Programme Hospitalier Recherche Clinique, Institut Pasteur, Inserm, French Public Health Agency.

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International audienceSince 2008, mass mortalities of 1-yr-old Crassostrea gigas associated with the ostreid herpesvirus OsHV-1 μVar have occurred along all the coasts of France when seawater temperature reaches 16 to 17°C. The present study aimed to characterize the effect of temperature on oyster survival in combination with OsHV-1 DNA quantification by standard real-time PCR and total vibrio population levels in oyster tissues. To examine the effect of seawater temperature on disease transmission and related mortality of oysters, cohabitation experiments were conducted between healthy naïve oysters and oysters previously exposed to field conditions in areas where mortalities were occurring. Oysters initially maintained in controlled conditions (free of mortality and negative for OsHV-1), and then transferred to an area where high mortalities were occurring among farmed stocks, became infected with OsHV-1 and exhibited high loads of vibrios followed by significant mortalities. When previously exposed oysters were maintained indoors at 13.0°C for 40 d and then at 20.6°C, they exhibited no mortality, were negative for OsHV-1 detection, and did not transmit the disease to healthy oysters. Survival of previously exposed oysters maintained indoors at 8 temperatures ranging from 13.4 to 29.0°C varied from 25 to 48% and was negatively correlated with holding temperature. Concomitantly, survival of naïve cohabiting animals (62 to 98%) decreased with increasing seawater temperature until a plateau was reached between 16.2 and 21.9°C, and increased at higher temperatures. Therefore, the optimal temperature range for disease transmission from field-exposed to naïve animals was between 16.2 and 21.9°C. Our results suggest that a long-term period (40 d) at low temperature (13°C) may offer a method of mitigating mortalities in oysters that have been exposed to an infective environment

Several studies have pointed out ethical shortcomings in the decision-making process for withholding or withdrawing life-supporting treatments. We conducted a study to evaluate the perceptions of all caregivers involved in this process in the intensive care unit. A closed-ended questionnaire was completed by 3,156 nursing staff members and 521 physicians from 133 French intensive care units (participation rate, 42%). Decision-making processes were perceived as satisfactory by 73% of physicians and by only 33% of the nursing staff. More than 90% of caregivers believed that decision-making should be collaborative, but 50% of physicians and only 27% of nursing staff members believed that the nursing staff was actually involved (p < 0.001). Fear of litigation was a reason given by physicians for modifying information given to competent patients, families, and nursing staff. Perceptions by nursing staff may be a reliable indicator of the quality of medical decision-making processes and may serve as a simple and effective tool for evaluating everyday practice. Recommendations and legislation may help to build consensus and avoid conflicts among caregivers at each step of the decision-making process.

A novel technique was developed for the flocculation of marine microalgae commonly used in aquaculture. The process entailed an adjustment of pH of culture to between 10 and 10.6 using NaOH, followed by addition of a non-ionic polymer Magnafloc LT-25 to a final concentration of 0.5 mg L−1. The ensuing flocculate was harvested, and neutralised giving a final concentration factor of between 200-and 800-fold. This process was successfully applied to harvest cells of Chaetoceros calcitrans, C. muelleri, Thalassiosira pseudonana, Attheya septentrionalis, Nitzschia closterium, Skeletonema sp., Tetraselmis suecica and Rhodomonas salina, with efficiencies ≥80%. The process was rapid, simple and inexpensive, and relatively cost neutral with increasing volume (cf. concentration by centrifugation). Harvested material was readily disaggregated to single cell suspensions by dilution in seawater and mild agitation. Microscopic examination of the cells showed them to be indistinguishable from corresponding non-flocculated cells. Chlorophyll analysis of concentrates prepared from cultures of ≤130 L showed minimal degradation after 2 weeks storage. Concentrates of T. pseudonana prepared using pH-induced flocculation gave better growth of juvenile Pacific oysters (Crassostrea gigas) than concentrates prepared by ferric flocculation, or centrifuged concentrates using a cream separator or laboratory centrifuge. In follow up experiments, concentrates prepared from 1000 L Chaetoceros muelleri cultures were effective as supplementary diets to improve the growth of juvenile C. gigas and the scallop Pecten fumatus reared under commercial conditions, though not as effective as the corresponding live algae. The experiments demonstrated a proof-ofconcept for a commercial application of concentrates prepared by flocculation, especially for use at a remote nursery without on-site mass-algal culture facilities.

Microalgae commonly used as feed for bivalves, Pavlova lutheri (P), Isochrysis affinis galbana (T) and Chaetoceros calcitrans forma pumilum (Cp), were fed to Pacific oyster Crassostrea gigas to assess their nutritional value for larval development and metamorphosis during two experiments. Monospecific, bispecific and trispecific diets were firstly evaluated during 3 weeks from D larvae to young postlarvae. Then bispecific diets, based on different T and Cp proportions, were assessed during a similar period. Concurrently, ingestion was studied through the whole larval and postlarval development for each diet and/or diet mixture. Because lipids are assumed to be a key nutrient for bivalves, biochemical analysis was undertaken on the second set of trials focused on fatty acids and sterols. Compared to the other diet mixtures (mono and plurispecific diet) TCp induced the best larval growth performance (13.2 μm day− 1), a high larval survival (98%) but did not result in higher metamorphosis (72%). In contrast, monospecific diet P was the poorest for larvae with low growth and low survival. When varying T and Cp proportions, best larval developments were induced with 25T/75Cp and 50T/50Cp diets, though quite similar to that obtained with 75T/25Cp. In contrast, unbalanced diets (95T/5Cp and 95Cp/5T) led to low larval performances. In addition, grazing experiences showed preferential uptake of microalgae with P < PT much less-than T much less-than Cp much less-than TCp = PCp = PTCp. For mixed diets a low daily consumption (< 10 000 microalgae per larvae) was noted during the first week followed by a second phase (next 8-10 days) with a sharp increase and regular intake, reaching 90 000 microalgae per larvae per day. Finally, a marked drop (40 000 microalgae per larvae) was observed at the beginning of metamorphosis from days 20 to 21. Principal component analysis between main fatty acids (19) and sterols (7) detected in larvae and postlarvae was used to discriminate profiles according to diets and/or metamorphosis competence. The correlation circle representation showed that the 26 variables are well explained by these combined variables (78%) with a repartition along the first principal component according to diets with a gradient from 5T/95Cp to 95T/5Cp. In contrast, postlarvae and larvae were discriminated on the second principal component while no relationships were found between competent and incompetent larvae.

Scallop Pecten maximus larvae have been cultured at the Argenton and Tinduff (Brittany, France) hatcheries with antibiotic treatment (chloramphenicol at 8 ppm) for 15 yr. Without treatment, outbreak of disease has normally occurred between Day 12 and Day 19 or sometimes earlier. A bacteriological study of larvae reared with and without antih~ot~c was performed over a 4 yr period. Among the collected strains, 2 clusters (C and F) of vibrios were present at high densities only in larvae cultured without treatment. One cluster (C) was routinely ~solated over the 4 yr of study, while the other (F) was collected only in the third year. Their virulence with respect to scallop larvae and their lack of infectivity with respect to oyster larvae were demonstrated in an exposure experiment. The vibrio F strain tended to lose its virulence after 5 subcultures, whereas the vibrio C strain retained the ability to kill scallop larvae in experimental infections. Three other vibrios isolated in moribund oyster larvae caused mortality in oyster larvae but not in scallop larvae. Different methods were used to determine the taxonomic position of these virulent bacteria. The phenotypic traits of bacterial isolates were determined with the Biolog GN microplate, the API 20E system and the reference method. Patterns of cytoplasmic proteins were identified by electrophoresis in SDS-PAGE. These different methods consistently confirmed the existence of 2 vibrio species pathogenic to scallop larvae. Affiliation of cluster F with Vibrio splendidus was assessed by Biolog tests and by analysis of 16s rRNA sequences. One pathogenic bacteria of oyster larvae was also very close to this second cluster, whereas the 2 others from moribund oyster larvae and cluster C may constitute 2 different species.