Human granulocytic ehrlichiosis (HGE) is an emerging tick-borne infection with a specific tropism for granulocytes. We previously isolated and cultivated the HGE agent in the promyelocytic leukemia cell line HL-60 and have also demonstrated the susceptibility of both granulocytic and monocytic human marrow progenitors. Circulating monocytes have not been observed to be infected, suggesting that cell susceptibility may be differentiation specific. To evaluate this hypothesis, HL-60 cells were differentiated towards granulocytes (with dimethyl sulfoxide or all-trans retinoic acid) or toward monocytes-macrophages (with 12-O-tetradecanoylphorbol-13-acetate [TPA], gamma interferon, or 1,25-dihydroxyvitamin D₃) and then challenged with HGE. HGE binding, internalization, and proliferation were compared in differentiated and untreated control HL-60 cells by immunofluorescence, electron microscopy, and Giemsa staining. Granulocytic differentiation resulted in a doubling of HGE binding and enhanced infection consistent with the agent's clinical tropism for neutrophils. Granulocytic cells were unable to kill internalized ehrlichiae even after activation induced by N-formyl-Met-Leu-Phe alone or together with tumor necrosis factor alpha. In contrast, monocyte-macrophage differentiation with TPA resulted in complete resistance to infection through at least two distinct mechanisms: (i) reduction in binding and uptake and (ii) killing of any internalized organisms. Diminished binding in TPA-treated cells correlated with their reduced expression of sialyl Lewis x (CD15s), a putative cellular receptor component for HGE. The degree of monocytic differentiation and activation induced (i.e., TPA > gamma interferon > vitamin D₃) correlated with resistance to HGE. Thus, HL-60 cells exhibit a striking differentiation-specific susceptibility to HGE. Differentiation-induced changes in bacterial adhesion and killing capacity underlie the tropism of HGE for granulocytic HL-60 cells and, conversely, the resistance of activated macrophages to infection.