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The cytological response of rainbow trout Oncorhynchus mykiss head kidney macrophages to ingested Mycobacterium spp. was examined in vitro. Mycobacterium marinum or Mycobacterium sp. TB267 isolated from snakehead fish Channa striata Bloch were opsonised with either fresh rainbow trout serum, serum which had been heat-inactivated, or rainbow trout antiserum against the extracellular products (ECP) of the 2 Mycobacterium spp. A monoclonal antibody against the ECP was also used as an opsonin. Suspensions of macrophages were prepared (1 ml of 1 x 10(7) cells ml-1), mixed with the opsonised bacteria (100 microliters of 2 x 10(9) ml-1), and incubated at 18 degrees C for 0.5, 1, 2, 4 or 6 h to allow phagocytosis to occur. A quantitative evaluation of the phagocytosis of the mycobacteria by the macrophages was carried out by electron microscopy. Macrophage phagosomes and their contents were examined and numbers of intact and partially degraded bacteria determined. Pre-labelling dense granules (secondary lysosomes) with ferritin enabled phagosome lysosome fusion to be identified and their frequency determined. Opsonisation of the mycobacteria was found to greatly enhance the phagocytic and killing activity of the rainbow trout macrophages.

Electron microscope studies of the in vitro phagocytosis of Mycobacterium spp. by rainbow trout Oncorhynchus mykiss head kidney macrophages