Project description:Background: While the luminal microbiome composition in the human cervicovaginal tract has been defined, the presence and impact of tissue-adherent ectocervical microbiota remain incompletely understood. Studies of luminal and tissue-associated bacteria in the gastrointestinal tract suggest that they may have distinct roles in health and disease. Here, we performed a multi-omics characterization of paired luminal and tissue samples collected from a clinically well-characterized cohort of Kenyan women. Results: We identified a tissue-adherent bacterial microbiome, with a higher alpha diversity than the luminal microbiome, in which dominant genera overall included Gardnerella and Lactobacillus, followed by Prevotella, Atopobium, and Sneathia. About half of the L. iners dominated luminal samples had a corresponding Gardnerella dominated tissue microbiome. Broadly, the tissue-adherent microbiome was associated with fewer differentially expressed host genes than the luminal microbiome. Gene set enrichment analysis revealed that L. crispatus-dominated tissue-adherent communities were associated with protein translation and antimicrobial activity, whereas a highly diverse microbiome was associated with epithelial remodeling and pro-inflammatory pathways. Communities dominated by L. iners and Gardnerella were associated with low host transcriptional activity. Tissue-adherent microbiomes dominated by Lactobacillus and Gardnerella correlated with host protein profiles associated with epithelial barrier stability, and with a more pro-inflammatory profile for the Gardnerella-dominated microbiome group. Tissue samples with a highly diverse composition had a protein profile representing cell proliferation and pro-inflammatory activity. Conclusion: We identified ectocervical tissue-adherent bacterial communities in all study participants. These communities were distinct from cervicovaginal luminal microbiota in a significant proportion of individuals. This difference could possibly explain that L. iners dominant luminal communities have a high probability of transitioning to high diverse bacterial communities including high abundance of Gardnerella. By performing integrative multi-omics analyses we further revealed that bacterial communities at both sites correlated with distinct host gene expression and protein levels. The tissue-adherent bacterial community is similar to vaginal biofilms that significantly impact women’s reproductive and sexual health.

Project description:Objective To characterise the vaginal microbiota, mucosal metabolome and host immune response in early pregnancy and investigate its relationship with adverse pregnancy outcomes, including ectopic pregnancy, within the context of pregnancies of unknown location (PUL). DesignProspective cohort study Setting Queen Charlotte’s and Chelsea Hospital, Imperial College Healthcare NHS Trust, London, UK. Population Ninety-one pregnancies of which 22 patients had a favourable outcome of a viable intrauterine pregnancy (VIUP). The remainder had an adverse outcome including 15 with a non-viable intrauterine pregnancy (NVIUP, i.e. miscarriage), 26 a failed PUL (FPUL), 20 an ectopic pregnancy (EP) and 8 a persistent PUL (PPUL). Methods Two matching pairs of vaginal swab samples were collected from women as early as four weeks gestation in pregnancies that resulted in an ectopic pregnancy, miscarriage or viable intrauterine pregnancy pregnancies matched for age, gestation and body mass index. Sequencing of the V1-V2 region of the 16S rRNA gene amplicon was used to characterize and compare vaginal bacterial compositions. The second of the pair of vaginal swabs was used sequentially first for DESI-MS direct-on swab untargeted metabolite profiling followed by extraction of the protein content for quantitative analysis of chemokine and cytokine levels using a 15-plex Luminex immune-profiling assay. Results Adverse final pregnancy outcomes were associated with reduced Lactobacillus spp. abundance (100% vs. 75.4%, p-value=9.83 x 10-3) and higher Shannon α-diversity (p-value=1.10 x 10-3) when compared to viable pregnancies. This association was independent of vaginal bleeding and observed prior to the diagnosis of the final pregnancy outcome (i.e. before the pregnancy was visible on ultrasound). Ectopic pregnancy had an even stronger association with Lactobacillus spp. depletion when compared to viable pregnancies (30% vs 0%, p-value =5.52 x 10-3). Although a strong immune mediator signature, which included MMP-1, IL-6, CCL-2/MCP-1, TNF-α, amongst others was observed in adverse pregnancy outcomes, vaginal bleeding was identified as a major confounder and adjustment of models for bleeding removed the association with outcome. Vaginal bleeding was also found to impact metabolic profiles, increasing the abundances of multiple lipid species. The prediction of outcome based on metabolic profiles was not possible but metabolic profiles were predictive with high accuracy of vaginal microbial composition at the genera level (L. dominant vs. L. deplete), and metabolic correlates of host immune activation were identified, mainly composed of lipids. Conclusions Early pregnancy vaginal microbiome communities dominated by L. crispatus or L. gasseri were observed in women with a PUL who go onto have a viable intrauterine pregnancy. Conversely, a vaginal microbiota deplete in Lactobacillus spp or dominated by L. iners is associated with a diagnosis of ectopic pregnancy in a PUL population. These findings suggest that vaginal microbiota composition is a risk factor for ectopic pregnancy. Vaginal bleeding is an inevitable cofounding factor that must be taken into consideration when performing multi-omic analysis of vaginal mucosal samples in similar clinical populations. Immune and metabolic profiles were particularly impacted by bleeding and bleeding could greatly impact the diagnostic usefulness of immune marker profiling. Further studies are required to clarify the role of microbes and infection in implantation and ectopic pregnancy, as well as determine the mechanistic pathways by which sub-optimal vaginal microbial composition increases risk. Through the integration of metataxonomics, metabolomic and immune profiling data obtained from corresponding samples, our findings demonstrate the robust predictive capacity of specific metabolome signatures. These signatures enable the simultaneous prediction of both the composition of the vaginal microbiome and the inflammatory status of the host, even in the presence of bleeding. The data derived from direct on-swab metabolic profiling using DESI-MS holds promise for swiftly stratifying the risk of early pregnancy loss by rapidly assessing the dynamics between the vaginal microbiota and host. Further validation, however, is essential for future studies to confirm this potential. Linked cross omic data sets:Meta-taxonomics data associated with this study are available in the European Nucleotide Archive (ENA): accession number PRJEB72306.

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Vaginal SynCom

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