flocculation. In turn, the flocs will cause a change in both the individual cell metabolism
and the environmental conditions.
The study of flocculation mechanisms has been centred on Saccharomyces cerevisiae
as it has a capital importance in the brewing industry (Dengis et al., 1995; Teixeira,
1988), although other microorganisms have been studied as well (Ananta et al., 1995;
Libicki et al., 1988; Moradas-Ferreira et al., 1994; Pereboom et al., 1990; Teixeira et al.,
1995). Despite the wealth of works published on yeast aggregate formation and although
it is accepted that flocculation is under genetic control, a fully explanatory interpretation
of the phenomenon has not been given yet. It is a very complex process, depending on
factors such as the microbial strain (growth, physiological state and metabolism), the
composition of the culture medium and the culture conditions (temperature, pH, agitation
and aeration) (Dengis et al., 1995; Scares et al., 1994).
Several mechanisms have been proposed to describe flocculation. Taylor and Orton
(1975) found that Ca2+ induced a conformational change in certain proteins, allowing
them to be recognised in the cell-cell adhesion process. Before, it was already known that
Ca2+ was necessary for yeast flocculation, but its role was just thought to be a bridge
between adjacent cells. Later on, Miki et al. (1982) suggested that the flocculation
mechanism should involve a lectin-like protein, as some specific sugar molecules
inhibited it. In fact, sugars like mannose and its derivatives were found to inhibit
flocculation of S. cerevisiae strains, where the mechanism involves mannan receptors’
recognition by lectins of an adjacent cell, requiring the presence of Ca2+ ions (Dengis et
al., 1995, Stratford, 1992). Other sugars (e.g. galactose and its derivatives) inhibit the
flocculation of other yeast species (Kluyveromyces bulgaricus and K.lactis), as the cell-
cell interactions involve a galactose-specific lectin (El-Behhari et al., 1998). Also found
in a flocculent bottom-fermenting brewer’s yeast strain is a mannose/glucose-specific
lectin-like protein; consequently the flocculation of this strain is inhibited by both glucose
and mannose (Kobayashi et al., 1998). In general, the mechanism above is a widely
accepted one and is presently believed to explain the phenomenon of flocculation in
yeast. It was shown, by studies on the interaction between flocculent and non-flocculent
cells of S. cerevisiae, that cell-cell interaction corresponds to a true stable binding and not
to a simple entrapment inside the floe matrix (Soares et al., 1992).
It is also clear that hydrophobic interactions play a crucial role in microbial adhesion
phenomena, having been demonstrated in e.g. Saccharomyces and Kluyveromyces strains
that an increase in flocculation is strongly correlated with an increase in cell wall surface
hydrophobicity (Azeredo et al., 1997; Straver et al., 1993; Teixeira et al., 1995; van der
Aar, 1996).
Although to a minor extent, other physicochemical properties such as cell walls’
isoelectric point (Dengis et al., 1995) or external medium properties such as pH
(Stratford, 1996; Yang and Choi, 1998), ionic concentration (Dengis et al., 1995) and
organic solvent concentration (Teixeira, 1988) are also important in flocculation. In fact,
all these factors together with the adequate Ca^2 + concentration will enable sufficient bond
strength between the cells (van Hamersveld et al., 1997), which is fundamental for floe
formation and stability.
When using flocculating microorganisms in bioreactors, the effects of physical
parameters on flocculation are also very important. Namely, the cell concentration must
be sufficient to provide an adequate number of collisions to form flocs (Glasgow, 1989;
Flocculation bioreactors 389