Primary Functions of Alcoholic Fermentation in Wine

  • Sugar Content Modification 
  • Decrease in acidity
  • Increase Phenolic Extraction
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Sugar Content Modification 
Winegrapes are predominantly composed of fructose and glucose. These sugars are converted into alcohol and, even if fermentation reaches completion, small quantities of sugars without sensory significance will remain. According to Jackson in Wine Science:

Winegrape’s sugar concentration at maturity: 19-25° Brix

Wine’s sugar concentration after vinification:
  • Fermentable sugars: preferably 1g/liter (< 0° Brix)
  • Non-fermentable sugars including arabinose, rhamnose, and xylose: (0.2 g/liter).

Higher quantities of residual sugar may be retained by prematurely stopping fermentation through chilling, centrifugation, distilled alcohol addition or filtration. Common examples include:
  • Sparkling wine, where sugar is reserved for secondary fermentation.
  • Dessert wines like port are made by the addition of distilled alcohol.
Decrease in acidity
Titratable acidity decreases and pH increases. Vilela (2017) in a literature review of deacidification by microbes noted:
  • Saccharomyces Cerevisiae metabolizes acetic acid during the fermentation process.
  • Other Saccharomyces strains like Schizosaccharomyces pombe and Saccharomyces paradoxus can deacidify wine through the metabolism of malic acid. However, the high fermentation temperatures of Schizosaccharomyces yeasts (relative to Saccharomyces cerevisiae) can adversely affect wine quality.
  • Malolactic Fermentation by selected strains of Oenococcus oeni or Lactobacillus plantarum can be used to lower acidity. Malolactic Fermentation will be discussed in depth in another issue.

For more on microbial deacidification read:
Increase Phenolic Extraction from Grape Solids
The extraction of color and aroma from grape phenolics occurs when grape skins, seeds and other solids are left in the fermentation vessel. And, as discussed in Hawaii Beverage Guide’s February issue, phenolic extraction can occur before, during and/or after fermentation.

In his PhD dissertation, Dr Patrick Setford developed a simulation of phenolic extraction during red wine fermentation. He outlined the phenolic extraction process as follows:

  • Immediately upon crushing, phenols leak from the edges of broken skin cells.

  • Concentration-driven diffusion from the grape solids into the liquid occurs throughout maceration by internal diffusion and then dissolved solute diffusion. The rate at which this occurs is limited by the plant's natural resistance to liquid penetration which limits the movement of the grape’s dissolved solids into the wine. More specifically in internal diffusion occurs in the following steps:
  1. Solvent diffusion into the porous solid
  2. Solute dissolution into the solvent
  3. Dissolved solute diffusion to the particle surface
  4. Dissolved solute diffusion from the particle surface to the surrounding solvent.

​Key factors that influence phenolic extraction
(as outlined in Setford et. al 2019):
Solvent Composition: Increased concentrations of ethanol and sulfur dioxide (SO2) increases the rate of internal diffusion; however, SO2 at levels normally associated with red wine fermentation appear to have a much lower effect.
  • Temperature: Extraction increases as temperature increases. This occurs because temperature influences the permeability of the cell membranes in the grape solids, the solubility of phenolic compounds, the rate of ethanol production during fermentation, and the viscosity and density of solvents.
  • Contact area between the grape liquid and the grape solids can impact the rate of extraction and the amount of extraction. This contact area, which is predominantly dictated by the cap management technique (e.g. manual punch-down, mechanical punch-down, pump-over and fermentation in a spiral rotor tank), influences the rate of anthocyanin extraction, however the quantity of extraction was dependent on the grape variety. Setford also noted that the amount of contact time has not been studied.
  • Reactions of extracted phenolics to the fermentation environment also influence phenolic extraction. The most notable are:
  • Oxidation, if increased during pre-fermentation, is found to decrease both the anthocyanin and total phenolic evolution.
  • Copigmentation, the process of anthocyanins being stabilized through non-covalent bonds forming between themselves or colorless cofactors, may account for up to 50% of the color observed in young red wines.  Tofalo et al (2021), in a literature review of how microorganism influence wine color, noted yeast can also:
    • Assist in pyranoanthocyanin (Anthocyanin + Hydroxycinnamate) development, especially Vitisins type A and B. This occurs through the secretion of pyruvic acid or acetaldehyde which can then bind to anthocyanins. These vitisins contribute 11-14 times more color than unmodified anthocyanins, shift the wine towards an orange-red hue, and affect the intensity and tonality of wine color by the action of β-glycosidase on anthocyanins or anthocyanidase enzymes.
    • Yeast cell walls can absorb tannins, anthocyanins and other volatile compounds that are released during maceration. The rate of absorption is influenced by temperature and ethanol concentration with higher temperatures increasing anthocyanin adsorption and higher ethanol concentrations having lower absorption.​
    • ​For more on copigmentation:  www.hawaiibevguide.com/wine-polyphenols 

Yeast produced aroma compounds

The multitude of yeast metabolic pathways that create additional aromatic compounds and what influences their development will be explored in-depth in another issue.

The following summarizes the following article which we highly recommend reading:
​Swiegers,Jan & Bartowsky, Eveline & Henschke, P.A. & Pretorius, Isak. (2005). Yeast and bacterial modulation of wine aroma and flavour. Australian Journal of Grape and Wine Research.11. 139-173. Source: www.researchgate.net/publication/280765129_Yeast_and_bacterial_modulation_of_wine_aroma_and_flavour
  • Higher Alcohols
  • Esters
  • Sulfur Compounds
  • Volatile Phenols
  • Other Compounds 
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Higher alcohols of:
  • Aliphatic alcohols: propanol, isoamyl alcohol, isobutanol and active amyl alcohol.
  • Aromatic alcohols: 2-phenylethyl alcohol and tyrosol.
  • Carbonyl compounds including acetaldehyde.
Esters:
  • Significant esters include: Ethyl acetate (fruity, solvent-like), isoamyl acetate (isopentyl acetate, pear-drops aromas), isobutyl acetate (banana aroma), ethyl caproate (ethyl hexanoate, apple aroma) and 2- phenylethyl acetate (honey, fruity, floral aromas)
  • Production includes both Saccharomyces and non-Saccharomyces strains including Hanseniaspora guilliermondii and Pichia anomala.​

Sulfur Compounds:
  • Thiols including:
  • Howell et al. (2004) found 4-mercapto-4-methylpentan-2-one (4 MMP), the varietal aromatic most notably in Sauvignon blanc, can be liberated from nonvolatile complexes with glycosides or cysteine by some yeast strains.
    This is done by the secretion of enzymes that liberate aroma compounds from glycosides (volatile aromatic compounds linked to a sugar moiety which render the aromatic compound nonvolatile/odorless). 5,6,7
  • For more on volatile thiols read:
    Heelan, A. (2015). Volatile Thiols. Waterhouse Lab. Retrieved February 20, 2022, from 
     https://waterhouse.ucdavis.edu/whats-in-wine/volatile-thiols
  • Sulfides like hydrogen sulfide (rotten egg), dimethyl sulfide (asparagus, corn molasses)
  • Sulfur-containing fusel alcohols: methionol or 3-methylthio-1-propanol (cauliflower, cabbage), 3-methylthiopropyl acetate (mushroom, garlic)
Volatile phenols
  • Produced by the decarboxylation of hydroxycinnamic acids can create barnyard-like off-odors which mask the fresh/floral characteristics of white and rosé wine.  These include
    • Ethylphenol: 4-ethylguaiacol and 4ethylphenol
    • Vinylphenol: 4-vinylguaiacol and 4-vinylphenol
​
Other influences on wine aroma by yeast include, according to Jackson in Wine Science:
  • Monoterpenoids
    • Produced by yeast species Kluyveromyces lactis, Torulaspora delbrueckii and Ambrosiozyma monospora
    • Found in hops and grapes
  • The production of fatty acids
  • Degradation of some grape aromatics, notably aldehydes. This may limit the expression of the herbaceous odor of C6 aldehydes and alcohols produced during the grape crush.
  • Aroma compounds produced from yeast autolysis, especially if the wine undergoes sur lie maturation (extended maturation on the lees).​

Wine Microbes Sources

  • Terminology of Yeast Origin
  • Laboratory Cultured Yeast
  • Epiphytic yeast and Spontaneous Fermentations
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Spontaneously Occurring Yeast vs Epiphytic Yeast vs Indigenous Yeast vs Endemic Yeast vs Native Yeast vs Wild Yeast
Though typically used interchangeably, these terms have different technical definitions. To clarify:
  • Spontaneously occurring yeast: Yeast that occurs without intentional inoculation in the wine. Used in reference to spontaneous fermentation.
  • Wild yeast: Generally used interchangeably with spontaneously occurring yeast, “wild yeast” typically refers to yeast that occurs on the grape prior to fermentation. Confusingly, wild yeast may also refer to a cultivated strain of non-Saccharomyces cerevisiae yeast because it is of the same non-Saccharomyces cerevisiae species as those found in a spontaneous fermentation. A better term for this cultivated wild yeast is wild type yeast.
  • Epiphytic: An epiphyte is an organism that grows on the surface of a plant and derives its moisture and nutrients from the air, rain, water or from debris accumulating around it. Therefore epiphytic yeast are those that grow on grapes in the vineyard.
  • Native: A species that has always been part of the particular environment.
  • Indigenous: A native species that is found in multiple locations.
  • Endemic: Native species only found in a specific and unique location.

For the purpose of this article and for Hawaii Beverage Guide’s future articles, we believe:
  • “Epiphytic yeast” is the most accurate term for yeast that are found in the vineyard on grapes because the determination of a yeast to be native, indigenous or endemic requires extensive study which is not typically performed.
  • “Spontaneously occurring yeast” is the most accurate term for yeast that generally occur in fermentation because identifying if the yeast came from the vineyard or winery equipment, then identifying whether the yeast is indigenous or endemic requires extensive study which is not typically performed.


Laboratory cultured yeast are the primary fermenters used by most commercial wineries.  Grape must is typically inoculated with laboratory cultivated yeast to make wine. Unlike the boiling wort in beer production, grape must is not sterilized before inoculation. This means epiphytic yeasts (those living on the grape in the vineyard) are typically present in a wine fermentation. Sulfur dioxide may be used to minimize undesired epiphytic yeast and its impact varies by cultivar, growing location, and winemaking style (including the duration of skin contact), clarification and other processing techniques. It should also be noted that using blends of inoculated strains or yeast species is commonplace.​
Sources of Wine Yeast

Epiphytic yeast As epiphytic and other spontaneously occurring yeast occur in all wine fermentations, there is a significant amount of academic research on what naturally occurring yeast exist and where they come from. The general answer is: There are a multitude of microbes initially present in fermented grape must, some of which have been identified. Only a few have known enological significance while the others are probably not metabolically active because of the acidic, generally anaerobic, and alcoholic conditions which are generally inhospitable to most yeasts, fungi, and bacteria (Jackson, Wine Science).

Some general trends on the sources of yeast that have known significance to winemaking according to Jackson in Wine Science are:
  • The winery building and winery equipment like crushers and presses are the major sources of yeast for spontaneous fermentation.10 This may be more prevalent in older equipment and buildings as modern wineries strive for conditions that limit unintended microbial development.
  • Damaged grapes have significantly larger, but still relatively small (compared to other microbes) populations, of epiphytic Saccharomyces cerevisiae yeasts, which may be brought to the berries by insects.11
  • Strains used in intentional inoculations of wine may spread over short distances outside the winery predominantly through water runoff, though they can spread further if the pomace is used as vineyard fertilizer.12
  • Strains can differ by region13 and vineyard management practices14 making spontaneous fermentation unique to the particular winery.

Spontaneous Fermentations
Spontaneous fermentations may result in wine that:
  • Showcases yearly variations in character because of the way the yeast act within the particular grape's chemistry
  • Showcases the wine’s “unique” terroir. However, given that Saccharomyces cerevisiae is predominantly from winery equipment rather than the vineyard, terroir’s definition needs to include the winery.
  • Has an extended lag period which, due to the low quantity of starting cells of Saccharomyces cerevisiae relative to that of inoculated fermentation, may be more susceptible to undesirable microbes. To mitigate uncertainty caused by the potential development of off-flavors, cultivated “wild yeast” can be used in inoculations.
"Wild" Wine Fermentation Microbes
Wine does not need to be inoculated.  Jackson notes in Wine Science  this is only necessary if:
  • The juice undergoes thermovinification or pasteurization.
  • To restart "stuck" fermentations.
  • To promote fermentation of juice containing a signification number of moldy grapes because these generally possesses fermentation inhibitors like acetic acid, that slow yeast growth and metabolism.
  • To create secondary fermentation in sparkling wine production.  However, the predominant reason for using a specific yeast strain is to avoid the production of undesirable flavors occasionally associated with spontaneous fermentation.​

General Microbial Development Cycle of
Inoculated Wine Fermentation (from Demuyter et al 2004)

For insight into the aroma impact of common wild yeast types:
Mateo, J. J., Jimenez, M., Huerta, T., & Pastor, A. (2002, November 13). Contribution of different yeasts isolated from musts of Monastrell grapes to the aroma of wine. International Journal of Food Microbiology. Retrieved February 20, 2022, from www.academia.edu/download/49589096/0168-1605_2891_2990102-u20161014-3577-msz7an.pdf

Selection of Yeast Strain [15]

​Yeast strain selection plays a significant role in the resulting wine aroma because yeast are responsible for the creation of aromatic compounds along with ethanol. As advancements in research by yeast procurement laboratories have allowed for winemakers to make better selections and produce more consistent wine, a particular strain’s performance is more general and unpredictable than it is definitive.  ​
  • Grape Chemistry Impact on Yeast Selection
  • Killer Factor Yeast
  • Fermentation Conditions
  • Additional Yeast Attributes
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Grape Chemistry Impact on Yeast Selection
Yeast selection is predominantly based upon cultivar and the chemical composition of the grape. Unlike the fermentation of grain, which does not have as significant a variation between aromatic compounds from year to year, grapes can vary significantly between vintages and even growing locations within a vineyard. This means the yeast used by a winery can change yearly, and though fermentation additives can be used to adjust grape must chemistry, many winemakers minimize these adjustments as they can create other challenges. According to Scott Labs and Lallemand, the grape chemical attributes to know for yeast selection are:

Fermentable sugar (Brix/Baum)
Yeast strains have varying ethanol tolerances, and the quantity of fermentable sugars is what dictates alcohol potential.

Yeast assimilable nitrogen (YAN)
Yeast strains vary in the amount of assimilable (usable) nitrogen they require for the synthesis of proteins, pyrimidine nucleotides, and nucleic acids.

The physical condition for every lot of fruit
On damaged fruit yeast may need to outcompete pre-existing microbes. There are also yeast that are more tolerant to fermentation in botrytis-infected grapes. Scott Labs annotates this trait as “Competitive Factor”.
Acidity: pH, titratable acidity (TA), and malic acid
Most yeast are tolerant to acidity within a certain pH. Additionally, yeast may be used to degrade malic acid or need to function in conditions ideal for malolactic fermentation by Oenococcus oeni.

(K+) Potassium concentration
Yeast require sufficient amounts of potassium to metabolize glucose and fructose as deficiencies may result in a stuck fermentation.16


Killer (Factor) Yeast
Yeasts possessing a “killer factor” have both a mycovirus and a satellite dsRNA which cause the secretion of a protein that kills cells not carrying the factor. ​​
Most killer proteins act only upon related yeast strains, however, some forms are known to be active against other yeast species, filamentous fungi and bacteria. Of the multiple killer factors identified, most do not affect Saccharomyces cerevisiae. The most significant that do are:
  • K1: The killer proteins produced by Saccharomyces cerevisiae act optimally at a pH above that normally found in wine, notably the K1 protein.
  • K2 killer factor: Most killer yeasts isolated from wine possess the K2 factor.

According to Jackson in Wine Science, the impact of killer yeast strains depends on juice pH, protein-binding substances like bentonite or yeast hulls, the presence of free ammonia nitrogen, and the rate of killer yeast strain growth and fermentation activity.

​A spontaneous occurrence of Saccharomyces, Kluyveromyces apiculata or Zygosaccharomyces bailii may replace the inoculated strains, cause sluggish or stuck fermentations, or create undesirable sensory attributes. To mitigate this situation Jackson in Wine Science suggests:
  • The usage of commercial yeast which can be developed, using gene replacement, to contain both K1 and K2.18,19
  • Sulfur dioxide used in combination with killer toxin resistant wine yeast strains.
  • Temperature control
  • Limiting prefermentative clarification (restrict nutrient loss), aeration (5 mg O2/liter) at the end of exponential cell growth.
  • The addition of ergosterol or long-chain unsaturated fatty acids (i.e., oleic, linoleic, or linolenic acids), yeast ghosts or other absorptive materials, such as bentonite, ammonium salts, absorptive substances like yeast hulls or bentonite to reduce the activity of killer proteins.
Fermentation Conditions
Yeast have different requirements regarding the conditions they need to survive and thrive. The nuances of this will be discussed in a later issue of Hawaii Beverage Guide. These conditions are generally:
  • Fermentation Vessel: Scott Labs, for example, includes “Suitable for Barrel Fermentation” as one of their data points for yeast.
  • Temperature control: Different yeast strains have different temperature tolerances. Ideally fermentation occurs near the middle of the temperature range in order to avoid stuck fermentations.
Additional Yeast Attributes
The more nuanced sensory attributes of yeast include its sensory impact. Scott Lab’s Yeast Choosing Guide also includes the following data points.
  • Enhanced mouthfeel (glycerin)
  • Aroma Characteristic Additions of: Varietal Enhancement, Esters (Fruit), Green (Thiols), Tropical (Thiols), Citrus (Esters and Thiols), Floral, Nutty, Mineral/Freshness, Spicy, No to Low Hydrogen Sulfide Production

​Wine Yeast Species 

Kingdom Fugi
SubKingdom: Dikarya
Division-Ascomycota
Class: ascomycete yeasts
These yeast are known as imperfect yeast/ imperfecti/ imperfect fungi as they do not fit into the common taxonomic classification system. Importantly, they have lost the ability to undergo sexual reproduction and instead asexually “clone” themselves via meiosis and cytoplasmic division. They also unusually use fermentative metabolism instead of aerobic respiration and have a high ethanol tolerance.

Over the past few decades yeast identification has changed from the usage of visual observation to using genetic mitochondrial DNA, PCR, and other molecular technologies to identify different yeasts.

Great list of wine yeast and insight into their enogolocal usage:
UC Davis Viticulture and Enology
wineserver.ucdavis.edu/industry-info/enology/wine-microbiology/yeast-mold

The Australian Wine Research
Institute's Available Microbial Strains
www.awri.com.au/research_and_development/wine-microorganism/winemaking-yeast-and-bacterial-strains/

​Primary Fermenters: Saccharomyces Strains

​Saccharomyces is preadapted to thriving in grape must and excludes other potential microbial competitors. [20] This is because it has a fermentative metabolism that produces as many ATP per second as is normally generated by respiration [21]; properties of osmotolerance; relative insensitivity to high acidity; acceptance of low oxygen concentrations; a higher tolerance for the byproducts of fermentation; and defensive strategies mediated by different mechanisms like cell-to-cell contact and secretion of antimicrobial peptides to combat other microorganisms.
  • Saccharomyces cerevisiae
  • S. bayanus and S. uvarum
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Saccharomyces cerevisiae
Though all S. cerevisiae possess the same enzymes, their rate of catalytic activities and fermentation by-products may vary in the same environmental conditions because of slight variations in regulation, or gene copy number. In wine fermentation, strains are typically categorized into those used to make white white wine and those used to make red wine. Most generally this is due to differences in the particular strain’s ideal fermentation temperature and pH range. According to Jackson:
  • Ethanol: 12-18%
  • Temperature 
    • White Wine
      • 10–15 ºC to encourage the production and retention of fruit esters
      • 15– 20 ºC to encourage the development of the varietal fragrance in certain cultivars.
    • Red Wine: 20–30 ºC is typical.
  • pH
    • White Wine (final pH): 3.0 - 3.4
    • Red Wine (final pH): 3.3 - 3.6
  • SO2: High Tolerance to SO2. This is what makes it an effective enological tool.


Saccharomyces bayanus and Saccharomyces uvarum
Origin: The origin of these two species, often considered strains, is unknown, as are their natural habitats. Wild strains have occasionally been isolated from the caddis fly, some mushroom species, and hornbeam tree exudate (Wine Science).

​Both of these Saccharomyces strains are capable of effective alcohol fermentation in place of Saccharomyces cerevisiae, and their usage is dependent on the desired winemaking technique. A study by Castellari et al., 1994 found that cryotolerant Saccharomyces cerevisiae p.r. uvarum and bayanus strains differ from ordinary non-cryotolerant strains in their ability to synthesize rather than decompose malic acid, their higher glycerol and succinic acid production, their lower acetic acid production and their lower ethanol yield.22

Saccharomyces bayanus var. bayanushas
Enological Significance
  • Ethanol: High tolerance
  • Temperature: Optimal temp: 29-30ºC
  • pH: Low pH tolerant
  • SO2: No (High tolerance)
  • Other
    • Well adapted to the production of sparkling wines and fino sherries (Wine Science).
    • Antonelli et al (1999) found Saccharomyces bayanus yeasts was characterized by the production of compounds such as 2-phenylethanol, 2-phenethyl acetate, ethyl lactate, 3-ethoxypropanol, and to a lesser extent, diethyl succinate and propionic acid while not producing undesirable compounds like acetic acid and sulfur anhydride. [23]

Saccharomyces bayanus var. uvarum
Enological Significance
  • Ethanol: High tolerance
  • Temperature: Ideal growth range of 25–30 °C, however it is more cryotolerant than mesophilic yeast like Saccharomyes cerivisae. This makes it useful for the cool fermentations (6-15 ºC) employed in tokaji, amarone, sauternes, and table wines. It is also found in cold climate regions like Alsace (Demuyter et al 2004).
  • pH: Low pH tolerant
  • SO2: No (High tolerance)
  • Other: Has the potential to synthesize desirable aroma compounds. For example Massoutier et al., 1998 found that regardless of temperature, cryotolerant yeasts (SY055 and 12233) produced twice as many isobutyl and isoamyl alcohols as mesophilic yeast and 2-phenethyl alcohol was produced by cryotolerant yeasts at levels 4 times as high as by mesophilic yeasts.

​Other Saccharomyces strains

  • S. pombe
  • S. paradoxus
  • S. ​cariocanus
  • Saccharomyces hybridization
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Schizosaccharomyces pombe
  • Enological Significance: Though usually considered a spoilage organism because of its production of undesirable compounds like hydrogen sulfide, Schizosaccharomyces pombe can decarboxylate malic acid to lactic acid.25 An experiment by Silva et al., 2003 showed that by encapsulating Schizosaccharomyces pombe in alginate beads, deacidification can occur while the aromatic compounds (hydrogen sulfide, acetaldehyde, methanol, isopropanol, amyl and iso-amyl alcohols) at the end of the alcoholic fermentation did not show a significant difference in comparison to the control fermentation.26
  • For a insightful literature review on Saccharomyces pombe read: Loira I, Morata A, Palomero F, González C, Suárez-Lepe JA. Schizosaccharomyces pombe: A Promising Biotechnology for Modulating Wine Composition. Fermentation. 2018; 4(3):70. https://doi.org/10.3390/fermentation4030070
  • Example Commercial Source: Scott Labs ProMalic®
Saccharomyces paradoxus
This species has the most genetic overlap with Saccharomyces cerevisiae27 and while not commonly found in vineyard environments it is widespread. There is even a strain of Saccharomyces paradoxus unique to Hawaii (72-145). [28]

Enological Significance
  • As fermentation starters because of their significant higher synthesis of glycerol and lower production of volatile acidity than Saccharomyces cerevisiae.29
  • To degrade malic acid. [30]
  • Example Commercial Source: Anchor Oenology’s Exotics Mosaic S. cerevisiae x S. Paradoxus https://hop.scottlab.com/exotics-mosaic-yeast-exotics
Saccharomyces cariocanus
Though no commercial strains exist of Saccharomyces cariocanus, a cross with Saccharomyces Cerevisiae called AWRI2794 has been developed by the Australian Wine Research Institute. In the production of a Tempranillo wine, the cerevisiae parent was described as ‘spicy’ with a ‘rich midpalate’ whereas the cariocanus hybrid was noted as being more complex with a softening of the palate.31 In the commercial product by Anchor Oenology called Exotics Novello, the strains is said to:
  • Have some pectinase activity.
  • Be a thiol revealer and ester producer in both white and red wines.
  • Reveal fresh fruit and floral esters while decreasing astringency and bitterness in whites.
  • Increase red and black fruits and spice while diminishing green and vegetal characters in reds.
  • Have improved mouthfeel and softness.
  • Produce low levels of Volatile Acidity and Hydrogen Sulfide.

Commercial Example: Anchor Oenology Exotics Novello
Saccharomyces hybridization
Though yeast will always reproduce asexually under winemaking conditions due to glucose catabolite repression, nutrient starvation can induce yeasts of the same genus (i.e. Saccharomyces) to reproduce sexually and form inter-species hybrids that can contain characterists of the parent strains.32 For more insight into the occurrence of natural hybrids read: González, S. S., Barrio, E., Gafner, J., & Querol, A. (2006). Natural hybrids from Saccharomyces cerevisiae, Saccharomyces bayanus and Saccharomyces kudriavzevii in wine fermentations. FEMS Yeast Research, 6(8), 1221-1234.

Source: https://academic.oup.com/femsyr/article/6/8/1221/570550

Common Non-Saccharomyces Yeast in Wine Fermentation

Non-Saccharomyces species which are spontaneously occurring or cultivated from “wild” origins can act as co-fermenters. Those cultivated and used as purposeful inoculants increases the probability that the specific strain and its features will be present in the wine rather than relying on a hope and prayer for the desired wild yeast to propagate.33 Common non-saccharomyces yeast used in winemaking include:

  • Aureobasidium spp.
  • Brettanomyces /Dekkera bruxellensis
  • Botrytis cinera (Noble Rot)
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​Aureobasidium spp. [34] It is found on grapes and can act as a natural biocontrol agent for Aspergillus and other grape vine pathogenic fungi and it can degrade Ochratoxin A. However, it does not survive in wine.
Brettanomyces /Dekkera bruxellensis35
Sources:
  • Grape surfaces (in the vineyard)
  • Contaminated (with Brettanomyces) winery equipment
  • Oak barrels if sanitation is neglected.
  • Spreads across wineries through contaminated wine and equipment and is difficult to control because it can go unnoticed till wine is permanently tainted.
  • Insects, especially fruit flies, are significant transmitters.

Enological Significance
  • Ethanol: Is fermentative up to 10-13% ABV
  • Heat: Thermal inactivation typically at temperatures over 50°C with heat sensitivity increasing in the presence of ethanol. 10-15°C inhibits growth
  • pH: Prefers acidic environments though values below 3.5 inhibit growth.
  • SO2: 0.8ppm of sulfur dioxide has been reported to be effective in inhibiting growth of the species.
  • Other:
    • Viewed as a contaminant because it can produce undesirable volatiles including:
    • 4-EP (ethyl phenol): Band-aid, elastoplastic
    • 4-EG (ethyl guaiacol): Smoky, spicy, cloves
    • 4-EC (ethyl catechol): Sweaty, horsey
    • Isovaleric acid: Rancid, cheesy, vomit
    • Combination of all the above: Horsey, barnyard, moldy
    • Competes with the Saccharomyces for the nutrients in the must thereby limiting its fermentation characteristics.
    • Principally a problem in red wines.
    • Prevention strategies include: improved hygiene, monitoring of nutrients and residual sugars during and at the end of fermentation, temperature control, use of sulfur dioxide, avoidance of old oak barrels, and any contaminated equipment or wine must be quarantined and contact with any other wine or equipment must be avoided.
Botrytis cinera (Noble Rot) [36]
Source: Common grape flora. Cold and wet climates can lead to the development of unwanted molds like Penicillium, Mucor, and Aspergillus sp. which can outcompete the botrytis cinerea.

Enological Significance:
Infected grapes are commonly used to produce Sauternes in France, and Trokenbeerenausleses and Tokay Asza wines in Hungary by over maturation.
  • Candida stellata
  • Candida krusei/Pichia kluyver
  • Candida pulcherrima/ Metschnikowia pulcherrima
  • Candida colliculosai
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Candida stellata [37]
Source: Normal grape/fermentation flora and typically found in highest numbers closer to the start of fermentation.

Enological Significance
  • Ethanol: Not sensitive
  • Heat: Growth at 37 °C, though variable, it can grow at higher pHs and flourish in wines fermenting at low temperatures. The population increases in cold soak wines and wines at low temperatures for solid settling.
  • pH: Sensitive to low pH
  • SO2: Somewhat sensitive
  • Other
    • C. stellata can appear as a white, cheese-like scum, on wine surfaces but does not seem to create substantial negative sensory or stability problems.
    • May enhance the sensory profile of wines with characteristics such as honey, apricot and sauerkraut.

Candida krusei/Pichia kluyveri38
Source: Normal grape/fermentation flora

Enological Significance
  • Ethanol: Tolerance up to 6.6% and can co-ferment a wine especially at low temperature however its presence in wine contributes to slime formation and ropiness.
  • Heat: 10-42°C optimum
  • pH: High tolerance
  • SO2: Sensitive
  • Other: Can be used in cacao fermentation.

​For a literature review on Candida krusei/Pichia kluyveri read:
Vicente, J., Calderón, F., Santos, A., Marquina, D., & Benito, S. (2021). High Potential of Pichia kluyveri and Other Pichia Species in Wine Technology. International journal of molecular sciences, 22(3), 1196. 
Source: www.ncbi.nlm.nih.gov/pmc/articles/PMC7866185/
Candida pulcherrima/ Metschnikowia pulcherrima39
Source: Normal grape/fermentation flora
Enological Significance
  • Ethanol: If present on the grapes or winery equipment at crush, it may be involved in the vinification of wine until 5% ABV. At concentrations in excess of 5%, Metschnikowia begins to die off.
  • ​Temperature: Maximum temperature for growth ~39 °C. The upper temperature limit for sporulation lies below 25 °C on all but the most favorable media.
  • pH: Can tolerate low pHs, but a pH over 3.6 allows for competition from bacteria and other microorganisms and a pH over 7 is inhibitory.
  • Other: Secretes acid proteases which may be effective at the degradation of wine proteins.
Torulaspora delbruecki/ Candida colliculosai [48,49]

Enological Significance (from CHR Hansen):
  • Consumes some sugar to alleviate osmotic (high sugar) stress on Saccharomyces. This makes it suitable for late harvest, icewine and high sugar fermentations where Volatile Acidity can be a challenge.
  • Produces polysaccharides, esters and complexity in white, rosé and red wines.

Commercial Strain:
  • Lallemand Biodiva
  • CHR-Hansen Prelude
  • ​Debaryomyces hansenii
  • Hansenula anomala
  • Hanseniaspora uvarum/Kloeckera apiculata
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>
​Debaryomyces hansenii [40]
Source: Normal grape/fermentation flora.

Enological Usage
  • Ethanol: Not Sensitive
  • Heat: Growth at 37-40 °C, none above 42 °C
  • pH: Not Sensitive (unless extreme)
  • SO2: Not Sensitive
  • Other: Produces high levels of β-glucosidases, which creates more monoterpenol in the wine, affecting sensory results. Sensory effects include higher terpenol flavors, especially linalool.

​For more read:
Garcia, A., Carcel, C., Dulau, L., Samson, A., Aguera, E., Agosin, E., & Günata, Z. (2006, July 20). Influence of a mixed culture with Debaryomyces vanriji and saccharomyces cerevisiae on the volatiles of a Muscat wine. Institute of Food Technologists. Retrieved February 20, 2022, from https://academic.oup.com/femsyr/article-pdf/10/2/123/18019536/10-2-123.pdf
Hansenula anomala [41]
Source: Normal grape/fermentation flora especially in Botrytis-infected fruit.

Ecological Significance
  • Ethanol: Can ferment glucose, sucrose, sometimes galactose, maltose and produces ethanol under anaerobiosis
  • Temperature: N/A
  • pH: N/A
  • SO2: N/A
  • Other:
    • May create: Excessive acetic acid and ethyl acetate production, pellicle formation, and acid metabolism (increasing pH).
    • An effective biocontrol agent of mold on grapes/grapevine and can produce killer factor effective against Dekkera/Brettanomyces spp.

Hanseniaspora uvarum/Kloeckera apiculata [42]
Source: Normal grape/fermentation flora and present early in the fermentation

Enological Significance:
  • Ethanol: H. uvarum: 3.4-6.7% K. appiculata: 9-12.5% temperature dependent
  • Heat: Growth: 8°C-36°C. Survives 20 min at 55°C, not 10 min at 60°C
  • pH: 1.5-7.5 K. appiculata more tolerant at low pHs
  • SO2: 100 mg/L
  • Other
    • Produces ethyl acetate
    • Secrets acid proteases which was found to effectively degrade wine proteins.​​
For more insight into non-Saccharomyces yeast utilization in wine read:
Scott Laboratories. (2021, June). Harnessing the Unique Powers of Non-Saccharomyces Yeasts. Scott Laboratories. Retrieved February 10, 2022, from https://scottlab.com/harnessing-the-unique-powers-of-non-saccharomyces-yeasts

Vilela, Alice. 2019. "Use of Nonconventional Yeasts for Modulating Wine Acidity" Fermentation 5, no. 1: 27. https://doi.org/10.3390/fermentation5010027

​Neil P. Jolly, Cristian Varela, Isak S. Pretorius, Not your ordinary yeast: non-Saccharomyces yeasts in wine production uncovered, FEMS Yeast Research, Volume 14, Issue 2, March 2014, Pages 215–237, https://doi.org/10.1111/1567-1364.12111​
  • Lachancea thermotolerans/Kluyveromyces thermotolerans
  • Metschnikowia fructicola
  • ​Metschnikowia pulcherrima
  • Pichia membranaefaciens
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>
Lachancea thermotolerans/Kluyveromyces thermotolerans [43]
Source: Normal grape/fermentation flora and present in some commercial yeast inoculates.

Enological Significance:
  • Ethanol:
    • Often found at the beginning of many fermentations but sensitive to ethanol.
    • Fermentation by Kluyveromyces thermotolerans may produce wine with different sensory properties including a different mouth feel due to production of lactic acid.
  • Heat: Growth at 37 °C
  • pH: Is sensitive to very low pH levels
  • SO2: Slightly sensitive
  • Other [44]
    • Converts glucose to lactic acid. This may help to acidify low acid musts while adding freshness and complexity.
    • May produce wines with increased concentrations of lactic acid, glycerol and 2-phenylethanol during mixed fermentations of grape musts.
    • The later a L. thermotolerans ferment is inoculated with S. cerevisiae the more lactic acid and glycerol the final wine will contain.

Commercial Strains:
  • CHR Hansen Concerto
  • CHR Hansen Octave
  • Lallemand Laktia
Metschnikowia fructicola45
  • Enological Significance: (of IOC Gaia from Scott Labs): Bioprotectant against volatile acidity producing native microflora. It can be added to red, white or rosé juices for protection during transportation or to protect red grapes during cold soak.
  • Commercial Strain: IOC Gaia

​Metschnikowia pulcherrima [46]
Source: Can be present on the grapes or winery equipment at crush.

Enological Significance:
  • Ethanol: Can be involved in fermentation until ethanol reaches a concentration in excess of 5%.
  • Temperature: Maximum temperature for growth ~ 39 °C.
  • pH: Can tolerate low pHs, but a pH over 3.6 allows for competition from bacteria and other microorganisms and a pH over 7 is inhibitory.
  • SO2: Sensitive
  • Other: Secretes acid proteases which may effectively degrade wine proteins.
  • Commercial Strains
    • Lallemand Flavia: Acts as a bioprotectant by utilizing oxygen as a growth factor and inhibiting volatile acidity producing native microflora. This oxygen scavenging protects white and rosé juice from oxidative damage and microbial spoilage.
    • Lallemand Initia: Produces enzymes which cleave aroma precursors to reveal the tropical, citrus and floral notes from terpenes and thiols in certain white and rosé wines.
Pichia membranaefaciens [47]
Can cause a yeast film to form and off aromas.

Wine Yeast Producers

Bacteria Fermentation

Purposeful inoculation with a lactic acid strain of bacteria, commonly Oenococcus oeni, is used to convert malolactic acid to lactic acid. This process and its nuanced impact on wine will be covered in a future issue of Hawaii Beverage Guide. However, most bacteria are unwanted and considered spoilage organisms. Often, these are acetic acid bacteria of the species Acetobacter and Glucanoacetobacter which turn ethanol into vinegar. Pediococcus damnosus is unwanted because it produces diacetyl, an unwanted buttery off-flavor.
UC Davis also has a great list of common wine bacteria which can be found here:
wineserver.ucdavis.edu/industry-info/enology/wine-microbiology/bacteria​

​Inoculation Process

Most wine is inoculated with cultured yeast to increase the likelihood that the desired yeast completes the fermentation and not another random yeast. To achieve this, a Saccharomyces cerevisiae population of about 105–106 cells/ml is desired, if using active dry yeasts, this corresponds to about 0.1–0.2 g/liter of must (active dry yeast often contains about 20–30 109 cells/g) (Wine Science).
  • Rehydration and Inoculation
  • Repeat use of yeasts in Wine fermentation
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>
Rehydration and Inoculation Process:
  1. Dry yeast is placed in water or dilute juice. For optimal rehydration of yeast, temperatures between 38 and 40 ºC for 20 min was found by Kraus et al 1981.50 Lallemand recommends 35-40 ºC water for 20 min in their Yeast Security Optimization process.
  2. To acclimate yeast to the grape must temperature, must is slowly added until the difference between yeast suspension temperature and must temperature does not exceed 10 °C. Steps 1 and 2 should not exceed 45 minutes.

    2a) If liquid yeast is used Step 1 is skipped. Liquid yeast, however, is more common in breweries than wineries.

  3. The yeast is then gradually integrated into the must.

  4. For a real example of the yeast hydration process for wine read Lallemand’s Yeast Security

Optimization (YSEO):
www.lallemandwine.com/en/north-america/expertise-document/yseo-process/

​Pies de Cuba inoculation [51]
Already fermenting must is added in a ratio of 2 to 10% of the total volume of liquid to inoculate clarified must. This speeds up the start of fermentation and simultaneously provides the opportunity to introduce a previously selected strain of yeast as a fermenting agent. Although the pie de cuba operation is sometimes carried out using spontaneous yeasts, more and more registered Denomination firms are opting to use indigenous yeasts selected to produce sherry wines of the best oenological and sensorial characteristics.


Repeat use of yeasts in Wine fermentation
Instead of rehydrating yeast, they can be isolated and used to inoculate successive fermentations by filtration, centrifugation, or spontaneous sedimentation.
Benefits, according to Jackson in Wine Science, include a reduction in:
  • Yeast inoculation costs, as the yeast does not have to be purchased from the supplier
  • The duration of fermentation and an increase in ethanol productivity and yield. [52]
  • Sulfur dioxide synthesis of sulfur dioxide in wine.

​Limitations
  • “Flavor drift”, caused by a change in the genetic characteristics of the general yeast population, results in a different outcome than that of the initial fermentation. This is seen more readily in beer which is often brewed in successive batches as many brewers will use yeast 7-15 generations before repitching. Wine is different, as batches are typically made at a rate of one per harvest, and there is typically only one harvest per year.
  • The rate of yeast multiplication overtime occurs at progressively reduced rates thereby increasing the potential for contamination by undesirable yeasts and bacteria.
  • Occasions when inoculation is necessary for alcoholic fermentation:
  • If the juice undergoes thermovinification or pasteurization.
  • To restart “stuck” fermentation.
  • To promote fermentation of juice containing a significant number of moldy grapes, because these generally possess fermentation inhibitors like acetic acid, that slow yeast growth and metabolism.
  • To create secondary fermentation in sparkling wine production. However, the predominant reason for using specific yeast strains is to avoid the production of undesirable flavors occasionally associated with spontaneous fermentation (Wine Science).

​Sources and Suggested Reading

1. Tofalo, R., Suzzi, G., & Perpetuini, G. (2021). Discovering the Influence of Microorganisms on Wine Color. Frontiers in Microbiology, 12, 790935-790935. https://doi.org/10.3389/fmicb.2021.790935

2. UC Davis Waterhouse Lab & Nguyen, T. (2016). Vitisins. Waterhouse Lab. Retrieved February 20, 2022, from https://waterhouse.ucdavis.edu/whats-in-wine/vitisins

3. Wine analysis. ETS Laboratories. (n.d.). Retrieved February 20, 2022, from www.etslabs.com/analyses/%234VP

4. Kate S. Howell, Jan H. Swiegers, Gordon M. Elsey, Tracey E. Siebert, Eveline J. Bartowsky, Graham H. Fleet, Isak S. Pretorius, Miguel A. de, Variation in 4-mercapto-4-methyl-pentan-2-one release by Saccharomyces cerevisiae commercial wine strains, FEMS Microbiology Letters, Volume 240, Issue 2, November 2004, Pages 125–129, https://doi.org/10.1016/j.femsle.2004.09.022

5. Liu, J., Zhu, X.-L., Ullah, N. and Tao, Y.-S. (2017), Aroma Glycosides in Grapes and Wine. Journal of Food Science, 82: 248-259. https://doi.org/10.1111/1750-3841.13598

6. Cavazza, A., Versini, G., Serra, A. D., & Romano, F. (1989, January 1). Characterization of six saccharomyces cerevisiae strains on the basis of their volatile compounds production, as found in wines of different aroma profiles: Semantic scholar. Semantic Scholar. Retrieved February 20, 2022, from https://www.semanticscholar.org/paper/Characterization-of-six-Saccharomyces-cerevisiae-on-Cavazza-Versini/6b99e01088fa6534b9337123e627fb2c02720ba6

7. Ferreira, A. M., Climaco, M. C., & Faia, A. M. (2000, September). The role of non-Saccharomycesspecies in releasing glycosidic bound fraction of grape aroma components – a preliminary study. The role of non-saccharomyces species in releasing glycosidic bound fraction of grape aroma components - a preliminary study. Retrieved February 20, 2022, from https://sfamjournals.onlinelibrary.wiley.com/doi/epdf/10.1046/j.1365-2672.2001.01348.x

8. Demuyter, C., Lollier, M., Legras, J. L., & Jeune, C. L. (2004, September 22). Predominance of saccharomyces uvarum during spontaneous alcoholic fermentation, for three consecutive years, in an Alsatian winery. Society for Applied Microbiology. Retrieved February 20, 2022, from https://sfamjournals.onlinelibrary.wiley.com/doi/abs/10.1111/j.1365-2672.2004.02394.x

9. Heard, G. and Fleet, G. (1988), The effects of temperature and pH on the growth of yeast species during the fermentation of grape juice. Journal of Applied Bacteriology, 65: 23-28. https://doi.org/10.1111/j.1365-2672.1988.tb04312.x

​10. Santamaría, P., Garijo, P., López, R., Tenorio, C., & Gutiérrez, A. R. (2005). Analysis of yeast population during spontaneous alcoholic fermentation: effect of the age of the cellar and the practice of inoculation. International journal of food microbiology, 103(1), 49-56. https://www.academia.edu/10252345/Analysis_of_yeast_population_during_spontaneous_alcoholic_fermentation_Effect_of_the_age_of_the_cellar_and_the_practice_of_inoculation

​11. Mortimer, R., & Polsinelli, M. (1999). On the origins of wine yeast. Research in microbiology, 150(3), 199-204. https://doi.org/10.1016/S0923-2508(99)80036-9
​12. Valero, E., Schuller, D., Cambon, B., Casal, M., & Dequin, S. (2005, July). Dissemination and survival of commercial wine yeast in the vineyard: A large-scale, three-years study. FEMS yeast research. Retrieved February 20, 2022, from https://academic.oup.com/femsyr/article/5/10/959/639836

​13. Khan, W., Augustyn, O. P. H., Van der Westhuizen, T. J., Lambrechts, M. G., & Pretorius, I. S. (1970, January 1). Geographic distribution and evaluation of saccharomyces cerevisiae strains isolated from vineyards in the warmer, inland regions of the Western Cape in South Africa. SUNScholar. Retrieved February 20, 2022, from https://scholar.sun.ac.za/handle/10019.1/101714

​14. Grangeteau, C., Roullier-Gall, C., Rousseaux, S., Gougeon, R. D., Schmitt-Kopplin, P., Alexandre, H., & Guilloux-Benatier, M. (2017). Wine microbiology is driven by vineyard and winery anthropogenic factors. Microbial biotechnology, 10(2), 354-370. https://sfamjournals.onlinelibrary.wiley.com/doi/pdfdirect/10.1111/1751-7915.12428

15. Scott Laboratories. (2021, July). Scott Labs Yeast Choosing Guide. Scott Labs. Retrieved February 25, 2022, from scottlab.com/scott-labs-yeast-choosing-guide

16. Thoukis, G., Ueda, M., & Wright, D. (1965). The formation of succinic acid during alcoholic fermentation. American Journal of Enology and Viticulture, 16(1), 1-8. https://
www.ajevonline.org/content/16/1/1.short

17. Magliani, W., Polonelli, L., Bertolotti, D., Gerloni, M., & Conti, S. (1997, July). Yeast Killer Systems. Clinical microbiology reviews. Retrieved February 20, 2022, from https://pubmed.ncbi.nlm.nih.gov/9227858

18. Boone, C., A-M. Sdicu, J. Wagner, R. Degre, C. Sanchez, and H. Bussey. "Integration of the yeast K1 killer toxin gene into the genome of marked wine yeasts and its effect on vinification." American Journal of Enology and Viticulture 41, no. 1 (1990): 37-42. https://
www.ajevonline.org/content/41/1/37.short

19. Baeza, M. B., Sanhueza, M. S., & Cifuentes, V. C. (2008, February). (PDF) occurrence of killer yeast strains in industrial and ... Research Gate. Retrieved February 20, 2022, from https://www.researchgate.net/publication/23411432_Occurrence_of_killer_yeast_strains_in_industrial_and_clinical_yeast_isolates

20. Pfeiffer, T., Schuster, S., & Bonhoeffer, S. (2001). Cooperation and competition in the evolution of ATP-producing pathways. Science, 292(5516), 504-507. www.researchgate.net/publication/12050915_Cooperation_and_Competition_in_the_Evolution_of_ATP-Producing_Pathways

21. Albergaria, H., & Arneborg, N. (2016, January 5). Dominance of saccharomyces cerevisiae in alcoholic fermentation processes: Role of physiological fitness and microbial interactions - applied microbiology and biotechnology. SpringerLink. Retrieved February 25, 2022, from https://link.springer.com/article/10.1007/s00253-015-7255-0

22. Castellari, L., Ferruzzi, M., Magrini, A., Giudici, P., Passarelli, P., & Zambonelli, C. (1994). Unbalanced wine fermentation by cryotolerant vs. non-cryotolerant Saccharomyces strains. Vitis, 33(1), 49-52. https://doi.org/
​10.5073/vitis.1994.33.49-52

23. Antonelli, A., Castellari, L., Zambonelli, C., & Carnacini, A. (1999). Yeast influence on volatile composition of wines. Journal of Agricultural and Food Chemistry, 47(3), 1139-1144. https://doi.org/10.1021/jf9807317

​24. Massoutier, C., Alexandre, H., Feuillat, M., & Charpentier, C. C. (n.d.). (PDF) isolation and characterization of cryotolerant ... ResearchGate. Retrieved February 20, 2022, from   www.researchgate.net/publication/256766647_Isolation_and_characterization_of_cryotolerant_Saccharomyces_strains

25. Rankine, B. C. (1968). The importance of yeasts in determining the composition and quality of wines. Vitis, 7(1), 22-49. Source:   https://ojs.openagrar.de/index.php/VITIS/article/view/7432
​26. Silva, S., Ramón-Portugal, F., Andrade, P., Abreu, S., de Fatima Texeira, M., & Strehaiano, P. (2003). Malic acid consumption by dry immobilized cells of Schizosaccharomyces pombe. American journal of enology and viticulture, 54(1), 50-55. https://www.ajevonline.org/content/54/1/50

27. Rokas, A., Williams, B. L., King, N., & Carroll, S. B. (2003). Genome-scale approaches to resolving incongruence in molecular phylogenies. Nature, 425(6960), 798-804.

28. Naumov, G. I. (1999). Divergent population of Saccharomyces paradoxus in the Hawaiian Islands: an in statu nascendi yeast species. Dokl. Biol. Sci.(Engl. Transl.), 364, 51-53.

29. Orlić, S., Arroyo-López, F., Huić-Babić, K., Lucilla, I., Querol, A. and Barrio, E. (2010), A comparative study of the wine fermentation performance of Saccharomyces paradoxus under different nitrogen concentrations and glucose/fructose ratios. Journal of Applied Microbiology, 108: 73-80. https://doi.org/10.1111/j.1365-2672.2009.04406.x

30. Redzepovic, S., S. Orlic, A. Majdak, B. Kozina, H. Volschenk, and M. Viljoen-Bloom. "Differential malic acid degradation by selected strains of Saccharomyces during alcoholic fermentation." International journal of food microbiology 83, no. 1 (2003): 49-61. www.academia.edu/download/42856797/Differential_malic_acid_degradation_by_s20160219-27802-142n5gn.pdf

31. Curtin, C., Australian Grape and Wine Authority, Australian Wine Research Institute, & Schmidt, S. (2017, September 22). Enhanced winemaking outcomes and wine style diversification through provision of fit for purpose yeast starter cultures. Wine Australia. Retrieved February 25, 2022, from www.wineaustralia.com/getmedia/2443715e-7bc3-43d9-b7a4-a94d93371efd/AWR-1301-Final-Report

32. Scottlabsltd.com. Scott Labs Canada. (n.d.). Retrieved February 25, 2022, from https://scottlabsltd.com/content/files/Documents/SLL/Handbooks/Scott%20CAN%202021%20Winemaking%20HB%20060121%20rev2.pdf

33. Scott Laboratories. (2021, June). Harnessing the Unique Powers of Non-Saccharomyces Yeasts. Scott Labs. Retrieved February 25, 2022, from https://scottlab.com/harnessing-the-unique-powers-of-non-saccharomyces-yeasts

34. UC Davis. (2018, March 21). Aureobasidium spp. Department of Viticulture and Enology. Retrieved February 25, 2022, from https://wineserver.ucdavis.edu/industry-info/enology/wine-microbiology/yeast-mold/aureobasidium-spp

35. UC Davis. (2018, March 20). Dekkera bruxellensis. Department of Viticulture and Enology. Retrieved February 25, 2022, from https://wineserver.ucdavis.edu/industry-info/enology/wine-microbiology/yeast-mold/dekkera-bruxellensis

36. UC Davis. (2018, March 9). Botrytis cinerea. Department of Viticulture and Enology. Retrieved February 20, 2022, from wineserver.ucdavis.edu/industry-info/enology/wine-microbiology/yeast-mold/botrytis-cinerea

37. UC Davis. (2018, March 9). Candida stellata. Department of Viticulture and Enology. Retrieved February 20, 2022, from wineserver.ucdavis.edu/industry-info/enology/wine-microbiology/yeast-mold/candida-stellata
​38. UC Davis (2018, March 9). Candida krusei. Viticulture and Enology. Retrieved February 25, 2022, from https://wineserver.ucdavis.edu/industry-info/enology/wine-microbiology/yeast-mold/candida-krusei

39. UC Davis (2018, March 20). Metschnikowia pulcherrima. Viticulture and Enology. Retrieved February 25, 2022, from https://wineserver.ucdavis.edu/industry-info/enology/wine-microbiology/yeast-mold/metschnikowia-pulcherrima

40. UC Davis. (2018, March 20). Debaromyces hansenii. Viticulture and Enology. Retrieved February 25, 2022, from https://wineserver.ucdavis.edu/industry-info/enology/wine-microbiology/yeast-mold/debaromyces-hansenii

41. UC Davis. (2018, March 20). Hansenula anomala. Department of Viticulture and Enology. Retrieved February 20, 2022, from https://wineserver.ucdavis.edu/industry-info/enology/wine-microbiology/yeast-mold/hansenula-anomala

42. U.C. Davis (2018, March 20). Hanseniaspora Uvarum. Viticulture and Enology. Retrieved February 25, 2022, from https://wineserver.ucdavis.edu/industry-info/enology/wine-microbiology/yeast-mold/hanseniaspora-uvarum

43. Hakim, S. (2018, March 20). Kluyveromyces thermotolerans. Viticulture and Enology. Retrieved February 25, 2022, from wineserver.ucdavis.edu/industry-info/enology/wine-microbiology/yeast-mold/kluyveromyces-thermotolerans

44. Neil P. Jolly, Cristian Varela, Isak S. Pretorius, Not your ordinary yeast: non-Saccharomyces yeasts in wine production uncovered, FEMS Yeast Research, Volume 14, Issue 2, March 2014, Pages 215–237, https://doi.org/10.1111/1567-1364.12111

45. Scott Laboratories. (n.d.). Gaia. Scott Laboratories. Retrieved February 20, 2022, from https://shop.scottlab.com/yeast/gaia-500g-015686
​46. UC Davis. (2018, March 20). Metschnikowia pulcherrima. Department of Viticulture and Enology. Retrieved February 25, 2022, from wineserver.ucdavis.edu/industry-info/enology/wine-microbiology/yeast-mold/metschnikowia-pulcherrima

47. Rankine, B. C. (1966). Pichia membranaefaciens, a yeast causing film formation and off-flavor in table wine. American Journal of Enology and Viticulture, 17(2), 82-86. https://www.ajevonline.org/content/17/2/82.short

48. Scott Laboratories. (n.d.). Biodiva. Scott Laboratories. Retrieved February 20, 2022, from https://shop.scottlab.com/fermentation-cellar/biodiva-500g-01569

49. van Breda, V., Jolly, N., & van Wyk, J. (2013). Characterisation of commercial and natural Torulaspora delbrueckii wine yeast strains. International Journal of Food Microbiology, 163(2-3), 80-88 source: https://doi.org/10.1016/j.ijfoodmicro.2013.02.011
source: digitalknowledge.cput.ac.za/bitstream/11189/2026/3/Van%20Breda_VM_Jolly_N_Van%20Wyk_J_AppSci_2013.pdf

50. Kraus, J. K., Scopp, R., & Chen, S. L. (1981). Effect of rehydration on dry wine yeast activity. American Journal of Enology and Viticulture, 32(2), 132-134. https://www.ajevonline.org/content/32/2/132

51. Vinos de Jerez. (n.d.). Vinification from Grape to Wine. Sherry Wines. Retrieved February 20, 2022, from www.sherry.wine/sherry-wine/production/vinification

52. Rosini, G. Wine-making by cell-recycle-batch fermentation process. Appl Microbiol Biotechnol 24, 140–143 (1986). https://doi.org/10.1007/BF00938785
Yeast Contribution to the Sensory Profile of Wine from Lallemand
www.lallemandwine.com/wp-content/uploads/2020/02/Cahier05-Lall.pdf