An outbreak of food poisoning in Canada during autumn 1987 was traced to cultured blue mussels (Mytilus edulis) from the Cardigan Bay region of eastern Prince Edward Island (P.E.I.). The toxin, identified as domoic acid, had not previously been found in any shellfish and this outbreak represents the first known occurrence of human poisoning by this neurotoxin. A plankton bloom at the time of the outbreak consisted almost entirely of the pennate diatom, Nitzschia pungens f. multiseries, and a positive correlation was found between the number of N. pungens cells and the concentration of domoic acid in the plankton. Nitzschia pungens f. multiseries isolated from Cardigan Bay produced domoic acid in culture at levels (1 to 20 pg∙cell−1) comparable with values estimated for N. pungens in the plankton samples. Isolates of several Cardigan Bay phytoplankton, including the closely related species Nitzschia seriata, failed to produce domoic acid. Other Nitzschia spp. and two Amphora coffeaeformis isolates also failed to produce domoic acid. We conclude that N. pungens was the major source of the domoic acid in toxic mussels in eastern P.E.I. The recurrence, in November 1988, of a monospecific bloom of N. pungens and the presence of domoic acid in plankton and mussels reinforced this conclusion.

Nitzschia pungens f. multiseries (clone NPARL) was grown in nonaxenic batch culture under a range of growth conditions. Domoic acid (DA) was not detected during exponential growth, but production promptly started at a rate of approximately 1 pg DA∙cell−1∙d−1 at the onset of the stationary phase, in this case induced by silicate limitation. Cellular DA reached a maximum of 7 pg∙cell−1; thereafter, DA production continued at the same rate, with cellular levels remaining relatively constant due to concurrent release of DA into the culture medium. DA production ceased in the absence of nitrogen during the stationary phase, but resumed when nitrate was added back to the medium. Low irradiance slowed the division rate and consequently delayed the attainment of the stationary phase, but DA production rates were comparable with the control once stationary phase was reached. Cells during the dark period of a light–dark cycle, or placed into darkness, or in the presence of the photosynthetic inhibitor DCMU promptly ceased DA production. We conclude that at least three conditions are required for DA production by clone NPARL: cessation of cell division, availability of nitrogen during the stationary phase, and the presence of light. Growth in medium f/2 fulfils these requirements.

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ABSTRACT. Dornoic acid (DA) production by Pseudo-nitzschia multiseries (Hasle) was studied at various silicate concentrations and under silicate perturbation. Both slowly dividing and non-dividing population~ produced DA, and the production rates were inversely correlated with the ambient silicate concentrations. Production of DA was significantly enhanced when overall cell metabolism (i.e growth rate) declined as a result of silicate stress. Following silicate starvation, cultures supplemented with silicate registered uptake, but suspended DA production. Results suggest that luxury uptake of Si by P. multisenes may happen only in phys~ologically active populations, i.e, the exponential phase, but not in the stationary phase. There were 2 stages of DA production The first stage corresponded to a decline in growth caused by moderately low lcvels of remaining s~licate in the medium, wh.ile the second stage was caused by severe silicate limitation. The production rate during the second stage (13.67 to 30.20 fg DA cell-' d-') was about an order of magnitude higher than during the first stage (0.97 to 4.98 fg DA cell-' d-l). Increases and decreases in cellular DA content corresponded to decreases and increases in growth rates.KEY WORDS: Domoic acid . Pseudo-nitzschia multiseries . Silicate limitation . Batch culture INTRODUCTIONPseudo-nitzschia multiseries, formerly known as Pseudonitzschia pungens f. multiseries (Hasle, 1995), produces domoic acid (DA), which has caused amnesic shellfish poisoning (ASP) in Atlantic Canada (Addison & Stewart 1989). The ASP problem has now been observed on both the Atlantic and Pacific coasts of North America (Fritz et al. 1992, Garrison et al. 1992. Studies on DA production have shown it to occur only during the stationary phase, coinciding with low levels of silicate in the medium (Subba Rao et al. 1990, Bates et al. 1991, Pan et al. 1991). These observations suggest a possible relationship between silicate limitation and DA production. However, direct linkage between DA production and silicon depletion is unlikely, as silicate is neither a component of DA nor apparently involved in its synthesis.The magnitude of a diatom bloom is often directly related to the availability of silicon in sea water. Silicon regulates the growth and frustule formation of diatoms. Decreases in silicate concentrations to low or undetectable levels in marine and freshwater habitats during diatom blooms have been well documented (Paasche 1973a, Sommer & Stabel 1983, Egge & Aksnes 1992, Harrison et al. 1993.In cultures, silicate concentrations in the medium may regulate the yield of diatom cells (Taguchi et al. 1987). Cessation of cell division, which may be due to cessation of DNA synthesis (Darley & Volcani 1969, Sullivan & Volcani 1973, was found to accompany depletion of silicon in the culture medium (Lewin 1955, Lewin & Chen 1968, Vaulot et al. 1987, Brzezinski et al. 1990. Bates et al. (1991) first showed a connection between silicate limitation and DA production. Two of their 5 treatments with low s...