NK dendritic cells (NKDC) are recently described immunologic cells that possess both lytic and Ag-presenting function and produce prolific quantities of IFN-gamma. The role of NKDC in innate immunity to bacterial infection is unknown. Because IFN-gamma is important in the immune response to Listeria monocytogenes (LM), we hypothesized that NKDC play a critical role during LM infection in mice. We found that LM increased the frequency and activation state of NKDC in vivo. Using in vivo intracellular cytokine analysis, we demonstrated that NKDC are a major source of early IFN-gamma during infection with LM. Adoptive transfer of wild-type NKDC into IFN-gamma-deficient recipients that were subsequently infected with LM decreased bacterial burden in the liver and spleen and prolonged survival. In contrast, NK cells were depleted early during LM infection, produced less IFN-gamma, and conferred less protection upon adoptive transfer into IFN-gamma-deficient mice. In vitro, LM induction of IFN-gamma secretion by NKDC depended on TLR9, in addition to IL-18 and IL-12. Our study establishes NKDC as innate immune responders to bacterial infection by virtue of their ability to secrete IFN-gamma.