Julie Overbaugh

Hormonal contraception, vitamin A deficiency, and other risk factors for shedding of HIV-1 infected cells from the cervix and vagina

BACKGROUND Factors that influence shedding of HIV-1 infected cells in cervical and vaginal secretions may be important determinants of sexual and vertical transmission of the virus. We investigated whether hormonal contraceptive use, vitamin A deficiency, and other variables were risk factors for cervical and vaginal shedding of HIV-infected cells. METHODS Between December, 1994, and April, 1996, women who attended a municipal sexually transmitted diseases (STDs) clinic in Mombasa, Kenya, and had previously tested positive for HIV-1, were invited to take part in our cross-sectional study. Cervical and vaginal secretions from 318 women were evaluated for the presence of HIV-1 infected cells by PCR amplification of gag DNA sequences. FINDINGS HIV-1 infected cells were detected in 51% of endocervical and 14% of vaginal-swab specimens. Both cervical and vaginal shedding of HIV-1 infected cells were highly associated with CD4 lymphocyte depletion (p = 0.00001 and p = 0.003, respectively). After adjustment for CD4 count, cervical proviral shedding was significantly associated with use of depot medroxyprogesterone acetate (odds ratio 2.9, 95% CI 1.5-5.7), and with use of low-dose and high-dose oral contraceptive pills (3.8, 1.4-9.9 and 12.3, 1.5-101, respectively). Vitamin A deficiency was highly predictive of vaginal HIV-1 DNA shedding. After adjustment for CD4 count, severe vitamin A deficiency, moderate deficiency, and low normal vitamin A status were associated with 12.9, 8.0, and 4.9-fold increased odds of vaginal shedding, respectively. Gonococcal cervicitis (3.1, 1.1-9.8) and vaginal candidiasis (2.6, 1.2-5.4) were also correlated with significant increases in HIV-1 DNA detection, but Chlamydia trachomatis and Trichomonas vaginalis were not. INTERPRETATION Our study documents several novel correlates of HIV-1 shedding in cervical and vaginal secretions, most notably hormonal contraceptive use and vitamin A deficiency. These factors may be important determinants of sexual or vertical transmission of HIV-1 and are of public health importance because they are easily modified by simple interventions.

Correlates of Mother-to-Child Human Immunodeficiency Virus Type 1 (HIV-1) Transmission: Association with Maternal Plasma HIV-1 RNA Load, Genital HIV-1 DNA Shedding, and Breast Infections

To determine the effects of plasma, genital, and breast milk human immunodeficiency virus type 1 (HIV-1) and breast infections on perinatal HIV-1 transmission, a nested case-control study was conducted within a randomized clinical trial of breast-feeding and formula feeding among HIV-1-seropositive mothers in Nairobi, Kenya. In analyses comparing 92 infected infants with 187 infants who were uninfected at 2 years, maternal viral RNA levels >43,000 copies/mL (cohort median) were associated with a 4-fold increase in risk of transmission (95% confidence interval [CI], 2.2-7.2). Maternal cervical HIV-1 DNA (odds ratio [OR], 2.4; 95% CI, 1.3-4.4), vaginal HIV-1 DNA (OR, 2.3; 95% CI, 1.1-4.7), and cervical or vaginal ulcers (OR, 2.7; 95% CI, 1.2-5.8) were significantly associated with infant infection, independent of plasma virus load. Breast-feeding (OR, 1.7; 95% CI, 1.0-2.9) and mastitis (relative risk [RR], 3.9; 95% CI, 1.2-12.7) were associated with increased transmission overall, and mastitis (RR, 21.8; 95% CI, 2.3-211.0) and breast abscess (RR, 51.6; 95% CI, 4.7-571.0) were associated with late transmission (occurring >2 months postpartum). Use of methods that decrease infant exposure to HIV-1 in maternal genital secretions or breast milk may enhance currently recommended perinatal HIV-1 interventions.

Human Immunodeficiency Virus Type 1 V1-V2 Envelope Loop Sequences Expand and Add Glycosylation Sites over the Course of Infection, and These Modifications Affect Antibody Neutralization Sensitivity

ABSTRACT Over the course of infection, human immunodeficiency virus type 1 (HIV-1) continuously adapts to evade the evolving host neutralizing antibody responses. Changes in the envelope variable loop sequences, particularly the extent of glycosylation, have been implicated in antibody escape. To document modifications that potentially influence antibody susceptibility, we compared envelope variable loops 1 and 2 (V1-V2) from multiple sequences isolated at the primary phase of infection to those isolated around 2 to 3 years into the chronic phase of infection in nine women with HIV-1 subtype A. HIV-1 sequences isolated during chronic infection had significantly longer V1-V2 loops, with a significantly higher number of potential N-linked glycosylation sites, than the sequences isolated early in infection. To assess the effects of these V1-V2 changes on antibody neutralization and infectivity, we created chimeric envelope sequences, which incorporated a subjects V1-V2 sequences into a common subtype A envelope backbone and then used them to generate pseudotyped viruses. Compared to the parent virus, the introduction of a subjects early-infection V1-V2 envelope variable loops rendered the chimeric envelope more sensitive to that subjects plasma samples but only to plasma samples collected >6 months after the sequences were isolated. Neutralization was not detected with the same plasma when the early-infection V1-V2 sequences were replaced with chronic-infection V1-V2 sequences, suggesting that changes in V1-V2 contribute to antibody escape. Pseudotyped viruses with V1-V2 segments from different times in infection, however, showed no significant difference in neutralization sensitivity to heterologous pooled plasma, suggesting that viruses with V1-V2 loops from early in infection were not inherently more neutralization sensitive. Pseudotyped viruses bearing chimeric envelopes with early-infection V1-V2 sequences showed a trend in infecting cells with low CD4 concentrations more efficiently, while engineered viruses with V1-V2 sequences isolated during chronic infection were moderately better at infecting cells with low CCR5 concentrations. These studies suggest that changes within the V1-V2 envelope domains over the course of an infection influence sensitivity to autologous neutralizing antibodies and may also impact host receptor/coreceptor interactions.

Selection for human immunodeficiency virus type 1 envelope glycosylation variants with shorter V1-V2 loop sequences occurs during transmission of certain genetic subtypes and may impact viral RNA levels.

ABSTRACT Designing an effective human immunodeficiency virus type 1 (HIV-1) vaccine will rely on understanding which variants, from among the myriad of circulating HIV-1 strains, are most commonly transmitted and determining whether such variants have an Achilles heel. Here we show that heterosexually acquired subtype A HIV-1 envelopes have signature sequences that include shorter V1-V2 loop sequences and fewer predicted N-linked glycosylation sites relative to the overall population of circulating variants. In contrast, recently transmitted subtype B variants did not, and this was true for cases where the major risk factor was homosexual contact, as well as for cases where it was heterosexual contact. This suggests that selection during HIV-1 transmission may vary depending on the infecting subtype. There was evidence from 23 subtype A-infected women for whom there was longitudinal data that those who were infected with viruses with fewer potential N-linked glycosylation sites in V1-V2 had lower viral set point levels. Thus, our study also suggests that the extent of glycosylation in the infecting virus could impact disease progression.

HIV vaccine research: the way forward.

The need to broaden research directed at answering fundamental questions in HIV vaccine discovery through laboratory, nonhuman primate (NHP), and clinical research has recently been emphasized. In addition, the importance of attracting and retaining young researchers, developing better NHP models, and more closely linking NHP and clinical research is being stressed. In an era of a level budget for biomedical research at the U.S. National Institutes of Health (NIH), HIV/AIDS vaccine research efforts will need to be carefully prioritized such that resources to energize HIV vaccine discovery can be identified. This article summarizes progress and challenges in HIV vaccine research, the priorities arising from a recent summit at NIAID, and the actions needed, some already under way, to address those priorities.

Longitudinal Analysis of Human Immunodeficiency Virus Type 1 RNA in Breast Milk and of Its Relationship to Infant Infection and Maternal Disease

Transmission of human immunodeficiency virus type 1 (HIV-1) via breast-feeding can occur throughout lactation. Defining both fluctuation in breast-milk virus level over time and how breast-milk virus correlates with mother-to-child transmission is important for establishing effective interventions. We quantified breast-milk HIV-1 RNA levels in serial samples collected from 275 women for up to 2 years after delivery. Higher maternal plasma virus load, lower maternal CD4 T cell count, and detection of HIV-1 DNA in maternal genital secretions were significantly associated with elevated breast-milk HIV-1 RNA. Within women who breast-fed, median virus load in colostrum/early milk was significantly higher than that in mature breast milk collected 14 days after delivery (P< or =.004). Breast-feeding mothers who transmitted HIV-1 to their infants had both significantly higher breast-milk viral RNA throughout lactation and more-consistent viral shedding, compared with mothers who did not transmit HIV-1. In breast-feeding women, a 2-fold-increased risk of transmission was associated with every 10-fold increase in breast-milk virus load (95% confidence interval, 1.3-3.0; P<.001). These results indicate that the risk of infant infection from breast-feeding is influenced by breast-milk virus load, which is highest early after delivery.

Neutralization Escape Variants of Human Immunodeficiency Virus Type 1 Are Transmitted from Mother to Infant

ABSTRACT Maternal passive immunity typically plays a critical role in protecting infants from new infections; however, the specific contribution of neutralizing antibodies in limiting mother-to-child transmission of human immunodeficiency virus type 1 is unclear. By examining cloned envelope variants from 12 transmission pairs, we found that vertically transmitted variants were more resistant to neutralization by maternal plasma than were maternal viral variants near the time of transmission. The vertically transmitted envelope variants were poorly neutralized by monoclonal antibodies biz, 2G12, 2F5, and 4E10 individually or in combination. Despite the fact that the infant viruses were among the most neutralization resistant in the mother, they had relatively few glycosylation sites. Moreover, the transmitted variants elicited de novo neutralizing antibodies in the infants, indicating that they were not inherently difficult to neutralize. The neutralization resistance of vertically transmitted viruses is in contrast to the relative neutralization sensitivity of viruses sexually transmitted within discordant couples, suggesting that the antigenic properties of viruses that are favored for transmission may differ depending upon mode of transmission.

The Effect of Treatment of Vaginal Infections on Shedding of Human Immunodeficiency Virus Type 1

To assess the effect of treatment of vaginal infections on vaginal shedding of cell-free human immunodeficiency virus type 1 (HIV-1) and HIV-1-infected cells, HIV-1-seropositive women were examined before and after treatment of Candida vulvovaginitis, Trichomonas vaginitis, and bacterial vaginosis. For Candida (n=98), vaginal HIV-1 RNA decreased from 3.36 to 2.86 log(10) copies/swab (P<.001), as did the prevalence of HIV-1 DNA (36% to 17%; odds ratio [OR], 2.8; 95% confidence interval [CI], 1.3-6.5). For Trichomonas vaginitis (n=55), HIV-1 RNA decreased from 3.67 to 3.05 log(10) copies/swab (P<.001), but the prevalence of HIV-1 DNA remained unchanged (22%-25%; OR, 0.8; 95% CI, 0.3-2.2). For bacterial vaginosis (n=73), neither the shedding of HIV-1 RNA (from 3.11 to 2.90 log(10) copies/swab; P=.14) nor the prevalence of DNA (from 21% to 23%; OR, 0.8; 95% CI, 0.3-2.0) changed. Vaginal HIV-1 decreased 3.2- and 4.2-fold after treating Candida and Trichomonas, respectively. These data suggest that HIV-1 transmission intervention strategies that incorporate diagnosis and treatment of these prevalent infections warrant evaluation.

Gender differences in HIV-1 diversity at time of infection

To develop an HIV-1 vaccine with global efficacy, it is important to identify and characterize the viruses that are transmitted, particularly to individuals living in areas of high incidence. Several studies have shown that virus from the blood of acutely infected adults was homogeneous, even when the virus population in the index case was genetically diverse. In contrast to those results with mainly male cohorts in America and Europe, in several cases a heterogeneous virus population has been found early in infection in women in Africa. Thus, we more closely compared the diversity of transmitted HIV-1 in men and women who became infected through heterosexual contact. We found that women from Kenya were often infected by multiple virus variants, whereas men from Kenya were not. Moreover, a heterogeneous virus was present in the women before their seroconversion, and in each woman it was derived from a single index case, indicating that diversity was most likely to be the result of transmission of multiple variants. Our data indicate that there are important differences in the transmitted virus populations in women and men, even when cohorts from the same geographic region who are infected with the same subtypes of HIV-1 are compared.

Association of Levels of HIV-1—Infected Breast Milk Cells and Risk of Mother-to-Child Transmission

Understanding how the level of human immunodeficiency virus type 1 (HIV-1)-infected breast milk cells (BMCs) affects HIV transmission via breast-feeding can shed light on the mechanism of infection and aid in establishing effective interventions. The proportion of infected cells to total cells was measured in serial breast milk samples collected from 291 HIV-1-infected women in Nairobi, Kenya, by use of real-time DNA polymerase chain reaction amplification of BMCs. The number of infected BMCs per million cells was associated with levels of cell-free viral RNA in breast milk (R=.144; P=.032), levels of cell-free virus in blood plasma (R=.365; P<.001), and the detection of proviral DNA in cervical and vaginal secretions (P<.001 and P = .030, respectively). The number of infected BMCs per million cells was lower in colostrum or early milk than in mature milk (P<.001). Previous studies demonstrated that the concentration of BMCs varies throughout lactation, and we used these data to transform infected BMCs per million cells to infected BMCs per milliliter. The estimated concentration of infected BMCs per milliliter was higher in colostrum or early milk than in mature milk (P<.001). Each log10 increase in infected BMCs per milliliter was associated with a 3.19-fold-increased risk of transmission (P=.002), after adjustment for cell-free virus in plasma (hazard ratio [HR], 2.09; P=.03) and breast milk (HR, 1.01; P=1.00). This suggests that infected BMCs may play a more important role in transmission of HIV via breast-feeding than does cell-free virus.