WINETECH Technical Yearbook 2020

ethanol , albei t over excessively long fermentation periods (e.g. 180 days) in monocultures (Vicente et al ., 2020; Du Plessis et al ., 2017). Consequently, the most preferred and efficient fermentation mode for this yeast is sequential inoculations with S. cerevisiae (Vicente et al ., 2020; Morata et al ., 2019) . EFFECT ON MALOLACTIC FERMENTATION Malic acid consumption has been reported in some strains of M. pulcherrima. Regarding inf luence on malolact ic fermentation, several strains have shown compatibility with Oenococcus oeni strains, however, this is dependent on the strain of both the yeast and the bacteria and the inoculation strategy. For instance, wines produced by inoculation of S. cerevisiae at 48 and 72 hours after M. pulcherrima were found to result in slower MLF than when S. cerevisiae was inoculated after 24 hours (Martín-García et al ., 2020). TRAITS OF OENOLOGICAL INTEREST ● Extracellular enzymes – effects on wine aroma/stability Metschnikowia pulcherrima strains produce a diversity of enzymes that are important in the release of varietal aromas. In particular, M. pulcherrima produces glycosidases, such as

the sugar is respired (Quirós et al ., 2014). Furthermore, M. pulcherrima strains show better growth, persistence and sugar consumption under aerobic conditions when in monoculture or in mixed culture fermentations with S. cerevisiae (Shekhawat et al ., 2017; Morales et al ., 2015) . ● Glycerol, VA, SO 2 and H 2 S production Metschnikowia pulcherrima generally produces moderate levels (0.3-0.4 g/L) of volatile acidity (expressed as acetic acid). When used in sequential fermentation with S. cerevisiae , it is known to reduce VA by 10-75% and increase glycerol by 4-40%, depending on the inoculation strategy, the strain and the persistence of the strain during fermentation (Vicente et al ., 2020). H 2 S production in M. pulcherrima is strain dependent, ranging from absent to high production, with most strains being H 2 S negative due to lack of sulphite reductase activity (Vicente et al ., 2020). ● Behaviour in pure culture fermentations Metschnikowia pulcherrima mainly displays low fermentative capacity with most strains producing up to 4.5% (v/v) ethanol. A few strains have been reported to be moderate fermenters able to produce 9-11.5%

β-glucosidase, β-xylosidase and in some strains (e.g. M. pulcherrima Flavia™) α-arabinofuranosidase, which release monoterpenes, mainly linalool and geraniol nerol (Morata et al ., 2019 & 2020). Furthermore, M. pulcherrima strains produce β-lyase which releases volatile thiols. The production of acid proteases is also widely distributed in this species and these enzymes have shown potential to combat haze formation in white wines. Regarding fermentat ion aromas, M. pulcherrima strains have been shown to consistently increase the production of 2-phenylethanol, fatty acids, acetoin, diacetyl and the total concentration of esters. Among the ethyl esters, ethyl octanoate (a pear-associated ester) is most notable (Vicente et al ., 2020). ● Ethanol reduction in wine Metschnikowia pulcherrima has been proposed as a suitable candidate yeast for lowering the ethanol content in wine. Indeed, several strains when used in sequential inoculation with S. cerevisiae 48 hours after M. pulcherrima or only after 50% of the sugar is consumed and with suitable oxygen input during fermentation were shown to reduce final ethanol levels by 0.6-1.6% v/v (Morata et al ., 2019; Hranilovic et al ., 2020; Puškaš et al .,

2020). However, this trait is more pronounced in synthetic grape juice fermentation and declines in real wine fermentation conditions, thus further investigation is required (Puškaš et al ., 2020). ● Chitin/mannoprotein content – effects on wine stability Studies have shown that some strains of M. pulcherrima display notable mannoprotein release ability which increases the mouthfeel properties of wine (Belda et al ., 2016). The release of mannoproteins has also been shown to improve protein stability in wine and lower haze potential (Snyman, 2019). Furthermore, recent studies have shown that M. pulcherrima strains including Level 2 Flavia, are able to reduce haze formation even when minimal protease activity is detected. This ability is thought to be due to high levels of chitin in the cell wall. The chitin has been shown to adsorb and subsequently remove grape chitinases which are major contributors to haze (Snyman, 2019; Ndlovu et al ., 2018). COMMERCIAL PRODUCTS AVAILABLE AND THEIR APPLICATIONS ACCORDING TO SUPPLIERS Only a couple of strains of M. pulcherrima have been commercialised. Amongst them

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