Identificador: TDX:428
Autores: Carreté Aixalà, Ramon
Resumen:
INFLUENCE OF WINE COMPONENTS OVER THE METABOLISM OF Oenococcus oeni AND OVER THE POPULATION DYNAMICS DURING MALOLACTIC FERMENTATIONMalolactic fermentation (MLF) in wine is carried out by certain lactic bacteria which decarboxylate L-malic acid to L-lactic and CO2. Oenococcus oeni is the most tolerant species under the unfavourable wine conditions (nutrients starvation, low pH, ethanol, fatty acids from yeasts, SO2, etc.) and the main responsible of driving the MLF. As a consequence of the MLF, the wine quality improves because the acidity decreases, the organoleptic characteristics are better and the microbiological stability of wine increases. Some biochemical parameters, indicators of metabolic cellular conditions, are the ATPase activity and the expression of stress proteins and exoproteases involved in the obtention of nitrogen. On the other hand, some unfavourable conditions for bacterial growth can modify the bacterial population dynamics during MLF by effects over species and strains of the same species. The studies performed in this work show that ATPase-membrane activity principally corresponds to an H+-ATPase, which is affected by cellular viability inhibitory concentrations of ethanol, SO2, copper and fatty acids. So, a direct relationship between growth inhibition of O. oeni in real cellar conditions and ATPase activity inhibition is stablished. The stress protein profile produced by ethanol is not significantly different from the control's one, whereas the incubation in acidic conditions or in the presence of C12 fatty acid inhibits the protein expression in general. Copper and low pH induce the overexpression of a proteinic band close to 40 kDa, as SO2 does, which N-terminal sequence shows a significant identity with the family of glyceraldehyde-3-phosphate dehydrogenases (GAPDH). In his turn, SO2 inhibits the cytoplasmatic GAPDH activity also. Using RT-PCR, it has been shown that gene expression levels of hsp18 (the gene of a heat-shock protein found in O. oeni) as a response to the heat shock are near fifty-fold greater than in normal conditions. On the other hand, in contact with inhibitory concentrations of C12, copper or SO2, it only appears weak induction as a response to copper. About the exoprotease activity, this one is not significative in the strains studied, except for final exponential growth phase in the presence of high ethanol levels (14 %). Under conditions of nitrogen starvation, it's observed a direct relationship between the bioavailability of α-amino nitrogen and growth rate of O. oeni. Different microvinification conditions were assayed using three different strains (CECT 4100 as the type strain; CR1, an strain isolated by our research group, and Vitilactic, a commercial strain). The type strain showed fewer ability for adaptation, whereas the isolated strain was the responsible for performing the FML the most times. To demonstrate that, we used Molecular Biology techniques, like specific PCR and RAPD-PCR. In cellar vinifications with normal and high levels of SO2 and lysozyme, it was observed a dominance of O. oeni species from the beginning of the alcoholic fermentation, and a different evolution of the populations in the presence of lysozyme. In all the cases, it's the same strain which performs FML and it coincides with that one imposed in other campaigns and isolated by our group as CR1.
Repositori URV