( Vitis vinifera ) cv Thompson seedless. Putative HIGS lines have been identified through both kanamycin-based selection and visual screening, by green fluorescent protein (GFP) detection, and molecularly characterized through PCR analysis. The efficacy of dsRNAs stably expressed in the transformed shoots has been preliminarily tested by inoculating detached leaves from in vitro elongated shoots of both HIGS grapevine and strawberry lines. Parallelly, dsRNAs molecules targeting cyp51 and sdhD of B. cinerea have been produced from E.coli and have been validated through SIGS approach on different organs and tissues of different plant species. The results achieved in this study have the main objective of confirming that RNAi systems are useful tools for the defence of plants from pathogens.