The challenge strain (89/4762) was cultured using an aerated 10-l carboy at room temperature (20 °C) in MSB. The bacteria were grown in 5 L of broth with aeration (120 l/h). After 48 h the cells were collected by centrifugation at 3000 ×g for 30 min at 4 °C and washed three times using 0.22 μm filtered seawater. Cell concentration was determined microscopically using a Helber counting chamber and adjusted to a final concentration of 1.0 × 106 cells/ml which was used as the challenge dose. This dose was calculated to give a 60%