A study on the yeast and mould biota associated with the production of Italian wines from post-harvest dried grapes

The wines obtained from post-harvest dried grapes are renowned wines, appreciated by consumers because of their complex sensorial notes due to the special vinification process, which involves the withering of grapes. The over-ripening phase is strongly influenced by the heterogeneous fungal biota present on the grape berry skin, and particularly by Botrytis cinerea. The development of the noble rot in berries affects the composition of the grape juice that presents high sugar content and inhibitory substances, as well. Therefore, grape juice fermentation proceeds hardly and is often accompanied by high production of volatile acidity. To better control the critical steps of Passito winemaking process, i.e. withering and fermentation, the use of suitable selected starter cultures could be applied as an innovative biotechnological strategy. Aims of this project were to increase knowledge about the diversity and evolution of indigenous eukaryotic microbiota associated with the production of some selected Italian wines (i.e. Amarone, Vin Santo di Gambellara, and Vino Santo Trentino) and to optimize the whole winemaking process through the use of selected indigenous strains of B. cinerea and yeasts. To these purposes, a large collection of moulds including B. cinerea was assembled; while a collection of yeasts (Saccharomyces and non-Saccharomyces) from wine aging in barrels and from the fermentation process of Garganega grape juice used for the production of Vin Santo di Gambellara was constituted. The isolates were identified using phenotypic and molecular methods, and then the collections were dereplicated by RAPD-PCR, PCR-RFLP, TEs, and microsatellite analysis. These techniques allowed the detection of a high genetic biodiversity among the B. cinerea and yeast isolates. The presence of distinctive strains could differentiate the withering of the studied grapes and the fermentation process, consequently selected strains of B. cinerea and yeasts were further characterized for several oenological properties (i.e. laccase activity and fungicide resistance for B. cinerea; resistance to stressful conditions and production of secondary metabolites for yeasts). In this way, strains of B. cinerea, S. cerevisiae and Zygosaccharomyces spp. were selected and tested in pilot-scale induction and fermentation trials, respectively. Considering the withering phase, at the end of the process the grapes were crushed and the grape juices were fermented. The gas-chromatographic analysis of the obtained wines indicated important differences between the inoculated and control grapes for several free volatile compounds. As expected, higher level of 1-octen-3-ol (mould flavor) was found for the wines obtained from the inoculated grape. These data suggest that the inoculum of selected B. cinerea strains on grapes influence the organoleptic characteristics of the must/wine, thus allowing a diversification of the final product. As regard the fermentation phase, the use of indigenous S. cerevisiae and non-Saccharomyces strains improved the fermentation process of Vin Santo di Gambellara. Indeed, failure to initiate fermentation and stuck fermentation were avoided, and a lower level of ethyl acetate was produced respect to the spontaneous fermentation. All selected S. cerevisiae yeasts produced flavors associated with white fruit, rose note, almond note and strawberry and caramel fragrance that emphasized the aromatic and flavor properties of this Passito wine. In conclusion, this research project suggests that the use of selected indigenous strains (moulds and yeasts) could improve the fermentation process of Passito wines minimizing risky conditions and improving the sensory quality of the final product. In other side, the use of valid alternative methods to SO2 addiction could improve the commercial impact of the wines free of chemical preservatives. In this way the modern consumer demand for wines that are safe and characterized by particular organoleptic flavour can be satisfied.