S-layer-mediated association of Aeromonas salmonicida with murine macrophages

The interaction of Aeromonas salmonicida with the murine macrophage (Mφ) cell line P388D₁ was used as a convenient model to study the involvement of the bacterial crystalline surface array (or A-layer) in the association with Mφs. A-layer-positive (A⁺) cells readily associated with Mφs in phosphate- buffered saline, whereas A^- mutants were unable to do so, even when the bacterium-Mφ interaction was forced by centrifugation. Mφs selectively interacted with A⁺ cells when challenged with mixtures of A⁺ and excess A^- cells. Electron microscopy indicated that in phosphate-buffered saline only A⁺ bacteria were readily internalized, although by a nonconventional mechanism, suggesting that efficient phagocytosis in the absence of opsonins was A-layer mediated. Latex beads coated with a partially assembled A-layer were more efficiently taken up than uncoated or A-protein-coated beads, indicating that an organized A-layer was essential for Mφ uptake. The reduced ability of Mφs plated on a substratum coated with the A-layer to bind A⁺ bacteria also suggested that association was both A-layer and receptor mediated. In the presence of tissue culture medium, competent Mφs interacted efficiently with A^- bacteria and internalized them through conventional phagocytosis. A⁺ cells were markedly cytotoxic to Mφs, whereas the A-protein or A-layer was not. A^- cells were cytotoxic to a lesser extent, suggesting that cytotoxicity was targeted.