Raphael Brizard

Study of genetic damage in the japanese oyster induced by an environmentally-relevant exposure to diuron: evidence of vertical transmission of dna damage

Pesticides represent a major proportion of the chemical pollutants detected in French coastal waters and hence a significant environmental risk with regards to marine organisms. Commercially-raised bivalves are particularly exposed to pollutants, among them pesticides, as shellfish farming zones are subject to considerable pressure from agricultural activities on the mainland. The aims of this study were to determine (1) the genotoxic effects of diuron exposure on oyster genitors and (2) the possible transmission of damaged DNA to offspring and its repercussions on oyster fitness. To investigate these points, oysters were exposed to concentrations of diuron close to those detected in the Marennes-Oleron Basin (two 7-day exposure pulses at 0.4 and 0.6 μg L(-1)) during the gametogenesis period. Genomic abnormalities were characterized using two complementary approaches. The Comet assay was applied for the measurement of early and reversible primary DNA damage, whereas flow cytometry was used to assess the clastogenic and aneugenic effect of diuron exposure. Polar Organic Chemical Integrative Samplers (POCIS) were used in exposed and assay tanks to confirm the waterborne concentration of diuron reached during the experiment. The results obtained by the Comet assay clearly showed a higher level of DNA strand breaks in both the hemocytes and spermatozoa of diuron-exposed genitors. The transmission of damaged genetic material to gamete cells could be responsible for the genetic damage measured in offspring. Indeed, flow cytometry analyses showed the presence of DNA breakage and a significant decrease in DNA content in spat from diuron-exposed genitors. The transmission of DNA damage to the offspring could be involved in the negative effects observed on offspring development (decrease in hatching rate, higher level of larval abnormalities, delay in metamorphosis) and growth. In this study, the vertical transmission of DNA damage was so highlighted by subjecting oyster genitors to short exposures to diuron at medium environmental concentrations. The analysis of POCIS showed that oysters were exposed to integrated concentrations as low as 0.2 and 0.3 μg L(-1), emphasizing the relevance of the results obtained and the risk associated to chemical contamination for oyster recruitment and fitness.

Acquisition of susceptibility to Vibrio penaeicida in Penaeus stylirostris postlarvae and juveniles

Vibriosis is a major disease problem in shrimp aquaculture, affecting all developmental stages, from larvae in hatchery tanks to juveniles and broodstock in growout ponds. However, bacterial strains responsible for vibriosis in the successive stages are usually considered to be different, and virulence specificity has been reported both at the species and at the stage levels. The so-called « Syndrome 93 » is a seasonal juvenile vibriosis caused by Vibrio penaeicida which affects Penaeus stylirostris in growout ponds and broodstock tanks in New Caledonia. This pathology does not cause any mortality in hatchery or nursery phases. An experimental infection design using balneation of postlarvae and early juveniles in V. penaeicida suspensions was used to evaluate the developmental stage at which shrimp become sensitive to this vibriosis. We demonstrated that the acquisition of susceptibility to this pathogen is very sudden and correlated with the acquisition of the definitive rostral formula, and from this draw conclusions regarding virulence mechanisms of V. penaeicida in P. stylirostris.

Changes in motility, ATP content, morphology and fertilisation capacity during the movement phase of tetraploid Pacific oyster (Crassostrea gigas) sperm

Changes in sperm features during the movement phase are especially interesting to study in external fertilization species whose sperm duration movement is long because this implies a significant adaptation of moving cells to the external medium. This study describes the changes in tetraploid Pacific oyster sperm characteristics in relation to time post activation. Sperm individually collected on three tetraploid males were activated in seawater. Their features were analysed over a 24h period and compared to a sperm pool collected on three diploid males as a reference. The percentage of motile spermatozoa, the intracellular ATP content, and the fine structure of spermatozoa were studied in relation to time post activation. Furthermore, the fertilisation capacity of sperm individually collected on five diploid males was assessed after 1 and 24h post activation. A forward progressive movement was maintained for at least a 20h duration. Compared to diploid males, the percentage of motile spermatozoa was lower in tetraploid males. The intracellular ATP concentration was higher in spermatozoa from tetraploid males than in spermatozoa from diploid males. A decrease in ATP content was observed in the first 6h post activation and severe alterations were observed in sperm morphology after 24h. Then, a lower fertilisation capacity of sperm from diploid males was observed at the end of the movement phase. The cessation of Pacific oyster sperm motility was unlikely caused by ATP consumption as ATP concentration was still high at the end of sperm movement but rather caused by drastic changes in sperm morphology. Compared to sperm collected on diploid males, the lower quality of sperm from tetraploid males was emphasized by a shorter movement duration and deeper morphological alterations at the end of the movement phase.

A white spot disease-like syndrome in the Pacific blue shrimp (Litopenaeus stylirostris) as a form of bacterial shell disease

In May 1997, some white lesions evoking the white spot syndrome disease were observed in Litopenaeus stylirostris broodstock in New Caledonia. The occurrence of these lesions was neither associated with mortality, nor with histological evidence of white spot syndrome baculovirus (WSBV). The evidence suggests that these lesions result from a form of bacterial disease and are associated with an increased bacterial flora on the outer surface of the cuticle, as well as an increased incidence throughout the molt cycle. A microscopic observation of these lesions allows them to be differentiated from WSBV-associated lesions.