Wine Spoilage Control: Impact of Saccharomycin on Brettanomyces bruxellensis and Its Conjugated Effect with Sulfur Dioxide

The yeast Brettanomyces bruxellensis is one of the most dangerous wine contaminants due to the production of phenolic off-flavors such as 4-ethylphenol. This microbial hazard is regularly tackled by addition of sulfur dioxide (SO2). Nevertheless, B. bruxellensis is frequently found at low levels (ca 103 cells/mL) in finished wines. Besides, consumers health concerns regarding the use of sulfur dioxide encouraged the search for alternative biocontrol measures. Recently, we found that Saccharomyces cerevisiae secretes a natural biocide (saccharomycin) that inhibits the growth of different B. bruxellensis strains during alcoholic fermentation. Here we investigated the ability of S. cerevisiae CCMI 885 to prevent B. bruxellensis ISA 2211 growth and 4-ethylphenol production in synthetic and true grape must fermentations. Results showed that B. bruxellensis growth and 4-ethylphenol production was significantly inhibited in both media, although the effect was more pronounced in synthetic grape must. The natural biocide was added to a simulated wine inoculated with 5 × 102 cells/mL of B. bruxellensis, which led to loss of culturability and viability (100% dead cells at day-12). The conjugated effect of saccharomycin with SO2 was evaluated in simulated wines at 10, 12, 13 and 14% (v/v) ethanol. Results showed that B. bruxellensis proliferation in wines at 13 and 14% (v/v) ethanol was completely prevented by addition of 1.0 mg/mL of saccharomycin with 25 mg/L of SO2, thus allowing to significantly reduce the SO2 levels commonly used in wines (150–200 mg/L).

Temporal Comparison of Microbial Community Structure in an Australian Winery

Most modern fermented foods and beverages are produced in fit-for-purpose facilities which are designed to ensure not only a reliable product, but also one safe for consumption. Despite careful hygiene, microorganisms can colonise these facilities and establish resident populations that can potentially contribute to the fermentation process. Although some microorganisms may not negatively affect the final product, spoilage microorganisms can be detrimental for quality, generating substantial economic losses. Here, amplicon-based phylotyping was used to map microbial communities within an Australian winery, before, during and after the 2020 vintage. Resident bacterial and yeast populations were shown to change over time, with both relative abundance and location within the winery varying according to sampling date. The bacterial family Micrococcaceae, and the genera Sphingomonas and Brevundimonas were the most abundant bacterial taxonomies, while Naganishia, Pyrenochaeta and Didymella were the most abundant fungal genera. Mapping the spatial distributions of the microbial populations identified the main locations that harboured these resident microorganisms, that include known wine spoilage yeasts and bacteria. Wine spoilage microorganisms, including the genefugura Lactobacillus, Acetobacter, Gluconobacter and Brettanomyces showed very low relative abundance and were found only in a couple of locations within the winery. Microbial populations detected in this facility were also compared to the resident microbiota identified in other fermented food facilities, revealing that microbial population structures may reflect the nature of the product created in each facility.

Assessing the Biofilm Formation Capacity of the Wine Spoilage Yeast Brettanomyces bruxellensis through FTIR Spectroscopy

Brettanomyces bruxellensis is a wine spoilage yeast known to colonize and persist in production cellars. However, knowledge on the biofilm formation capacity of B. bruxellensis remains limited. The present study investigated the biofilm formation of 11 B. bruxellensis strains on stainless steel coupons after 3 h of incubation in an aqueous solution. FTIR analysis was performed for both planktonic and attached cells, while comparison of the obtained spectra revealed chemical groups implicated in the biofilm formation process. The increased region corresponding to polysaccharides and lipids clearly discriminated the obtained spectra, while the absorption peaks at the specific wavenumbers possibly reveal the presence of β-glucans, mannas and ergosterol. Unsupervised clustering and supervised classification were employed to identify the important wavenumbers of the whole spectra. The fact that all the metabolic fingerprints of the attached versus the planktonic cells were similar within the same cell phenotype class and different between the two phenotypes, implies a clear separation of the cell phenotype; supported by the results of the developed classification model. This study represents the first to succeed at applying a non-invasive technique to reveal the metabolic fingerprint implicated in the biofilm formation capacity of B. bruxellensis, underlying the homogenous mechanism within the yeast species.

Brettanomics I: The Cost of Brettanomyces in California Wine Production

The yeast, Brettanomyces bruxellensis (Brett) is a significant cause of quality defects associated with red wine spoilage. At least some wine producers spend significant resources to prevent, detect, and mitigate damage from Brett, and many express concern about it, but some producers and consumers say they like it in small doses. Brett damage is especially of concern in premium red wine and has become more of a concern to producers in recent years as consumers have become better informed about it. We combine information from diverse sources to develop an initial understanding of the economics of Brettanomyces and management practices to mitigate its consequences. An analysis of detailed confidential data from three wineries in California reveals that at least some wineries are incurring significant costs to reduce the risk of infection with Brettanomyces. Some other wineries that opt not to spend so much on prevention are incurring higher costs in treating infected wines and in lost value from wines being downgraded to lower-valued blends. Results from an online survey of industry participants reinforce the analysis of the detailed data from the three wineries and suggest that the findings may be indicative of conditions more generally across the industry. (JEL Classifications: D22, D24, L66)

Effect of Candida intermedia LAMAP1790 Antimicrobial Peptides against Wine-Spoilage Yeasts Brettanomyces bruxellensis and Pichia guilliermondii

Wine spoilage yeasts are one of the main issues in the winemaking industry, and the control of the Brettanomyces and Pichia genus is an important goal to reduce economic loses from undesired aromatic profiles. Previous studies have demonstrated that Candida intermedia LAMAP1790 produces antimicrobial peptides of molecular mass under 10 kDa with fungicide activity against Brettanomyces bruxellensis, without affecting the yeast Saccharomyces cerevisiae. So far, it has not been determined whether these peptides show biocontroller effect in this yeast or other spoilage yeasts, such as Pichia guilliermondii. In this work, we determined that the exposure of B. bruxellensis to the low-mass peptides contained in the culture supernatant of C. intermedia LAMAP1790 produces a continuous rise of reactive oxygen species (ROS) in this yeast, without presenting a significant effect on membrane damage. These observations can give an approach to the antifungal mechanism. In addition, we described a fungicide activity of these peptides fraction against two strains of P. guilliermondii in a laboratory medium. However, carrying out assays on synthetic must, peptides must show an effect on the growth of B. bruxellensis. Moreover, these results can be considered as a start to develop new strategies for the biocontrol of spoilage yeast.

Chitooligosaccharide as A Possible Replacement for Sulfur Dioxide in Winemaking

Sulfur dioxide (SO2) has been used for centuries as a preservative in winemaking. However, the addition of SO2 is associated with allergic reactions and can negatively affect wine quality. In our work, chitooligosaccharide (COS) was applied as an alternative to SO2 in winemaking, and its antimicrobial activity during winemaking was investigated in comparison with the action of SO2. The optimal concentration of COS was identified as 500 mg/L. The antimicrobial effect of COS was evaluated using known and our own separated wine spoilage organisms. The antimicrobial effect of 500 mg/L COS was found to be comparable with that of 100 mg/L SO2. Furthermore, using 500 mg/L COS as an additive during winemaking did notinfluence the cell growth of Saccharomyces cerevisiae. Therefore, COS can be used as an additive in winemaking.