The innate immune response to pathogens is critical to host control of infection. Trypanosoma cruzi induces a robust type I interferon (IFN) response in a wide variety of cell types. We demonstrate that T. cruzi engages a novel signaling pathway that converges on TBK1 and IRF3 to induce IFNβ in isolated macrophages and fibroblasts. Additionally, T. cruzi triggers a type I IFN response in vivo, at a local infection site. Robust induction of type I IFN-stimulated genes is observed in excised skin 24 hours post-infection where the level of induction is parasite strain-dependent, with a low virulence strain triggering a muted IFN response. Significant induction of a type I IFN response both in vitro and at the site of infection in vivo suggests a role for this family of cytokines in T. cruzi infection. Although previous attempts to characterize the role of type I IFNs in T. cruzi using mice deficient in type I IFN signaling (IFNAR ^-/-) have yielded conflicting results, a more systematic examination of the parameters of strain and dose on T. cruzi infection in WT and IFNAR^-/- mice demonstrates that the effects of type I IFN in the context of T. cruzi infection are dependent on infective dose. Specifically, mice deficient in type I IFN signaling are protected from lethal infection while the mice of the same genetic background infected with a sub-lethal dose of T. cruzi show no significant differences in morbidity or mortality. Although the mechanism for this detrimental effect of type I IFNs is not clear, gross differences in pathology, infiltrating cell types, levels of apoptosis, or cytokine profiles are not observed. Taken together, these results suggest a complex immunomodulatory role for type I IFNs that could influence infection and pathology during T. cruzi infection. Further characterization of the signaling pathways upstream of T. cruzi-induced IFNβ production and the mechanisms by which type I IFNs are detrimental during T. cruzi infection will provide valuable insight into the complex signaling networks and immunomodulatory effects of this family of cytokines in the control of intracellular pathogens.