Antigenic and biochemical study of PKX, the myxosporean causative agent of proliferative kidney disease of salmonid fish

This work aimed at developing techniques to enable further investigations on the epidemiology and immunology of proliferative kidney disease (PKD) and of its causative agent, PKX, a probable myxosporean parasite. An improved PKX cell concentration technique resulted in preparation of an enriched proliferative kidney cell (EPKC) antigen (Ag) that offered a suitable material for rabbit and mouse immunizations, electrophoretical analysis and antigenicity studies of parasitic materials. Rabbit sera and a panel of 11 monoclonal antibodies (MAbs) specific for PKX were obtained. They reacted in ELISA, immunofluorescence tests, immunohistochemistry, immunodot blot and Western blot with EPKC Ag, live or fixed parasitic cells and infected kidney extracts, depending on the Abs and immunological test used. These MAbs were divided into 4 groups according to their reactivity in the above tests and led to the study of the possible localization of putative epitopes on parasite cells. At least 4 major parasitic proteins were visualized by SDS-PAGE electrophoresis and a fifth one, with a relative mobility of 13 kDa, was immunostained in Western blot by at least 2 MAbs and rabbit antiserum. The membrane fluorescence reactivity of the epitope suggested this protein was external and could be of immunological importance. The antigenicity detected by histoimmunochemistry was that of primary cells and was apparently specific for PKX since the screening of kidney tissue of 9 non-salmonid fish species harbouring developmental stages of 3 myxosporean genera remained negative. EPKC Ag was also antigenic for trout undergoing experimental PKD, thus providing evidence for a humoral response to this infection. Our investigations result in the establishment of a novel set of immunological probes suitable for monitoring the course of PKD, checking antigenicity of actinosporeans, purifying parasitic proteins and screening a PKX expression Library.