Background: Women with elevated vaginal mucosal inflammation are at an increased risk of acquiring HIV. While host genetic variation influences protein levels in circulation, its effects on the cervicovaginal proteome and HIV susceptibility have not been previously characterized.

Methods: We conducted the first cervicovaginal protein quantitative trait loci (pQTL) study to assess genetic regulation of protein expression using samples from 589 South African women enrolled in the CAPRISA004 tenofovir gel trial. Proteomic data of cervicovaginal lavage samples, genome-wide genotyping data, vaginal microbiome profiles, and epidemiological data were previously generated from trial participants.

Results: The rs143864957 (AG/-) deletion allele was associated with reduced genital A2ML1 levels (β = -0.53; 95% confidence interval (CI): -0.71- -0.35; p = 1.5x10-8) and 2.1 times higher odds of HIV acquisition (95% CI: 1.1-4.4; p = 0.036). This genetic effect on A2ML1 displayed heterogeneity across vaginal microbiome community state types (CSTs) (Q = 8.1; p = 0.043; I² = 61%), with greater A2ML1 reduction observed in inflammatory CSTs with higher bacterial loads. Higher A2ML1 levels were associated with reduced genital inflammation across all participants (incident rate ratio (IRR) = 0.79; 95% CI: 0.72-0.90; p = 7.1x10-7). However, this A2ML1-inflammation relationship interacted with bacterial abundance, A2ML1's anti-inflammatory efficiency decreased above a bacterial load threshold of 110.1 units (95% CI: 96.9-120.0). This threshold corresponded closely with transitions in vaginal CST structure.

Conclusion: We demonstrate that vaginal microbiome composition amplifies host genetic effects on mucosal immunity. The impact of rs143864957 on A2ML1, an antiprotease previously linked to HIV susceptibility, is potentiated in inflammatory, high-bacterial-load mucosal environments where reduced A2ML1 fails to control inflammation, compromising mucosal barrier integrity. This gene-microbiome interaction contributes to understanding individual variation in HIV acquisition risk at the genital mucosa.

Introduction
Lipid metabolites form the structural foundation of the cervicovaginal epithelium, regulate immune response, and fuel host cells and the microbial community. To date, the impact of hormonal contraceptives (HC) on vaginal lipidome has not been studied. Previously, injectable HC, depot medroxyprogesterone acetate (DMPA), was shown to be associated with increased microbial diversity and inflammation, potentially increasing susceptibility to HIV-1. Here, we investigated the effect of injectable and oral HCs on vaginal lipid metabolism in healthy Kenyan sex workers.
Methods
Forty-five sex workers were recruited out of a total of 370 screened from the Sex Workers Outreach Programme (SWOP) Program in Nairobi, Kenya, based on the inclusion criteria for this study. Cervicovaginal lavage (CVL) samples were collected from sex workers who were using DMPA, oral contraceptive (OCP), or no hormone (NH). High throughput liquid chromatography coupled with mass spectrometry (LC-MS)-based lipidomics and bioinformatics approaches were used to analyze vaginal lipid species.
Results
Using untargeted lipidomics we found that DMPA significantly downregulated major lipid classes such as phospholipids, lysophospholipids, sphingolipids (ceramide, sphingomyelin, & glycosphingolipids), and fatty acids (p<0.001, FDR<0.05). CVL from DMPA group also showed significant elevation in triglycerides (TG) and steroid lipids when compared with OCP or NH groups. In contrast, OCP consistently lowered vaginal triglycerides and increased the fatty acid and diacylglycerol substrates compared to NH group. Interestingly, such DMPA associated signatures of vaginal lipidomic alterations were greatly contributed from microbes, revealing the microbiome-lipidome interplay.
Conclusion
DMPA significantly downregulated structural and signaling lipids and upregulated storage lipids in the vaginal microenvironment. Such dysregulation in lipid metabolism leading to low ceramide, sphingomyelin, phospholipids, and elevated TG has been implicated in bacterial imbalances, poor epithelial membrane integrity, and inflammation. These findings underscore the need to address cervicovaginal lipidomic dysregulation associated with HCs while designing interventions to improve women’s reproductive health.

Background: The targets of HIV infection are CD4+ T helper (Th) cells, which exist in distinct subsets, including Th1, Th2, Th17, and Th1/Th17, each with unique functions. These Th subsets vary in their susceptibility to HIV. Depot medroxyprogesterone acetate (DMPA), an injectable contraceptive, has been associated with increased HIV acquisition, though underlying mechanisms remain unclear. This study examines how DMPA and the menstrual cycle affect Th subsets and their potential effects on HIV susceptibility.
Methods: Peripheral blood was collected from female sex workers (FSW) in Nairobi, Kenya. The case group comprised DMPA users (n=52), while controls were non-hormonal contraceptive users sampled during the follicular (n=43) or luteal (n=25) phases of the menstrual cycle, with some participants contributed sample from both phases. Cells were identified by flow cytometry using chemokine receptor profiles: Th1 (CCR6⁻CXCR3⁺), Th2 (CCR6⁻CXCR3⁻CCR4⁺), Th17 (CXCR3⁻CCR6⁺CD161⁺), and Th1/17 (CCR6⁺CXCR3⁺), along with activation markers (CD38, HLA-DR, CD69). Cytokine production was also assessed following PMA/ionomycin stimulation.
Results: CD4+ T cell activation was significantly higher in the follicular compared to the luteal phase, indicated by increased % of CD4⁺HLA-DR⁺CD38⁻ (p=0.014) cells and mean fluorescent intensity (MFI) CD4+ CD38+ cells (p=0.041). Activated Th1 (%HLA-DR⁻CD38⁺, p=0.036), Th2 (%HLA-DR⁺CD38⁻, p=0.002), and Th17 (HLA-DR+ MFI, p=0.048) subsets were significantly higher in the follicular phase. Similarly, CD4+ T cells during the follicular phase showed higher levels of IFN-γ (p=0.097) and IL-4 (p=0.006) following stimulation compared to the luteal phase (p<0.05). Interestingly, CCR5 MFI was elevated on CD4⁺HLA-DR⁺CD38⁻ (p=0.033) cells and Th2⁺HLA-DR⁺CD38⁻ (p=0.004) cells among DMPA users compared to follicular and on Th1/Th17⁺HLA-DR-CD38+ cells (p=0.07) relative to luteal.
Conclusion: Differential Th cell activation was detected across menstrual cycle phases, with the follicular phase exhibiting higher levels of HIV target cells. DMPA users also showed higher CCR5 levels on activated T cells, which may contribute to increased HIV risk.

Introduction:In Kenya, sexually transmitted infection (STI) management is symptom-based. Consequently, the prevalence of chlamydia and gonorrhea is unclear, and circulating strain genetic diversity and the presence of antimicrobial resistance (AMR) are not characterized.

Methods:This project will characterize chlamydia and gonorrhea prevalence, incidence and molecular characteristics among female sex workers in Kenya. Participants were recruited in Nairobi, with samples collected bi-monthly over 12 months and a subset (n=380) were selected for STI testing. Urine was tested by rRNA amplification using a commercial assay. Chlamydia-positive samples were genotyped by Sanger sequencing of the ompA gene and gonorrhea-positive samples were typed by Neisseria gonorrhoeae Multi-Antigen Sequence Typing (NG-MAST), targeting porB and tbpB genes, and tested by RT-PCR to predict AMR.

Results:STI testing was performed on screening, enrollment, month-4, -8 and -12 visit samples. Chlamydia prevalence was 9.9%(95%CI 8.5-11.5), peaking during month-4[10.9%(95%CI 8.1-14.4)], and gonorrhea prevalence was 2.8%(95%CI 2.1-3.7), peaking during enrollment[3.7%(95%CI 2.2-6.1)]. Annual incidence was 61.1(95%CI 36.4-85.9) chlamydia infections per 1000 person-years and 19.4(95%CI 5.2-33.7) gonorrhea infections per 1000 person-years. Notably, incidence rates peaked between enrollment and month-4, corresponding with condom shortages. Chlamydia typing revealed types F[34.0% (95%CI 26.5-41.5)] and E[21.6% (95%CI 15.1-28.1)] as the most prevalent. No LGV-types were detected despite claims it is endemic across Africa. Gonorrhea characterization found 81.8%(95%CI 68.9-95.0) of infections had novel PubMLST types, while RT-PCR predicted 97.4%(95%CI 92.3-100.0) of infections were ciprofloxacin resistant, 5.6%(95%CI 0.0-13.0) were azithromycin resistant and 8.1%(95%CI 0.0-16.9) had decreased susceptibility to cephalosporins.

Conclusion:Chlamydia and gonorrhea prevalence in this population exceeds WHO estimates for women in the African region. The observed STI incidence increase during the condom shortage underscores the consequences of STI prevention funding cuts. Diverse assortment of chlamydia and gonorrhea types is evident. Further identification of types and AMR will impact Kenyan STI prevention and treatment policies, contributing to global STI control.

Background: Female sex workers (FSWs) experience an elevated risk of acquiring HIV. As women age, they transition into menopause. Sex work and menopause are both associated with increased inflammation. Yet how menopausal status modifies the immune response remains poorly understood.
Methods: Peripheral blood mononuclear cells and cervical mucosal cells were collected from 37 FSWs from the Sex Worker Outreach Program (SWOP) in Nairobi, Kenya. Blood and mucosal cytokine and T cell profiles were analyzed. A general-linear model controlling for duration of sex work was used to determine differences between women living with (WLWH) and without HIV and by menopausal status.
Results: In FSWs not living with HIV, menopause increased systemic and mucosal inflammation, including higher plasma levels of IL-6 (p=0.044), TNF-α (p=0.01), MCP-1 (p=0.012), and IL-1RA (p=0.010) and increased cervicovaginal TNF-α (p=0.017) and IL-1β (p=0.017) compared to premenopausal women. At the cellular level, they displayed a lower proportion of regulatory T cells (p=0.026) and higher levels of activated proinflammatory Th17 CCR5+ cells (p=0.011) in the blood, and a higher mucosal proportion of HIV target cells, CD4+CD69+CCR5+ (p<0.001), compared to premenopausal FSWs. Premenopausal FSWs LWHIV had higher blood levels of activated T cells (CD4+CD69+, p=0.042) compared to postmenopausal FSWs LWHIV. In premenopausal FSWs LWHIV, we observed higher IFN-γ (p=0.026) and IL-8 (p=0.004) in the blood and mucosal IFN-γ (p=0.001), IL-15 (p=0.026), IL-17A (p=0.032), and TNF-α (p=0.001), while postmenopausal FSWs LWHIV exhibited elevated plasma IL-28A (p=0.007).
Conclusion: We observed that menopause affects the inflammatory profile in FSWs regardless of HIV status. However, in FSWs not living with HIV, the changes in T cell activation are more profound, which could affect susceptibility to HIV infection. More studies are needed to fully understand how menopause affects women and to provide better policies for aging women and HIV risk.

Introduction: People living with HIV (PLWH) are exposed to higher incidence and risk of comorbidities compared to the general population, including neurocognitive dysfunction. Notably, approximately 50% of anti-retroviral-treated individuals have been reported to experience HIV-associated neurocognitive disorders (HAND), with a cumulative burden on health services and overall decrease in quality of life. This suggests the need for sustained refinement of existing models and/or the development of newer models to better our understanding of HAND.
Methods: We used the humanized bone marrow liver thymus (huBLT) mouse model, with NSG and NSG-SGM3 mice, to investigate the subclinical and clinical features of HAND. HuBLT mice, which reconstitute with a human immune system at biologically relevant compartments, were infected intraperitoneally with HIV-1. Cellular and viral neuroinvasion were measured by flow cytometry, immunohistochemistry and immunofluorescence. Markers of inflammation, blood-brain barrier permeability, and neuronal injury were assessed through qPCR, immunohistochemistry and immunofluorescence, while overt neurological changes were evaluated through behavioral testing.
Results: Monocyte-macrophages colocalized with HIV-1 p24 during human immune cell and virus neuroinvasion in huBLT mice. Infected mice displayed distinct profiles of interferon-responsive genes and significantly decreased expression of occludin and claudin-5 in the brain, marking blood-brain barrier permeability. Similarly, infected mice had significantly poorer performance in the accelerating rotarod test, indicating impaired motor coordination and balance, characteristic symptoms of HAND. Infection also associated with hyperactive behaviors in the open field test, reminiscent of mania-like behaviors observed in some PLWH. Finally, preliminary immunofluorescence indicated pathologic features of RNA metabolism and reduced neuronal cell health during infection.
Conclusions: While mouse models have previously been used to evaluate HAND, none have characterized the clinical manifestations in huBLT mice yet. Our results demonstrate the huBLT mouse model as a suitable tool for the characterization of HIV-1-related motor and mood manifestations in the context of neuroinflammation and neuronal injury.