Through complex interplay with APCs, subsets of NK cells play an important role in shaping adaptive immune responses. Bovine tuberculosis, caused by Mycobacterium bovis, is increasing in incidence and detailed knowledge of host–pathogen interactions in the natural host is essential to facilitate disease control. We investigated the interactions of NK-cell sub-populations and M. bovis-infected DCs to determine early innate mechanisms in the response to infection. A sub-population of NK cells (NKp46+CD2−) selectively expressing lymphoid homing and inflammatory chemokine receptors were induced to migrate towards M. bovis-infected DCs. This migration was associated with increased expression of chemokines CCL3, 4, 5, 20 and CXCL8 by M. bovis-infected DCs. Activation of NKp46+CD2− NK cells and secretion of IFN-γ was observed, a response reliant on localised IL-12 release and direct cellular interaction. In a reciprocal manner, NKp46+CD2− cells induced an increase in the intensity of cell surface MHC class II expression on DCs. In contrast, NKp46+CD2+ NK cells were unable to secrete IFN-γ and did not reciprocally affect DCs. This study provides novel evidence to demonstrate distinct effector responses between bovine NK-cell subsets during mycobacterial infection.