PROJECT SUMMARY Chlamydia trachomatis (Ct) is the pathogen that causes the world’s most common bacterial sexually transmitted infection, with over 129 million cases in 2020. Untreated urogenital Ct can have severe sequelae, including pelvic inflammatory disease, ectopic pregnancy, tubal factor infertility, epididymitis, urethritis, and prostatitis. Untreated anorectal Ct can lead to proctitis, ulcerations, abscesses, fissures, and fistulas. Anorectal infections likely also act as a reservoir for the bacterium, in which urogenital infection occurs via autoinoculation. Most Ct research has focused on female urogenital infections, as these infections more commonly cause severe sequelae. However, there is a large gap and an urgent need to study Ct infections in males. Although male urogenital tract infections are less likely to lead to severe sequelae, males can transmit infection to the female reproductive tract, which is more vulnerable to complications. Additionally, anorectal infections occur in men who have sex with men, men who have sex with women and in women. Current treatment is curative antibiotics, but these do not prevent reinfection, do not robustly clear infection at all anatomic sites, and can have poor patient compliance. Identifying strategies to reduce and prevent transmission of Ct, as well as limit infection, is very urgent. Our lab has developed vaccines against Chlamydia using highly immunogenic bacteriophage virus- like particles (VLPs) as a platform displaying short peptide epitopes of Ct adhesion factors in an immunogenic manner. In particular, our Qb-VD4-MOMP vaccine, targeting Ct adhesion factor Major Outer Membrane Protein, provides protection in female mice. However, the anorectal tract and male urogenital tract have distinct immunological environments, and Ct infections at these sites are not well studied. This F31 proposal aims to investigate the ability of our Qb-VD4-MOMP vaccine to protect against Chlamydia infection in male mice. The overall goal of this proposed research is to determine the immunogenicity and protective ability of these vaccines in male mice, utilizing urogenital, anorectal, and sexual transmission infection models. The outcome of this research will highlight the populations that will benefit from vaccination by preventing both infection and transmission. Aim 1 will investigate vaccine efficacy in male mice urogenitally or anorectally infected with Chlamydia. Additionally, vaccine efficacy in the context of immunization of either or both partners will be investigated in mouse sexual transmission models of Chlamydia (Aim 2). These vaccines may or may not elicit the same protection in males as in females, and the use of adjuvants, mixed vaccines, or alternative vaccine targets may be necessary to provide protection in males. Overall, this research will reveal important aspects of vaccine-mediated protection and advance these vaccines closer to human clinical trials.