Investigation of the SO2 stress response in Brettanomyces/Dekkera bruxellensis using RNA-seq

Sulphur dioxide (SO2) is the most common additive worldwide used to prevent and/or control the microbial contamination in wine. Its use should be limited due to the detrimental effect to human health and the intolerance shown by a number of wine consumers. Brettanomyces/Dekkera bruxellensis, the main spoilage yeast of wine, is able to growth and produce volatile phenols even when SO2 is present. The SO2 resistance is a strain-dependent characteristic that needs to be further investigated. The two completely sequenced strains of B./D. bruxellensis, AWRI1499 (Curtin et al., 2012) and CBS2499 (Piškur et al., 2012), were used for batch fermentations carried out in synthetic wine medium. When cells reached the exponential phase of growth, SO2 was added (0.35 mg/L as molecular SO2). Samples for transcriptomic analysis were collected before SO2 pulse (T0), 5 hours after the addition (T5h) and at cell growth recovery (Tr). The results showed that more genes were significantly differentially expressed within the comparison Tr-vs-T0 than in the T5h-vs-T0 in both strains, thus indicating that mainly an adaptive response is activated. Among genes affected by SO2 addition and shared by the two strains, upregulated genes participating to carbohydrate, acyl-CoA and monocarboxylic acid metabolism were found. Interestingly, AWRI1499-0080, the gene homolog to SSU1, encoding the enzyme involved in the main mechanism of sulphite detoxification in Saccharomyces cerevisiae, increased its expression in the long term response up to 4 and 47 folds in CBS2499 and AWRI1499, respectively, thus highlighting the importance of this detoxification mechanism in B./D. bruxellensis as well. This study confirms the great ability of B./D. bruxellensis yeasts to survive and growth in extreme environmental conditions. In conclusion, due to the capacity of this yeast to counteract the stress induced by the addition of SO2, a proper management of the winemaking, including a deep cleaning of the winery equipment, is advisable for preventing, or at least minimizing the Brett contamination in wine.