Design of pesticide exposition and experimental viral infection.

<p>Pesticide contamination. Pacific oysters were exposed with the mixture of 14 pesticides during 24h. Oysters were not fed during the pesticide contamination. Three tanks were used in each experiment: 2 tanks with pesticides (60 oysters per tank, 120 oysters in total) and 1 tank without pesticides (90 oysters) receiving the same quantity of solvent used for preparing pesticides (the solvent was added in the water; negative controls). Experimental viral infection. After pesticide exposition during 24 hours, experimental OsHV-1 infection was carried out by intramuscular injection of a viral suspension. Experimental design included also oysters injected with an OsHV-1 suspension treated with pesticides. Two hundred and ten oysters were “anaesthetized” during 4 h. One hundred μL of an OsHV-1 (μVar genotype) suspension at 1 × 10⁴ copies of viral DNA/μL or sterile artificial water were injected. Oysters were then placed in tanks containing 3 L of filtered seawater (1 μm) at 22°C without food supply (ten oysters per tank) using the following design: (A) oysters injected with sterile artificial seawater (3 tanks with seawater for oysters without pesticides, 6 tanks with seawater for oysters with pesticide exposure), (B) oysters injected with OsHV-1 suspension (3 tanks with seawater for oysters without pesticides, 6 tanks with seawater for oysters with pesticide exposure), (C) oysters injected with OsHV-1 suspension treated with pesticides 10X (3 tanks with seawater for oysters with pesticide exposure). Survival was monitored during nine days after injection.