Mary Shepherd

Male circumcision and risk of HIV-1 and other sexually transmitted infections in India

Circumcised men have a lower risk of HIV-1 infection than uncircumcised men. Laboratory findings suggest that the foreskin is enriched with HIV-1 target cells. However, some data suggest that circumcision could simply be a marker for low-risk behaviours. In a prospective study of 2298 HIV-uninfected men attending sexually transmitted infection clinics in India, we noted that circumcision was strongly protective against HIV-1 infection (adjusted relative risk 0.15; 95% CI 0.04-0.62; p=0.0089); however, we noted no protective effect against herpes simplex virus type 2, syphilis, or gonorrhoea. The specificity of this relation suggests a biological rather than behavioural explanation for the protective effect of male circumcision against HIV-1.

The etiology of genital ulcer disease by multiplex polymerase chain reaction and relationship to HIV infection among patients attending sexually transmitted disease clinics in Pune, India

OBJECTIVES To determine the etiology of genital ulcer disease (GUD) among patients attending sexually transmitted disease (STD) clinics in Pune, India, and to examine the relationship to HIV infection and compare the clinical diagnosis of GUD with the results of a multiplex polymerase chain reaction (M-PCR) assay for Treponema pallidum, herpes simplex virus (HSV), and Hemophilus ducreyi infection. METHODS Between June 20, 1994, and September 26, 1994, 302 patients with a genital ulcer were evaluated. Clinical etiology of GUD was based on physical appearance and microbiologic evaluations which included darkfield microscopy and serology for syphilis. Swabs of each genital ulcer were tested for HSV antigen by enzyme immunoassay (Herpchek; Dupont, Wilmington, DE) and processed in a multiplex PCR assay (M-PCR; Roche, Branchburg, NJ) for simultaneous detection of HSV, Treponema pallidum, and Hemophilus ducreyi. RESULTS Two hundred seventy-seven men and 25 women with a median age of 25 were evaluated. The seroprevalence of HIV was 22.2%. The etiology of GUD as determined by M-PCR was HSV (26%), H. ducreyi (23%), T. pallidum (10%), and multiple infections (7%); no etiology was identified in 34%. HIV seroprevalence was higher among those patients positive for HSV compared with other etiologies (OR = 2.1, CI: 1.2-3.7; p = 0.01). When compared with M-PCR, the Herpchek test was 68.5% sensitive and 99.5% specific. Darkfield detection for T. pallidum was 39% sensitive and 82% specific, in contrast to rapid plasma reagin and fluorescent treponemal antibody absorption test, which was 66% sensitive and 90% specific. Clinical diagnosis alone or in combination with basic laboratory tests showed poor agreement with M-PCR.

Feasibility of pooling sera for HIV-1 viral RNA to diagnose acute primary HIV-1 infection and estimate HIV incidence.

ObjectiveTo develop a pooling method for detection of viral RNA for diagnosis of acute HIV infection and estimation of HIV-1 incidence. MethodsSera from 700 consecutive seronegative patients attending sexually transmitted disease clinics in Pune, India, were screened individually for p24 antigen, and pooled into seven pools of 100 for detection of HIV-1 RNA by reverse transcriptase-polymerase chain reaction. HIV-1 incidence was calculated by the traditional cohort method, the p24 antigen method, and a multistage pooling method in which RNA-positive pools were re-analyzed in smaller pools. ResultsSera from 700 individuals were grouped into seven pools of 100, of which four were positive. These four positive pools were subdivided into eight pools of 50, of which seven were positive. The seven positive pools were subdivided into 35 pools of 10, of which 10 were positive. Based on the 10 RNA-positive pools, the point estimate of HIV-1 incidence was 19.9% per year [95% confidence interval (CI), 7.3–31.8%]. Of the 700 samples analyzed for p24 antigen, eight were positive, resulting in a point estimate of incidence of 18.5%/year (8.0–36.5%). In contrast, the incidence rate based on the traditional cohort method of follow-up was lower at 9.4%/year (4.8–16.4%) due to a low follow-up rate. Testing of individual samples from the 10 RNA-positive pools identified 10 individuals with acute primary HIV-1. ConclusionThe multistage pooling method for detection of HIV-1 RNA was more sensitive than the p24 antigen method, and was five-fold less expensive than the p24 antigen assays. Pooling samples for RNA detection was effective in estimating current incidence rates with cost savings that would be practical for use in developing countries.

HIV testing and counseling among men attending sexually transmitted disease clinics in Pune, India: Changes in condom use and sexual behavior over time

Objectives:To describe changes in sexual behavior and condom use among male heterosexual clients at two public sexually transmitted disease (STD) clinics in Pune, after exposure to HIV testing, counseling, and condom promotion. Design:From 13 May 1993 to 11 April 1997, 6819 heterosexual men were screened for HIV infection as part of the HIV Network for Prevention Trials study. A total of 1628 HIV-seronegative men agreed to return at 3-month intervals for repeated HIV counseling and testing. Counseling at each visit focused on reinforcing messages of monogamy, condom use with sexual partners, and provision of government-provided condoms. Methods:Data were collected at baseline and at subsequent 3-month intervals, on demographics, previous STD diagnoses, medical history, sexual behavior, knowledge of HIV/AIDS, and practices related to the prevention of HIV. STD were assessed through physical examination and specimen collection, and blood was drawn for HIV-1 and HIV-2 antibody testing. Results:The level of consistent condom use with sex workers increased proportionately with follow-up time: at 6 months men were 2.8 times more likely to consistently use condoms (P < 0.001), at 18 months they were 3.6 times more likely (P < 0.001), and after 24 months they were 4.7 times more likely to be using condoms every time. The risk of HIV seroconversion in men was lowest for those who reported ‘always’ using condoms (adjusted relative risk, 0.68; P = 0.42; HIV incidence, 4.0) compared with those who reported ‘never’ using a condom (adjusted relative risk, 2.94; P < 0.001; HIV incidence, 14.0). Conclusion:Ongoing counseling and testing was positively associated with risk-reduction behaviors amongst a large proportion of men recruited for this study.

Alcohol and other substance use in STD clinic patients: relationships with STDs and prevalent HIV infection.

Background and Objectives Sexually transmitted diseases have been epidemiologically linked to a variety of high-risk behaviors, including substance abuse. However, the relationship between alcohol consumption and risk for specific STDs has not been explored previously. Goal of this Study We hypothesized that alcohol use is a risk factor for STD acquisition. Methods Cross-sectional analysis of enrollment visits in a prospective cohort study evaluating behavioral and biological risk factors for STD in an inner-city STD clinic population. Subjects were administered a comprehensive behavioral and questionnaire which included a detailed sexual history and evaluation for substance use. Alcohol users were classified as “frequent drinkers” (>2 times/week) and “infrequent” drinkers. Clinical exam included laboratory evaluation for syphilis, gonorrhea, chlamydia, and human immunodeficiency virus infection (HIV). Results In 1990–1992, 1,145 patients were enrolled; 245 (21%) were “frequent” drinkers. “Frequent” drinkers were more likely in the past month to have had more than 2 sex partners (OR = 3.3; 95% confidence interval [95%CI] 2.4–4.7), and use cocaine or injecting drugs. Frequent drinkers were more likely to be diagnosed with HIV (OR = 2.5; 95%CI 1.6–4.0) and syphilis (OR = 1.74,95%CI 1.1–2.8), but not with gonorrhea or chlamydia. Similar patterns were seen in cocaine and injecting drug users. In a multivariate analysis controlling for gender, syphilis, injecting drug, cocaine use, and sexual orientation, there was a borderline association between frequent alcohol use and HIV infection (OR = 1.63; 95%CI 0.95–2.8; P = .07). Conclusions Alcohol is frequently used in individuals at risk for STD, and is associated with other risk variables for HIV infection. Use of multiple drugs, including alcohol, is common. Nevertheless, these data suggest that alcohol use may have an independent behavioral effect that would increase the risk for HIV infection.

High rates of syphilis among STI patients are contributing to the spread of HIV-1 in India

Background: Recent syphilis outbreaks have raised concern regarding the potential enhancement of HIV transmission. The incidence of syphilis and its association with HIV-1 infection rates among a cohort of sexually transmitted infection (STI) clinic attendees was investigated. Methods: 2732 HIV-1 seronegative patients attending three STI and one gynaecology clinic, were enrolled from 1993–2000 in an ongoing prospective cohort study of acute HIV-1 infection in Pune, India. At screening and quarterly follow up visits, participants underwent HIV-1 risk reduction counselling, risk behaviour assessment and HIV/STI screening that included testing for serological evidence of syphilis by RPR with TPHA confirmation. Patients with genital ulcers were screened with dark field microscopy. Results: Among 2324 participants who were HIV-1 and RPR seronegative at baseline, 172 participants were found to have clinical or laboratory evidence of syphilis during follow up (5.4 per 100 person years, 95% CI 4.8 to 6.5 per 100 person years). Independent predictors of syphilis acquisition based on a Cox proportional hazards model included age less than 20 years, lack of formal education, earlier calendar year of follow up, and recent HIV-1 infection. Based on a median follow up time of 11 months, the incidence of HIV-1 was 5.8 per 100 person years (95% CI 5.0 to 6.6 per 100 person years). Using a Cox proportional hazards model to adjust for known HIV risk factors, the adjusted hazard ratio of HIV-1 infection associated with incident syphilis was 4.44 (95% CI 2.96 to 6.65; p<0.001). Conclusions: A high incidence rate of syphilis was observed among STI clinic attendees. The elevated risk of HIV-1 infection that was observed among participants with incident syphilis supports the hypothesis that syphilis enhances the sexual transmission of HIV-1 and highlights the importance of early diagnosis and treatment of syphilis.

HIV-1-specific cytolytic T-lymphocyte activity correlates with lower viral load, higher CD4 count, and CD8+CD38-DR- phenotype: Comparison of statistical methods for measurement

OBJECTIVES The objective of this study was to use novel statistical methods to determine the correlation between HIV-1-specific cytolytic T-lymphocyte (CTL) activity and HIV-1 plasma viral load, in a blinded study of HIV-infected patients at various stages of clinical disease. METHODS Peripheral blood mononuclear cells (PBMC) were collected and stored at enrollment and 2 weeks later, from 15 HIV-infected individuals who were receiving stable antiretroviral therapy for the previous 6 weeks and during the study period. HIV-1-specific CTL activity was measured using an antigen-specific PBMC in vitro stimulation method. Measurements of plasma viral load, as well as CD4+ and CD8+ T lymphocytes expressing T-cell activation markers (DR and CD38) were also performed at each time point. CTL activity was quantified using three separate statistical methods: area under the net HIV-specific lysis curve (AUC), lytic units (LU20), and linear regression (LR) of net HIV-specific lysis. RESULTS HIV-1 nef-, pol- and gag-specific CTL activity (AUC method) was significantly higher in subjects with a plasma viral load < or = 30,000 RNA copies/ml, than in those with viral load >30,000 RNA copies/ml. When plasma viral load was analyzed as a continuous variable, there was a strong correlation between higher CTL activity and lower viral load for nef (r2 = .77; p < .001), pol (r2 = .63; p < .001) and gag (r2 = 0.75; p < .001) targets by the AUC, but not for the LU20 analysis. Using the LR analysis, which is less dependent on in vitro PBMC growth than the AUC analysis, an independent association was demonstrated between nef- and gag-specific CTL activity and lower viral load. Measurement of CTL activity was also significantly correlated with a higher percentage of circulating CD8+DR-CD38- T lymphocytes. CONCLUSIONS In this blinded study using an in vitro stimulation of frozen PBMC, higher HIV-1 nef-, pol-, and gag-specific CTL activity correlated with lower plasma viral load, particularly in patients with a CD4 count <500 cells/mm3. Two new statistical methods for estimating CTL activity, AUC and LR analyses, were superior to the standard lytic unit (LU20) method for demonstrating this correlation. These data also demonstrated that higher circulating CD8+ T lymphocytes with a DR-CD38-phenotype, correlate with a lower plasma viral and load and higher HIV-specific CTL activity. This suggests that lymphocytes with this double-negative phenotype may include circulating HIV-specific CD8+ CTL.

Low carotenoid concentration and the risk of HIV seroconversion in Pune, India.

Summary: Low vitamin A and carotenoid levels could increase the risk of sexual HIV acquisition by altering the integrity of the genital epithelium or by immunologic dysfunction. We addressed this issue by measuring serum vitamin A and carotenoid levels in patients who were at risk of subsequent HIV infection. In a nested casecontrol study in individuals attending two sexually transmitted disease (STD) clinics in Pune, India, serum micronutrient levels were measured in 44 cases with documented HIV seroconversion (11 women and 33 men) and in STD patients matched for gender and length of follow‐up with no subsequent HIV seroconversion (controls). STD patients in Pune had low vitamin A and carotenoid levels, and low serum &bgr;‐carotene levels were independently associated with an increased risk of subsequent HIV seroconversion. STD patients with &bgr;‐carotene levels less than 0.075 &mgr;mol/L were 21 times more likely to acquire HIV infection than those with higher levels (adjusted odds ratio = 21.1;/p = .01). No such association was observed in case of other non‐provitamin A carotenoids. This study reports the first evidence of an association between low serum provitamin A carotenoid levels and an increased risk for heterosexual HIV acquisition in STD patients in Pune, India.

Definitions of genital ulcer disease and variation in risk for prevalent human immunodeficiency virus infection

Objectives: Although genital ulcer disease (GUD) has been associated with human immunodeficiency virus (HIV) infection in a number of studies, definitions of genital ulceration have varied. The authors hypothesized that the association of GUD with prevalent HIV infection may vary according to the definition of GUD that is used. Methods: As part of a prospective cohort study, 863 patients were interviewed and examined who presented to a sexually transmitted disease (STD) clinic for new symptom evaluation and who agreed to HIV testing to determine demographic and behavioral risk associated with prevalent HIV infection. To determine the association between GUD and prevalent HIV, the following definitions of GUD were used: observed ulcers, history of syphilis, serologic evidence of syphilis, observed culture‐proven genital herpes, and serologic evidence of herpes simplex virus type II (HSV‐2) infection. Results: Of 481 men and 382 women enrolled, prevalent HIV infection was detected in 12.5% and 5.2%, respectively. In multivariate analyses controlling for known HIV risk behaviors, prevalent HIV infection was associated with observed GUD (odds ratio [OR] = 2.0, 95% confidence intervals (CI) = 1.0–3.9), a history of syphilis (OR = 6.0, CI = 2.8–12.7), and serologic evidence of syphilis (OR = 3.7, CI = 1.9–7.0), but not with serologic evidence of HSV‐2 (OR = 1.2, CI = 0.7–2.1), nor with observed HSV‐2 culture‐positive genital ulcerations (OR = 1.0, CI = 0.4–4.2). Factors contributing to different strengths of association between HIV infection and a history of syphilis or serologic evidence of syphilis included the presence of under‐diagnosed syphilis infection in people with reactive serologic tests and the absence of serologic reactivity in people with a positive history. Conclusions: Although GUD is strongly associated with prevalent HIV, the strength of the association depends on the definition of GUD used. For accurate evaluation of people at risk for HIV, clinicians and researchers should use multiple definitions of GUD.