minutum compared to control oysters. At the same time, many oysters exposed to the harmful microalgae presented a strong inflammatory response in different tissues of the digestive gland: in intestine as well as in digestive ducts and tubules. Spermatozoa in oysters exposed to A. minutum were morphologically and functionally modified compared to spermatozoa of control oysters. Indeed, spermatozoa were less motile and had lower ATP content in oysters exposed to A. minutum. Meanwhile, spermatozoa produced by control oysters showed higher 2 percentage of mortality and relative DNA content than those produced by A. minutum exposed oysters. Finally, the characteristics of the mitochondria of spermatozoa also appeared to be modified upon A. minutum exposure. The results of this study suggests that an exposure of oysters to A. minutum, reducing energy status and motility of spermatozoa associated to morphological changes at the cellular and sub-cellular levels, can have consequences on spermatozoa fertility and reproduction success.

The possible effect of Alexandrium spp. containing paralytic shellfish poisoning (PSP) toxins on the hemocyte parameters of oysters was tested experimentally. In separate experiments, eastern oysters, Crassostrea virginica, were exposed to bloom concentrations of the sympatric dinoflagellate, Alexandrium fundyense, alone and in a mixture with a non-toxic diatom, Thalassiosira weissflogii, and Pacific oysters, Crassostrea gigas, were exposed to a mixed suspension of the sympatric, toxic species Alexandrium catenella, with T. weissflogii. Measurements of numbers of oyster hemocytes, percentages of different cell types, and functions (phagocytosis, reactive oxygen species-ROS-production, and mortality) were made using flow-cytometric methods. During and after exposure, no significant effect of Alexandrium upon hemocyte numbers, morphology, or functions were detected, despite observations of adductor-muscle paralysis in eastern oysters and measured toxin accumulation in C. gigas. Significant effects of physical displacement of oysters into the experimental conditions and of temperature (repeated at 12 and 18°C) upon immune status were detected in Pacific oysters. Correlations between hemocyte numbers and function were consistent with previous studies and were representative of normal, unstressed oysters. The finding of no effect of toxic Alexandrium spp. upon oyster hemocytes is consistent with the knowledge that PSP toxins interfere specifically with sodium-channel function in neural tissues and supports the expectation that sodium-channel physiology has no importance in hemocyte functions in oysters. Finally, we found no evidence of bioactive compounds, other than PST, in the two species of Alexandrium studied.

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Toxigenic saprophytic fungi were isolated from samples of shellfish, sediment and seawater obtained from marine shellfish farming areas. The 456 strains identified included 12 different genera, with a clear predominance (68%) of Penicillium, Aspergillus, Trichoderma and Cladosporium. To assess the risk of poisoning due to the presence of these fungi in shellfish farming areas, the strains were cultured in liquid medium, filtered, and tested on larvae of Artemia salina, a small crustacean highly sensitive to mycotoxins. Thirty-five point five percent of the strains proved active with this test. This study confirms the existence of fungi in shellfish farming areas, as suggested by our earlier work showing that filter-feeding shellfish accumulate toxic metabolites of fungal origin. The presence of fungi in the marine environment represents a real risk of poisoning through the consumption of contaminated shellfish.

Several experiments using a self-regulated system were conducted to define the factors likely to influence the uptake of paralytic shellfish poison (PSP) by oysters in the Penzé estuary (France, Brittany). Each 4-day experiment was carried out in a recirculated sea water system using 15 Pacific oysters (Crassostrea gigas) separated from each other and supplied with unfiltered natural seawater containing alternatively toxic (Alexandrium minutum) or non-toxic (Skeletonema costatum) algal diets. The food supply and exposure times to toxic diets were determined according to field studies of the upstream and downstream movement of patches containing A. minutum. The experimental parameters corresponded roughly to the hydrological conditions generally observed in June when tidal coefficients are lowest and blooms occur: (i) A. minutum concentrations in sea water of 200, 5000 and 10 000 cell ml −1 ; (ii) inorganic matter consisting of 5 and 15 mg L −1 of calcinated muddy sediments; and (iii) low and high tide salinities of 25 and 35% , respectively. Significant experimental contamination (greater than the 80 µg STX equiv. 100 g −1 sanitary threshold) occurred after 4 days of exposure for the monospecific A. minutum diet (20−200 cell ml −1) and alternated A. minutum and S. costatum diets (5000 and 20 000 cell ml −1 , respectively). Contamination levels were less than the sanitary threshold for alternated A. minutum/S. costatum diets of 200 and 20 000 cell ml −1 , respectively, and for a monospecific A. minutum diet (1000−10 000 cell ml −1). In the last case, the accumulation rate was quite low, possibly because of inhibition of the filtration rate related to a lower biodeposit production rate and decreased feeding time activity. The addition of inorganic matter appeared to play a significant role in the observed increase of toxin uptake, whereas salinity was not a determining factor for toxin accumulation rates. These last observations were corroborated by statistical analysis and stepwise multiple linear regressions integrating all or some of the experimental parameters.

The effects of toxic and non-toxic strains of Alexandrium sp. on the initial feeding responses of the Pacific oyster Crassostrea gigas were determined by continuous monitoring of the clearance rate of individual oysters. This was determined fluorometrically by loss of fluorescence of live microalgae introduced as food. Feeding oysters with toxic or non-toxic A. tamarense and toxic A. fundyense resulted in stop/start clearance behaviour. In contrast, control oysters fed a reference microalga, Zsochrysis sp., known to support their growth, did not exhibit this behavior. Pacific oysters acclimated to Isochrysis sp., fed mixtures of Alexandrium/ Isochrysis, showed further evidence of stop/start clearance. Exposure for periods up to 48 hours with non-toxic A. tamarense did not enhance clearance of either Alexandrium or Zsochrysis. 0 Ifremer-Elsevier, Paris Crassostrea gigas / feeding physiology I toxic microalgae I dinoflagellate I paralytic shellfish poisoning. Risumk-Effet d'un dinoflagelle producteur de PSP, Alexandrium sp., sur la rCponse alimentaire prkcoce de Crassostrea gigas. Les effets de souches toxiques et de souches non toxiques d'Alexandrium sp. sur la reponse alimentaire precoce de l'huitre creuse, Crassostrea gigas, ont et6 determines grace a un contrble continu du taux de filtration individuel des huitres. Ce taux a et6 Cvalue par la perte de fluorescence, mesuree par spectrofluorimetrie de flux, des microalgues introduites comme nourriture. Qu'elles soient aliment&es avec A. tamarense toxique ou non toxique, ou avec A. fundyense toxique, les huitres ont montre dans tous les cas une activite de filtration du type s ouverture/fermeture B. En revanche, les huitres temoins nounies avec une algue de reference, Isochrysis sp., connue par ailleurs pour ameliorer leur croissance, ne montraient pas ce type d'activite. Les huitres creuses preacclimatees a une alimentation a base de Isochrysis sp. et nourries avec differents melanges Alexundrium/lsochrysis, ont Cgalement montre une filtration de type ouverture/fermeture. Des expositions de 48 heures au regime constitue d'A. tamarense non toxique n'ont pas ameliore la filtration, que ce soit avec Alexundrium ou Isochrysis. 0 Ifremer-Elsevier, Paris Crassostrea gigas / physiologie alimentaire / microalgues toxiques / dinoflagellks / paralytic shellfish poisoning

In October and November 2003 a bloom of the toxic dinoflagellate Alexandrium catenella was observed in the Northeast zone of Thau lagoon (French Mediterranean coast). Sea water samples were collected every hours to evaluate time-related variations of phytoplankton concentrations and to compare the relative ratio of A. catenella versus other phytoplankton species during the outbreak. From these observations, trials using recirculated sea water systems were performed to: i) evaluate the physiological effects on oyster of increasing proportions of A. catenella within a mixed microalgal diet where the non-toxic diatom Thalassiosira weissflogii was present at concentration: 1500 cells ml −1 , ii) compare the effect of two temperatures (12 • C and 18 • C) on paralytic toxin accumulation rates in oyster flesh by ion-pairing high performance liquid chromatography (IP-HPLC) detection, iii) analyse toxin biotransformation during the contamination process, iv) evaluate the role of the different types of oyster tissue on the bioaccumulation mechanism. The results showed: i) a significant effect of temperature increase on clearance rate and toxin uptake, ii) no detectable time-related effects of toxic algal food on pre-ingestion sorting or on toxin profiles in oyster flesh, iii) either negative or positive effects of A. catenella concentrations on toxin uptake ("threshold" effect), iv) high amounts of toxins in the digestive gland, accounting for more than 80% of overall shellfish toxicity. The daily amount of toxins (Q tox) taken up by each oyster was evaluated by means of a global one-compartment model. However a two-compartment model finally gave the best match with real contamination kinetics, since it integrated both toxins sequestered in oyster tissues and toxins moving in through the digestive tract.

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