Background: Semen induces an immune response at the female genital tract (FGT) to promote conception. It is also the primary vector for HIV transmission to women during condomless sex. Since genital inflammation and immune activation increase HIV susceptibility in women, semen-induced alterations at the FGT may have implications for HIV risk. Here we investigated the impact of semen exposure, as measured by self-reported condom use and Y-chromosome DNA (YcDNA) detection, on biomarkers of female genital inflammation associated with HIV acquisition. Methods: Stored genital specimens were collected biannually (mean 5 visits) from 153 HIV-negative women participating in the CAPRISA 008 tenofovir gel open-label extension trial. YcDNA was detected in cervicovaginal lavage (CVL) pellets by RT-PCR and served as a biomarker of semen exposure within 15 days of genital sampling. Protein concentrations were measured in CVL supernatants by multiplexed ELISA, and the frequency of activated CD4+CCR5+ HIV targets was assessed on cytobrush-derived specimens by flow cytometry. Common sexually transmitted infections (STIs) and bacterial vaginosis (BV)-associated bacteria were measured by PCR. Multivariable linear mixed models were used to assess the relationship between YcDNA detection and biomarkers of inflammation over time. Results: YcDNA was detected at least once in 69% (106/153) of women during the trial (median 2, range 1–5 visits), and was associated with marital status, cohabitation, the frequency of vaginal sex, and Nugent Score. YcDNA detection but not self-reported condom use was associated with elevated concentrations of several cytokines: IL-12p70, IL-10, IFN-γ, IL-13, IP-10, MIG, IL-7, PDGF-BB, SCF, VEGF, β-NGF, and biomarkers of epithelial barrier integrity: MMP-2 and TIMP-4; and with reduced concentrations of IL-18 and MIF. YcDNA detection was not associated with alterations in immune cell frequencies but was related to increased detection of P. bivia (OR = 1.970; CI 1.309–2.965; P = 0.001) at the FGT. Conclusion: YcDNA detection but not self-reported condom use was associated with alterations in cervicovaginal cytokines, BV-associated bacteria, and matrix metalloproteinases, and may have implications for HIV susceptibility in women. This study highlights the discrepancies related to self-reported condom use and the need for routine screening for biomarkers of semen exposure in studies of mucosal immunity to HIV and other STIs.

Introduction Semen induces mucosal changes in the female reproductive tract to improve pregnancy outcomes. Since semen‐induced alterations are likely short‐lived and genital inflammation is linked to HIV acquisition in women, we investigated the contribution of recent semen exposure on biomarkers of genital inflammation in women at high HIV risk and the persistence of these associations. Methods We assessed stored genital specimens from 152 HIV‐negative KwaZulu‐Natal women who participated in the CAPRISA 008 trial between November 2012 and October 2014. During the two‐year study period, 651 vaginal specimens were collected biannually (mean five samples per woman). Cervicovaginal lavage (CVL) was screened for prostate‐specific antigen (PSA) by ELISA, whereas Y‐chromosome DNA (YcDNA) detection and quantification were conducted by RT‐PCR, representing semen exposure within 48 hours (PSA+YcDNA+) and semen exposure within three to fifteen days (PSA−YcDNA+). Soluble protein concentrations were measured in CVLs by multiplexed ELISA. T‐cell frequencies were assessed in cytobrushes by flow‐cytometry, and vulvovaginal swabs were used to detect common vaginal microbes by PCR. Linear mixed models adjusting for factors associated with genital inflammation and HIV risk were used to assess the impact of semen exposure on biomarkers of inflammation over multiple visits. Results Here, 19% (125/651) of CVLs were PSA+YcDNA+, 14% (93/651) were PSA−YcDNA+ and 67% (433/651) were PSA−YcDNA−. Semen exposure was associated with how often women saw their partners, the frequency of vaginal sex in the past month, HSV‐2 antibody detection, current gonorrhoea infection and Nugent Score. Both PSA detection (PSA+YcDNA+) and higher cervicovaginal YcDNA concentrations predicted increases in several cytokines, barrier‐related proteins (MMP‐2, TIMP‐1 and TIMP‐4) and activated CD4+CCR5+HLA‐DR+ T cells (β = 0.050; CI 0.001 to 0.098; p = 0.046) and CD4+HLA‐DR+ T cells (β = 0.177; CI 0.016 to 0.339; p = 0.032) respectively. PSA detection was specifically associated with raised pro‐inflammatory cytokines (including IL‐6, TNF‐α, IP‐10 and RANTES), and with the detection of BVAB2 (OR = 1.755; CI 1.116 to 2.760; p = 0.015), P. bivia (OR = 1.886; CI 1.102 to 3.228; p = 0.021) and Gardnerella vaginalis (OR = 1.815; CI 1.093 to 3.015; p = 0.021). Conclusions More recent semen exposure was associated with raised levels of inflammatory biomarkers and the detection of BV‐associated microbes, which declined by three to fifteen days of post‐exposure. Although transient, semen‐induced alterations may have implications for HIV susceptibility in women.

Genital inflammation (GI) undermines topical HIV pre-exposure prophylaxis (PrEP) efficacy through unknown mechanisms. Here, associations between activated endocervical CD4+ T cell numbers and higher deoxyadenosine triphosphate (dATP) concentrations suggest that competition for intracellular metabolites within HIV target cells may reduce the efficacy of antiretroviral-based PrEP in women with GI.