A better understanding of the regulatory role of genital tract T cells is much needed. In this study, we have analyzed the phenotype, distribution, and function of T lymphocytes in the female genital tract of naive, pregnant, or Chlamydia trachomatis-infected C57BL/6 mice. Unexpectedly, we found that the dominant lymphocyte population (70–90%) in the genital tract was that of CD3+ αβTCR int CD4− CD8− T cells. Moreover, these cells were CD90 low but negative for the classical T cell markers CD2 and CD5. The CD3+ B220 low cells were NK1. 1 negative and found in nude mice as well as in mice deficient for MHC class II, β 2-microglobulin, and CD1, indicating extrathymic origin. They dominated the KJ126+ Vβ8. 2+ population in the genital tract of DO11. 10 OVA TCR-transgenic mice, further supporting the idea that the CD3+ B220 low cells are truly T cells. The function of these T cells appeared not to be associated with immune protection, because only CD4+ and CD8+ T cells increased in the genital tract following chlamydial infection. Notwithstanding this, the infected, as well as the uninfected and the pregnant, uterus was dominated by a high level of the CD3+ CD4− CD8− B220 low cells. Following in vitro Ag or polyclonal stimulation of the CD3+ CD4− CD8− B220 low cells, poor proliferative responses were observed. However, these cells strongly impaired splenic T cell proliferation in a cell density-dependent manner. A large fraction of the cells expressed CD25 and produced IFN-γ upon anti-CD3 plus anti-CD28 stimulation, arguing for a strong regulatory role of this novel T cell population in the mouse female genital tract.