Paralytic shellfish posisoning (PSP) is a life-threatening affection that results from comsumption of seafood contaminated with paralytic shellfish toxins (PSTs), which are potent neurotoxins comprising of saxitoxin (STX) and more than tweenty other chemically related derivatives. PSTs blocks sodium channels, an action which in humans leads to several neurological affections and in some cases death. So far, PSTs have been detected in laboratory cultures of dinoflagellates mainly belonging to the genera Alexandrium, Pyrodinium and Gymnodinium. However, controversy surrounds suggestions that bacteria associated with these dinoflagellates are either directly or indirecly involved in PST’s producction in dinoflagellates. Bacterial influence on dinoflagellates toxicity was first postulated in 1960’s due to the ocurrence of bacteria-like particles within dinoflagellates cells. Since then there have been several publications for and against these theories. The present work aimed to examine the role of bacteria in dinoflagellates toxicity, by defining their bacterial microbiology using microbiological clasical techniques for bacterial isolation and molecular techniques such as DNA extraction, 16S rRNA clonning, DGGE or TSA-FISH, as well as confocal laser scanning microscopy (CLSM). In this study information is provided on Alexandrium minutum clones in laboratory cultures from different origins. They were used to compare bacterial profiles with the aim to correlate their toxicity differences with their bacterial composition profiles. During this study some methods were either developed or modified for use specifically to study bacterial-toxic dinoflagellates interactions. A new bacteria specie isolated from the toxic dinoflagellate Alexandrium minutum was also proposed. The results of clonning, DGGE and TSA-FISH showed that the bacterial phylogenetic diversity was limited to two bacteria Phyla, the Proteobacteria (mainly belonging to α and γ classes) and Bacteriodetes (Sphingobacteria class, Cytophaga- Favobacter-Bacteroidetes). Results display a species-specific association between some bacteria and the dinoflagellates clones in culture. Confocal microscopy using Bacteria, Arquea and class specific oligonucleotide probes demonstrated internal bacteria in Alexandrium minutum cells as well as bacteria attached to the theca. PST production in bacteria-free dinoflagellates compared to axenic cultures led to the conclusion that bacteria could somehow influence dinoflagellate cell growth, as well as some bacteria could directly influence PST’s concentrations in Alexandrium minutum cultures. The mechanisms by which bacteria have this influence are unknown altohugh the production of co-factors, signalling molecules, nutrient availability and bacterial transformation of toxins was suggested. Several researchers have reported specific interactions between bacteria and phytoplankton, which has led them to propose that bacteria may play a role in controlling phytoplankton dynamics. This study provide infomation from a range of potentially algal-lytic tested bacteria isolated from Alexandrium minutum cultures. These algal-lytic bacteria could play an important role in regulating the onset and development of harmful algal blooms.

Tesis doctoral inédita leída en la Universidad Autónoma de Madrid, Facultad de Ciencias, Departamento de Biología Molecular. Fecha de lectura: 2-03-2007

Peer reviewed