IHNV challenge of rainbow trout. At 30 days postvaccination (dpv), fish (mean
weight, 1.33 g) were challenged by immersion with the IHNV-RA strain. Fish
were challenged with the viral doses 103 or 105 PFU/ml. Two tanks (50 fish per
tank) of each group were exposed to each IHNV-RA dose in 2 liters of water at
15°C for 5 h. The fish were monitored for clinical signs of disease, such as a
distended abdomen, petechial hemorrhaging, and a whirling form of swimming.
Postchallenge fish mortality in each tank was recorded, and dead fish were
removed daily for 30 days. Plaque assays were performed to confirm that the fish
were infected with IHNV (data not shown). The relative percent survival (RPS)
was calculated for each vaccination and challenge. RPS was calculated by the
following formula, as described by Johnson et al. (23): RPS 5 [1 2 (% mortality
of vaccinated fish/% mortality of control fish)] 3 100, where pcDNA3-injected
fish were used as the control fish for these calculations.