Diversity of Vaginal Microbiota Affects Epithelial Barrier Permeability Among African Pregnant Women

Abstract BackgroundWomen with diverse genital anaerobic bacterial communities, are at over 4-fold higher risk of HIV acquisition than women with a Lactobacillus-rich vaginal microbiome. The mechanisms underlying this are poorly understood. We set out to examine how vaginal microbiota diversity affects epithelial integrity in HIV sero-negative pregnant women. We also investigated how HIV infection alters mucosal integrity within the prevalent genital microbiome diversity.MethodsWe assessed epithelial permeability by measuring the concentrations of tight junction proteins, claudin-1 and zonula occludens- (ZO)-1, in cervico-vaginal lavages (CVL) by enzyme-linked immunosorbent assay (ELISA). Cytokines in the vaginal fluids were measured by multiplex magnetic bead assay to establish the inflammatory state. Bacterial cell-free supernatants were used to treat vaginal epithelial cells and tissues, mimicking the in-vivo vaginal milieu. Gene and protein expression levels of tight junctions of vaginal epithelial cells and tissues in response to treatment were quantified by QuantiGene™ Plex Gene Expression Assay and by western blot respectively. The cytokine response of vaginal epithelial cells, VK2 (E6/E7, ATCC® CRL-2616™), to bacterial cell-free supernatants was measured by ELISA method.Results Among women with CT3 cervicotype, HIV sero-negative pregnant women had significantly higher claudin-1 in their CVL than the HIV-infected pregnant women (p=0.0011). IL-8 (p=0.0028), IL-1beta (p<0.0001) and TNF-alpha (p=0.0283) were significantly higher among HIV-negative pregnant women with a non-Lactobacillus dominant vaginal microbiota than those with a Lactobacillus-dominant vaginal microbiota. Bacterial cell-free supernatants from Lactobacillus elicited low levels of pro-inflammatory cytokines IL-1alpha, IL8, IL-6 and IL-1 beta in comparison to media and G. vaginalis. Treatment with G. vaginalis supernatant lowers claudin-1 and claudin-4 expression yet presence of either L. crispatus or L. iners mitigates this effect of G. vaginalis as observed by immuno-staining of treated vaginal cells.Conclusions Pregnant women with a non-Lactobacillus dominant microbiome had a disrupted epithelial barrier and elevated pro-inflammatory cytokines, making them at a higher risk of HIV acquisition than women with a Lactobacillus-dominant microbiome. Targeting vaginal microbiota and/or its effects on the vaginal epithelium and cervico-vaginal milieu can potentially lower rates of HIV acquisition and transmission.