Analysis of Transcriptomic Response to SO2 by Oenococcus oeni Growing in Continuous Culture.

To successfully complete malolactic fermentation (MLF), Oenococcus oeni must overcome wine stress conditions of low pH, high ethanol, and the presence of SO2. Failure to complete MLF may result in detrimental effects to the quality and stability of the resulting wines. Research efforts to date have focused on elucidating the mechanisms and genetic features that confer the ability to withstand low pH and high ethanol concentrations on O. oeni; however, the responses to SO2 stress are less well defined. This study focused on characterizing the transcriptional response of O. oeni to SO2 challenge during cultivation in a continuous system at wine-like pH (3.5). This experimental design allowed the precise discrimination of transcriptional changes linked to SO2 stress from responses associated with growth stage and cultivation parameters. Differential gene expression analysis revealed major transcriptional changes following SO2 exposure and suggested that this compound primarily interacts with intracellular proteins, DNA, and the cell envelope of O. oeni. The molecular chaperone hsp20, which has a demonstrated function in the heat, ethanol, and acid stress response, was highly upregulated, confirming its additional role in the response of this species to SO2 stress. This work also reports the first nanopore-based complete genome assemblies for O. oeni. IMPORTANCE Malolactic fermentation is an indispensable step in the elaboration of most wines and is generally performed by Oenococcus oeni, a Gram-positive heterofermentative lactic acid bacterium species. While O. oeni is tolerant to many of the wine stresses, including low pH and high ethanol concentrations, it has high sensitivity to SO2, an antiseptic and antioxidant compound regularly used in winemaking. Understanding the physiological changes induced in O. oeni by SO2 stress is essential for the development of more robust starter cultures and methods for their use. This study describes the main transcriptional changes induced by SO2 stress in the wine bacterium O. oeni and provides foundational understanding on how this compound interacts with the cellular components and the induced protective mechanisms of this species.

Whole transcriptome RNAseq analysis of Oenococcus oeni reveals distinct intra-specific expression patterns during malolactic fermentation, including genes involved in diacetyl metabolism.

We report the first whole transcriptome RNAseq analysis of the wine-associated lactic acid bacterium Oenococcus oeni using a combination of reference-based mapping and de novo transcript assembly in three distinct strains during malolactic fermentation in Cabernet Sauvignon wine. Two of the strains (AWRIB551 and AWRIB552) exhibited similar transcriptomes relative to the third strain (AWRIB419) which was dissimilar by comparison. Significant intra-specific variation for genes related to glycolysis/gluconeogenesis, purine metabolism, aminoacyl-tRNA biosynthesis, ABC transporters and phosphotransferase systems was observed. Importantly, thirteen genes associated with the production of diacetyl, a commercially valuable aroma and flavour compound, were also found to be differentially expressed between the strains in a manner that correlated positively with total diacetyl production. This included a key strain-specific gene that is predicted to encode a l-lactate dehydrogenase that may enable l-lactic acid to be utilised as a precursor for the production of diacetyl. In conjunction with previous comparative genomic studies of O. oeni, this study progresses the understanding of genetic variations which contribute to the phenotypes of this industrially-important bacterium.

Saccharomyces cerevisiae-Oenococcus oeni interactions in wine: current knowledge and perspectives.

Winemaking can be summarized as the biotransformation of must into wine, which is performed principally by Saccharomyces cerevisiae strains during the primary or alcoholic fermentation. A secondary fermentation, the so-called malolactic fermentation (MLF) is a biodeacidification that is often encouraged, since it improves wine stability and quality. Malolactic fermentation usually occurs either spontaneously or after inoculation with selected bacteria after alcoholic fermentation. The main organism responsible for MLF, the lactic acid bacterium Oenococcus oeni, develops in physicochemically harsh conditions, which may lead to MLF failure. Furthermore, yeast that ferment must before or together with O. oeni can prevent or stimulate the progress of MLF. These phenomena are part of the interactions observed between yeast and bacteria. The mechanisms that govern yeast bacteria interaction are reviewed and the consequences for winemaking are discussed. In the light of recent advances, future prospects are also presented.

Mousy off-flavor of wine: precursors and biosynthesis of the causative N-heterocycles 2-ethyltetrahydropyridine, 2-acetyltetrahydropyridine, and 2-acetyl-1-pyrroline by Lactobacillus hilgardii DSM 20176.

The N-heterocyclic bases, 2-ethyltetrahydropyridine (1), 2-acetyl-1-pyrroline (2), and 2-acetyltetrahydropyridine (3) are associated with the occurrence of mousy off-flavor in wine. The biosynthesis of these N-heterocycles by the wine lactic acid bacterium, Lactobacillus hilgardii DSM 20176, was studied by high-cell-density incubation in combination with a minimal chemically defined N-heterocycle assay medium. The key components of the defined N-heterocycle assay medium included D-fructose, ethanol, L-lysine, L-ornithine, and mineral salts. N-heterocycle formation was quantitatively determined by gas chromatography-mass spectrometry. The formation of 2 and 3 required the concomitant availability of a fermentable carbohydrate (D-fructose), ethanol, and iron (Fe(2+)). In addition, L-ornithine stimulated the formation of 2 and repressed 3 formation, whereas L-lysine stimulated the formation of 3 and repressed 2 formation. Incorporation of d(6)-ethanol into the acetyl side chain of 2 and 3, and of d(4)-acetaldehyde into the acetyl side chain of 3, confirmed that ethanol and acetaldehyde could serve as major side chain precursors. A pathway for the formation of 2 and 3 by heterofermentative lactic acid bacteria is proposed involving the interaction of accumulated C-2 intermediates from the heterolactic pathway and N-heterocyclic intermediates derived from the metabolism of L-ornithine and L-lysine.