Herpes simplex virus type 2 (HSV-2) infection enhances human immunodeficiency virus (HIV) susceptibility and may increase sexual transmission, but few data on HSV-2 prevalence are available from West Africa. The main objective of this study was to describe the prevalence and determinants of HSV-2 among adults in urban and rural Burkina Faso. We conducted (i) a 2-stage clustered population-based survey among adults aged 15 to 49 years in Ouagadougou (N = 883 women and 791 men), the capital city of Burkina Faso, and (ii) a cross-sectional study among attendees (N = 2018) of all 98 antenatal clinics from 4 provinces of the country. HSV-2 infection was diagnosed using a specific IgG2 enzyme-linked immunosorbent assay test (Kalon). The prevalence of HSV-2 among pregnant women was 18.0% (95% confidence interval (CI): 16.3%-19.7%), with a trend for lower rates in rural (17.3%; CI: 15.5%-19.2%) versus semiurban areas (21.4%; CI: 17.2%-26.2%) (P = 0.08), whereas the prevalence of HSV-2 was 23.7% (CI: 20.9%-26.6%) among women in Ouagadougou (N = 883), and 15.3% (CI: 12.9%-18.0%) among men (N = 791). Using multivariable logistic regression analysis among women, older age, being married, contraceptives, bacterial vaginosis (BV), and HIV infection were significantly associated with HSV-2 infection. Among men, only increased age and HIV were independently associated with HSV-2. The prevalence of HSV-2 infection was high among young women, including in rural areas, with a strong association with BV. Therefore, rural areas should not be neglected in prevention programs. New potent BV, hormonal contraceptives, and HSV control strategies may be helpful for HIV control.