HSV-2 Seronegative Research Articles

Low prevalence of bacterial vaginosis in Kenyan adolescent girls and rapid incidence after first sex

Bacterial vaginosis (BV) is a risk factor for sexually transmitted infections (STI) including HIV. Adult African women have high prevalence of BV, but it is not known when first BV occurs. We sought to describe BV in younger African women, before and after first sex, and to determine incidence of BV and significant correlates of BV incidence and recurrence. In a prospective observational cohort study enrolling adolescents with limited sexual experience, young women ages 16-21 were recruited in Thika, Kenya. Eligible participants were HIV and HSV-2 seronegative and reported zero or one lifetime sexual partner. Nugent score was determined at quarterly visits from vaginal Gram stains. Trends in BV were described over time; hazard ratios were calculated using Cox regression and relative risk of BV was estimated using generalized estimating equations and Poisson regression. We enrolled 400 participants with a median age of 18.6 years (IQR 16-21). The majority (322, 80.5%) reported no history of sex, while 78 (19.5%) reported sex with 1 partner. At enrollment, BV (Nugent score ≥7) was uncommon (21 of 375, 5.6%). Overall, 144 participants had BV at least once, for an incidence rate of 16.5 cases per 100 person-years. Prior to first sex, BV was present at 2.8% of visits, compared to 13.7% of visits post-first sex. An adjusted model of BV incidence observed that first sex was associated with more than twofold increased BV risk (aHR 2.44, p=0.009, CI 1.25-4.76). Chlamydia diagnosis (aHR 1.73, p=0.02, CI 1.1-2.8) and HSV-2 seropositivity (aHR 2.88, p=0.021, CI 1.17-7.09) were both associated with incident BV. A multivariate GEE model including all episodes of BV demonstrated risk factors including first sex, STIs, urban residence, recent sex and no income; the most important risk factor was first sex (aRR 1.92, p=0.018, CI 1.12-3.31). Probability of BV increased with each subsequent episode; mean Nugent scores increased after each BV episode. Using detailed longitudinal observation, we found that Kenyan adolescents have almost no BV prior to first sex, and that initiation of sexual activity was the strongest risk factor for both prevalent and incident BV.

Incidence of Herpes Simplex Virus Type 2 Infection Among African Women Using Depot Medroxyprogesterone Acetate, a Copper Intrauterine Device, or a Levonorgestrel Implant for Contraception: A Nested Randomized Trial

Background: Globally, women have higher herpes simplex type 2 (HSV-2) prevalence than men; data from observational studies suggest a possible association of HSV-2 acquisition with use of certain contraceptive methods, particularly intramuscular depot medroxyprogesterone acetate (DMPA-IM). Methods: Within a randomized trial of the effect of three contraceptive methods – DMPA-IM, a copper intrauterine device (IUD), and a levonorgestrel (LNG) implant – on HIV acquisition, we assessed HSV-2 acquisition. We followed women who were HSV-2 and HIV seronegative, aged 16-35 years, and seeking effective contraception for 12-18 months at 12 sites in Eswatini, Kenya, South Africa, and Zambia from 2015-2018. HSV-2 serologic testing was done at enrollment and final study visits according to a standardized algorithm. Intention–to-treat analysis using Poisson regression with robust standard errors compared HSV-2 incidence by contraceptive method. Findings: At baseline, 4062 randomized women were HSV-2 seronegative, of whom 3898 (96.0%) had a conclusive HSV-2 result at their final study visit. Of these, 614 (15.8%) acquired HSV-2, at an incidence of 12.4/100 person-years (p-y): 10.9/100 p-y among women assigned DMPA-IM, 13.7/100 p-y the copper IUD, and 12.7/100 p-y the LNG implant. Incidence rate ratios (IRR) for HSV-2 acquisition were 0.80 (95% confidence interval [CI] 0.65-0.97) for DMPA-IM compared with copper IUD, 0.86 (95% CI 0.71-1.05) for DMPA-IM compared with LNG implant, and 1.08 (95% CI 0.89-1.30) for copper IUD compared with LNG implant. HSV-2 acquisition risk was significantly increased among women who also acquired HIV during follow-up (IRR 3.55, 95% CI 2.78-4.48). Interpretation: In a randomized trial, DMPA-IM was not associated with increased risk of HSV-2 acquisition. HSV-2 incidence was >10% per year, emphasizing the need for development of novel HSV-2 prevention strategies for African women. Clinical Trial Registration Details: ClinicalTrials.gov number NCT02550067. Funding Information: The ECHO Trial was made possible by the combined generous support of the Bill & Melinda Gates Foundation, the American people through the United States Agency for International Development and the President’s Emergency Plan for AIDS Relief, the Swedish International Development Cooperation Agency as part of the EDCTP2 programme supported by the European Union, the South African Medical Research Council, and the United Nations Population Fund. Contraceptive supplies were donated by the Government of South Africa and United States Agency for International Development. Additional funding for testing for herpes simplex type 2 infection was provided in part by the US Centers for Disease Control and Prevention (U48 DP 005013 SIP 14-023) and the University of Washington/Fred Hutch Center for AIDS Research (US National Institutes of Health P30 AI057757). Declaration of Interests: None. Ethics Approval Statement: Ethics review committees at each study site approved the study protocol. All participants provided written informed consent, which included counseling about randomization, each study contraceptive method, and their rights as research participants.

Sexually Transmitted Infections Among Kenyan Adolescent Girls and Young Women With Limited Sexual Experience.

Objectives: Globally, the highest rates of sexually transmitted infections (STIs) are among the 15-24 age group. Studying adolescent girls and young women (AGYW) pre-sexual debut could identify risk factors for STI acquisition.Methods: We recruited a prospective cohort of low-risk AGYW aged 16-20 in Kenya. Participants were HIV and HSV-2 seronegative and reported no history of sexual intercourse or reported sex with one partner. Participants underwent genital exams, nucleic acid testing of vaginal swabs for Neisseria gonorrhea (NG), Chlamydia trachomatis (CT), Trichomonas vaginalis (TV), and vaginal gram stains for vaginal dysbiosis by Nugent score. STI correlates were described using χ2 test and t-test.Results: We enrolled 400 AGYW, of which 322 (80.5%) reported never having had sex, while 78 (19.5%) reported prior sex with 1 partner. Among the 78 participants reporting prior sex, 20 (25.6%) reported contraception use in the last 3 months, with 60% using only emergency contraceptive pills. Despite self-reported history, of 373 subjects who underwent STI testing, 49 subjects (13.1%) tested positive for STIs, with 41 CT, 5 GC, and 3 TV cases. Of these 49 subjects, 33 (67.3%) reported no prior sexual intercourse. Bacterial vaginosis was rare and 90% of subjects had a normal Nugent score (0–3).Conclusions: Upon baseline evaluation of a cohort of low risk AGYW, we found high numbers of STIs, especially CT, which is not routinely screened for in Kenyan settings. Interventions to address STIs and unintended pregnancy should target girls pre-sexual debut, including those who do not self-identify as at risk.

HSV-2 infects T follicular helper cells to promote HIV reactivation

Abstract HSV-2 is one of the most significant factors fueling the global HIV epidemic and coinfection is associated with higher HIV plasma viral loads and increased rates of HIV transmission and HSV-2 recurrences. The mechanisms underlying this syndemic likely reflect local and systemic effects. To explore potential systemic effects, we characterized PBMC from a cohort of HIV-1+ women who were or were not HSV-2 seropositive (HSV-2+) and identified phenotypic changes in CD4 T cells including increased frequencies of activated cells and cells expressing co-receptors for HIV entry in HIV+/HSV-2+ vs. HIV+/HSV-2 seronegative (HSV-2−) women. Surprisingly, IL-32, a proinflammatory cytokine, was lower in CD4 T cells from HSV-2+ women. Addition of IL-32γ to CD4 T cells from HIV+ women blocked HIV reactivation following stimulation with phytohemagglutinin, suggesting that HSV-2 induces phenotypic changes in CD4 T cells to promote HIV reactivation. Subsequent RNAseq analysis of CD4 T cells infected ex vivo with HSV-2 also showed lower levels of IL-32, as well as elevated bcl6, itch, and irf4 transcripts along with decreased ccr7, psgl1, and s1pr1, suggesting a transcriptional landscape consistent with T follicular helper (TFH) cells. Flow cytometry studies using CD4 T cells isolated from PBMC from HIV− or HIV+ donors, as well as from tonsils, showed an increased frequency of TFH cells in HSV-2 infected CD4 T cells, suggesting preferential infection of this subpopulation, which is known to harbor latent HIV. HSV-2 infection of CD4 T cells isolated from HIV+ donors or latently infected T cell lines triggered HIV reactivation. These findings demonstrate that HSV-2 directly infects CD4 T cell subpopulations including TFH cells to promote HIV reactivation.

128. Sexually Transmitted Infections Among Adolescent Girls in Thika, Kenya

BackgroundGlobally, the rates of sexually transmitted infections (STIs) are highest among 15- to 24-year-old girls, especially in Sub-Saharan African countries where the adolescent sexual health is poor. Recruiting girls presexual debut could identify risk factors for STI acquisition.MethodsWe recruited a prospective cohort of “low-risk” adolescent girls aged 16–20 in Kenya. To be eligible, girls were HIV and HSV-2 seronegative and reported no history of sexual intercourse or reported having sex with only one partner. Demographic data were collected, and girls had nucleic acid testing of vaginal swabs for Neisseria gonorrhea, Chlamydia trachomatis, and Trichomonas vaginalis, and vaginal gram stains for bacterial vaginosis (BV).ResultsWe enrolled 400 girls, with a median age of 18.6 years. In this cohort, 322 (80.5%) girls reported never having had sex, while 78 (19.5%) reported prior sex with 1 partner. Of those reporting prior sex, only 20 (25.6%) reported contraception use in the last 3 months, with 60% using only emergency contraceptive pills. The median age of sexual partners was 22 (IQR 19–25). Of the 373 participants with an STI swab result, 49 participants (13.1%) tested positive for STIs at entry into the study, with 41 chlamydia, 5 gonorrhea, and 3 trichomonas cases. Of these 49 participants, 33 (67.3%) had denied prior sexual intercourse. Testing positive for STIs was, however, significantly different among those reporting prior sexual intercourse vs. reporting never having had sex, 21.1% vs. 11.1% (P = 0.02). BV was rare (5.6%) in the cohort, with 90% of participants with a normal Nugent score of 0–3.ConclusionIn the initial testing of a sexually inexperienced cohort of girls, we found unexpectedly high numbers of prevalent STIs, especially chlamydia which is not routinely screened for in Kenyan settings. Additionally, lack of sexual activity appeared overreported. BV was rare, with much lower prevalence than in adult women in Africa. Our data suggest that prior to initiation of sexual activity, most girls in this Kenyan cohort have vaginal microflora that is dominated by Lactobacillus. Interventions to address STIs, including pre-exposure prophylaxis for HIV, should be targeted at girls at a young age, presexual debut, and in nonmedical settings where girls can be reached who do not selfidentify as at risk for STI.Disclosures All authors: No reported disclosures.

Performance of Commercial Enzyme-Linked Immunoassays for Diagnosis of Herpes Simplex Virus-1 and Herpes Simplex Virus-2 Infection in a Clinical Setting.

US Food and Drug Administration-approved enzyme-linked immunoassays (EIA) for determining type-specific herpes simplex virus (HSV) serostatus are widely used in clinical practice. We compared the performance of such assays with the University of Washington Western blot (UW WB) in patients who sought confirmation of their HSV serology result. We reviewed charts of all persons evaluated at the Westover Heights Clinic in Portland, Oregon, from July 2010 through September 2015, who had a HSV EIA, followed by UW WB. Of 864 persons, 47% were women. The median age was 36 years (range, 18-73 years). Using UW WB to define infection status, 286 (33%) persons were HSV-1 seropositive only, 104 (12%) were HSV-2 seropositive only, 134 (16%) were both HSV-1 and HSV-2 seropositive, 235 (27%) were HSV seronegative, and 105 (12%) had indeterminate results. Compared with the UW WB as the criterion standard, EIA was 70.2% sensitive and 91.6% specific for HSV-1, and 91.9% sensitive and 57.4% specific for HSV-2.Among 278 persons who were HSV-1 seropositive by EIA, 255 were confirmed by the UW WB (positive predictive value [PPV], 91.7%). Of the 360 persons that were HSV-1 seronegative by the EIA, 252 were seronegative by UW WB (negative predictive value [NPV], 70.0%). Among 381 persons with HSV-2 EIA seropositivity, 193 tested HSV-2 seropositive by the UW WB (PPV, 50.7%). Of the 270 persons HSV-2 seronegative by EIA, 17 were seropositive with the UW WB (NPV, 93.7%). Among 261 persons with an EIA HSV-2 index value = 1.1-2.9, 39.8% of results were confirmed by UW WB, compared with 78.6% of the 70 persons with an EIA index value of 3 or greater (P < 0.001). The risk of false-positive HSV-2 EIA results was higher in those with HSV-1 antibody (47.1% vs 37.1%, P = 0.036). US Food and Drug Administration-approved EIAs have poor PPV for HSV-2 and poor NPV for HSV-1 in clinical practice. More accurate rapid type-specific HSV antibody tests are needed.

A direct quantitative PCR-based measurement of herpes simplex virus susceptibility to antiviral drugs and neutralizing antibodies

Herpes simplex viruses (HSV) are common human pathogens that can cause painful but benign manifestations and recurrent complaints, but can also cause significant morbidity and mortality on infection of the eye or brain and with disseminated infection of an immunosuppressed patient or a neonate. HSV growth inhibition measurement by plaque or yield reduction is a key task in the development of novel antiviral compounds but the manual methods are very labour intensive. The sensitive and specific PCR technology could be an effective method for quantitation of HSV DNA related to virus replication; however the currently described PCR approaches have a major limitation, namely the requirement of purification of DNA from the infected cells. This limitation makes this approach unfeasible for high-throughput screenings. The monitoring of HSV specific antibody titre is essential in vaccination trials and in the improvement of HSV-based oncolytic virotherapy. Usually, conventional cytopathic effect-based and plaque reduction neutralization tests are applied to measure the neutralization titre, but these methods are also time-consuming. To overcome this, we developed a quantitative PCR (qPCR) method for the detection of HSV-2 DNA directly from the infected cells (direct qPCR) and the method was further adapted to measure the titre of HSV specific neutralizing antibody in human sera. The conditions of direct qPCR assay were optimized to measure the antiviral activity of known and novel antiviral substances. Using HSV-2 seronegative and seropositive patients’ sera, the validity of the direct qPCR neutralization test was compared to traditional cytopathic effect-based assay.The direct qPCR method was able to detect the HSV-2 DNA quantitatively between multiplicity of infection 1/64 and 1/4194304, indicating that the dynamic range of the detection was approximately 65,500 fold with high correlation between the biological and technical replicates. As a proof of the adaptability of the method, we applied the direct qPCR for antiviral inhibitory concentration 50 (IC50) measurements of known and novel antiviral compounds. The measured IC50 of acyclovir was ∼0.28μg/ml, similar to the previously published IC50 value. The IC50 of novel antiviral candidates was between 1.6–3.1μg/ml. The direct qPCR-based neutralization titres of HSV positive sera were 1:32–1:64, identical to the neutralization titres determined using a traditional neutralization assay. The negative sera did not inhibit the HSV-2 replication in either of the tests.Our direct qPCR method for the HSV-2 growth determination of antiviral IC50 and neutralization titre is less time-consuming, less subjective and a more accurate alternative to the traditional plaque titration and growth reduction assays.

Obstetric outcomes of human herpes virus-2 infection among pregnant women in Benin, Nigeria.

This study investigated the obstetric outcomes of herpes simplex virus (HSV-2) infection among pregnant women. In this prospective cohort study, a total of 674 consenting pregnant women attending ante-natal clinic in the University of Benin Teaching Hospital and Central Hospital Benin were recruited between November 2011 and December, 2012. The women were screened for HSV-1, and HSV-2 using glycoprotein-G-based type-specific Enzyme Linked Immunosorbent Assay on archived blood samples; and were followed up to the delivery period and postnatal clinic. The HSV-2-seronegative participants underwent second blood sampling for HSV-2 IgG and IgM assay during the delivery period. The patients were thus categorized into "HSV-2 seropositive", "HSV-2-seronegative," and "incident HSV-2 infection" cohorts. The pregnancy outcomes were assessed by review of hospital records. Data analysis was with SPSS version 16 software. Of 674 pregnant women surveyed, 312 (46.3%) were HSV-2 seropositive; while 362 (56.7%) were HSV-2 seronegative. Comparing the "HSV-2 seropositive" and "HSV-seronegative" groups, there were no significant differences in occurrence of low birth weight (LBW), prematurity, spontaneous abortions, and stillbirth events (P=0.96; 0.95; 1.0; and 0.77, respectively). Comparing the "incident HSV-2 infection" with the "HSV-2 seronegative" groups, the relative risks of occurrence of LBW deliveries, preterm deliveries, and stillbirths were 12.6, 25.1, and 4.5, respectively. First episode HSV-2 infection among pregnant women in Benin, Nigeria is associated with an increased risk of occurrence of spontaneous abortion, LBW delivery, stillbirths, and preterm delivery.

P5.072 The Performance of Two IgG ELISA Methods to Detect HSV-2 Infections Among South-African Women Who Are at Higher Risk of Becoming HIV Infected

BackgroundSpecimens collected in Pretoria for the FEM-PrEP study, a phase III trial on pre-exposure prophylaxis for HIV prevention among African women, were tested for Herpes Simplex Virus type II (HSV-2)....

P3.228 HSV-2 Seroincidence and Its Association with Medical Male Circumcision, HIV, Genital Ulcer Disease, and Penile Epithelial Trauma

BackgroundWe estimated the 72-month efficacy of medical male circumcision (MMC) against HSV-2 seroincidence among men in the randomised trial of MMC in Kisumu, Kenya.MethodsFrom 2002–2005, 2,784 men aged 18–24 were...

Changes in the Soluble Mucosal Immune Environment During Genital Herpes Outbreaks

Genital tract secretions provide variable inhibitory activity against herpes simplex virus (HSV) ex vivo. We hypothesize that the anti-HSV activity may prevent the spread of virus from the more commonly affected sites, such as the external genitalia, to the upper genital tract. The antimicrobial activity of cervicovaginal lavage (CVL) and concentrations of mucosal immune mediators were measured in 10 HIV-seronegative women with an active external herpetic lesion and compared with 10 HIV-seronegative women who were HSV-1 and HSV-2 seronegative. Samples were obtained at the time of a symptomatic external lesion (day 0), after 1 week of oral acyclovir (day 7), and 1 week after completing treatment (day 14). Controls were evaluated at parallel intervals. The anti-HSV activity was higher in CVL obtained from cases compared to controls at presentation (day 0) (54.3% vs. 28%), fell to similar levels on day 7, and then rebounded on day 14 (69% vs. 25%). The anti-HSV activity correlated positively and significantly with the concentrations of several inflammatory proteins; the concentrations of these proteins tended to be higher in cases compared with controls and followed a similar temporal pattern. Increases in inflammatory immune mediators and anti-HSV activity were detected in CVL at the time of clinical outbreaks and after completion of a short course of acyclovir. These mucosal responses may protect against HSV spread but could facilitate HIV infection and contribute to the clinical observation that, independent of clinical lesions, HSV-2 is a risk factor for HIV acquisition.

Uptake of Voluntary Counselling and Testing among Young People Participating in an HIV Prevention Trial: Comparison of Opt-Out and Opt-In Strategies

BackgroundHIV voluntary counselling and testing (VCT) is an integral component of HIV prevention and treatment programmes. However, testing coverage in sub-Saharan Africa is still low, particularly among young people. As treatment becomes more widely available, strategies to expand VCT coverage are critically important. We compare VCT uptake using two delivery strategies (opt-in and opt-out) within the MEMA kwa Vijana trial in 20 communities in northwest Tanzania.MethodsWe analysed data from 12,590 young persons (median (IQR) age 22 years (20–23)) to assess the effect of delivery strategy on VCT uptake. Ten communities used an opt-in approach and 10 used opt-out, balanced across intervention and control. Conditional logistic regression was used to examine factors associated with uptake within each strategy.ResultsVCT uptake was significantly higher with the opt-out approach (90.9% vs 60.5%, prevalence ratio = 1.51, CI = 1.41–1.62). Among females, uptake in the opt-out approach was associated with decreased knowledge of HIV acquisition, sex with a casual partner, and being HSV-2 seronegative; among males, uptake was associated with lower education and increasing lifetime partners. In contrast, uptake using the opt-in approach varied by ethnic group, religion and marital status, and increased with increasing knowledge of STI acquisition (males) or pregnancy prevention (females).ConclusionVCT uptake among young people was extremely high when offered an opt-out strategy. Sociodemographic and knowledge factors affected uptake in different ways depending on delivery strategy. Increased knowledge may increase young persons' self-efficacy, which may have a different impact on testing uptake, depending on the approach used.

P1-S5.25 Acyclovir and transmission of HSV-2 from HSV-2/HIV-1 dually infected persons

BackgroundDaily suppressive therapy with valacyclovir reduces the risk of sexual transmission of HSV-2 in healthy HSV-2 serodiscordant heterosexual couples by 48%. Whether suppressive therapy reduces HSV-2 transmission from persons who...

The prevalence, incidence and risk factors of herpes simplex virus type 2 infection among pregnant Zimbabwean women followed up nine months after childbirth

BackgroundHerpes simplex virus type 2 (HSV-2) is the leading cause of genital ulcer disease worldwide. The virus can be transmitted to neonates and there are scarce data regarding incidence of HSV-2 among women in pregnancy and after childbirth. The aim of this study is to measure the incidence and risk factors for HSV-2 infection in women followed for 9 months after childbirth.MethodsPregnant women were consecutively enrolled late in pregnancy and followed at six weeks, four and nine months after childbirth. Stored samples were tested for HSV-2 at baseline and again at nine months after childbirth and HSV-2 seropositive samples at nine months after childbirth (seroconverters) were tested retrospectively to identify the seroconversion point.ResultsOne hundred and seventy-three (50.9%) of the 340 consecutively enrolled pregnant women were HSV-2 seronegative at baseline. HSV-2 incidence rate during the 10 months follow up was 9.7 (95% CI 5.4-14.4)/100 and 18.8 (95% CI 13.9-26.1)/100 person years at risk (PYAR) at four months and nine months after childbirth respectively. Analysis restricted to women reporting sexual activity yielded higher incidence rates. The prevalence of HSV-2 amongst the HIV-1 seropositive was 89.3%. Risk factors associated with HSV-2 seropositivity were having other sexual partners in past 12 months (Prevalence Risk Ratio (PRR) 1.8 (95% CI 1.4-2.4) and presence of Trichomonas vaginalis (PRR 1.7 95% CI 1.4-2.1). Polygamy (Incidence Rate Ratio (IRR) 4.4, 95% CI 1.9-10.6) and young age at sexual debut (IRR 3.6, 95% CI 1.6-8.3) were associated with primary HSV-2 infection during the 10 months follow up.ConclusionsIncidence of HSV-2 after childbirth is high and the period between late pregnancy and six weeks after childbirth needs to be targeted for prevention of primary HSV-2 infection to avert possible neonatal infections.

The Acceptance of HSV-Testing Partners of HSV-2 Seronegative Pregnant Women

To estimate the acceptance of HSV testing partners of HSV seronegative pregnant women and identify associated factors. Consecutive women identified as HSV-2 seronegative during routine testing in pregnancy were asked to invite their partners for HSV testing to identify serodiscordance. Logistic regression identified factors associated with partner testing. Between 2001 to 2006, 315 women enrolled (28% of those approached) and 242 (77%) partners were tested. Married couples were most likely to be tested [adjusted odds ratio (aOR) 7.72, 95% CI: 2.47-24.15]. Partners of black women (aOR 0.17, 95% CI: 0.04-0.71), and those with at least a college degree (aOR 0.43, 95% CI: 0.19-0.98) were less likely to be tested. In this population, partner testing among HSV-2 seronegative women was feasible which supports further study to determine if identification of partners who pose a potential risk of HSV infection during pregnancy is an effective approach to reduce HSV acquisition in pregnant women.