Secondary Logo

Journal Logo

Article

Use of a Glycoprotein G-Based Type-Specific Assay to Detect Antibodies to Herpes Simplex Virus Type 2 Among Persons Attending Sexually Transmitted Disease Clinics

WHITTINGTON, WILLIAM L. H. AB*‡; CELUM, CONNIE L. MDMPH,*‡; CENT, ANNE MS† AND; ASHLEY, RHODA L. PhD

Author Information
  • Free

Abstract

GENITAL HERPES, most often caused by herpes simplex virus type 2 (HSV-2), is one of the most prevalent sexually transmitted diseases (STDs). The seroprevalence of HSV-2 infection in the United States has increased 30% between 1979 and 1990, and 22% of adults are now estimated to be infected. 1 Although the spectrum of disease caused by HSV-2 is broad, the majority of HSV-2 genital infections are subclinical and unrecognized because of the mild, atypical, or nonexistent nature of symptoms. 1,2 However, approximately 60% of persons who are HSV-2 seropositive and without a history of genital herpes are able to identify symptoms of genital herpes after receiving symptom-recognition counseling. 2 More than 80% of asymptomatic HSV-2–seropositive women shed HSV intermittently from the genital tract. 3 Furthermore, HSV-2 is often acquired from persons with such unrecognized infections. 4,5

In the absence of an effective HSV-2 vaccine, prevention strategies may potentially include counseling about symptom recognition, asymptomatic HSV-2 shedding, and the use of antiviral agents to reduce infectivity. 6–9 A randomized trial is currently assessing the effect of daily suppressive antiviral therapy on HSV-2 shedding and transmission to uninfected partners. To be effective, a prevention strategy that includes counseling or suppressive treatment would necessitate the identification of persons with subclinical HSV-2 infection. Therefore, serologic testing for HSV-2 infection is likely to be a central component of genital herpes prevention. 7,10

A recent study suggested that most patients attending STD clinics are interested in knowing their HSV-2 serostatus. 11 However, identification of such persons has been limited by the absence of accurate and affordable tests to detect HSV type-specific antibodies. 12 Reliable assays have been available only through specialized research laboratories, and when available their cost has been prohibitive for widespread screening purposes (up to US $126 for an HSV Western blot). Recently developed type-specific enzyme-linked immunosorbent assays (ELISA) have shown promise 12,13 and are likely to be much less expensive than HSV Western blot tests. In this study, we assessed the rates of HSV-2 antibody test acceptance with and without a charge for testing. We also further defined the performance characteristics of the Gull/Meridian HSV-2 and HSV type 1 (HSV-1) type-specific glycoprotein G (gG) assays relative to Western blot analysis in serum specimens prospectively collected from persons attending STD clinics.

Methods

Study Population and Design

Herpes serologic screening was offered to clients attending the Public Health, Seattle and King County STD clinics during two periods. From January to April 1998, the Gull/Meridian HSV-1 and HSV-2 gG ELISA were offered with Western blot confirmation of reactive ELISA results at a charge of US $15 per patient. From June to July 1998, the gG ELISA and HSV Western blot assays were offered without charge as part of a research protocol to further assess performance characteristics of the gG-based ELISA relative to Western blot analysis. 13 For both periods, demographic, behavioral, laboratory, and clinical data were obtained from the computerized STD clinic record. Clinicians approached patients in a similar manner during both study periods with a brief script that described the Gull/Meridian assay and the benefits of knowing one’s HSV-2 serostatus. However, during the second period, informed consent was obtained to allow for the collection of additional data about history of signs or symptoms compatible with genital and oral herpes and regarding the repeat testing of persons with initial atypical Western blot results or unresolved equivocal ELISA results at 4 to 6 weeks. The study protocol was reviewed and approved by the University of Washington Human Subjects Review Committee.

Laboratory Methods

Sera were tested at the University of Washington Diagnostic Virology Laboratory. Serum specimens were stored at 2 °C to 8 °C before testing. The Gull/Meridian HSV-2 and HSV-1 type-specific gG assays were provided by the manufacturer (Gull/Meridian, Salt Lake City, UT) and were performed according to the manufacturer’s instructions. 13 The University of Washington Western blot HSV assay was performed as previously described. 14

The testing of samples initially reported as equivocal by the gG ELISA was repeated in accordance with the manufacturer’s instructions; results of repeat tests were used for all analyses. For persons with atypical Western blot results or unresolved equivocal ELISA results who returned for repeat testing during the research study, the Western blot results from the second sample were used for analysis. For the small number of remaining specimens with atypical Western blot results that were not resolved by repeat testing, atypical Western blots results were considered negative.

Statistical Methods

Sensitivity, specificity, positive predictive value, and negative predictive value (with 95% CI) of the gG ELISA were calculated by established methods, 15 with results of the University of Washington HSV Western blot analysis used as the reference standard. 14 Frequencies of dichotomous variables were compared using the chi-square test; continuous variables were compared by parametric or nonparametric tests as appropriate. Logistic regression techniques were used to control for covariates in analyses of correlates of HSV-2 seropositivity and HSV-2 test acceptance.

Results

HSV-2 Serologic Test Acceptance

During the 4 months in which HSV antibody tests were offered to STD clinic attendees for a fee of $15, 558 of 3099 persons (18.0%) accepted HSV-2 serologic testing (Table 1). In comparison, HSV testing was accepted by 756 of 1477 persons (51.6%) seen during the 2-month research study when HSV-2 serologies were offered at no cost (P < 0.0001). Both men and women, all age and racial groups, heterosexual and gay or bisexual men, and persons with and without bacterial STDs were more likely to be tested when HSV-2 testing was free during the research period compared to fee-for-service testing (P < 0.01, each group). In multivariate analyses, acceptance of HSV serologic testing was associated with free testing (odds ratio, 7.5; 95% CI, 6.0–9.9) and increasing age (Table 2). Persons of color were less likely to request testing (odds ratio, 0.3; 95% CI, 0.27–0.40; for comparison of blacks with whites).

T1-7
Table 1:
Characteristics of Patients Accepting HSV Serologic Testing at the STD Clinic: Comparison of a $15/Test Charge (January–April 1998) With Free Testing (June–July 1998)
T2-7
Table 2:
Multivariate Correlates of HSV Serologic Test Acceptance by STD Clinic Attendees

HSV-1 and HSV-2 gG ELISA Performance

During the research study, 759 persons agreed to participate. Because of the mislabeling of two specimens and the inadvertent dual enrollment of one person, three specimens were excluded from analyses. By Western blot analysis, HSV-2 prevalence was 29.9% (226 of 756 persons). Compared with HSV-2 Western blot analysis, sensitivity of the HSV-2 gG ELISA was 80.5% (95% CI, 74.1–85.7%) and specificity was 98.5% (95% CI, 96.9–99.3%). Positive and negative predictive values of the HSV-2 gG ELISA were 95.8% and 92.2%, respectively (Table 3). By Western blot analysis, HSV-1 prevalence was 55.8% (422 of 756 persons). Compared with HSV-1 Western blot analysis, sensitivity of the HSV-1 gG ELISA was 91.0% (95% CI, 87.7–93.5%) and specificity was 96.4% (95% CI, 93.6–98.0%). Positive and negative predictive values of the HSV-1 gG ELISA were 97.0% and 89.4%, respectively (Table 3). Apparent false-positive and false-negative gG ELISA results were not associated with a history of genital or oral lesions, current lesions, gender, or age of the participant (data not shown).

T3-7
Table 3:
Comparison of gG ELISA With Western Blot Analysis

Characteristics of HSV-2–Seropositive Persons

During the research period, among participants without a history of genital herpes, HSV-2 seroprevalence was higher among women (35.0%) than among men (21.4%) and among blacks (44.8%) than among whites (20.6%), and HSV-2 seroprevalence was greater in persons 30 years or older (P < 0.01 for each comparison) (Table 4). There was no significant difference in HSV-2 seropositivity by sexual orientation among men, median number of sexual partners in the past 2 months or 12 months, or diagnosis of gonorrhea or chlamydia at that clinic visit. In multivariate analysis, female gender (odds ratio, 3.3; 95% CI, 2.1–5.3), black race (odds ratio, 3.7; 95% CI, 2.2–6.0), and older age (30–39 years, odds ratio 3.2; ≥ 40 years, odds ratio 3.0 relative to < 30 years) were independently associated with HSV-2 infection.

T4-7
Table 4:
HSV-2 Infection by Selected Characteristics Among Men and Women Without History of Genital Herpes—Seattle STD Clinics (1998)

Discussion

Among persons attending Seattle STD clinics, HSV antibody testing was accepted by approximately half of patients during a research study that provided testing without charge. However, persons of color and younger persons were less likely to accept testing. Possible explanations for lower rates of test acceptance by these groups include social and psychological factors associated with less interest in HSV-2 testing among blacks and youth, economic barriers for which young age and minority status are proxy measures, and perceived risk of HSV-2 infection by the patient or the clinician who offered testing. Data regarding reasons that almost half of STD clinic attendees did not accept free HSV-2 serologic testing are not available. Optimal approaches and effective patient messages about genital herpes epidemiology and serologic testing should be the focus of future studies.

During the research study, HSV-1 and HSV-2 infections were identified in 55.8% and 29.9% of tested persons, respectively, by Western blot analysis, which is somewhat higher then HSV-2 seroprevalence in the general US adult population. 1 In this high HSV-1 and HSV-2 prevalence population, the positive predictive value of the Gull/Meridian ELISA for HSV-1 was 97.0% (95% CI, 94.6–98.4%) and the positive predictive value for HSV-2 was 95.8% (95% CI, 91.6–98.0%). Apparent false-positive Gull/Meridian HSV-1 and HSV-2 ELISA results were not associated with history of clinical disease (oral or genital herpes) or age of the patient. Positive predictive values of 95% or greater for both the HSV-1 and HSV-2 ELISA suggest that STD clinic patients with a reactive HSV-1 or HSV-2 Gull/Meridian ELISA assay can be counseled that there is a very high likelihood that they have been infected with HSV-1 or HSV-2, respectively, even without Western blot confirmation.

The negative predictive values for HSV-1 and HSV-2 were 89.4% (95% CI, 85.7–92.3%) and 92.2% (95% CI, 89.6–94.2%), respectively. Apparent false-negative Gull/Meridian ELISA results were not associated with a previous diagnosis of oral or genital herpes or compatible symptoms, patient age, or current clinical presentation (e.g., primary HSV infection). Previous studies of the Gull/Meridian ELISA have indicated that it may take as long as several months for detection of antibodies during seroconversion. 15 Therefore, persons with recent history of symptoms suggestive of HSV or known recent exposure to HSV should be counseled that subsequent repeat testing is indicated. However, in the absence of a history of possible primary HSV or recent HSV exposure, negative predictive values of 89% or greater appear adequate for a screening test.

A recent population-based seroprevalence study reported an overall HSV-2 prevalence of 22% with increased prevalence of HSV-2 associated with older age, minority ethnicity, and female gender, the same correlates found in the current STD clinic population. 1 Given the ability to reliably screen for HSV-2 antibodies, the relatively high HSV-2 prevalence in the general population, and interest in and potential benefits of HSV-2 serologic testing, 11 it is likely that affordable, type-specific HSV serologic tests would be used by primary care providers and be acceptable to patients. However, this study indicates that there is price sensitivity about the cost of HSV-2 serologic testing, at least among low-income STD clients; acceptance of testing among STD clinic clients increased from 18% during the period in which a modest fee ($15) was charged to 52% during the research period in which the Gull/Meridian HSV gG ELISA with confirmatory Western blot testing was offered for no charge. However, these findings about price sensitivity of HSV-2 serologic testing must be cautiously interpreted in the context of a small proportion of Seattle STD clinic clients being charged for other STD services.

Now that the Gull/Meridian HSV gG ELISA and other type-specific serologic tests are becoming more widely available, clinicians must be prepared to interpret test results and provide counseling about HSV-2 infection. Indications for HSV-2 serologic testing may include pregnancy and, if these women are seronegative for HSV-2, counseling and HSV serologic testing of their partners. Discordant couples in which the male partner is HSV-2 seropositive and the pregnant woman is HSV-2 seronegative should be counseled about safe sex to avoid the acquisition of HSV-2, particularly in the third trimester, to reduce the risk of neonatal HSV after primary HSV in the mother. 16 A second group for possible HSV-2 serologic screening would be HIV-infected persons without a history of anogenital herpes, particularly those with lower CD4 counts (e.g., < 200) among whom recurrence rates and HSV shedding are more frequent. 17,18 The recent findings of high HIV viral copy numbers in swab specimens obtained from HIV-positive men with genital HSV lesions 19 indicate that counseling about HSV-2 symptom recognition, asymptomatic shedding, risk reduction, and potentially antiviral suppressive therapy may help reduce the transmission of HSV and HIV by sexually active persons with HIV and HSV-2. However, the performance of the Gull/Meridian ELISA has not been assessed in persons with HIV, and previous work has suggested that HSV-2 serologic test performance may be less satisfactory in such populations. 20

The limitations of this study include insufficient data about factors related to HSV-2 antibody test acceptance. This study was not designed to develop and validate selective screening criteria or to study the sensitivity of the Gull/Meridian HSV gG ELISA in persons with primary genital HSV infection. Finally, we did not enroll persons with known HIV infection and, therefore, cannot assess the sensitivity and specificity of the Gull/Meridian HSV gG ELISA in HIV immunodeficiency.

In summary, selective HSV-2 serologic screening of high-prevalence populations, such as STD clinic attendees, may identify a considerable number of persons with unrecognized genital HSV-2 infection. The performance of the Gull/Meridian HSV-1 and HSV-2 ELISA was sufficient to recommend its use for screening in high-prevalence populations, assuming that quality assurance and performance characteristics of the test are maintained during routine manufacture. However, selective screening criteria, counseling messages, impact of HSV-2 testing on quality of life and sexual behavior, and effective prevention strategies need to be identified and validated before widespread HSV serologic screening programs are implemented. Lastly, the cost and impact of such programs need to be assessed relative to other clinical and public health priorities.

References

1. Fleming DT, McQuillan GM, Johnson RE, et al. Herpes simplex virus type 2 in the United States, 1976 to 1994. N Engl J Med 1997; 337: 1105–1111.
2. Langenberg A, Benedetti J, Jenkins J, et al. Development of clinically recognizable genital lesions among women previously identified as having “asymptomatic” herpes simplex virus type 2 infection. Ann Intern Med 1989; 110: 882–887.
3. Wald A, Zeh J, Selke S, et al. Reactivation of genital herpes simplex virus type 2 infection in asymptomatic seropositive persons. N Engl J Med 2000; 342: 844–850.
4. Mertz GJ, Benedetti J, Ashley R, et al. Risk factors for the sexual transmission of genital herpes. Ann Intern Med 1992; 116: 197–202.
5. Mertz GJ, Schmidt O, Jourden JL, et al. Frequency of acquisition of first-episode genital infection with herpes simplex virus from symptomatic and asymptomatic source contacts. Sex Transm Dis 1985; 12: 33–39.
6. Whittington WL, Cates WJ. Acyclovir therapy for genital herpes: enthusiasm and caution in equal doses. JAMA 1984; 251: 2116–2117.
7. Corey L. Raising the consciousness for identifying and controlling viral STDs: Fears and frustrations. Sex Transm Dis 1997; 25: 58–69.
8. Blower SM, Porco TC, Darby G. Predicting and preventing the emergence of antiviral drug resistance in HSV-2. Nat Med 1998; 4: 673–678.
9. Wald A, Zeh J, Barnum G, et al. Suppression of subclinical shedding of herpes simplex virus type 2 with acyclovir. Ann Intern Med 1996; 124: 8–15.
10. Barton SE. The practical application of serological testing for HSV infection. Herpes 1998; 5: 39–41.
11. Fairley T, Monteiro EF. Patient attitudes to type specific serological tests in the diagnosis of genital herpes. Genitour Med 1997; 73: 259–262.
12. Ashley, RL. Type-specific antibodies to HSV-1 and-2: review of methodology. Herpes 1998; 5: 33–38.
13. Ashley RL, Wu L, Pickering JW, Tu MC, Schnorenberg L. Premarket evaluation of a commercial glycoprotein G-based enzyme immunoassay for herpes simplex virus type-specific antibodies. J Clin Microbiol 1998; 36: 294–295.
14. Ashley RL, Militoni J, Lee F, et al. Comparison of Western blot (immunoblot) and glycoprotein G-specific immunodot enzyme assay for detecting antibodies to herpes simplex virus types 1 & 2 in human sera. J Clin Microbiol 1988; 26: 662–667.
15. Hennekens CH. Epidemiology in Medicine. 1st ed. New York: Little, Brown, 1987: 331.
16. Brown ZA, Selke S, Zeh J, et al. The acquisition of herpes simplex virus during pregnancy. N Engl J Med 1997; 337: 509–515.
17. Schacker T, Zeh J, Hu HL, Hill E, Corey L. Frequency of symptomatic and asymptomatic herpes simplex virus type 2 reactivations among human immunodeficiency virus-infected men. J Infect Dis 1998; 178: 1616–1622.
18. Rompalo AM, Astemborski J, Schoenbaum E, et al. Comparison of clinical manifestations of HIV infection among women by risk group, CD4+ cell count, and HIV-1 plasma viral load. J Acquir Immun Defic Syndr Hum Retrovirol 1999; 20: 448–454.
19. Schacker T, Ryncarz AJ, Goddard J, et al. Frequent recovery of HIV-1 from genital herpes simplex virus lesions in HIV-1 infected men. JAMA 1998; 280: 61–66.
20. Safirin S, Arvin A, Mills J, Ashley R. Comparison of the Western immunoblot assay and a glycoprotein G enzyme immunoassay for detection of serum antibodies to herpes simplex virus type 2 in patients with AIDS. J Clin Microbiol 1992; 30: 1312–1314.
© Copyright 2001 American Sexually Transmitted Diseases Association