Calcium carbonate precipitation by cyanobacterial polysaccharides - Tufas and Speleothems: Unravelling the Microbial and Physical Controls - page 51

(Schultze-Lam et al. 1996; Ferris & Lowson 1997).
Once all EPS binding places are occupied, the sol-
ution saturation state increases if there is a continued
rise in local concentration of dissolved Ca 2þ and
HCO 3 2 (Arp et al. 2003). The decomposition of
EPS causes a release of HCO 3 2 and Ca 2þ ions into
the local environment, thus increasing the saturation
state regarding relation to calcium carbonate and
promotes precipitation. Continued precipitation
will reduce the saturation state as Ca 2þ and HCO 3 2
are removed from the solution (Arp et al. 2003).
Uncharacterized EPS produced by Desulfovibrio
desulfuricans G20, (a strain of sulphate reducing
bacteria, SRB), altered the CaCO 3 mineral mor-
phology (Bosak & Newman 2005). The influence
of EPS on calcium carbonate precipitation is sup-
posed to be based on their calcium binding capacity.
The EPS of three different SRB strains have recently
been characterized and their calcium binding
capacity has been estimated (Braissant et al. 2007).
Cyanobacteria have been observed to precipi-
tate CaCO 3 in a range of environments (Dittrich
et al. 2004; Lee et al. 2004). As shown by many
researchers, different cyanobacterial species exhibit
different calcification fabrics (e.g. Pentecost 1991;
Merz 1992). Furthermore, cyanobacteria have
been known as potential EPS producers for a
long time (De Philippis et al. 1991). This has
highlighted the potential of cyanobacterial EPS
from strains such as Cyanospira capsulata and
Aphanothece halophytica GR02 for biotechnologi-
cal applications (see for a review (De Philippis
et al. 2001).
It is assumed that acidic EPS probably play an
important role in crystal nucleation, although the
effect of Ca 2þ -binding by acidic EPS on sustaining
CaCO 3 precipitation is minor in freshwater biofilms
(Shiraishi et al. 2008). Therefore, EPS can more
than likely influence the formation of tufa fabrics
by providing nucleation sites, as can the cell
surfaces of heterotrophic bacteria (e.g. Ferris &
Beveridge 1984; Bosak & Newman 2003).
Picocyanobacteria are small unicellular cyano-
bacteria with a cell diameter of 0.2 to 2 mm, com-
monly found in soils and freshwater. They
contribute significantly to the overall primary pro-
duction in ecosystems of all climatic zones
(Agawin et al. 2000; Stockner et al. 2000; Bell &
Kalff 2001). Picocyanobacteria have also been
observed in mats, biofilms in hot springs, as well
as in hypersaline ponds (Ferris et al. 1996; Garcia-
Pichel et al. 1998; Ward et al. 1998; Miller &
Castenholz 2000). Robbins & Blackwelder hypoth-
esized that calcium carbonate crystals can be
nucleated on both the organics and cell membranes
of picoplankton cells (Robbins & Blackwelder
1992). Interestingly, picocyanobacteria from both
the pelagic and biofilms in the euphotic zone of

temperate-zone lakes belong to the same evolution-
ary lineage of cyanobacteria (Becker et al. 2004).
Knowledge about EPS compositions of cyano-
bacteria is crucial in order to understand biofilm
formations, cell attachment to surfaces and cell-
mineral interactions (de Winder et al. 1999; Hirst
et al. 2003). Until now, the functional groups of
extracellular polysaccharides of picocyanobacteria
of Synechococcus-type have not been investigated
in that respect. Cyanobacterial extracellular poly-
mers are characterized by a presence of different
proteins, uronic acids, pyruvic acid, and sulphate
groups (Parikh & Madamwar 2006). The total buf-
fering capacity plays an extremely important role
in this respect as it reflects the binding capacity of
polymers. Previous work has shown that EPS in cya-
nobacterial mats probably plays an important role in
carbonate nucleation (Shiraishi et al. 2008). This
important geochemical attribute of cyanobacteria
has not been assessed in cyanobacterial cultures
obtained from freshwater.
Despite EPS ubiquitous distribution, there is still
a great lack of knowledge concerning the diversity
of extracellular polysaccharides of different pico-
cyanobacterial strains and about those EPS com-
ponents that may be responsible for calcium
carbonate precipitation. The aim of this study is
three-fold: to determine the total buffering capacity
of the extracellular polysaccharides of three differ-
ent strains of picocyanobacteria using potentio-
metric acid-base titrations; to characterize the
functional groups by infrared spectroscopy; and to
investigate their potential to precipitate calcium
carbonate using batch precipitation experiments.

The isolation of extracellular
polysaccharides

PCC 7942, Syn. Green and Syn. Red picocyanobac-
teria Synechococcus-type strains were used in all
experiments presented here. The PCC 7942 strain
was obtained from the Pasteur Institute in Paris,
France. The Syn. Green and Red strains were iso-
lated from the water column of two stratified
lakes: the Pl¨ner See and Lago Maggiore (courtesy
of C. Callieri). Cells were grown as a batch culture
using modified Z/10 medium, under a 14 h/10 h
light/dark condition, with a light intensity of
c.10mEm 22 s 21 (Dittrich & Sibler 2005). Differ-
ent growth conditions and physical parameters are
known to affect the production and properties of
extracellular polymeric substances in algae and
cyanobacteria (De Philippis et al. 1991). In order
to generate reproducible experimental results that
reflect the environmental conditions in biofilms,
cyanobacterial cells in the stationary growth phase
were used for the polysaccharides isolations.