Directed Evolution of Oenococcus oeni: optimising yeast-bacteria interactions for improved malolactic fermentation

The present study was performed in a traditional winery located in the viticultural area of Brunello di Montalcino, Siena, Italy, in the vintage 2015. Actually, in this winery Sangiovese grape musts are fermented in large oak barrels by a single strain of Saccharomyces cerevisiae previously isolated in the same winery. Pumping over operations are carried out once or twice a day until the end of alcoholic fermentations. The aim of this work was to investigate on the oenological properties of Sangiovese wine produced with the traditional winemaking process adopted by the winery under study obtained from the fermentation of whole berries compared to that from crushed grape must. In particular, two lots of 65q of Sangiovese grapes from the same 3ha vineyard were vinified in 150hL oak barrels.

Climate change and harvest date decisions have led to the evolution of must quality over the last decades. Increases in must sugar concentrations are among the most obvious consequences, quantitatively. Saccharomyces cerevisiae is a robust and acid tolerant organism. These properties, its sugar to ethanol conversion rate and ethanol tolerance make it the ideal production organism for wine fermentations. Unfortunately, high sugar concentrations may affect S. cerevisiae and lead to growth inhibition or yeast lysis, and cause sluggish or stuck fermentations. Even sublethal conditions cause a hyperosmotic stress response in S. cerevisiae which leads to increased formation of fermentation by-products, including acetic acid, which may exceed legal limits in some wines.

Fermentative aromas (especially esters and higher alcohols) highly impact the organoleptic profile of young and white wines. The production of these volatile compounds depends mainly on temperature and Yeast Available Nitrogen (YAN) content in the must. Available dynamic models predict the main reaction
(bioconversion of sugar into ethanol and CO2 production) but none of them considers the production kinetics of fermentative aroma compounds during the process of fermentation. We determined the production kinetics of the main esters and higher alcohols for different values of initial YAN content and temperature, using an innovative online monitoring Gas Chromatography device.

The aromatic complexity of a wine results from the perception of the association of volatile molecules and each aroma can be categorized into different families. The “green” aromas family in red wines has retained our attention by its close link with the fruity perception. In that study, the “green” olfactory concept of red wines was considered through a strategy combining both sensory analysis and hyphenated chromatographic techniques including HPLC and MDGC (Multidimensional Gas Chromatography). The aromatic space of this concept was specified by lexical generation through a free association task on 22 selected wines by a panel of wine experts. Then, 70 French red wines were scored on the basis of the intensity of their “green” and “fruity” attributes.

Among red wines ethyl esters, those from short hydroxylated and branched-chain aliphatic acids constitute a family with a particular behavior and sensory importance. They have been previously discussed in the literature [1] and recent studies have established that some of them were strongly involved in of red wines’ fruity aroma [2]. As some among them have an asymmetrical carbon atom, it seemed important to separate their different enantiomers to obtain an accurate assessment of their organoleptic impact. Three chiral esters have been identified, presenting alkyl and/or hydroxyle substituants: ethyl 2-hydroxy-4-methylpentanoate, ethyl 2-methylbutanoate, and ethyl 3-hydroxybutanoate.