Mary K. Grabowski

The Rates of HIV Superinfection and Primary HIV Incidence in a General Population in Rakai, Uganda

BACKGROUND Human immunodeficiency virus (HIV) superinfection has been documented in high-risk individuals; however, the rate of superinfection among HIV-infected individuals within a general population remains unknown. METHODS A novel next-generation ultra-deep sequencing technique was utilized to determine the rate of HIV superinfection in a heterosexual population by examining two regions of the viral genome in longitudinal samples from recent HIV seroconverters (n=149) in Rakai District, Uganda. RESULTS The rate of superinfection was 1.44 per 100 person years (PYs) (95% confidence interval [CI], .4-2.5) and consisted of both inter- and intrasubtype superinfections. This was compared to primary HIV incidence in 20 220 initially HIV-negative individuals in the general population in Rakai (1.15 per 100 PYs; 95% CI, 1.1-1.2; P= .26). Propensity score matching (PS) was used to control for differences in sociodemographic and behavioral characteristics between the HIV-positive individuals at risk for superinfection and the HIV-negative population at baseline and follow-up. After PS matching, the estimated rate of primary incidence was 3.28 per 100 PYs (95% CI, 2.0-5.3; P = .07) controlling for baseline differences and 2.51 per 100 PYs (95% CI, 1.5-4.3; P = .24) controlling for follow-up differences. CONCLUSIONS This suggests that the rate of HIV superinfection in a general population is substantial, which could have a significant impact on future public health and HIV vaccine strategies.

Previously Transmitted HIV-1 Strains Are Preferentially Selected During Subsequent Sexual Transmissions

BACKGROUND A genetic bottleneck is known to exist for human immunodeficiency virus (HIV) at the point of sexual transmission. However, the nature of this bottleneck and its effect on viral diversity over time is unclear. METHODS Interhost and intrahost HIV diversity was analyzed in a stable population in Rakai, Uganda, from 1994 to 2002. HIV-1 envelope sequences from both individuals in initially HIV-discordant relationships in which transmission occurred later were examined using Sanger sequencing of bulk polymerase chain reaction (PCR) products (for 22 couples), clonal analysis (for 3), and next-generation deep sequencing (for 9). RESULTS Intrahost viral diversity was significantly higher than changes in interhost diversity (P < .01). The majority of HIV-1-discordant couples examined via bulk PCR (16 of 22 couples), clonal analysis (3 of 3), and next-generation deep sequencing (6 of 9) demonstrated that the viral populations present in the newly infected recipient were more closely related to the donor partners HIV-1 variants found earlier during infection as compared to those circulating near the estimated time of transmission (P = .03). CONCLUSIONS These findings suggest that sexual transmission constrains viral diversity at the population level, partially because of the preferential transmission of ancestral as opposed to contemporary strains circulating in the transmitting partner. Future successful vaccine strategies may need to target these transmitted ancestral strains.

Molecular tools for studying HIV transmission in sexual networks

Purpose of reviewPhylogenetics is frequently used for studies of population-based HIV transmission. The purpose of this review is to highlight the current utilities and limitations of phylogenetics in HIV epidemiological research from sample collection through to data analysis. Recent findingsStudies of HIV phylogenies can provide critical information about HIV epidemics that are otherwise difficult to obtain through traditional study design such as transmission of drug-resistant virus, mixing between demographic groups, and rapidity of viral spread within populations. However, recent results from empirical and theoretical studies of HIV phylogenies challenge some of the key assumptions and interpretations from phylogenetic studies. Recent findings include lack of transmission bottlenecks in MSM and injection drug use epidemics, evidence for preferential transmission of HIV virus in heterosexual epidemics, and limited evidence that tree topologies correlate directly with underlying network structures. Other challenges include a lack of a standardized definition for a phylogenetic transmission cluster and biased or sparse sampling of HIV transmission networks. SummaryPhylogenetics is an important tool for HIV research, and offers opportunities to understand critical aspects of the HIV epidemic. Like all epidemiological research, the methods used and interpretation of results from phylogenetic studies should be made cautiously with careful consideration.

Heterogeneity of the HIV epidemic in agrarian, trading, and fishing communities in Rakai, Uganda: an observational epidemiological study

BACKGROUND Understanding the extent to which HIV burden differs across communities and the drivers of local disparities is crucial for an effective and targeted HIV response. We assessed community-level variations in HIV prevalence, risk factors, and treatment and prevention service uptake in Rakai, Uganda. METHODS The Rakai Community Cohort Study (RCCS) is an open, population-based cohort of people aged 15-49 years in 40 communities. Participants are HIV tested and interviewed to obtain sociodemographic, behavioural, and health information. RCCS data from Aug 10, 2011, to May 30, 2013, were used to classify communities as agrarian (n=27), trading (n=9), or lakeside fishing sites (n=4). We mapped HIV prevalence with Bayesian methods, and characterised variability across and within community classifications. We also assessed differences in HIV risk factors and uptake of antiretroviral therapy and male circumcision between community types. FINDINGS 17 119 individuals were included, 9215 (54%) of whom were female. 9931 participants resided in agrarian, 3318 in trading, and 3870 in fishing communities. Median HIV prevalence was higher in fishing communities (42%, range 38-43) than in trading (17%, 11-21) and agrarian communities (14%, 9-26). Antiretroviral therapy use was significantly lower in both men and women in fishing communities than in trading (age-adjusted prevalence risk ratio in men 0·64, 95% CI 0·44-0·97; women 0·53, 0·42-0·66) and agrarian communities (men 0·55, 0·42-0·72; women 0·65, 0·54-0·79), as was circumcision coverage among men (vs trading 0·48, 0·42-0·55; vs agrarian 0·64, 0·56-0·72). Self-reported risk behaviours were significantly higher in men than in women and in fishing communities than in other community types. INTERPRETATION Substantial heterogeneity in HIV prevalence, risk factors, and service uptake in Rakai, Uganda, emphasises the need for local surveillance and the design of targeted HIV responses. High HIV burden, risk behaviours, and low use of combination HIV prevention in fishing communities make these populations a priority for intervention. FUNDING National Institute of Mental Health, the National Institute of Allergy and Infectious Diseases, the National Institute of Child Health and Development, and the National Institute for Allergy and Infectious Diseases Division of Intramural Research, National Institutes of Health; the Bill & Melinda Gates Foundation; and the Johns Hopkins University Center for AIDS Research.Summary Background Understanding the extent to which HIV burden differs across communities and the drivers of local disparities is critical for an effective and targeted HIV response. We assessed community-level variations in HIV prevalence, risk factors, and treatment and prevention service uptake in Rakai, Uganda. Methods The Rakai Community Cohort Study (RCCS) is an open, population-based cohort surveying persons aged 15–49 in 40 communities. Participants are HIV tested and interviewed to obtain sociodemographic, behavioral, and health information. RCCS data from August 2011 to May 2013 were used to classify communities as agrarian (n=27), trading (n=9), or lakeside fishing sites (n=4). HIV prevalence was mapped using Bayesian methods, and variability across and within community classifications was characterized. Differences in HIV risk factors and uptake of antiretroviral therapy and male circumcision between community types were assessed. Findings 17,119 individuals were included; 9215 (54%) were female. 9931 participants resided in agrarian, 3318 in trading, and 3870 in fishing communities. There was large variation in HIV prevalence, ranging from 9% to 43% across communities. Fishing communities had a higher median HIV prevalence (41%, range: 37–43%) compared to trading (17%, range: 11–22%) and agrarian communities (14%, range: 9–26%); ART and male circumcision coverage were significantly lower in fishing communities. Self-reported risk behaviors were significantly higher in men compared to women and in fishing communities compared to other community types. Interpretation There is substantial heterogeneity in HIV prevalence, risk factors, and service uptake across communities within one region of Uganda. These findings underscore the need for local surveillance and have important implications for the design of targeted HIV responses. In particular, the extremely high HIV burden and risk behaviors, and low use of combination HIV prevention in fishing communities make these areas a priority for intervention.

Reactivation of herpes simplex virus type 2 after initiation of antiretroviral therapy.

BACKGROUND The association between initiation of antiretroviral therapy (ART) for human immunodeficiency virus (HIV) infection and possible herpes simplex virus type 2 (HSV-2) shedding and genital ulcer disease (GUD) has not been evaluated. METHODS GUD and vaginal HSV-2 shedding were evaluated among women coinfected with HIV and HSV-2 (n = 440 for GUD and n = 96 for HSV-2 shedding) who began ART while enrolled in a placebo-controlled trial of HSV-2 suppression with acyclovir in Rakai, Uganda. Monthly vaginal swabs were tested for HSV-2 shedding, using a real-time quantitative polymerase chain reaction assay. Prevalence risk ratios (PRRs) of GUD were estimated using log binomial regression. Random effects logistic regression was used to estimate odds ratios (ORs) of HSV-2 shedding. RESULTS Compared with pre-ART values, GUD prevalence increased significantly within the first 3 months after ART initiation (adjusted PRR, 1.94; 95% confidence interval [CI], 1.04-3.62) and returned to baseline after 6 months of ART (adjusted PRR, 0.80; 95% CI, .35-1.80). Detection of HSV-2 shedding was highest in the first 3 months after ART initiation (adjusted OR, 2.58; 95% CI, 1.48-4.49). HSV-2 shedding was significantly less common among women receiving acyclovir (adjusted OR, 0.13; 95% CI, .04-.41). CONCLUSIONS The prevalence of HSV-2 shedding and GUD increased significantly after ART initiation, possibly because of immune reconstitution inflammatory syndrome. Acyclovir significantly reduced both GUD and HSV-2 shedding and should be considered to mitigate these effects following ART initiation.

Male circumcision decreases high‐risk human papillomavirus viral load in female partners: A randomized trial in Rakai, Uganda

Male circumcision (MC) reduces high‐risk human papillomavirus (HR‐HPV) infection in female partners. We evaluated the intensity of HR‐HPV viral DNA load in HIV‐negative, HR‐HPV‐positive female partners of circumcised and uncircumcised men. HIV‐negative men and their female partners were enrolled in randomized trials of MC in Rakai, Uganda. Vaginal swabs were tested for HR‐HPV genotypes by Roche HPV Linear Array which provides a semi‐quantitative measure of HPV DNA by the intensity of genotype‐specific bands (graded:1–4). We assessed the effects of MC on female HR‐HPV DNA load by comparing high intensity linear array bands (3–4) to low intensity bands (1–2) using an intention‐to‐treat analysis. Prevalence risk ratios (PRR) of high intensity bands in partners of intervention versus control arm men were estimated using log‐binomial regression with robust variance. The trial included 335 women with male partners in the intervention arm and 340 in the control arm. At enrollment, the frequency of HR‐HPV high intensity linear array bands was similar in both study arms. At 24 months follow‐up, the prevalence of high intensity bands among women with detectable HR‐HPV was significantly lower in partners of intervention arm (42.7%) than control arm men (55.1%, PRR = 0.78, 95%CI 0.65–0.94, p = 0.02), primarily among incident HR‐HPV infections (PRR = 0.66, 95% CI 0.50–0.87, p = 0.003), but not persistent infections (PRR = 1.02, 95% CI 0.83–1.24). Genotypes with high HR‐HPV band intensity were more likely to persist (adjHR = 1.27 95% CI 1.07–1.50), irrespective of male partner circumcision status. MC reduces HR‐HPV DNA load in newly infected female partners.

Investigation of pre-diagnostic virological markers for progressive multifocal leukoencephalopathy in human immunodeficiency virus-infected patients

Progressive multifocal leukoencephalopathy (PML) is a severe neurological disorder due to JC virus (JCV) infection. Pre‐diagnostic biological markers and risk factors for PML are not well understood. We conducted a case–control study nested within the Multicenter AIDS Cohort Study to examine the association between JCV viruria and viremia and serum antibody to JCV capsids, in relation to subsequent PML diagnoses, 5 months to 12 years later. Other demographic and immunologic factors were also examined. The study population included 28 incident cases of PML, 26 matched HIV‐positive controls, and 50 HIV‐negative controls. Prevalence of JCV viruria was 37% in cases, 42% in HIV‐positive controls, and 28% in HIV‐negative controls (P = 0.43). Among persons with JCV viruria, persistent viruria was more common in cases (89%) than in HIV‐positive controls (33%) (P = 0.02). Presence of JCV viruria was not related to the time to PML diagnosis (OR: 1.03, 95% CI: 0.8–1.4); however, the urinary concentration of JCV DNA increased with proximity to the date of PML diagnosis in cases. JCV seropositivity did not differ between cases or controls (P = 0.42). Four cases tested JCV seronegative, including one case only 5 months prior to diagnosis with PML. JCV DNA was detected in the serum of one HIV‐positive control. Smoking was the only demographic variable analyzed associated with an increased risk for PML (MOR: 9.0, 95% CI: 1.2–394.5). The results suggest that persistent JCV viruria and increasing urinary concentration of JCV DNA may be predictive of PML for some patients. J. Med. Virol. 81:1140–1150, 2009.

Human Papillomavirus Clearance Among Males Is Associated With HIV Acquisition and Increased Dendritic Cell Density in the Foreskin

BACKGROUND The association between human papillomavirus (HPV) infection and the risk of human immunodeficiency virus (HIV) seroconversion is unclear, and the genital cellular immunology has not been evaluated. METHODS A case-control analysis nested within a male circumcision trial was conducted. Cases consisted of 44 male HIV seroconverters, and controls were 787 males who were persistently negative for HIV. The Roche HPV Linear Array Genotype Test detected high-risk HPV (HR-HPV) and low-risk HPV (LR-HPV) genotypes. Generalized estimating equations logistic regression was used to estimate adjusted odds ratios (aORs) of HIV seroconversion. In addition, densities of CD1a(+) dendritic cells, CD4(+) T cells, and CD8(+) T cells were measured using immunohistochemistry analysis in foreskins of 79 males randomly selected from participants in the circumcision trial. RESULTS HR-HPV or LR-HPV acquisition was not significantly associated with HIV seroconversion, after adjustment for sexual behaviors. However, HR-HPV and LR-HPV clearance was significantly associated with HIV seroconversion (aOR, 3.25 [95% confidence interval {CI}, 1.11-9.55] and 3.18 [95% CI, 1.14-8.90], respectively). The odds of HIV seroconversion increased with increasing number of HPV genotypes cleared (P < .001, by the test for trend). The median CD1a(+) dendritic cell density in the foreskin epidermis was significantly higher among males who cleared HPV (72.0 cells/mm(2) [interquartile range {IQR}, 29.4-138.3 cells/mm(2)]), compared with males who were persistently negative for HPV (32.1 cells/mm(2) [IQR, 3.1-96.2 cells/mm(2)]; P = .047), and increased progressively with the number of HPV genotypes cleared (P = .05). CONCLUSIONS HPV clearance was associated with subsequent HIV seroconversion and also with increased epidermal dendritic cell density, which potentially mediates HIV seroconversion.

High-risk human papillomavirus viral load and persistence among heterosexual HIV-negative and HIV-positive men

Objectives High-risk human papillomavirus (HR-HPV) viral load is associated with HR-HPV transmission and HR-HPV persistence in women. It is unknown whether HR-HPV viral load is associated with persistence in HIV-negative or HIV-positive men. Methods HR-HPV viral load and persistence were evaluated among 703 HIV-negative and 233 HIV-positive heterosexual men who participated in a male circumcision trial in Rakai, Uganda. Penile swabs were tested at baseline and 6, 12 and 24 months for HR-HPV using the Roche HPV Linear Array, which provides a semiquantitative measure of HPV shedding by hybridisation band intensity (graded: 1–4). Prevalence risk ratios (PRR) were used to estimate the association between HR-HPV viral load and persistent detection of HR-HPV. Results HR-HPV genotypes with high viral load (grade:3–4) at baseline were more likely to persist than HR-HPV genotypes with low viral load (grade: 1–2) among HIV-negative men (month 6: adjPRR=1.83, 95% CI 1.32 to 2.52; month 12: adjPRR=2.01, 95% CI 1.42 to 3.11), and HIV-positive men (month 6: adjPRR=1.33, 95% CI 1.06 to 1.67; month 12: adjPRR=1.73, 95% CI 1.18 to 2.54). Long-term persistence of HR-HPV was more frequent among HIV-positive men compared with HIV-negative men (month 24: adjPRR=2.27, 95% CI 1.47 to 3.51). Persistence of newly detected HR-HPV at the 6-month and 12-month visits with high viral load were also more likely to persist to 24 months than HR-HPV with low viral load among HIV-negative men (adjPRR=1.67, 95% CI 0.88 to 3.16). Conclusions HR-HPV genotypes with high viral load are more likely to persist among HIV-negative and HIV-positive men, though persistence was more common among HIV-positive men overall. The results may explain the association between high HR-HPV viral load and HR-HPV transmission.

Use of injectable hormonal contraception and women's risk of herpes simplex virus type 2 acquisition: a prospective study of couples in Rakai, Uganda

BACKGROUND The injectable hormonal contraceptive depo-medroxyprogesterone acetate (DMPA) has been associated with increased risk of HIV acquisition, but findings are inconsistent. Whether DMPA increases the risk of other sexually transmitted viral infections is unknown. We assessed the association between DMPA use and incident herpes simplex virus type 2 (HSV2) infection in women. METHODS In this prospective study, we enrolled HIV-negative and HSV2-negative women aged 15-49 years whose HIV-negative male partners were concurrently enrolled in a randomised trial of male circumcision in Rakai, Uganda. We excluded women if either they or their male partners HIV seroconverted. The primary outcome was HSV2 seroconversion, assessed annually. The male circumcision trial was registered with ClinicalTrials.gov, number NCT00425984. FINDINGS Between Aug 11, 2003, and July 6, 2006, we enrolled 682 women in this study. We noted HSV2 seroconversions in 70 (10%) women. Incidence was 13·5 per 100 person-years in women consistently using DMPA (nine incident infections per 66·5 person-years), 4·3 per 100 person-years in pregnant women who were not using hormonal contraception (18 incident infections per 423·5 person-years), and 6·6 per 100 person-years in women who were neither pregnant nor using hormonal contraception (35 incident infections per 529·5 person-years). Women consistently using DMPA had an adjusted hazard ratio for HSV2 seroconversion of 2·26 (95% CI 1·09-4·69; p=0·029) compared with women who were neither pregnant nor using hormonal contraception. Of 132 women with HSV2-seropositive partners, seroconversion was 36·4 per 100 person-years in consistent DMPA users (four incident infections per 11 person-years) and 10·7 per 100 person-years in women who were neither pregnant nor using hormonal contraception (11 incident infections per 103 person-years; adjusted hazard ratio 6·23, 95% CI 1·49-26·3; p=0·012). INTERPRETATION Consistent DMPA use might increase risk of HSV2 seroconversion; however, study power was low. These findings should be assessed in larger populations with more frequent follow-up than in this study, and other contraceptive methods should also be assessed. Access to a wide range of highly effective contraceptive methods is needed for women, particularly in sub-Saharan Africa. FUNDING Bill and Melinda Gates Foundation, Doris Duke Charitable Foundation, US National Institutes of Health, and Fogarty International Center.