Project Summary/Abstract The overarching goal of this proposal is to advance the understanding of the immune response to and protective effects of rectal Chlamydia trachomatis (CT) infections. CT is the most common bacterial sexually transmitted disease in the world and is associated with substantial reproductive tract morbidity. Rectal CT is increasingly recognized as a common infection among STI clinic patients, yet despite its high prevalence, there is no clear data describing how rectal infection affects the immune response to CT or the risk of reinfection. A better understanding of how the immune system responds to CT infection is urgently needed to inform effective chlamydia vaccine strategies. Animal studies in mice and nonhuman primates (NHP) have demonstrated rectal CT infections as non- pathogenic and persistent, similar to humans infected with non-lymphogranuloma venereum (LGV) serovars in the rectum. Furthermore, studies in the murine model have shown that rectal infection confers trans-mucosal protection against urogenital challenge, suggesting that rectal CT infection causes enhanced systemic and transmucosal immune responses to CT. It is possible the gastrointestinal tract (GIT) will be an optimal site for attenuated mucosal CT vaccines. However, more data on immune responses and potentially protective effects of rectal infection in NHPs and humans are urgently needed and will be provided in the proposed studies. Mouse and human studies provide evidence that secretion of IFN-γ by CD4+ T cells is essential for any protective immunity. The role for antibodies is less clear but appears to rely not on direct neutralization of bacteria, but rather on secondary functions of the Fc region of antibodies in activating innate and effector cells. Except for a few mouse model studies, the Fc-dependent functions of anti-CT antibodies remain largely unexplored. This project will provide informative data describing functional CT protective immunity in NHP (controlled exposures) and human (cross-sectional sampling) studies. We will conduct complementary studies in the pig-tailed macaque model and in clinical studies to address the hypothesis that rectal CT infection induces robust anti-CT immune responses that may protect from genital disease. We will directly test whether rectal infection in macaques protects against subsequent genital infection. In both macaques and humans, we will define anti-CT immune responses that may correlate with protection from reinfection and disease: secretion of IFN-γ by CD4+ T cells, presence of anti-CT antibodies in the mucosa, and functional antibody responses, using new methods to test for Fc-receptor-mediated functions of anti-CT antibodies. This project is the first step to demonstrate that rectal infection modifies anti-CT immunity in NHPs and humans, and will provide important evidence as to whether the GIT could serve as a mucosal delivery site for future vaccination strategies.