TOWARDS A METABOLOMIC CHARACTERISATION OF THE GRAPEVINE RESPONSE TO FLAVESCENCE DORÉE INFECTION BY NMR AND LC-MS PROFILING

Flavescence dorée (FD) is a quarantine disease of grapevine, involving interactions between the plants, leafhopper vectors (phytophagous insect Scaphoideus titanus Ball, Cicadellidae family), and Flavescence dorée phytoplasma. This disease is a major threat to vineyard survival in different European grapegrowing areas. Differences in FD susceptibility between grapevine varieties exist, in terms of multiplication of the associated phytoplasma and its spread in the plant. Cabernet Sauvignon (CS) is highly susceptible to FD, whereas Merlot (M) is less susceptible. The objective of this project is to determine which primary and specialised metabolites are altered in grapevines following infection by FD phytoplasma under controlled conditions. Plants of CS and M were infected with FD phytoplasma on a single leaf via the natural insect vector S. titanus in a high containment greenhouse. Sampling took place 7-day, or 7-week post-inoculation. The leaves were harvested and immediately frozen in liquid N2 to quench metabolism. Phytoplasma titers were determined on fresh powder of each leaf. Metabolomic analyses were performed on a selection of the leaf sample extracts by 1 H-NMR, LC-LTQ/Orbitrap-HRMS, and LC-QqQ-MS. NMR spectra were acquired at 500 MHz and processed using NMRProcFlow (nmrprocflow.org). For both LC-MS, metabolites were separated by reverse-phase chromatography with a H2O/MeCN gradient. Untargeted experiments were performed on a UHPLC-ESI-LTQ/Orbitrap-MS with an ESI probe operated in positive mode and processed using MS Dial v4.7 8. Targeted experiments were performed on an LC-QqQ-MS system with a MRM method in positive or negative mode and the quantification by using calibration curves built with standards.The phytoplasma titre data, metabolite quantification data or metabolomic signatures of leaf samples were managed with ODAM tool