BACKGROUND: The impact of antiretroviral therapy (ART) on herpes simplex virus type-2 (HSV-2) replication is unclear. The aim of this study was to assess factors associated with cervicovaginal HSV-2 DNA shedding and genital ulcer disease (GUD) in a cohort of women living with human immunodeficiency virus type-1 (HIV-1) in Burkina Faso. METHODS: Participants were screened for cervicovaginal HSV-2 DNA, GUD, cervicovaginal and systemic HIV-1 RNA, and reproductive tract infections every 3-6 months over 8 years. Associations with HSV-2 shedding and quantity were examined using random-effects logistic and linear regression, respectively. RESULTS: Of the 236 women with data on HSV-2 shedding, 151 took ART during the study period. Cervicovaginal HSV-2 DNA was detected in 42% of women (99 of 236) in 8.2% of visits (151 of 1848). ART was associated with a reduction in the odds of HSV-2 shedding, which declined for each year of ART use (odds ratio [OR], 0.74; 95% confidence interval [CI], .59-.92). In the multivariable model, the impact of ART was primarily associated with suppression of systemic HIV-1 RNA (adjusted OR, 0.32; 95% CI, .15-.67). A reduction in the odds of GUD was also observed during ART, mainly in those with HIV-1 suppression (adjusted OR, 0.53; 95% CI, .25-1.11). CONCLUSIONS: ART is strongly associated with a decrease in cervicovaginal HSV-2 shedding, and the impact was sustained over several years.