Major surface protein 2 (MSP2) of the bovine rickettsial pathogen Anaplasma marginale is an abundant, serologically immunodominant outer membrane protein. Immunodominance partially results from numerous CD4+ T cell epitopes in highly conserved amino and carboxy regions and the central hypervariable region of MSP2. However, in long-term cultures of lymphocytes stimulated with A. marginale, workshop cluster 1 (WC1)+ gammadelta T cells and CD4+ alphabeta T cells proliferated, leading to a predominance of gammadelta T cells. As gammadelta T cells proliferate in A. marginale-stimulated lymphocyte cultures, this study hypothesized that gammadelta T cells respond to the abundant, immunodominant MSP2. To test this hypothesis, gammadelta T cell clones were isolated from MSP2 vaccinates and assessed for antigen-specific proliferation and interferon-gamma secretion. Seven WC1+ gammadelta T cell clones responded to A. marginale and MSP2, and three of these proliferated to overlapping peptides from the conserved carboxy region. The gammadelta T cell response was not major histocompatibility complex-restricted, although it required antigen-presenting cells and was blocked by addition of antibody specific for the T cell receptor (TCR). Sequence analysis of TCR-gamma and -delta chains of peripheral blood lymphocytes identified two novel TCR-gamma chain constant (Cgamma) regions. It is important that all seven MSP2-specific gammadelta T cell clones used the same one of these novel Cgamma regions. The TCR complementarity-determining region 3 was less conserved than those of MSP2-specific CD4+ alphabeta T cell clones. Together, these data indicate that WC1+ gammadelta T cells recognize A. marginale MSP2 through the TCR and contribute to the immunodominant response to this protein.