Mycobacterium (M.) vaccae is a fast-growing species of saprophytic bacteria that is widely distributed. To understand the host immune responses induced by M. vaccae isolated from bovine submaxillary lymph nodes, C57BL/6 mice were infected with reference strain M. vaccae Bacillus Calmette-Guérin (BCG) and isolated M. vaccae using intraperitoneal injections. Comparison of the bacterial replication and organ pathology between M. vaccae and M. vaccae BCG revealed that M. vaccae was more malignant than M. vaccae in mice. We also demonstrated that serum from the M. vaccae-infected mice contained a higher expression level of gamma-interferon (IFN-γ), tumor necrosis factor alpha, monocyte chemoattractant protein-1, interleukin (IL)-4, IL-12, IL-10 and transforming growth factor beta than did the other groups, especially after week 4. Furthermore, when the numbers of CD3âºCD4âºIFN-γ⺠and CD3âºCD4âºIL4⺠cells in the infected mice were observed by flow cytometry, we found that a powerful T helper 1 (Th1) response was induced by M. vaccae infection, which was associated with the emergence of CD3âºCD4âºIFN-γ⺠cells. However, the Th1 response declined over time, which was associated with appearance of the CD4âºCD25âºFoxP3⺠and CD4âºCD25âºCD152âºTreg cell reaction. In addition, a strong Th2 response was found. Finally, we found that M. vaccae infection increased the production of type I IFNs, which was associated with a reduced Th1 response.