PROJECT SUMMARY Chlamydia trachomatis is the leading cause of sexually transmitted infections of bacterial origin worldwide. No vaccines are available, infections are often asymptomatic, and if left untreated, long-term sequelae have an adverse effect on women reproductive health. The primary site of infection is the endocervical epithelium, where the bacteria replicates in the host cell cytosol within a membrane-bound compartment called the inclusion. Interventions to prevent the establishment of the Chlamydia replicative niche would reduce vaginal shedding (i.e. transmission) and ascension to the upper reproductive tract (i.e. long-term pathology). It is more and more appreciated that the state of the tissue at the time of infection most likely dictates the outcome of infection; however, little is known about intrinsic and extrinsic factors that collectively prime individual cells within the tissue. Human epidemiological studies, murine and cell culture models of infection, have been used to gain a better understanding of these processes; however, there are substantial limitations with these models. To address this gap, in close collaboration with the Biomimetic Models Core, we will use a human three-dimensional biomimetic cervicovaginal model of chlamydial infection and investigate the effects of sex hormones on C. trachomatis infection. We will assess bacterial growth (Aim 1), tissue integrity and physiology (Aim 2) and the immune response (Aim 3). Altogether, the exposure of ex vivo reconstituted cervicovaginal tissue in its physiological environment to infectious chlamydiae will offer a unique model system to characterize if and how sex hormones influence susceptibility to C. trachomatis infection. In the long term, this model will be applicable to the study of fundamental processes related to any sexually transmitted pathogen and will offer a relatively simple and cost- effective way for testing novel therapies to control the spread of sexually transmitted infections.