Beschreibung: The overall objective of the NORCOHAB cruise was to study the coastal oceanographic processes and mechanisms underlying the dynamics of key toxic bloom species and the biogeographical distribution of their toxins in the water column of different North Sea coastal waters. The cruise was endorsed by SCOR-IOC Programme GEOHAB (Global Ecology and Oceanography of Harmful Algal Blooms), under the auspices of the Core Research Project on HABS in Fjords and Coastal Embayments. Following the prescribed format of GEOHAB research the studies were international, multidisciplinary and comparative with the ultimate aim of modeling dynamics and behavior.We assessed and compared key genotypic and phenotypic properties of HAB species in relation to grazing and toxic and allelochemical interactions in Scottish and Norwegian/Danish (Skagerrak) coastal areas. For the first time, on board measurements of known phycotoxins collected directly from the plankton were subjected to analysis by high resolution tandem mass-spectrometry coupled with liquid chromatography (LC-MS/MS). These measurements included domoic acid and a large variety of lipophilic toxins associated with marine plankton. We also performed grazing experiments with protists and copepods on selected key toxin algal species in on-board incubations. The following main goals of the cruise programme were successfully addressed: 1) the phenotypic and genotypic characteristics of populations of selected HAB species were compared, including that of a new dinoflagellate identified as the proximal source of azaspiracid poisoning (AZP) in the North Sea; 2) the toxin profile and content of HAB species and toxin transfer and metabolism to the next trophic level were assessed and compared; and 3) insights into the relative importance of grazing by both metazoa and protists as a potential “top down” regulatory mechanism for population dynamics of HAB species was achieved.

Beschreibung: Molluscan safety monitoring programs search for control systems able to give an estimation of the concentration levels of marine toxins present in water, which could represent a risk for the health of consumers of shellfish products. In this study a deployment of passive sampling devices consisting of resin filled sachets was carried out weekly during two years in Alfacs Bay, a coastal embayment of the Ebro Delta in the NW Mediterranean, Spain. Lipophilic marine toxin concentration profiles were obtained analyzing the content of the polymeric resins after extraction and analysis by liquid chromatography coupled to a quadrupole ion trap mass spectrometer.

Beschreibung: Coupling Physiological Responses of the Toxic Haptophyte Prymnesium parvum to Patterns in Gene ExpressionMichael F. Freitag1, Uwe John1 & AD Cembella11Alfred Wegener Institute for Polar and Marine research, 27576 Bremerhaven, Germany 2Sára Beszteri, Alfred Wegener Institute, Bremerhaven GermanyA normalized cDNA library2 from the toxic haptophyte Prymnesium parvum was used to design an oligonucleotide-based microarray platform. Allelopathic interactions were investigated between P. parvum strain RL10 and the heterotrophic dinoflagellate Oxyhrris marina. Cultures of Prymnesium were incubated together with both whole cells and cell-free filtrate from Oxyhrris cultures. During incubation with Prymnesium, the presence of Oxyhrris cells, compared to cell-free culture medium, showed no differential effect on the gene expression profile of the haptophyte. A bioassay measuring the toxicity of Prymnesium against the sensitive cryptophyte Rhodomonas baltica was additionally performed. In this case, Rhodomonas viabilities following incubation with Prymnesium and Prymnesium previously incubated with O. marina medium were virtually identical. This suggests no correlation between the presence of O. marina medium, and induced toxicity in P. parvum. Similar experiments, involving the dinoflagellate Heterocapsa rotunda and an unclassified toxic crooccal cyanobacteria strain: NIVA CYA 331, are currently underway. All tentative unique genes (TUGs) identified from the microarray hybridization will be further classified through the use of a Prymnesium fosmid library prepared from axenic cultures.

Beschreibung: Marine dinoflagellates of the genus Alexandrium are well known for their ability to produce the potent neurotoxin saxitoxin and other analogues responsible for paralytic shellfish poisoning (PSP). Dinoflagellates are the only known eukaryotes confirmed to produce these neurotoxins, but they are also synthesized by certain brackish and freshwater cyanobacteria giving rise to speculations about their prokaryotic origin. Thus far, no distinct enzymes or genes involved in the actual synthesis of the toxins have been unequivocally identified in either dinoflagellates or cyanobacteria. However, detailed proposals for such a pathway exist, and the requirements for biosynthesis have been examined in vitro. In order to contribute to the search for genes involved in PSP toxin production, we developed a DNA microarray based upon an expressed sequence tag (EST) library from A. minutum. This species tends to produce a limited sub-set of PSP toxins, typically dominated by the sulfated carbamate derivatives known as gonyautoxins. The array was used in gene expression studies involving both toxic and non-toxic A. minutum strains, as well as different physiological conditions within one strain. We identified genes associated with the ability to produce PSP toxins in an inter-strain comparison. Here we compare these data to gene expression correlated with differences in toxicity under different physiological conditions.

Beschreibung: Cylindrospermopsis raciborskii is a filamentous, potentially toxic bloom-forming cyanobacterium in tropical and temperate freshwaters. We compared the cylindrospermopsin (CYN) producing C. raciborskii strain CS-505 (Australia) against a PSP toxin producing strain D9 (Brazil), originally identified as conspecific. Cylindrospermopsis raciborskii forms a monophyletic group comprising identities 〉99% for the 16S rRNA gene, whereas strains D9 and CS-505 are 99.7% similar. We sequenced the complete genome of these strains, the first for any PSP- or CYN-toxin producing cyanobacterium. The genome sizes are the smallest known for filamentous cyanobacteria (3.3 and 3.9 Mbp for D9 and CS-505, respectively), but exhibit an apparently large size difference. Genome comparison yielded 471 and 801 strain-specific genes for D9 and CS-505, respectively, and revealed remarkable differences between strains. These differences are mainly related to signatures for high genomic plasticity in CS-505, gene loss in D9 (such as the lack of nitrogen fixation genes), and genes for secondary metabolite production, specifically in the STX and CYN clusters. Functional analysis via gene expression studies using microarrays and qPCR have supported the function of the identified toxin-related gene clusters as well as indicating fundamental differences in phenotypic expression between the strains.

Beschreibung: Cyanobacterial toxins as microcystins (MCs), nodularins (NODs), Paralytic Shellfish Poisoning (PSP) toxins, anatoxins (ANAs), and cylindrospermopsins (CYNs) with over 100 known varieties, occur worldwide associated with human and animal lethal poisoning.In contrast to all analytical methods for toxin determination are based on LC/MS-MS measurements with Multiple Reaction Monitoring (MRM) the application of Precursor Ion mode allows the coverage of all these structural variants. Although in MRM mode enables a higher sensitivity, a lot of information regarding structural changes is missed. The new generation of Q-TRAPs combines the advantages of quadrupoles having a high selectivity with the high sensitivity of ion-trap systems. Here we published results, showing the suitability of Precursor Ion mode for detection of cyanobacterial toxins extracted from phytoplankton.