Examples exist in the literature that demonstrate that treatment with immunostimulatory cytosine-phosphate-guanosine (CpG)-DNA can protect mice against infection by intracellular pathogens. There are, however, few studies reporting that CpG-DNA offers similar disease protection in other species. In this study, we assessed the potential of a class A and class B CpG oligodeoxynucleotide (ODN) to induce innate immune responses in sheep, an outbred species. Using peripheral blood mononuclear cells, we have for the first time demonstrated CpG-ODN-induced innate immune responses, including natural-killer-like activity [non-major histocompatibility complex (MHC)-restricted cytotoxicity], interferon-alpha secretion and 2'-5'A oligoadenylate synthetase activity, that could contribute to immune protection in sheep. The type and magnitude of these responses were dependent on ODN class and non-MHC-restricted killing was not associated with interferon-gamma production. The latter observation is in contrast with observations reported for mice and humans. These observations support the conclusion that differences in CpG-ODN-induced responses exist among species and that specific ODN sequences can significantly influence innate immune responses.