CHANGES IN METABOLIC FLUXES UNDER LOW PH GROWTH CONDITIONS: CAN THE SLOWDOWN OF CITRATE CONSUMPTION IMPROVE OENOCOCCUS OENI ACID-TOLERANCE?

Abstract

Oenococcus oeni is the main Lactic Acid Bacteria responsible for malolactic fermentation, converting malic acid into lactic acid and carbon dioxide in wines. Following the alcoholic fermentation, this second fermentation ensures a deacidification and remains essential for the release of aromatic notes and the improvement of microbial stability in many wines. Nevertheless, wine is a harsh environment for microbial growth, especially because of its low pH (between 2.9 and 3.6 depending on the type of wine) and nutrient deficiency. In order to maintain homeostasis and ensure viability, O. oeni possesses different cellular mechanisms including organic acid metabolisms which represent also the major pathway to synthetize energy in wine. Indeed, uptake and consumption of malate and citrate by this bacteria enables to activate a proton motive force (PMF) hence maintaining an intracellular pH by proton consumption1,2.

Citrate is found in wine at small concentrations (0.13 to 0.90 g/L). It can be metabolized by O. oeni into acetate, pyruvate and then aromatic compounds such as diacetyl, acetoin and 2,3-butanediol. The ability of citrate metabolism to activate the PMF could play a central role in the acid-tolerance of this bacterium. Nevertheless, a previous study has described an inhibition of O. oeni growth at low pH in presence of high amounts of citrate3. This toxic effect could come from the synthesis of one of the citrate metabolites as acetate.

In order to understand how citrate metabolism can be linked to acid tolerance of this bacterium, consumption of citrate was investigated in a great diversity of O. oeni strains. In addition, malate and sugar consumptions were also followed, as they can be impacted by citrate metabolism. These experiments enabled to draw metabolic fluxes in O. oeni according to the pH of the medium. In most cases, ma- late is first metabolized, then citrate and sugars, sequentially, proving that the priority is given to organic acid consumption at the expense of sugars in this bacterium. However, this experiment revealed different citrate consumption profiles which may be correlated to a greater or lesser acid tolerance according to the strain. Furthermore, a genomic comparison demonstrated the presence of mutations in the citrate operon of acid-tolerant industrial strains. Hence, acid tolerance could be linked to a change in the rate of citrate consumption in O. oeni.