Functional characterization of grapevine MLO genes to define their roles in Powdery mildew susceptibility by CRISPR/Cas9 genome editing

Ochratoxin A (OTA) is a mycotoxin produced by several filamentous fungi infecting grape bunches (Penicillium and Aspergillus spp.), this toxin pass to must when grapes are crushed and later it is found in wine. Following the evaluations of the toxicity of OTA, European Commission Regulations have been promulgated introducing upper limits for OTA concentrations in various commodities (cereals, cereal products, dried vine fruit, coffee, wine, grape juice, baby foods and dietary foods for special medical purposes).

Recent studies have demonstrated the role of certain lactones, particularly 2-nonen-4-olide, and volatile thiols (3-sulfanylhexan-1-ol) in the over ripped aromas of noble rot sweet wines (Stamatopoulos et al. 2014ab). These compounds are partly formed during the maturation and under the activity of B. cinerea on grapes. This research was carried out in the vineyard of Sauternes with aim to better understand their genesis depending on the grape over-ripening on two different soil types during 3 vintages. Thus, the study was conducted, with the Sémillon grape, during vintages 2012, 2014 & 2015, at 4 stages of over-maturation of the grapes (healthy, pourri plein, pourri roti, pourri roti + 15 days) considering two vineyard plots with different soil characteristics (calcosol & peyrosol) planted with the 315 Sémillon clone and grafted on 101-14 rootstock respectively in 1981 and 1980 and cultivated with the same vineyard management. Volatile lactones were assayed by liquid-liquid extraction followed by GC/MS analysis and the precursors of 3-sulfanylhexanol by an adaptation of the method by Capone et al. 2010 (SPE-
UPLC/FTMS).

Sunburn results from a combination of excessive photosynthetically active radiation (PAR) and UV radiation and temperature that can be exacerbated by other stress factors such as water deficit. Sunburn is a physiological disorder that affects the visual and organoleptic properties of grapes. The appearance of brown and necrotic spots severely affects the commercial value of the fruit, and in extreme cases, significantly decreases yield. This damage occurs with some frequency in sensitive varieties such as Graciano.

Herbicides are commonly sprayed in the vine row to prevent competition with vines for water and minerals and to keep weeds from growing into the bunch zone. Sprays are applied before budbreak and reapplied multiple times during the season to keep the undervine bare. There is growing concern about the negative effects of herbicides on humans and the environment, and weeds in New Zealand have developed resistance to herbicides. Therefore, it is imperative that we reduce our reliance on herbicides in viticulture and incorporate methods that do not engender resistance.

Today the knowledge in terms of molecular composition of the colloidal matrix is ​​not enough in order to control its stability, according to the number of winemaking and wine stabilization processes. The physico-chemical processes during the winemaking change the composition and quantity of wine macromolecules. The goal today is to determine which analytical techniques will allow to discriminate these winemaking processes in order to better understand their impact on colloidal matrix stability as well as which molecules are responsible for its instabilities. METHODS: Wines obtained after conventional winemaking were subjected to different fining and chemical stabilization treatments. Different methods were used to investigate the wine macromolecular composition and stability after chemical stabilization, including quantitative and qualitative analyzes of total soluble polysaccharides by extraction under acidified ethanol, and by size exclusion separation as well as qNMR metabolomics. RESULTS: Observation of a slight difference at the quantitative level using classical analysis between the winemaking processes was observed as well as a strong discrimination by qNMR metabolomics.