Human herpesvirus 8 infection of the genital tract of HIV-seropositive and HIV-seronegative women at risk of sexually transmitted diseases

Epidemiological studies support the sexual transmission of human herpesvirus 8 (HHV8) between homosexual men [1,2]. HHV8 is shed in the urogenital tract of men [3] and HHV8 DNA sequences have been detected in the genital tracts of a few HHV8-seropositive women [4,5]. In Honduran women, risk factors for HHV8 seropositivity included HIV infection and having multiple sex partners [6]. Although anti-HHV8 antibodies have been detected in up to 28% of HIV-seropositive or HIV-seronegative women [6–9], the prevalence of HHV8 infection in the genital tracts of women is unknown. We conducted a cross-sectional study to estimate the prevalence of HHV8 infection as measured by polymerase chain reaction (PCR) in genital specimens collected from HIV-infected women and HIV-seronegative women at risk of sexually transmitted diseases (STD) as previously defined [10,11]. Genital specimens (n = 303), consisting of 72 vaginal tampons and 231 cervicovaginal lavages, from 300 women (200 HIV-seropositive, 100 HIV-seronegative women at risk of STD) participating in the Canadian Women's HIV Study [10,11] were selected consecutively. Study inclusion criteria and procedures for sample collection and processing have been described in previous publications [10–12]. All samples tested positive for β-globin DNA [12]. Processed samples were amplified for HHV8 with a standardized competitive PCR assay [13,14]. Each PCR run included negative, weak positive (12.5 HHV8 DNA copies), and strong positive (5000 HHV8 DNA copies) controls. Samples and controls were spiked with 500 copies of an heterologous internal control (IC) containing HHV8 primer sequences at both ends [14]. Amplified products were run on two separate 1.5% agarose gels followed by Southern blot hybridization with specific radiolabelled HHV8 and IC probes [13] and analysis with a phosphor imager (Fig. 1).Fig. 1.: Differential hybridization of polymerase chain reaction products was done with an internal control (IC) probe and a human herpesvirus 8 (HHV8)-specific probe. Lanes 1–4, DNA from cervicovaginal cells (all negative for HHV8) in the presence of 500 copies of the IC; lanes 5–9, increasing concentrations of the HHV8 plasmid (6.25, 12.5, 25, 50, and 5000 copies) in the presence of 500 copies of the IC; lane 10, blank.The average age of HIV-seropositive and HIV-seronegative women was 32.3 ± 7.3 years (median, 31 years; age range, 17–63 years) and 27.8 ± 6.5 years (median, 28 years; age range, 16–43 years), respectively. Thirteen (4.6%) and 76 (27.1%) women had sex with bisexual men or men born in Africa or the Caribbean, respectively. Having at least five lifetime male sexual partners was reported by 105 (54.7%) HIV-seropositive women and 83 (83%) HIV-seronegative women. Among participating HIV-seropositive women, 46 (24.3%) had CD4 cell counts below 200 × 106 cells per litre, 98 (51.9%) had CD4 cell counts between 200 and 499, and 45 (23.8%) had CD4 cell counts above 500. Eighty-four (43.8%) HIV-seropositive women had an AIDS-defining condition. However, only two HIV-seropositive women had Kaposi's sarcoma (KS). Because the IC was not amplified for nine specimens and insufficient specimens precluded PCR analysis for another 10 samples, 284 samples (195 from HIV-seropositive women, 89 from HIV-seronegative women) were included in the study. The competitive PCR assay reliably detected 12.5 HHV8 DNA copies in multiple experiments using serial dilutions of a HHV8 plasmid in the presence of 500 copies of IC (data not shown). HHV8 DNA was not detected in any genital samples, including the two women with KS. An example of a typical HHV8 PCR run is shown in Fig. 1. The 95% confidence interval for the prevalence rate of genital HHV8 infection in HIV-seropositive women was 0.0–2.0%. As found by other authors [5,15], we were unable to detect HHV8 DNA sequences in the genital specimens from sexually active women in Canada even in the presence of KS. However, HHV8 DNA has been detected in a limited number of genital specimens from women from endemic regions for HHV8 infection (Italy and Africa), suggesting that geographical differences in the prevalence of HHV8 infection could account for differences in the HHV8 detection rates [4,5]. Our PCR test was sensitive and consistently detected 12.5 copies of HHV8 DNA. Women recruited in our study had risk factors associated with HHV8 seropositivity in women [6]. Testing only HHV8-seropositive women would have increased our chances of detecting HHV8 in the genital tract [4], but serum samples were not collected from our participants. Including participants irrespective of HHV8 serology results provided a less biased evaluation of the prevalence of genital HHV8 infection. These findings, from the largest group of HIV-infected women tested for HHV8 DNA to date, demonstrate that HHV8 could be detected in at most 2.0% of genital specimens from HIV-seropositive Canadian women, even if they are exposed to men who were themselves at risk of HHV8 infection. Although this may reflect the low prevalence of HHV8 in our cohort or the intermittent genital shedding of HHV8, it could also suggest that heterosexual transmission of HHV8 is not an effective means of viral transmission. These results concur with the low prevalence rate of KS in HIV-infected women in contrast to HIV-infected men who have sex with men. Guy Boivina Catherine Hankinsbc Normand Lapointede Sharon Walmsleyf Annie Gaudreaua Pierre Forestg the Canadian Women's HIV Study Group,* François Coutléecdg

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