Bactericidal activity of brook trout (Salvelinus fontinalis) peritoneal macrophages against avirulent strains of Aeromonas salmonicida

Abstract

The bactericidal activity of glycogen-elicited peritoneal macrophages of brook trout (Salvelinus fontinalis) against avirulent strains of Aeromonas salmonicida was assayed using the tetrazolium dye, MTT. Peritoneal macrophages effectively killed all avirulent strains of A. salmonicida tested during a 5 h incubation period when bacterial concentrations of 4 × 103-6·25 × 104 cfu were added to tissue culture plate wells containing 5 × 105 macrophages. Bacteria were not killed when RTG-2 cells were used in lieu of macrophages and there was a clear dose response between the number of macrophages seeded and the efficacy of killing, indicating that macrophages were the effector cells. Bacterial killing was confirmed by the plate count method which showed that greater than 80% of the bacteria added to macrophages were killed within the first hour of exposure. Killing was demonstrated with peritoneal macrophages obtained from juvenile and adult brook trout as well as with peritoneal macrophages isolated from Atlantic salmon (Salmo salar) smolts. This study demonstrated that the MTT bactericidal assay can be used to detect bacterial killing. However, the plate count method described in this study gives reproducible results, and demonstrates the rapid and efficient killing of avirulent strains of A. salmonicida by brook trout and Atlantic salmon peritoneal macrophages.