The HIV pandemic disproportionately affects women, with most infections acquired through receptive vaginal sex. Although the target cells by which HIV establishes infection in the female genital tract remain poorly defined, it is known that immune activation results in CD4+ T cells with enhanced susceptibility, as does expression of the mucosal integrin α4β7 and the HIV coreceptor CCR5. Blood and cervical cytobrush specimens were collected from female sex workers (FSWs) in Nairobi, Kenya. Genital infection diagnostics were performed, T cell populations were defined by multiparameter flow cytometry based on their expression of surface receptors relevant to mucosal homing and/or HIV acquisition, and cytokine production was assayed by intracellular cytokine staining. The integrin α4β7 was expressed on 26.0% of cervical CD4+ T cells, and these cells were more likely to express both the HIV coreceptor CCR5 (p < 0.0001) and the early activation marker CD69 (p < 0.0001) but not CXCR4 (p = 0.34). Cervical Th17 frequencies were enhanced compared with blood (7.02 versus 1.24%; p < 0.0001), and cervical IL-17A+ CD4+ T cells preferentially coexpressed α4β7 and CCR5. Expression of IFN-γ and IL-22 was greater in cervical Th17 cells than in blood Th17 cells. In keeping with the hypothesis that these cells are preferential HIV targets, gp120 preferentially bound CCR5+ cervical T cells, and cervical Th17 cells were almost completely depleted in HIV+ FSWs compared with HIV− FSWs. In summary, a subset of Th17 CD4+ T cells in the cervical mucosa coexpresses multiple HIV susceptibility markers; their dramatic depletion after HIV infection suggests that these may serve as key target cells during HIV transmission.

BackgroundFunctional analysis of mononuclear leukocytes in the female genital mucosa is essential for understanding the immunologic effects of HIV vaccines and microbicides at the site of HIV exposure. However, the best female genital tract sampling technique is unclear.Methods and FindingsWe enrolled women from four sites in Africa and the US to compare three genital leukocyte sampling methods: cervicovaginal lavages (CVL), endocervical cytobrushes, and ectocervical biopsies. Absolute yields of mononuclear leukocyte subpopulations were determined by flow cytometric bead-based cell counting. Of the non-invasive sampling types, two combined sequential cytobrushes yielded significantly more viable mononuclear leukocytes than a CVL (p<0.0001). In a subsequent comparison, two cytobrushes yielded as many leukocytes (∼10,000) as one biopsy, with macrophages/monocytes being more prominent in cytobrushes and T lymphocytes in biopsies. Sample yields were consistent between sites. In a subgroup analysis, we observed significant reproducibility between replicate same-day biopsies (r = 0.89, p = 0.0123). Visible red blood cells in cytobrushes increased leukocyte yields more than three-fold (p = 0.0078), but did not change their subpopulation profile, indicating that these leukocytes were still largely derived from the mucosa and not peripheral blood. We also confirmed that many CD4+ T cells in the female genital tract express the α4β7 integrin, an HIV envelope-binding mucosal homing receptor.ConclusionsCVL sampling recovered the lowest number of viable mononuclear leukocytes. Two cervical cytobrushes yielded comparable total numbers of viable leukocytes to one biopsy, but cytobrushes and biopsies were biased toward macrophages and T lymphocytes, respectively. Our study also established the feasibility of obtaining consistent flow cytometric analyses of isolated genital cells from four study sites in the US and Africa. These data represent an important step towards implementing mucosal cell sampling in international clinical trials of HIV prevention.

HIV prevalence and incidence were extremely high in Nairobi MSM SW; a combination of interventions including increasing condom use, pre-exposure prophylaxis and access to effective treatment is urgently needed to decrease HIV transmission in this key population.

With two million new HIV infections annually, ongoing investigations of risk factors for HIV acquisition is critical to guide ongoing HIV prevention efforts. We conducted a prospective cohort analysis of HIV uninfected female sex workers enrolled at an HIV prevention clinic in Nairobi (n = 1640). In the initially HIV uninfected cohort (70 %), we observed 34 HIV infections during 1514 person-years of follow-up, i.e. an annual incidence of 2.2 % (95 % CI 1.6-3.1 %). In multivariable Cox Proportional Hazard analysis, HIV acquisition was associated with a shorter baseline duration of sex work (aHR 0.76, 95 % CI 0.63-0.91), minimum charge/sex act (aHR 2.74, 0.82-9.15, for low vs. intermediate; aHR 5.70, 1.96-16.59, for high vs. intermediate), N. gonorrhoeae infection (aAHR 5.89, 95 % CI 2.03-17.08), sex with casual clients during menses (aHR 6.19, 95 % CI 2.58-14.84), Depo Provera use (aHR 5.12, 95 % CI 1.98-13.22), and estimated number of annual unprotected regular partner contacts (aHR 1.004, 95 % CI 1.001-1.006). Risk profiling based on baseline predictors suggested that substantial heterogeneity in HIV risk is evident, even within a key population. These data highlight several risk factors for HIV acquisition that could help to re-focus HIV prevention messages.

These data support the model of initial CD4 T-cell depletion followed by overall T-cell influx in response to infection and concomitant increases in immune activation, inflammation, and regulatory markers. These data are among the earliest characterization of the cellular milieu in the female genital tract following male-to-female HIV transmission.