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.

Evidence for high prevalence & rapid transmission of HIV among individuals attending STD clinics in Pune, India.

Prevalence and incidence of HIV-1 infection among persons attending two STD clinics in Pune between May 1993 and October 1995 are reported. On screening 5321 persons, the overall prevalence of HIV-1 infection was found to be 21.2 per cent, being higher in females (32.3%) than in males (19.3%). Analysis of behavioural and biological factors showed that old age, sex work, lifetime number of sexual partners, receptive anal sex, lack of circumcision, genital diseases and lack of formal education were related to a higher HIV-1 seroprevalence. The observed incidence rate of 10.2 per cent per year was very high, much higher in women than in men (14.2% and 9.5% per year respectively) and over three times higher among the sex workers. Females in sex work, males having recent contacts with female sex workers (FSWs) and living away from the family and persons with previous or present genital diseases had a higher risk of seroconversion. Condom usage was shown to have a protective effect in seroprevalence and seroincidence analysis. With limited available resources and lack of a suitable vaccine or a drug, long-term prevention policy of creating awareness in the community must be supplemented by strengthening STD control measures and promotion of condom use and safe sex. Factors related to availability and utilization of condoms must be carefully investigated.

Risk factors for HIV infection in people attending clinics for sexually transmitted diseases in India.

OBJECTIVE: To investigate the risk factors for HIV infection in patients attending clinics for sexually transmitted diseases in India. DESIGN: Descriptive study of HIV serology, risk behaviour, and findings on physical examination. SUBJECTS: 2800 patients presenting to outpatient clinics between 13 May 1993 and 15 July 1994. SETTING: Two clinics and the National AIDS Research Institute, in Pune, Maharashtra State, India. MAIN OUTCOME MEASURE: HIV status, presence of sexually transmitted diseases, and sexual behaviour. RESULTS: The overall proportion of patients infected with HIV was 23.4% (655/2800); 34% (184) of the women and 21% (459) of the men were positive for HIV infection. Of the 560 women screened, 338 (60%) had a reported history of sex working, of whom 153 (45%) were infected with HIV-1. The prevalence of HIV-1 infection in the 222 women who were not sex workers was 14%. The significant independent characteristics associated with HIV infection based on a logistic regression analysis included being a female sex worker, sexual contact with a sex worker, lack of formal education, receptive anal sex in the previous three months, lack of condom use in the previous three months, current or previous genital ulcer or genital discharge, and a positive result of a Venereal Disease Research Laboratory test. CONCLUSIONS: In India the prevalence of HIV infection is alarmingly high among female sex workers and men attending clinics for sexually transmitted diseases, particularly in those who had recently had contact with sex workers. A high prevalence of HIV infection was also found in monogamous, married women presenting to the clinics who denied any history of sex working. The HIV epidemic in India is primarily due to heterosexual transmission of HIV-1 and, as in other countries, HIV infection is associated with ulcerative and non-ulcerative sexually transmitted diseases.

PIP: During May 1994-July 1995 in India, health workers at two clinics for sexually transmitted diseases (STDs) in Pune provided HIV serological screening and a physical examination to 2800 patients who completed a questionnaire, so that researchers could identify risk factors for HIV-1 infection in this high-risk population. 60% of the female STD patients were sex workers. 90% of the men had contact with a sex worker within the last 3 months. 14% of all STD patients were women who were not sex workers. 82% of these were married and monogamous. 13% of the married and monogamous women tested positive for HIV-1. Overall, 23.4% of all STD patients tested positive for HIV-1. Only 24% of all STD patients had used condoms during the last 3 months. The leading clinical diagnosis was chancroid (33% for men and 20% for women). The logistic regression analysis showed that the risk factors for HIV-1 infection were: being a sex worker (odds ratio [OR] = 3.7; p 0.001), contact with a sex worker (OR = 1.71; p = 0.01), receptive anal sex (OR = 3.52; p 0.001), tattooed after 1985 (OR = 1.31; p = 0.01), current or previous genital ulcer (OR = 1.29; p = 0.01), and a positive result of a Venereal Disease Research Laboratory test (OR = 1.33; p = 0.01). Protective factors were some formal education (OR = 0.76; p = 0.02), and condom use (sometimes/always, OR = 0.75; p = 0.03). These findings indicate a need for comprehensive and national STD services in India to control STDs and a health education campaign on HIV/AIDS to reduce high risk behavior.

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.

Rapid differentiation of the major causative agents of bacterial meningitis by use of frequency-pulsed electron capture gas-liquid chromatograph: analysis of acids.

The major causative agents of bacterial meningitis, Haemophilus influenzae serogroup B, Neisseria meningitidis serogroups B and C, Klebsiella pneumoniae, Streptococcus pneumoniae, and two types of Escherichia coli, were cultured in a modified chemically defined Catlin medium and in a commercial version of the unmodified Catlin medium. The spent media were extracted under acidic conditions, and electron-capturing derivatives were prepared by derivatization with trichloroethanol or haptafluorobutyric anhydride. The derivatives were analyzed on a gas chromatograph equipped with a frequency-pulsed electron capture detector and a PEP-2 computer. The data obtained from the study show that these organisms can be easily distinguished from each other on the basis of metabolic products detected in either type of medium. Three different metabolic groups were detected within two serogroups of N. meningitidis. The methods are practical, and the new technique should offer clinical laboratories and hospitals a better method for rapid identification of this important group of pathogens.

Rapid differentiation of the major causative agents of bacterial meningitis by use of frequency-pulsed electron capture gas-liquid chromatography: analysis of amines.

The major causative agents of bacterial meningitis (Haemophilus influenzae serogroup B, Neisseria meningitidis serogroups B and C, Klebsiella pneumoniae, Steptococcus pneumoniae, and two types of Escherichia coli) were cultured in a chemically defined medium, and selected strains were further studied in Todd-Hewitt medium. After acidic extraction of the spent media with chloroform, a basic extraction was made with chloroform to obtain amines. A third extraction was performed on re-acidified Todd-Hewitt medium with ethyl ether to obtain hydroxyacids. The extracts were derivatized with heptafluorobutyric anhydride-ethanol to form electron-capturing derivatives, and the derivatives were analyzed on a frequency-pulsed electron capture gas-liquid chromatograph (FPEC-GLC) equipped with a PEP-2 computer. The data obtained from the study showed that amines were produced by these organisms that formed characteristic patterns. Different serotypes of K. pneumoniae and the two serogroups of N. meningitidis produced different types of FPEC-GLC profiles within serotypes. E. coli produced several hydroxy acids on Todd-Hewitt medium that made it unique among the organisms studied. The methods used are practical and the techniques have potential for use in clinical laboratories and hospitals as a valuable aid for the rapid identification of the major causative agents of bacterial meningitis.

Treatment of necrolytic migratory erythema in glucagonoma syndrome.

The glucagonoma syndrome is characterized by elevated serum glucagon, a pancreatic alpha-cell tumor, anemia, hypoaminoacidemia, and necrolytic migratory erythema. Necrolytic migratory erythema may cause marked morbidity and is frequently misdiagnosed. A 42-year-old white woman with a 1 1/2-year history of refractory dermatitis (most severe on the lower extremities) had the glucagonoma syndrome. Her severe morbidity was markedly relieved with the administration of intravenous amino acids. This therapy was successful in controlling the necrolytic migratory erythema through recurrences after somatostatin (SMS 201-995), surgical debulking, and chemotherapy proved inadequate.

Gout masquerading as rheumatoid vasculitis.

A 30-year-old Mexican woman had rash, deep ulcerations of her lower extremities, and debilitating polyarthritis. Her disorder simulated rheumatoid vasculitis, but serum rheumatoid factor was absent. The diagnosis of gout was confirmed by uric acid crystals in joint fluid and skin biopsy specimens and by x-ray crystallography. The age and sex were unusual for a patient with gout, and she had none of the commonly associated metabolic defects. This unique presentation for urate arthropathy needs further study.

Metabolic analysis of serologically defined Neisseria meningitidis isolates by frequency-pulsed electron capture gas-liquid chromatography.

Eight serogroups [A, B, C, D, X, Y, 29E [Z'], and W135] along with nongroupable and nontypable strains of Neisseria meningitidis were cultured in a defined liquid medium. The whole-culture spent medium was extracted, derivatized, and analyzed by frequency-pulsed electron capture gas-liquid chromatography. The frequency-pulsed electron capture gas-liquid chromatography profiles were then used to group the organisms on the basis of their metabolic profiles. First, two basic groups were formed which consisted of elevated spent-medium compounds and reduced spent-medium compounds. Then, these two basic groups were further subdivided on the basis of metabolites and cellular fatty acids detected in the spent medium to form a total of seven metabolic groups. The nongroupable strains (with the exception of two strains) fell within one of these metabolic groups. The elevated spent-medium compounds group contained all of the A, B, Y, and Z' strains, and the reduced spent-medium compounds group contained most of the C and all of the W135 and D strains. The potential usefulness of the metabolic group scheme is discussed.

Synthetic peptides of human lysosomal cathepsin G with potent antipseudomonal activity.

Enzymatically active and inactive (diisopropylfluorophosphate-treated) cathepsin G exerted antibacterial action in vitro against Staphylococcus aureus, whereas only enzymatically active cathepsin G displayed bactericidal action against Pseudomonas aeruginosa. In order to further test the requirement for protease activity for the antipseudomonal action of cathepsin G, synthetic peptides spanning the full-length mature protein were prepared and examined for antibacterial action. Surprisingly, three structurally distinct peptides that correspond to residues 61 to 80, 117 to 136, and 198 to 223 within the full-length protein were found to exert potent antipseudomonal action (> 4.5 logs of killing at 500 micrograms/ml) against P. aeruginosa ATCC 27853 and four mucoid clinical isolates. Only the peptide (CG117-136) corresponding to residues 117 to 136 (117-RPGTLCTVAGWGRVSMRRGT-136) within cathepsin G exerted antibacterial action against the gram-positive pathogen S. aureus. The antipseudomonal action of CG117-136 was rapid and could be inhibited either by increasing concentrations of NaCl or by 0.5 mM MgCl2 plus 0.5 mM CaCl2, and these conditions appeared to reduce binding of the peptide to whole bacteria. Variants of peptide CG117-136 lacking either a hydrophobic N-terminal domain or a positively charged C-terminal domain were found to have significantly less antipseudomonal action than CG117-136. The antibacterial capacity of the all-D-enantiomeric form of peptide CG117-136 was found to be identical to that of the all-L-peptide, suggesting that the mechanism of killing does not require the recognition of a target site possessing a chiral center.

Thoracic pedicle screw placement guided by computed tomographic measurements.

Cadaveric pedicle screw placement guided by the measurements from axial computed tomography (CT) scans in the thoracic spine was assessed in this study. Axial CT scans were performed on four cadaveric thoracic spines, and the measurements included the pedicle transverse angle, inner pedicle width, and distance between the midline of the vertebra and the pedicle axis on the dorsal aspect of the lamina. With utilization of the data from CT scans, screws were directly placed into the thoracic pedicle from T1 to T10. Screw penetration of the pedicle was determined by gross examination. The results showed that the largest pedicle transverse angle was found at the levels of T1-2, and the smallest occurred at the T3 through T8 levels. The value of the pedicle inner width was quite different between specimens with a minimum of 3.0 mm at T4 and a maximum of 9.2 mm at T10. Gross examination of the pedicle showed that 13 (16.3%) of 80 screws penetrated the pedicle wall, with a Grade I penetration in 11 pedicles and a Grade II penetration in 2 pedicles. Screw penetration of the medial wall was found in four pedicles and penetration of the lateral wall was noted in nine pedicles. No screw penetration of the superior and inferior walls of the pedicle was identified in any of the four specimens. Thoracic pedicle screw placement guided by the measurements from axial CT scans significantly reduced the incidence of pedicle penetration. Axial CT measurements of the pedicle inner diameter and transverse angle as well as the starting point for screw insertion are recommended if pedicle screw fixation is intended in the thoracic spine.

Evaluation of an enzyme immunoassay for detection of Chlamydia trachomatis in urine of asymptomatic men.

In a study of 1,486 men attending two sexually transmitted disease clinics, of whom 891 had no symptoms of urethritis, we compared an enzyme immunoassay (EIA) (Baxter-Bartels, formerly Northumbria AntigEnz) of urine sediment to urethral culture for the detection of Chlamydia trachomatis. C. trachomatis prevalence by culture alone was 7.7% in asymptomatic men and 10.9% in symptomatic men. Discrepant results between EIA of urine and urethral culture were evaluated by direct fluorescent-antibody staining (DFA) for elementary bodies in urine sediment or in culture transport media. When chlamydial infection was defined as either a positive urethral culture or positive EIA confirmed by DFA, chlamydia prevalence increased to 8.9% in asymptomatic men and 11.6% in symptomatic men. The urine EIA sensitivity, specificity, and positive and negative predictive values for chlamydia detection in asymptomatic men were 84.8, 99.3, 91.8, and 98.5%, respectively, with nearly identical results for symptomatic men. The sensitivities of urethral culture alone compared with the combination of urethral culture and urine EIA (with DFA confirmation) were 87.3 and 94.3% for asymptomatic and symptomatic men, respectively. The present EIA of urine sediment is both highly sensitive and specific for the detection of C. trachomatis in asymptomatic men, thus providing a noninvasive screening method for chlamydia infection in asymptomatic men attending sexually transmitted disease clinics.

Impact of Chlorine and Heat on the Survival of Hartmannella vermiformis and Subsequent Growth of Legionella pneumophila.

Hartmannella vermiformis, a common amoebal inhabitant of potable-water systems, supports intracellular multiplication of Legionella pneumophila and is probably important in the transportation and amplification of legionellae within these systems. To provide a practical guide for decontamination of potable-water systems, we assessed the chlorine and heat resistance of H. vermiformis. H. vermiformis cysts and trophozoites were treated independently with chlorine at concentrations of 2.0 to 10.0 ppm for 30 min and then cocultured with L. pneumophila. Both cysts and trophozoites were sensitive to concentrations between 2.0 and 4.0 ppm and above (trophozoites somewhat more so than cysts), and 10.0 ppm was lethal to both forms. Hartmannellae treated with chlorine up to a concentration of 4.0 ppm supported the growth of legionellae. To determine whether heat would be an effective addendum to chlorine treatment of amoebae, hartmannellae were subjected to temperatures of 55 and 60 degrees C for 30 min and alternatively to 50 degrees C followed by treatment with chlorine at a concentration of 2 ppm. Fewer than 0.05% of the amoebae survived treatment at 55 degrees C, and there were no survivors at 60 degrees C. Pretreatment at 50 degrees C appeared to make hartmannella cysts more susceptible to chlorine but did not further reduce the concentration of trophozoites.

Risk factors and clinical presentation of acute primary HIV infection in India.

CONTEXT: Most previous studies of clinical presentation and risk factors in early human immunodeficiency virus (HIV) infection have relied on retrospective analyses and referred seroconverters, and thus were subject to possible bias. OBJECTIVES: To apply a method based on measurement of prevalent HIV-1 p24 antigenemia for identification of risk factors for newly acquired HIV infection and to describe the signs and symptoms of acute HIV infection. DESIGN AND SETTING: Nested case-control study in Pune, India. PARTICIPANTS: HIV antibody-negative persons attending 2 sexually transmitted disease (STD) clinics between May 1993 and June 1996. OUTCOME MEASURES: Prevalent p24 antigenemia, risk factors for HIV infection, and clinical symptoms of acute primary HIV infection. RESULTS: Of 3874 HIV antibody-negative persons tested, 58 (1.5%) were p24 antigen positive at initial presentation to the clinics. Unprotected sexual contact with a commercial sex worker (CSW) was reported by 39 (77%) of the 51 p24 antigenemic men, compared with 131 (51 %) of 255 control men (adjusted odds ratio [AOR], 3.4; 95% confidence interval [CI], 1.2-9.6; P=.02). The presence of an active genital ulcer at the time of screening was found in 46 (79%) of the 58 p24 antigenemic men and women, compared with 137 (47%) of the 290 control subjects (AOR, 4.2; 95% CI, 2.0-9.0; P<.001). Signs and symptoms independently associated with p24 antigenemia in HIV antibody-seronegative persons included fever, which was reported by 28 (48%) of the 58 p24 antigenemic subjects, but only 52 (18%) of the 290 control subjects (AOR, 4.7; 95% CI, 2.4-9.0; P<.001). Joint pain was reported by 10% of subjects recently HIV infected, compared with 2% of the control subjects (AOR, 6.5; 95% CI, 1.7-24.8; P=.006). Night sweats were reported by 9% of the p24 antigenemic, but only 1% of the control subjects (AOR, 9.1; 95% CI, 1.7-47.6; P=.009). Overall, fever, joint pain, and/or night sweats were reported in 27 (47%) of the 58 subjects with recent HIV infection. CONCLUSIONS: This systematic case-control study of p24 antigen screening in HIV-seronegative patients attending STD clinics in India identified unprotected sex with a CSW and a genital ulcer as independent risk factors associated with newly acquired HIV infection. In addition, p24 antigen positivity identified recent fever, night sweats, and arthralgias as symptoms that may be predictive of recent HIV infection. In a study of patients attending STD clinics in India, screening for p24 antigen in HIV antibody-negative persons was found to be a reliable and effective research method for determining recent risk behavior and identifying clinical signs of acute primary HIV infection.

Spread of HIV infection in married monogamous women in India.

CONTEXT: A high prevalence of human immunodeficiency virus (HIV) infection in female sex workers (FSWs) and men who attend sexually transmitted disease (STD) clinics poses a risk for spread of infection to other populations. OBJECTIVE: To examine spread of HIV to a low-risk population by comparing prevalence of, and risk factors for, HIV and STDs in FSWs and non-FSWs. METHODS: Women attending STD clinics in Pune, India, were assessed for STDs and HIV from May 13, 1993, to July 11, 1996. Demographic and behavioral information was collected, and clinical and laboratory assessment was performed. MAIN OUTCOME MEASURE: Prevalence and risk determinants of HIV infection. RESULTS: Of 916 women enrolled, 525 were FSWs and 391 were non-FSWs. Prevalence of HIV in FSWs and non-FSWs was 49.9% and 13.6%, respectively (P<.001). In multivariate analysis, inconsistent condom use and genital ulcer disease or genital warts were associated with prevalent HIV in FSWs. History of sexual contact with a partner with an STD was associated with HIV in non-FSWs. CONCLUSIONS: Infection with HIV is increasing in non-FSWs, previously thought to be at low risk in India. Since history of sexual contact with their only sex partner was the only risk factor significantly associated with HIV infection, it is likely that these women are being infected by their spouses. This underscores the need for strengthening partner-notification strategies and counseling facilities in India.

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

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 case-control 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 beta-carotene levels were independently associated with an increased risk of subsequent HIV seroconversion. STD patients with beta-carotene levels less than 0.075 micromol/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.

Incidence and predictors of human immunodeficiency virus type 1 seroconversion in patients attending sexually transmitted disease clinics in India.

The first estimates of the seroincidence of human immunodeficiency virus type 1 (HIV-1) and of the risk factors for seroconversion in a cohort of high-risk patients attending sexually transmitted disease (STD) clinics in India are reported. Between 1993 and 1995, 851 HIV-1-seronegative persons were evaluated prospectively every 3 months for HIV infection and biologic and behavioral characteristics. The overall incidence of HIV-1 was 10.2/100 person-years (95% confidence interval, 7.9-13.1). The incidence among commercial sex workers (CSWs) was 26.1/100 person-years, compared with 8.4 among non-CSWs. Recurrent genital ulcer disease and urethritis or cervicitis during the follow-up period were independently associated with a 7- (P < .001) and 3-fold (P = .06) increased risk of HIV-1 seroconversion, respectively. Because of the association of recurrent ulcerative and nonulcerative STDs with HIV-1 seroconversion in this setting, behavioral and biologic measures directed at the prevention and control of STDs would be expected to greatly reduce the transmission of HIV-1 infection in similar high-risk groups.

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.

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.

PIP: The etiology of genital ulcer disease (GUD) and the relationship between GUD and HIV infection were investigated in 302 patients presenting to a sexually transmitted disease clinic in Pune, India, in a 3-month period in 1994. Swabs of each genital ulcer were tested for herpes simplex virus (HSV) antigen by enzyme immunoassay and processed in a multiplex polymerase chain reaction (M-PCR) assay for simultaneous detection of HSV, Treponema pallidum, and Haemophilus ducreyi. The seroprevalence of HIV in this series was 22.2%. Clinical diagnosis of GUD was undermined when HIV infection was present. The etiology of GUD according to M-PCR was HSV in 26%, chancroid in 23%, primary syphilis in 10%, and multiple infections in 7%; no etiology could be identified in the remaining 34% of cases. Attempts to differentiate the etiology of GUD based solely on clinical grounds resulted in many inaccurate diagnoses. Chancroid was the most common clinical diagnosis (40%), followed by HSV (24%), syphilis (20%), and multiple infections (3%). HIV seroprevalence was significantly higher in patients with HSV compared with other etiologies (odds ratio, 2.1; 95% confidence interval, 1.2-3.7), presumably as a result of HIV-induced immunosuppression and consequent HSV reactivation. Until rapid, inexpensive, and sensitive assays become available, syndromic treatment with antibiotics should be provided to patients with GUD in order to reduce the risk of acquiring HIV infection.