Yeast populations viable

WL is the acronym for Wallerstein Laboratories, which originally marketed the medium. WL, sometimes called WL-Nutritional (WLN), does not contain cycloheximide and so is used for determination of total viable yeast populations. This WL medium contain a pH indicator, brom-cresol green, which permits rapid screening of acid-producing colonies. In addition, this medium contains casitone, a specially prepared pancreatic digest of casein available from Difco. 

Viable yeast populations can also be estimated directly by counting under the microscope using specific coloration or epifluorescence techniques. Viable populations can also be determined with ATP measurements using bioluminescence. Bouix et al. (1997) proposed the use of immunofluorescence to detect bacterial contaminations during winemaking. 

Fig. 3.2. Yeast growth cycle and fermentation kinetics of grape mnst containing high sugar concentrations (320 g/1) (Lafon-Lafourcade, 1983). (I) Total yeast population. (II) Viable yeast population. (HI) Fermented sugar...

Initial sugar concentration 260 g/1 initial viable yeast population 10 cells/ml dry yeast Saccharomyces cerevisiae-, fermentation temperature 19°C. Yeast populations are counted at the end of fermentation. 

As alcoholic fermentation takes place, the alcohol concentration increases in the medium. The negative effects of yeast metabolism are compensated in the end by the positive ones. When the yeast population enters the stationary phase, the situation is not static in reality, the viable population count is composed of cells that actively multiply while others are lyzed. The latter cells play an important role vis-a-vis the bacteria—they liberate vitamins, nitrogen bases, peptides and amino acids. All of these components act as growth factors for the bacteria. 

In practice, stability is satisfactory if the viable yeast population is less than 1 cell/ml. It might be preferable to set a lower limit (for example less than 1 cell per 100 ml) but in this case the sample would have to be filtered for the germ count. This operation can be very difficult, if not impossible—for example, with new sweet botry-tized wines. The number of pasteurization units required to sterilize a wine in terms of its constitution is given in Table 9.5. The heating time directly depends on the industrial pasteurization flow rate. From the graph in Figure 9.1, the required temperature can be predicted. 

Regular monitoring of the microbiological state of the wine to verify sterility. If the viable yeast population increases exaggeratedly and attains 1000 cells/ml, an additional sterilization is effected. This increase of the yeast population (Table 9.6) can be explained by the presence of an excessive residual population in the wine (>1 cell/ml) preventing a sufficient sterihty, or by subsequent contaminations. 

Detection of Malo-Lactic Fermentation. It is imperative that the winemaker, to control malo-lactic fermentation, has a satisfactory method for its detection. Disappearance of malic acid is the indication of the fermentation, but the formation of lactic acid is not sufficient evidence since it might also be formed by yeast and by bacteria from other carbohydrate sources. The rate of conversion of malic acid is expected to reflect bacterial metabolism and growth. In New York State wines, Rice and Mattick (41) showed bacterial growth (as measured by viable count) to be more or less exponential to 106-107 cells/ml, preceding disappearance of malic acid. The rate of loss of malic acid is probably also exponential. Malic acid seems to disappear so slowly that its loss is not detected until a bacterial population of about 106-107 cells/ml is reached then it seems to disappear so rapidly that its complete loss is detected within a few days (41). Rice and Mattick (41) also showed a slight increase in bacterial population for a few days following this. 

We have found that turbidity measurements can be used as a routine test to keep track of the yeast cell population. Fermentations routinely were examined microscopically for cell count and viability. We use a 1% methylene blue stain to differentiate viable and dead yeast cells. The dead cells take the stain. 

The success of an alcoholic fermentation depends on maintaining the population of viable yeast at sufficient levels until all the fermentable sugars have been fully consumed (Bisson 1999 Zamora 2004). Otherwise, the winemaker is faced with the serious problem of stuck and sluggish fermentations. The causes and the ways to avoid stuck and sluggish fermentations are discussed later (Bisson and Butzke 2000). 

The model was adapted from Kontoravdi et al. (2005) for cell growth/death, nutrient uptake, and major metabolism. The model was further developed to include description of cell cycle sub-populations. The cell cycle representation was based on the yeast model of Uchiyama Shioya (1999) and the tumour cell model of Basse et al. (2003). Eq.(l)-(4) express viable cell concentration(Xv[cell L" ]) in terms of cells in Gq/Gi, S, and G2/M phases. As a simplification in notation, Gq/Gi cells will be indicated as G unless otherwise stated. Xoi, Xs, X02/M [cell L" ] are concentrations of viable cells in Gq/Gi, S, and G2/M phase, respectively, whereas Fo ,[L h" ] is the outlet flowrate. F[L] is the cell culture volume b, ki, k [h" ] are the transition rates of cells from Gi to S, S to G2, and M to Gi respectively and /[Pg.110]

It is generally accepted that in the case of sound, undamaged grapes, the viable population of yeasts ranges from 10 to 10 CFU/mL (Parish and Carroll, 1985 Fleet and Heard, 1993). The most frequently isolated native species is Kloeckera apiculata, which may account for more than 50% of the total yeast flora recovered from fruit. Lesser numbers of other yeasts, such as species of Candida, Cryptococcus, Debaryomyces, Hansenula, Issatchenkia, Kluyveromyces, Metschnikowia, Pichia, and Rhodotorula, have also been reported (Heard and Fleet, 1986 Holloway et al., 1990 Longo et al., 1991 Fleet and Heard, 1993 Sabate et al., 2002). If grape juice or concentrate... 

Bioluminescence is the process by which a molecule in the excited state emits light, which is then measured (Hartman et al., 1992). This process can be used to measure the amount of ATP produced by microorganisms as part of their metabolism. In theory, measurement of this compound should provide an estimate of viable cell numbers because higher populations of microorganisms produce more ATP. Because a single yeast cell will have generally more ATP than a bacterial cell, the detection limit for yeast could be as low as 10 cells (Hartman et al., 1992). These authors further suggested that a practical limit for bacteria is closer to 1000 to 10,000 cells. 

This phenomenon is not exclusive to lactic bacteria, but certainly applies to many other microorganisms. It is easily demonstrated for acetic bacteria in winemaking. As soon as they are deprived of oxygen, the difference between the viable and VNC populations increases rapidly, then disappears completely as soon as the wine is aerated (Millet and Lonvaud-Funel, 2000). The same experiments showed that yeast and bacteria in VNC state decrease in size and some of them may pass through filters intended to eliminate them. 

Table 8.16. Sulflting to inhibit yeasts in a sweet wine at the end of fermentation (values are number of viable cells, capable of producing colonies in Petri dishes, per ml initial population 58 x 10 /ml) (Ribdreau-Gayon et al., 1977)...

---