An enzyme linked immunosorbent assay for the detection of Kudoa thyrsites in Atlantic salmon Salmo salar

An antigen capture enzyme linked immunosorbent assay (ELISA) using high-titre rabbit polyclonal antiserum was developed to detect soluble Kudoa thyrsites antigen in skeletal muscle of Atlantic salmon. Raw serum provided capture antibodies and biotinylated polyclonal immunoglobulins served to detect the captured antigen. The optical density (OD) regressed significantly with the number of purified, glutaraldehyde-fixed K. thyrsites spores. Higher OD values were obtained with antigen extracted from the supernatant of an infected muscle homogenate using cold buffer containing Tween 20 compared with Triton X-100 or without detergent. OD values were also higher from cold supernatants compared with boiled supernatants and with uncentrifuged homogenates. OD values from unexposed, freshwater-reared salmon were negligible. Agreement between ELISA OD values and the results of polymerase chain reactions derived from 25 exposed Atlantic salmon was good compared with the agreements between ELISA or PCR and histological analysis. The relationship between a histologically determined K. thyrsites plasmodium score and ELISA OD value suggested the presence of histologically undetectable extrasporogonic stages in some samples. Serological detection of soluble K. thyrsites antigen may form the basis of a quantitative, field-based diagnostic assay.