Submitted to: Infection and Immunity
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 3/29/2026
Publication Date: N/A
Citation: N/A

Interpretive Summary: Chlamydia trachomatis, the bacterium that causes chlamydia, can lead to infertility in women if it spreads upwards from the cervix into the uterus and Fallopian tubes. Infected women have very different amounts of Chlamydia in the cervix, and higher amounts are linked to a higher risk of the infection spreading upwards. The factors that influence how much Chlamydia grows and how well the body controls it, are not fully understood. However, studies show that bacterial vaginosis (BV), an imbalance of vaginal bacteria and the most common vaginal infection, can make it harder for the body to fight off Chlamydia. BV significantly changes both the local immune response and nutrients available in the genital tract, which we hypothesized would influence Chlamydia burden. To investigate this, we used advanced multi-omics methods to analyze the type of vaginal bacteria (the microbiome), levels of tryptophan and its breakdown products (important nutrients used by both Chlamydia and BV-associated bacteria), immune signals (cytokines), in addition to Chlamydia load. When we grouped women based on their tryptophan-related metabolites, we identified three distinct patterns called metabolome state types (MSTs). MST I, seen mostly in women with healthy, Lactobacillus-dominant vaginal bacteria, had higher tryptophan and normal tryptophan metabolism. MST II and MST III, which were both linked to BV-associated bacterial communities, showed lower tryptophan levels. MST II showed broadly reduced levels of most metabolites, while MST III had higher levels of certain tryptophan-derived metabolites made by microbes. We also found that certain bacteria, nutrient patterns, and immune signals tend to appear together. Using a type of network analysis, we identified distinct combinations of bacteria, metabolites, and immune signals that could predict whether a woman had a high or low Chlamydia load. Important predictors included specific immune molecules, such as CXCL9, CXCL10, and IL-17, BV-associated bacteria, and certain indole-related tryptophan metabolites. Overall, our results show that cervical Chlamydia burden depends on the combined state of the vaginal bacteria, nutrient availability, and immune responses, not just a single factor. This helps explain how BV may influence the risk of Chlamydia growing and spreading in the reproductive tract.

Technical Abstract: Chlamydia trachomatis (Ct) remains a major public health concern and causal agent of upper reproductive tract pathology. There is a broad spectrum of cervical Ct loads in infected women, and higher loads are associated with upper tract ascension. Factors that modulate host-Ct interactions and their outcomes are poorly defined, but emerging studies indicate that bacterial vaginosis (BV), the most common vaginal dysbiosis, significantly modulates the genital immune and nutritional milieu. We performed an integrated multi-omics analysis of the cervico-vaginal microbiome, tryptophan metabolome, and cervical cytokines to identify ecological features associated with BV status and variation in Ct load. Cervico-vaginal samples were analyzed by 16S rRNA gene sequencing, targeted UPLC-MS/MS quantification of tryptophan metabolites, and multiplex cytokine profiling. Ordination analyses showed that microbiome, tryptophan metabolome, and cytokine profiles each separated strongly by BV status, whereas Ct load separated only by cytokine profiles. K-means clustering of tryptophan metabolites defined three metabolome state types (MSTs). MST I, associated primarily with Lactobacillus crispatus-dominated community state type (CST) I, exhibited high tryptophan availability, abundant indole-3-lactic acid, and complete kynurenine-pathway activity. Both MST II and MST III associated with BV-associated CST IV and showed marked tryptophan depletion. MST II was broadly depleted of most tryptophan metabolites, while MST III was enriched in late-stage, microbially derived indole metabolites and kynurenic acid. Hierarchical all-against-all association testing revealed coordinated relationships linking clusters of bacterial taxa, tryptophan metabolites, and cytokines. Multi-omic network analyses using MintTea identified integrated microbial-metabolic-immune feature modules that predicted high verses low Ct load, highlighting CXCL9, CXCL10, IL-17, BV-associated taxa, and indole metabolites as key discriminative features. These results demonstrate that cervical Ct load reflects integrated microbial-metabolic-immune ecological states rather than any single factor, defining multi-omic pathways through which BV may influence Ct infection dynamics.