Dinoflagellates and Toxin Production 217
(Gallardo-Rodríguez et al. 2012a) have greatly hampered development of toxin-derived leads for
pharmacores. There are at present many difficulties to grow dinoflagellates in laboratory cultures (using
conventional reactors), and several attempts have failed to obtain the intended compounds (Wynn et al.
2010; Gallardo-Rodríguez et al. 2012a). Nevertheless, production of such potent bioactive compounds
in relatively high quantities (and in a safe mode) is an important issue before pharmacological studies
and pre-clinical trials can be developed (Glaser and Mayer 2009; Zittelli et al. 2013). The production of
toxin by dinoflagellate cultures in photobioreactors is still the preferred approach—despite the underlying
constraints regarding fastidious growth and extreme sensitiveness to shear stress (Gallardo-Rodríguez et
al. 2009). Most cultivation systems grow cells in suspension (i.e., vessels or classical tubular reactors),
with conventional operation conditions (i.e., continuous light supply, or CO 2 supply turbulent bubbling);
this can lead to cell damage due to high shear stress, thus raising a difficulty for dinoflagellate cultivation
at a large scale. Stirring is necessary to avoid gradients of CO 2 and O 2 , while enhancing mass transfer to
cells for performance of photosynthesis. A uniform rate of supply of light should also be assured—thus
preventing photoinhibition or photoxidation. It is believed that such susceptibility is caused by a complex
cell organization, and the nature of stimuli that can affect (in a transient or permanent way) the ability of
biomass to divide and grow.
Several improvements have been done in this area of dinoflagellate cultivation, yet the biotoxin titers
remain very low (in the order of picogram). This chapter includes a brief revision on the diversity and main
features of dinoflagellates, ecology, and HABs; how and why toxins are synthesized, and their putative
pharmacological applications; the major difficulties encountered to produce toxins via synthetic routes;
the ongoing efforts pertaining to genetic and metabolic engineering; and discuss the main cause affecting
dinoflagellate growth, that is, shear stress, arising from turbulence and agitation throughout cultivation
in conventional photobioreactors. The main advances anticipated as necessary for dinoflagellate culture
in bioreactor will be discussed last, with the goal of producing such biotoxins in a more efficient manner.
Diversity and features
Dinoflagellates (Phylum: Dinoflagellata; Division: Phyrrophyta) are a large group of eukaryotic,
biflagellate organisms exhibiting great diversity in morphology, cellular organization, and behavior (Taylor
1987; Gomez 2012). Ubiquitous in all types of ecosystems (i.e., marine, freshwater, benthic, brackish,
ice sea), dinoflagellate populations distribute according to temperature, salinity, and depth (Taylor et al.
2008). Ecologically speaking, dinoflagellates can occur both in the water column, as a component of the
plankton (ca. 90% responsible for primary productivity), and at the bottom of water bodies, as part of
benthos (Gomez 2012). The unique diversity of this taxon reflects most trophic types: from photosynthetic
and pure autotrophically-growing species (Gaines and Elbrachter 1987; Schnepf and Elbrächter 1992),
to mixotrophic species that acquire nutrients from both photosynthesis and dissolved organic matter
or organic particulates (phagotrophy); and from pure heterotrophy to parasitic or symbiotic ways of
living (Bralewska and Witek 1995; Gomez 2012). Roughly half of the species possess photosynthetic
pigments, and thus play a major role as primary producers in freshwater and marine habitats (Hickman
et al. 2008). Their relatively slow proliferation among unicellular algae may justify why these organisms
are particularly well-suited for symbiotic relationships (Hackett et al. 2004). They can indeed be found
associated with several marine organisms, for example, sea anemones, protozoa, certain invertebrates,
and stony corals (Coffroth and Santos 2005). Interestingly, only corals with symbiotic dinoflagellates–
in with Symbiodinium genus accounting for the most representative group, can form coral reefs, thus
conveying bright colors (Fensome et al. 1995; Wong and Kwok 2005).
Belonging to the ancient eukaryotic lineage Alveolata, dinoflagelaltes are composed by unicellular
microalgae species with complex and unique morphological features (Leander and Keeling 2004).
The Alveolata is in fact one of the most biologically vast supergroups of single-celled eukaryotic
microorganisms, consisting of ciliates, dinoflagellates, and apicomplexans (Kellmann et al. 2010).
Typically exhibiting between 10 and 100 µm in length (or, in more extreme situations, 2 µm–2 mm),
dinoflagellates differ from other two groups for possessing two dissimilar flagella (at some stage of their
life) (Taylor 1987; Lin 2011). The major difference on flagella lies on their relative insertion into the
(Gallardo-Rodríguez et al. 2012a) have greatly hampered development of toxin-derived leads for
pharmacores. There are at present many difficulties to grow dinoflagellates in laboratory cultures (using
conventional reactors), and several attempts have failed to obtain the intended compounds (Wynn et al.
2010; Gallardo-Rodríguez et al. 2012a). Nevertheless, production of such potent bioactive compounds
in relatively high quantities (and in a safe mode) is an important issue before pharmacological studies
and pre-clinical trials can be developed (Glaser and Mayer 2009; Zittelli et al. 2013). The production of
toxin by dinoflagellate cultures in photobioreactors is still the preferred approach—despite the underlying
constraints regarding fastidious growth and extreme sensitiveness to shear stress (Gallardo-Rodríguez et
al. 2009). Most cultivation systems grow cells in suspension (i.e., vessels or classical tubular reactors),
with conventional operation conditions (i.e., continuous light supply, or CO 2 supply turbulent bubbling);
this can lead to cell damage due to high shear stress, thus raising a difficulty for dinoflagellate cultivation
at a large scale. Stirring is necessary to avoid gradients of CO 2 and O 2 , while enhancing mass transfer to
cells for performance of photosynthesis. A uniform rate of supply of light should also be assured—thus
preventing photoinhibition or photoxidation. It is believed that such susceptibility is caused by a complex
cell organization, and the nature of stimuli that can affect (in a transient or permanent way) the ability of
biomass to divide and grow.
Several improvements have been done in this area of dinoflagellate cultivation, yet the biotoxin titers
remain very low (in the order of picogram). This chapter includes a brief revision on the diversity and main
features of dinoflagellates, ecology, and HABs; how and why toxins are synthesized, and their putative
pharmacological applications; the major difficulties encountered to produce toxins via synthetic routes;
the ongoing efforts pertaining to genetic and metabolic engineering; and discuss the main cause affecting
dinoflagellate growth, that is, shear stress, arising from turbulence and agitation throughout cultivation
in conventional photobioreactors. The main advances anticipated as necessary for dinoflagellate culture
in bioreactor will be discussed last, with the goal of producing such biotoxins in a more efficient manner.
Diversity and features
Dinoflagellates (Phylum: Dinoflagellata; Division: Phyrrophyta) are a large group of eukaryotic,
biflagellate organisms exhibiting great diversity in morphology, cellular organization, and behavior (Taylor
1987; Gomez 2012). Ubiquitous in all types of ecosystems (i.e., marine, freshwater, benthic, brackish,
ice sea), dinoflagellate populations distribute according to temperature, salinity, and depth (Taylor et al.
2008). Ecologically speaking, dinoflagellates can occur both in the water column, as a component of the
plankton (ca. 90% responsible for primary productivity), and at the bottom of water bodies, as part of
benthos (Gomez 2012). The unique diversity of this taxon reflects most trophic types: from photosynthetic
and pure autotrophically-growing species (Gaines and Elbrachter 1987; Schnepf and Elbrächter 1992),
to mixotrophic species that acquire nutrients from both photosynthesis and dissolved organic matter
or organic particulates (phagotrophy); and from pure heterotrophy to parasitic or symbiotic ways of
living (Bralewska and Witek 1995; Gomez 2012). Roughly half of the species possess photosynthetic
pigments, and thus play a major role as primary producers in freshwater and marine habitats (Hickman
et al. 2008). Their relatively slow proliferation among unicellular algae may justify why these organisms
are particularly well-suited for symbiotic relationships (Hackett et al. 2004). They can indeed be found
associated with several marine organisms, for example, sea anemones, protozoa, certain invertebrates,
and stony corals (Coffroth and Santos 2005). Interestingly, only corals with symbiotic dinoflagellates–
in with Symbiodinium genus accounting for the most representative group, can form coral reefs, thus
conveying bright colors (Fensome et al. 1995; Wong and Kwok 2005).
Belonging to the ancient eukaryotic lineage Alveolata, dinoflagelaltes are composed by unicellular
microalgae species with complex and unique morphological features (Leander and Keeling 2004).
The Alveolata is in fact one of the most biologically vast supergroups of single-celled eukaryotic
microorganisms, consisting of ciliates, dinoflagellates, and apicomplexans (Kellmann et al. 2010).
Typically exhibiting between 10 and 100 µm in length (or, in more extreme situations, 2 µm–2 mm),
dinoflagellates differ from other two groups for possessing two dissimilar flagella (at some stage of their
life) (Taylor 1987; Lin 2011). The major difference on flagella lies on their relative insertion into the