**2. Why the surveillance on cyanobacteria?**

60 Novel Approaches and Their Applications in Risk Assessment

Cyanobacteria are autotrophs and possess all the photosynthetic pigment (chlorophyll *a*, carotenoids, allophycocyanin, phycobilins, phycoerythrins) except chlorophyll *b* (Castenholz, 2001). Prochlorophytes are also cyanobacteria that contain chlorophyll *a* and *b*, but, opposing to other cyanobacteria, lack phycobiliproteins (Castenholz, 2001). Cyanobacteria have the ability to use low light intensities effectively, since they are able to produce the accessory pigments needed to adsorb light most efficiently in the habitat in which they are present, providing them a great advantage for the colonization of a wide range of ecological niches (van den Hoek et al., 1995; WHO, 1999). Phycobiliprotein synthesis is particularly susceptible to environmental influences, especially light quality. The chromatic adaptation is largely attributable to a change in the ratio between phycocyanin and phycoerythrin in the phycobilisomes. The photosynthetic pigments are located in thylakoids that are free in the cytoplasm near the cell periphery (Fig. 2). Cell colours vary from blue-green to violet-red due to the chlorophyll *a* masking by the carotenoids and accessory pigments. The pigments are involved in phycobilisomes, which are found in rows on the outer surface of the thylakoids (Fig. 2) (WHO, 1999). Cyanobacteria are also able of storing essential nutrients and metabolites within their cytoplasm. Prominent cytoplasmic inclusions such as glycogen and cyanophycin granules (polymers of the amino acids arginine and asparagine), polyphosphate bodies, carboxysomes (containing the primary enzyme for photosynthetic CO2 fixation, ribulose 1,5-bisphosphate carboxylase-oxygenase: RuBisCO) and gas vacuoles (Fig. 2) can be observed by electron microscopy. The occurrence of fimbriae (pili) is abundant in many cyanobacteria with varying patterns. Some filamentous forms are also able of gliding (sliding) (van den Hoek et al., 1995; WHO, 1999; Castenholz, 2001).

Fig. 2. Cyanobacteria cell structure. (A)Transmission electron micrographs showing the ultrastructure of an *Anabena circinalis* vegetative cell; (B) Schematic diagram of a cyanobacterial vegetative cell. S: external 4-layered cell wall; OM: outer membrane; PL: peptidoglycan layer; CM: cytoplasmic membrane; CW: cell wall; E: cell envelope; TH: thylakoid; PB: phycobilisome; CY: cytoplasm; GV: gas vesicle; GG: glycogen granules; N: nucleoplasmic region; C: carboxysome; PP: polyphosphate granule; CP: cyanophycin granule; LP: lipid droplets (adapted from van den Hoek et al., 1995; Castenholz, 2001).

Cyanobacteria are common constituents of the phytoplankton in aquatic environments. In optimal conditions these phytoplanktons can develop massively and form blooms, becoming the dominant organism in the water column and creating serious problems in water quality (Cood, 2000; Vasconcelos, 2006). The water quality deterioration produced by cyanobacterial blooms includes foul odours and tastes, deoxygenation of bottom waters (hypoxia and anoxia), fish kills, food web alterations and toxicity. Other threatening characteristic of these organisms is their ability to produce toxins that affects other living organisms and humans (Carmichael, 2001). The capacity of mass development together with the ability to produce potent toxins enlightens the importance of implementing regular monitoring programs for cyanobacteria and cyanotoxins in freshwater environments, in order to minimize potential health risks to animal and human populations that results from exposure through drinking and recreational activities. The implementation of surveillance programs on cyanobacteria involves understanding the ecophysiology of cyanobacteria, bloom dynamics, conditions that promote blooms, production of toxins and their impact in human and animal health (McPhail & Jarema, 2005).

Cyanobacteria possess some ecostrategies that allows them to overcome other organism and become dominant. In general there are four constraints on cyanobacteria growth as prerequisites for bloom enhancement: light, nutrients, temperature and stability of the water column. Cyanobacteria requires low light intensities for growth, compared with algae, which provides competitive advantages in lakes which are turbid due to growth of other phytoplankton. They also have a higher affinity for uptake phosphorous and nitrogen than many other photosynthetic organisms and they have a substantial storage capacity for phosphorous (Mur et al., 1999). Some genera like *Anabaena, Aphanizomenon*, *Cylindrospermopsis*, *Nodularia* and *Nostoc* have specialized cells (heterocysts) (Fig. 1) for nitrogen fixation and blooms of these genera can often be related with periodic nitrogen limitation. This means that they can compete other phytoplankton under conditions of phosphorous and nitrogen limitation (Briand et al., 2003; Sunda et al., 2006).

The success of some cyanobacteria is also due to the presence of gas vacuoles that provide buoyancy regulation. During water stratification conditions cyanobacteria can migrate in the water column, accessing light in the surface layers and nutrients near the sediment. During photosynthesis, carbohydrates are accumulated which makes them heavy and sinking away from light and when the carbohydrates are respired, buoyancy is restored. As large colonies sink faster than small ones or single cells, genera like *Microcystis*, *Anabaena*, *Aphanizomenon* and *Nodularia* have scum-forming strategies (Vance, 1965; Mur et al., 1999). Cyanobacteria also produce active substances that inhibits the growth of competing algae and grazers that feed upon them, which can also promote cyanobacteria proliferation (Briand et al., 2003; Granéli & Hansen, 2006; Sunda et al., 2006; van Apeldoorn et al., 2007; Figueredo et al. 2007). As a consequence of the characteristics mentioned above the cyanobacterial cells numbers in water bodies vary seasonally. In temperate regions, seasonal successions of organisms belonging to different phytoplankton taxa are often observed. Whereas at the beginning of


Risk Assessment of Cyanobacteria and Cyanotoxins,

the Particularities and Challenges of *Planktothrix* spp. Monitoring 63

Table 1. Cyanotoxins detected and correspondent taxa from which have been isolated, as well as their primary target in mammals. Based on the information from Chorus et al., 2000; Charmichael, 2001; Codd et al., 2005; Stewart et al., 2006; van Apeldoom et al., 2007; Bláha et al., 2009; Valério et al., 2010; Mihali et al., 2009. \* - the dose needed to kill 50% of exposed

animals.


Table 1. Cyanotoxins detected and correspondent taxa from which have been isolated, as well as their primary target in mammals. Based on the information from Chorus et al., 2000; Charmichael, 2001; Codd et al., 2005; Stewart et al., 2006; van Apeldoom et al., 2007; Bláha et al., 2009; Valério et al., 2010; Mihali et al., 2009. \* - the dose needed to kill 50% of exposed animals.

Risk Assessment of Cyanobacteria and Cyanotoxins,

(reviewed in Pouria et al, 1998).

et al. 2001).

the Particularities and Challenges of *Planktothrix* spp. Monitoring 65

Example 2 – Symptoms after exposure during heamodialysis treatment: weakness, muscular pain, nauseas, vomiting, neurologic symptoms (head pain, vertigo, deafness, blindness and seizures), increase of hepatic damage biomarkers, hepatomegaly, hepatic failure and death

Fig. 3. Organizational chart of the steps involved in risk assessment (adapted from Dolah

supplied with untreated- or ineffective-treated water.

Besides the acute effects mentioned above, few papers reports the association between the ingestion of water contaminated with microcystins and the increase of hepatocarcinoma (Yu, 1995; Ueno et al., 1996) and colorectal cancer (Zhou et al, 2002) in human populations

Laboratorial studies have demonstrated that, in fact, microcystins, nodularins and cylindrospermopsin are genotoxic (reviewed in Zĕgura et al., 2011) and the carcinogenic

the summer a great variety of microalgae and cyanobacteria usually co-exist in the same water body, towards the end of summer this diversity may drop drastically as the result of the mass development of the cyanobacterial communities (blooms) (Sze, 1986). These blooms may be formed by a consortium of cyanobacteria producing different amounts of toxins at different rates, with the same bloom-forming species having both toxigenic and non-toxigenic strains, indistinguishable by morphological examination. Cyanobacterial blooms are complex and can develop in a rather sudden and unpredictable way.
