Flow cytometry as a tool for assessment of the fish cellular immune response to pathogens

Flow cytometry provides rapid and reproducible methods for analysing fish leucocyte subpopulations and cellular immune functions. Assays devoted to the study of non-specific cellular defence mechanisms of rainbow trout were developed. In addition to the assessment of these defence mechanisms (phagocytic activity, oxidative burst and natural cytotoxicity assays), the modulations of cell responses under pathological conditions were analysed. Flow cytometric profiles of cell populations are representative of the organ from which the leucocytes originated. Diseases induce modifications of cell number and cell morphology. Study of cytograms provides helpful information on the fish reaction to pathogens. Thus the kidney inflammatory response characterizing proliferative kidney disease is mostly due to the proliferation of the kidney lymphocytes. Flow cytometric study of the cell cycle has allowed the demonstration of the in situ proliferation of the pronephros lymphocytes. Impairment of phagocytic activity occurred very early after VHSV experimental infection. Despite wide inter-fish variability, a reduced phagocytic activity was demonstrated in PKX-infected trout. The oxidative burst was not affected by either of these pathogens. Non-specific cytotoxic activity was observed in blood, pronephros, spleen and thymus. The levels of cytotoxicity varied considerably between individual fish. Even though the large inter-fish variability hampered the interpretation of the data, flow cytometry provides useful techniques for the study of immunological and clinical aspects of fish pathology.