Exploring Lachancea thermotolerans biodiversity: insights into lactic acid production and carbon metabolism in response to oxygen availability

Abstract

ABSTRACT Lactic acid production by Lachancea thermotolerans offers a promising biological strategy to restore wine acidity often compromised by climate change, yet this metabolic trait remains highly variable and poorly controlled. This study systematically evaluated the impact of controlled oxygenation regimes on fermentation performance and primary metabolism in three genetically distinct L. thermotolerans strains. Oxygen availability drastically altered metabolic fluxes. Under continuous aerobic conditions, carbon was redirected toward biomass formation (9- to 13-fold increase), with reduced ethanol and lactic acid yields, as well as increased succinic acid, acetoin, and 2,3-butanediol, whereas fully anaerobic conditions lowered fermentation performance but substantially increased lactic acid. These results confirmed that L. thermotolerans is a weakly Crabtree-positive yeast with mixed respiratory-fermentative metabolism. Carbon redistribution under aerobic conditions was not fully compensated by measured metabolites, resulting in substantially decreased carbon recovery (~70%) and suggesting undetermined respiratory CO2 production alongside diversion of carbon toward undetermined intracellular components. This imbalance was supported by a decline in cell density following peak growth, consistent with potential redox imbalance or oxidative stress. Lactic acid production occurred primarily under oxygen limitation, independent of ethanol formation, indicating a regulated, facultative adaptive response rather than a constitutive fermentation product. Strain-dependent differences further revealed distinct carbon allocation strategies. While oxygen strongly influenced metabolic behavior, pulse strategies showed limited, condition-dependent effects, highlighting the importance of timing and intensity for modulating lactic acid production. Overall, oxygen is a powerful lever to modulate metabolic fluxes in L. thermotolerans, but its application for predictable bio-acidification requires further optimization. IMPORTANCE Wine acidity is increasingly difficult to maintain under climate change, creating a need for reliable biological solutions. Some strains of Lachancea thermotolerans can produce lactic acid during fermentation, but this metabolic trait is inconsistent and difficult to control. This study shows that oxygen availability is a key factor shaping how this yeast uses carbon, shifting it between growth and lactic acid production. Under the continuous aerobic condition, carbon flux was redirected toward biomass, whereas the fully anaerobic condition favored lactic acid production but reduced fermentation performance. Importantly, pulsed oxygen additions did not provide consistent control of lactic acid production, but they revealed trends that highlight the importance of timing and intensity. These findings show that lactic acid production is a flexible response to environmental conditions rather than a constitutive trait. This work provides a foundation for developing strategies to better control lactic acid production through timed oxygen addition during fermentation. Wine acidity is increasingly difficult to maintain under climate change, creating a need for reliable biological solutions. Some strains of Lachancea thermotolerans can produce lactic acid during fermentation, but this metabolic trait is inconsistent and difficult to control. This study shows that oxygen availability is a key factor shaping how this yeast uses carbon, shifting it between growth and lactic acid production. Under the continuous aerobic condition, carbon flux was redirected toward biomass, whereas the fully anaerobic condition favored lactic acid production but reduced fermentation performance. Importantly, pulsed oxygen additions did not provide consistent control of lactic acid production, but they revealed trends that highlight the importance of timing and intensity. These findings show that lactic acid production is a flexible response to environmental conditions rather than a constitutive trait. This work provides a foundation for developing strategies to better control lactic acid production through timed oxygen addition during fermentation.