Immunohistochemical studies of Aeromonas salmonicida infection in freshwater pomfret (Colossoma macropomum)

Abstract

Aeromonas salmonicida cause furunculosis disease in both freshwater and marine fish, which represents a great threat in fish farming. The aim of this research to develop and produce polyclonal A. salmonicida antibody from rabbits to strengthen diagnostic method of furunculosis through immunohistochemistry method. A total of 15 samples of freshwater pomfret (Colossoma macropomum) was divided into 3 groups which intramuscularly injected with 10⁹ cell/mL of Aeromonas salmonicida antigen, then sacrificed on the third, sixth, and ninth day post infection for histological and immunohistochemical purpose. Polyclonal primary antibody against A. salmonicida infection was obtained from rabbit’s serum which was previously immunized with the O antigen of A. salmonicida containing 10⁹ cells/mL, with weekly increased volume (0.5; 1; 2; 3 mL). Anti-Aeromonas salmonicida rabbit serum was then purified to obtain its immunoglobulin G by precipitating it first using ammonium sulfate and filtered using dialysis membrane.  Immunoreactivity against the bacterial antigen was evidenced in the skin, spleen, liver, and kidney starting from third day post infection and further distributed until ninth day post infection. Septicaemia in challenged fish was evidenced by histology examination characterized by vascular and necrotic changes. Polyclonal antibody against Aeromonas salmonicida and immunohistochemistry was found to be useful for identifying the pathogen from the first stage of the disease and differentiate A. salmonicida infection from others bacterial septicemic disorders in freshwater pomfret.