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The interaction of Legionella micdadei with human polymorphonuclear neutrophils (PMNs) and serum was investigated to determine their roles in host defense against this organism. Serum and PMNs from normal donors having no antibody to L micadei were used. PMNs phagocytized once-agar-passaged (74.6% +/- 6.5%) and twice-agar-passaged (87.3% +/- 1.0%) L micdadei less (P less than 0.05) than L micdadei passaged multiple times on agar (97.5% +/- 1.0%) or Staphylococcus aureus (98.3% +/- 0.5%). Under the same conditions, no phagocytosis of Legionella pneumophila occurred. Use of heat-inactivated serum abolished phagocytosis. PMN killing of once-agar-passaged (1.5% +/- 1.5%) and twice-agar-passaged (0) L micdadei was less (P less than 0.001) than that of L micdadei passaged multiple times on agar (88.6% +/- 4.0%) or S aureus (96.8% +/- 0.5%). L micdadei was not killed by fresh serum, although, in contrast to L pneumophila, it was opsonized by C3. Thus virulent L micdadei is phagocytized, but not killed, by human PMNs, with complement being the major opsonin.

We have recently shown that Legionella micdadei is ingested, but not killed, by human neutrophils. Herein we investigate the role of human monocytes in defense against this organism. Serum and monocytes from normal donors having no detectable antibody to L. micdadei were used. Egg-passaged L. micdadei organisms multiplied inside these monocytes with a peak growth of 2 log units within 12 h. No growth occurred when monocytes were omitted or when sonicated monocytes were used. Electron microscopy 18 h after infection revealed these organisms to be intracellular in normal-appearing phagosomes. When the input multiplicity of L. micdadei was greater than 1 CFU per monocyte, no intracellular growth occurred. When egg-passaged Legionella pneumophila organisms were used, intracellular organisms were found in phagosomes studded with ribosomes at the same time period. Peak intracellular growth of L. pneumophilia occurred by 48 h. L. micdadei activated the complement system and was opsonized by C3. However the use of complement-depleted (heat-inactivated) serum as the opsonic source had no effect on the bacterium's ingestion or growth in the monocyte. Thus, L. micdadei multiples in human monocytes. This entry and growth is independent of antibody or complement. The intracellular locations of L. micdadei and L. pneumophila differ, suggesting different mechanisms for the survival of these two organisms in the monocyte.

The roles of the classical and alternative pathways of complement activation and of antibody in the phagocytosis of Legionella micdadei by polymorphonuclear leukocytes were studied. Normal serum was treated with the appropriate chelators or with heat to inactivate the classical, alternate, or both pathways of complement activation. Normal and complement-depleted sera with or without antibody were employed as opsonins for L. micdadei in phagocytosis assays. There was no difference in the phagocytosis of L. micdadei promoted by normal serum and either C4-deficient serum or serum in which the classical pathway had been inactivated. Both normal and classical pathway-deficient sera promoted significantly greater phagocytosis than did sera in which the alternate pathway or both the alternate and classical pathways had been inactivated. Thus, polymorphonuclear leukocyte phagocytosis of L. micdadei in the absence of antibody required an intact alternate pathway. Specific antibody partially restored opsonization to sera deficient in the alternate or both complement pathways, but phagocytosis was still significantly less than that with the alternate pathway intact.