Mother-to-child transmission remains the main cause of global pediatric HIV infections, especially in sub-Saharan Africa. Many interventions based on single-dose antiretroviral therapy have been implemented to reduce the mother-to-child transmission of HIV. In resource-limited settings, highly active antiretroviral therapy (HAART) has only been recommended for HIV-infected pregnant women requiring treatment for their own health. Here, we assessed the efficacy over 18 months of maternal HAART versus peripartum short-course antiretroviral therapy (SCART) regimens for the prevention of mother-to-child transmission (PMTCT) of HIV. We conducted a retrospective cohort study of patients from two medical centers in Ouagadougou, Burkina Faso. The PMTCT files and registers from 1 January 2003 to 31 December 2006 were obtained from routine data collected at these sites. The main assessment criterion was the rate of HIV-1 positivity in children born to HIV-positive mothers as measured with HIV-1 rapid tests at 18 months. A total of 586 pregnant HIV-1-infected women in PMTCT programs were selected. Among these women, 260 were undergoing HAART and 326 received single-dose nevirapine (91.3%) or single-dose zidovudine (8.7%) at delivery. HIV-1 serological tests were performed on 454 children at 18 months old. The rate of HIV-1 vertical transmission was 0% (0/195) in the HAART group and 4.6% (12/259) in the single-dose monotherapy group (P<0.01). Eight infants in the HAART cohort and 30 in the SCART cohort were breastfed; three in the SCART group were HIV-positive. A total of 62 children died, 19 in the HAART group and 43 in the single-dose monotherapy group. Our study confirms that HAART for mothers effectively reduces the risk of infant HIV infection while preserving the breastfeeding option for mothers.