CORNELL
U N I V E R S I T Y 		11Food Science 430
Wine is chemistry, biology and psychology!

Calendar

yeast ecology
starter cultures
sllides
Reading: Jackson pp. 241-246, 254-259;

ecology | starter cultures | Slides03 | Slides04 |
YEAST ECOLOGY
1 Feb. 96 Thomas Henick-Kling

  • Vineyard Site
  • Cultivar Selection
  • Vineyard Management
  • Grape Maturity & Harvest
  • Juice Extraction
  • Alcoholic Fermentation- yeast ecology, fermentation pathway
  • Malolactic Fermentation
  • Wine Aging (Storage)

Yeast Ecology
    where do the yeasts come from?
    vineyard & winery environment: grapes, leaves, soil, hands, equipment surfaces, air
      on grapes: 103 to 105 CFU/mL
    on grapes:
    Kloeckera apiculata (Hanseniaspora)
    Metschnikowia
    Candida stellata, C. pulcherrima
    50-75%

    Cryptococcus
    Rhodotorula
    Pichia membranefaciens
    Kluyveromyces
    Hansenula
    Torulaspora delbrueckii.
    Saccharomyces cerevisiae

    1-10%

    on equipemnt surfaces & in wine during alcoholic fermentation: Saccharomyces cerevisiae
    Saccharomyces cerevisiae (bayanus)
    Candida, Pichia, Hansenula, Brettanomyces
    after alcoholic fermentation: Brettanomyces/Dekkera
    Zygosaccharomyces bailii,

Wine Yeasts
    Saccharomyces cerevisiae
    synonyms: Saccharomyces cerevisiae (bayanus)
      S. uvarum, S. oviformis, S. carlsbergiensis, S. logo, S. chevalieri, S. diastaticus, S. fructuum, S. italicus, S. hispanica, S. oxydans, S. pasteurianus, S. prostoserdovii, S. sake, S. sterineri, S. vini
    (Barnett et al. Yeasts. Characteristics and identification. Cambridge University Press. 1983)

    Yeast Growth During Fermentation
    Succession of different yeasts, finally dominance of Saccharomyces cerevisiae.

    Initial rapid growth until oxygen is depleted, alcohol content becomes inhibitory, nutrients become limited.

    Yeast Growth

    1. increase in cell size (volume), cell mass
    2. increase in cell density (budding)

      sugar in grapes: glucose & fructose 160-240 g/L other sugars, usually not fermented by yeast, <1 g/L
    ethanol 8-14% in table wines

    CO2, heavy, suffocating danger, fermentation rooms must be well ventilated

    Heat must be dealt with:
    in general for every Birx (10g/L) sugar consumed the temperature increases by 1.3 degree C.

Natural Fermentation
Spontaneous Fermentation
Wild Fermentation

    Spontaneous fermentation of wine is carried out by indigenous yeasts.

    indigenous= native = wild yeasts = natural yeasts

    Selected starter culture yeasts - are natural yeast that have been selected from spontaneous fermentations purified to single strains (single cell) and then cultured outside the wine for reinoculation as a pure culture.

    The term 'natural' fermenation should not be used. All fermentation is natural (=carried out by the one or several of a group of yeasts) whether a yeast culture was added or not.

    We should distinguish between spontaneous and inoculated fermentations.

    Wine-related yeasts:
    Saccharomyces cerevisiae     are generally called wine yeasts Metchnikowia, Kloeckera, Hanseniaspora, Hansenula, Pichia, Schizosaccharomyces, Zygosaccharomyces, Brettanomyces and Dekkera are usually called spoilage yeast.

    All of these yeasts (and a few more) are wine related yeasts. The spoilage yeasts become spoilage organisms when they are present in high cell densities and for extended amount of time. There are also stron differences between strains. Some strains of Kloeckera may be harmless or even beneficial to the wine flavor, others are very damaging (prducing much acetic acid and ethyl acetate).

Methods for Differentiation of Yeast
Isolated from Must and Wine

    With classical tools of microbiology the various yeasts isolated from grape must and wine have been identified using colony morphology, cell morphology, observation of spores, and biochemical tests. These tests are very time consuming and not always reliable.

    Molecular biology has given new tools for characterization of various yeasts.

    i.e.
    Saccharomyces cerevisiae
    Kloeckera apiculate
    genus species

    Karyotyping The various yeasts isolated from wine differ very strongly in the number and size of chromosomes they carry. In yeast karyotyping the yeast cells are lyzed and the isolated chromosomes are separated according to size on an agarose gel. Different numbers and different sizes of chromosomes give in different banding patterns. This method allows a very good gross separation of the most important groups (genera) of wine related yeasts.

    RFLP For more detailed examination of the various yeasts, for species and strain identification, a more detailed look at the yeast DNA is necessary. Restriction Fragment Length Polymorphism is a very helpful method for this purpose. Various enzymes can be used to cut the chromosomal DNA in fragments of different sizes. These fragments can be separated again by size and the bands obtained can be characteristic of a specific yeast. Yet there can be much variability in the arrangemnt of these short fragments.

    PCR Polymerase Chain Reaction avoids some of the variability of RFLP by using specific segments of DNA and amplifying (making many thousands of copies) them. The copies again separated by size also yield characteristic patterns. PCR and RFLP can be combined. PCR fragment produced can be cut by selected enzymes to give more detailed information allowing better separation between different yeast strains.

    The difficulty in PCR analysis is to find 'characteristic' sequences that can be amplified. Sequences which are highly conserved (don't vary much from one generation of yeast to the next) and sequences that are different from yeast species to yeast species must be selected.

    The difficulty in RFLP analysis is that too many small fragments are produced and that all strains of yeast look the same. Some detail must be produced but not too much.

FS430 Revised 2.04.97