Herpes Simplex Virus Seroprevalence and Risk Factors in 2 Canadian Sexually Transmitted Disease Clinics

Singh, Ameeta E. BMBS, MSc*†; Romanowski, Barbara MD*; Wong, Tom MD, MPH‡; Gourishankar, Sita MD, MSc*; Myziuk, Linda MLT*; Fenton, Jayne MLT§; Preiksaitis, Jutta K. MD*§

Sexually Transmitted Diseases:

Objectives: The objectives of this study were to determine the seroprevalence and risk factors for herpes simplex virus (HSV) types 1 and 2 in patients attending 2 Canadian sexually transmitted disease (STD) clinics.

Study: Stored sera were tested for the presence of IgG class antibodies to HSV-1 and HSV-2 and results linked to that obtained from a risk behavior questionnaire.

Results: Overall prevalences for HSV-1 and -2 were 56% and 19%, respectively. HSV-1 and -2 seropositivity was associated with increasing age, female gender, nonwhite ethnicity, and a history of STD. HSV-2 seropositivity was also associated with a history of genital herpes, presence of genital sores, and coinfection with either human immunodeficiency virus (HIV) or hepatitis C (HCV).

Conclusions: Herpes simplex infection is common in this high-risk Canadian population. Our finding that HCV seropositivity was a significant predictor for HSV-2 seropositivity emphasizes the overlap between pathogens that are primarily thought to be bloodborne pathogens and sexually transmitted infections and the need to target prevention in these areas concurrently.

In Brief

This study in patients attending 2 Canadian sexually transmitted disease clinics showed that a number of factors were associated with herpes simplex virus (HSV) type 1 and 2 seropositivity and emphasized the association between hepatitis C, a pathogen which is usually bloodborne, and HSV-2, a sexually transmitted pathogen.

Author Information

From the * University of Alberta, Alberta, Canada; † Alberta Health and Wellness, Capital Health STD Centre, Alberta, Canada; the ‡ Universities of Ottawa and Toronto, Health Canada, Ottawa, Canada; and the § Provincial Laboratory for Public Health, Alberta, Canada

The authors thank E. Francis Cook, ScD, for review of statistical analysis and manuscript and the staff of Edmonton and Calgary STD clinics for facilitating data collection.

Correspondence: Ameeta E. Singh, BMBS, MSc, 23rd Floor, Telus Plaza North Tower, P.O. Box 1360 Stn Main, 10025 Jasper Avenue, Edmonton AB T6J 2C3, Alberta, Canada. E-mail: ameeta.singh@gov.ab.ca.

Received for publication March 1, 2004, and accepted August 12, 2004.

Article Outline

HERPES SIMPLEX INFECTIONS MAY be the result of herpes simplex virus (HSV) types 1 or 2.1 HSV-2 may cause genital or orolabial vesicular and ulcerative lesions in adults and severe systemic disease in neonates and immunocompromised patients. In addition to the significant morbidity in adults, and also mortality in neonates, genital herpes may result in significant psychologic distress. Genital ulcers secondary to HSV-2 may also facilitate the acquisition or transmission of human immunodeficiency virus (HIV)2–8 Although both viruses may cause either genital or oropharyngeal infection,9–11 most infections with HSV-2 affect the genitals, whereas HSV-1 commonly causes oropharyngeal infection.1 It is estimated that between 17% and 78% of genital herpes is the result of HSV-1.9–12

Clinical diagnosis and viral isolation alone significantly underestimate the prevalence of infection because the majority of persons infected with HSV are asymptomatic.13–18 Seroprevalence studies are therefore the only accurate way to determine the level of infection in a particular community. Two U.S. studies showed that HSV-2 seroprevalence rose a dramatic 30% from 16.4% in 1976–1980 to 21.7% between 1988 and 1994 in persons aged 12 or older.19,20 The seroprevalence of HSV-2 antibodies varied from <1% in persons under 15 years of age to 20% to 60% over the age of 30.19 Furthermore, seroprevalence varied not only by age, but also gender and ethnicity, with higher rates in women and persons of nonwhite origin.19,21 Not surprisingly, seroprevalence rates have been demonstrated to vary with the population studied, with higher rates in persons attending sexually transmitted disease (STD) clinics.15,22 Worldwide seroprevalence rates of HSV-2 infection are variable, with lower reported rates in Europe and Australia1 and seroprevalence rates of 61% to 95% in high-risk populations in developing countries,.1

Significant progress in our understanding of HSV-1 and -2 infection has occurred over the last decade, in part related to the development of sensitive and specific HSV type-specific laboratory tests. Very few HSV seroprevalence studies have been performed in Canada. One small Ontario study in a small general population sample23 carried out between 1978 and 1980 assessed HSV-2 seroprevalence and associated sexual and socioeconomic factors. This study showed a higher seroprevalence rate in females, those of lower socioeconomic status, younger age at first sexual intercourse, those with multiple sexual partners, and individuals who do not own their own residence. A recent study of 1215 antenatal women in British Columbia demonstrated an age-adjusted seroprevalence rate of 60% for HSV-1 and 17% for HSV-2.24 However, little is known regarding HSV seroprevalence or its associated factors in STD clients in the current era.

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Materials and Methods


Ethics approval for the study was obtained from the Heath Research Ethics Board at the University of Alberta. We performed an anonymous, unlinked cross-sectional study evaluating the seroprevalence of HSV-1 and -2 and associated risk factors in persons attending the 2 main STD clinics in Calgary and Edmonton (Alberta, Canada) between May 1994 and May 1995. The clinics provide free, confidential medical care, testing, and treatment for approximately 20% of all STD cases reported in Alberta. Serologic testing was conducted between January and March 2001 on leftover stored sera from 7266 samples. All patients who had blood drawn for syphilis, hepatitis, or HIV between those dates at the 2 clinics and had sufficient remaining sera for testing were eligible for inclusion into the study. Individuals were invited to participate in the study only once.

Information from a risk behavior questionnaire was obtained during the original study,25 and informed consent was obtained before administration of the questionnaires. The following information was included in the questionnaires and were the variables examined for association with seropositivity with HSV-1 and/or -2: demographic data (eg, age and gender), ethnic/racial background, highest level of education attained, sexual behavior, and history of STDs. Questionnaires were not administered to those with poor understanding of English or impaired as a result of alcohol or drugs.

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Serologic Testing

Laboratory testing was performed at the Northern Alberta Provincial Laboratory on sera, which was previously tested for anti-HIV and antihepatitis C antibodies.25,26 The samples were analyzed by enzyme-linked immunosorbent assay (ELISA) for the qualitative detection of human IgG class antibodies to HSV-1 and HSV-2 (HerpeSelect ELISA; Focus Technologies, Cypress, CA, formerly MRL Diagnostics). This test is U.S. Food and Drug Administration (FDA)-approved and has demonstrated sensitivities and specificities of 89% and 89% and 96% and 97% for HSV-1 and HSV-2, respectively. Sample optical density (OD) readings were compared with reference cutoff OD readings to determine positive, negative, or equivocal status. The ELISA was repeated for the equivocal samples. If the results were again equivocal, an immunoblot was performed (HerpeSelect Immunoblot; Focus Technologies). The immunoblot was reported as positive, negative, or equivocal based on the presence and intensity of specific bands. The HerpeSelect Immunoblot is also FDA-approved and was chosen because of its excellent performance when compared with Western blot (sensitivities and specificities: HSV-1 99.3% and 95.1%, HSV-2 97.3% and 98.1%). Data entry included rechecking of 10% of all entries.

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Statistical Analysis

Data from the questionnaires was previously double-entered in a D-base program. The HSV result was entered into a Microsoft Excel spreadsheet. The 2 databases were merged using a unique identifier before statistical analysis.

Statistical analysis was carried out using SPSS (Statistical Package for Social Sciences, Chicago, IL) 11.5 for Windows. Univariate and multivariate analyses were performed separately for each of 2 outcomes, HSV-1 seropositivity and HSV-2 seropositivity, but did not exclude individuals who were seropositive for HSV-1 and HSV-2 simultaneously. Analysis was carried out using Student t test, chi square, Fisher exact test, and logistic regression. Unadjusted and adjusted odds ratios (ORs) and 95% confidence intervals (CIs) were calculated. The level of statistical significance was set at 0.05 (2-tailed). Although a number of variables were present in the original dataset, variables chosen for this analysis were selected based on possible causal pathways for HSV-1 or -2 seropositivity. All variables reaching significance in univariate analyses and gender were entered into a forward logistic regression model.

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Study Population

In the original study,25 blood was drawn on 9506 persons who were asked to participate in the study. A total of 1768 declined and 1070 were ineligible. Of the eligible 6668 samples, sufficient sera were available for HSV-1 and -2 testing in 6555 specimens. Some of the characteristics of the individuals tested for HSV-1 and -2 are shown in Table 1. Approximately half were tested at each of the 2 study sites. Fifty-seven percent were male and the mean age was 27.6 years (range, 14–70 years). Eighty-two percent were white and 6% were Aboriginal. The majority was heterosexual (84%) and had a fixed address (88%). Almost half of the study population had only completed high school. The mean age at initiation of sexual activity was 16.4 years (range, 1–53 years). The mean number of sexual partners in the preceding 2 months was 1.7 (range, 0–600). Only 16% of the study population used condoms always and 2% were sex trade workers in the 12 months preceding the study. Although 44% had used illicit drugs in the past, only 6% had used injection drugs. A third had a history of STD (any self-reported STD other than HIV) and 6% had a history of genital herpes. Ninety-four (1.4%) tested positive for HIV and 201 (3%) for hepatitis C.

Overall, 3652 (56%) were seropositive for HSV-1 and 1250 (19%) for HSV-2; these included 744 (11%) who were seropositive for both.

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Factors Associated With Seropositivity to Herpes Simplex Virus Type 1

Table 2 shows the unadjusted and adjusted odds ratios, 95% confidence intervals, and P values for variables associated with HSV-1 seropositivity in multivariate analysis. Although a number of variables were initially associated with HSV-1 seropositivity in unadjusted analyses, only a few factors were independent predictors by forward logistic regression.

In multivariate analysis, increasing age was a significant predictor for HSV-1 seropositivity. Persons of Aboriginal ethnicity had an adjusted odds ratio for HSV-1 seropositivity of 2.8 (95% CI, 2.1–3.7) and those who were nonwhite, non-Aboriginal were 2.1 times more likely to be seropositive than whites (95% CI, 1.7–2.6). Persons with sexual debut <14 years of age were also more likely (adjusted OR, 1.4; 95% CI, 1.1–1.7) to be seropositive as were individuals with a history of STD (adjusted OR, 1.3; 95% CI, 1.1–1.4). Seropositivity increased with an increase in number of sexual partners in the preceding 2 months.

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Factors Associated With Seropositivity to Herpes Simplex Virus Type 2

The unadjusted and adjusted odds ratios, 95% confidence intervals, and P values for each variable associated with HSV-2 seropositivity in multivariate analysis are shown in Table 3. Numerous variables were associated with HSV-2 seropositivity in unadjusted analyses but only a few factors remained as independent factors after adjusted forward logistic algorithm.

When compared with individuals <19 years old, the odds of HSV-2 seropositivity increased progressively with increasing age, with persons aged 20 to 29 years having an odds ratio of 2.7 (95% CI, 2.0–3.8), those aged 30 to 39 years having an odds ratio of 7.2 (95% CI, 5.2–10.1), and those aged 40 to 70 having an odds ratio of 8.3 (95% CI, 5.8–11.8). Females were 2.2 (95% CI, 1.9–2.6) times more likely to be seropositive. Other factors predictive for HSV-2 seropositivity were aboriginal ethnicity (adjusted OR, 2.6; 95% CI, 2.0–3.4) and other nonwhite, non-Aboriginal ethnicities (adjusted OR, 1.8; 95% CI, 1.4–2.3), history of genital herpes (adjusted OR, 8.6; 95% CI, 6.4–11.4), history of STD (adjusted OR, 1.9; 95% CI, 1.6–2.2), and presence of genital sores on examination (adjusted OR, 2.3; 95% CI, 1.8–3.0). Of the 1250 individuals seropositive for HSV-2, 29.8% had a history of genital herpes. Persons who were coinfected with HIV (adjusted OR, 2.6; 95% CI, 1.6–4.2) and hepatitis C (adjusted OR, 2.6; 95% CI, 1.8–3.6) were also more likely to be seropositive.

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We have performed the largest study to date examining the seroepidemiology of HSV-1 and -2 infection among persons attending STD clinics. The 56% seroprevalence of HSV-1 in our study is consistent with the previously reported seroprevalence of 50% to 90% in other populations.27,28 In contrast, the HSV-2 seroprevalence of 19% in this study is lower than the range reported in other STD clinic settings, where seroprevalence has ranged from 23% to 75%.22,28–32 This low rate is surprising and may reflect an overall lower seropositivity rate in the general Alberta and Canadian population. This is partly supported by the limited available data on HSV-2 seroprevalence in Canada such as the rate of 17% in antenatal women tested in British Columbia.24 Canadian figures on HSV prevalence are limited, and this study provides 1 of the few estimates of point prevalence that exists in the literature.

Similar to other studies,16,29,31,33 our study found that seropositivity for HSV-1 and -2 was greater among women. This is postulated to be the result of the higher efficiency of HSV transmission to women.16 Our study also confirms the increase in seropositivity for HSV-1 and -2 with increasing age.20,21,29,31,33–37 Similar to other studies, our study did not show a difference in seropositivity by sexual orientation.38,39

Interestingly, we found the seroprevalence of HSV-1 to be significantly higher in Canadian Aboriginals and nonwhite, non-Aboriginal persons. Although some studies have shown higher HSV-1 seropositivity with nonwhite race,21,37 others11 have shown an association only with white race.

Similarly, the prevalence of HSV-2 was greater in nonwhite ethnic groups as previously reported in other studies.20,21,32 The seropositivity for HSV-2 was even higher for Canadian Aboriginals, a group who has previously been reported to be disproportionately affected by other sexually transmitted infections and HIV in Canada.40

In our study, like in previously published studies, only a minority of persons with HSV-2 antibody reported symptoms resulting from genital herpes.20–23,34 In this study, almost 30% of HSV-2-seropositive individuals reported a history of genital herpes. This is at the upper end of the range of 2.6% to 29% reported in other studies of HSV-2-seropositive persons.20,23,34,41 The recognition that a minority of individuals with genital herpes are aware of their diagnosis has raised the issue of offering HSV-2 serologic screening as a potential prevention strategy. This may enable infected individuals to be taught to recognize clinical outbreaks, thereby decreasing further sexual transmission.

In this Alberta study, we confirmed the finding of previous studies that HSV-1 behaves similarly to other sexually transmitted infections with seropositivity associated with increasing number of sexual partners in the preceding 2 months, a history of STDs, and age at sexual debut less than 14 years.29,42,43 However, it is recognized that only a portion of HSV-1-seropositive persons have genital herpes.

Although other studies have shown a correlation of HSV-2 with number of sexual partners,29,34,35 our study did not demonstrate this correlation. This lack of significance may be related to the fact that we only captured information on the number of sexual partners in the preceding 2 and 12 months and not their lifetime number of sexual partners. However, a history of STDs as a surrogate marker of unsafe sexual behavior was predictive for HSV-2 seropositivity as has been shown in other studies.20,29,34,35 The apparent lack of effect of commercial sex trade work or contact, or exchange of drugs or sex for money, is probably related to the relatively small sample sizes of these subgroups.

Our study likewise did not demonstrate the effect of condoms in preventing transmission of HSV-2 as previously demonstrated by Wald et al. in HSV-2-discordant heterosexual couples.44 This is likely related in part to the cross-sectional design of our study as compared with the longitudinal design of the Wald study. In addition, our questionnaire design differed in that it asked about condom use over a 2- or 6-month period rather than with each sexual act like in the Wald study.

Our study demonstrated a correlation among HSV-2 seropositivity, HIV, and hepatitis C (HCV) infections. This has been previously described for both HIV and HCV and is thought to be related to increased transmission of these agents with subclinical genital erosions.36,45,46 HCV seropositivity is a marker of injection drug use with the majority of newly reported infections in Alberta attributed to acquisition through injection drug use.47 Previous studies have reported that persons who use injection drugs are at higher risk for STD, HIV, and HCV, findings that are in part related to the exchange of drugs for sex.48,49

Our study has the advantage of a large sample size and its anonymous unlinked design ensuring that there is no participation bias. Furthermore, the administration of a detailed risk behavior questionnaire at the time of the original study and testing of sera for HIV and HCV allowed for detailed evaluation of the impact of risk behavior on seropositivity. Potential limitations of this study include recall and information-reporting biases, which are always problems in studies involving sexual histories; the retrospective nature of the study; and generalizability and applicability because it was performed in a high-risk STD clinic population and not in the general Canadian population.

In summary, our study is the largest Canadian study to examine the seroprevalence and associated risk factors for HSV-1 and -2 in Canada. Like with other sexually transmitted infections and bloodborne pathogens (BBP), Canadian Aboriginals appear to be at higher risk of developing infection with both HSV-1 and HSV-2. Our finding that HCV seropositivity was a significant predictor for HSV-2 seropositivity emphasizes the overlap between pathogens that are primarily thought to be BBP and sexually transmitted infections and the need to target prevention in these areas concurrently. This has implications for programs that need to address not only the more traditional aspects of prevention of sexually transmitted infections through behavior modification to promote safer sex practices, but also prevention campaigns designed to modify behavior related to injection drug use and its associated high-risk sexual activity. The high rate of asymptomatic genital HSV infection, which contributes to further sexual transmission, raises the question of introducing serologic HSV screening programs.

Repeat cross-sectional studies of HSV-1 and HSV-2 seropositivity in the same clinics and other populations in Alberta and Canada are warranted to determine if we are observing a rise in seropositivity as reported in other countries, including the United States and Sweden.1,50,52

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