BACKGROUND: Many HIV-infected children in sub-Saharan Africa reside in rural areas, yet most research on treatment outcomes has been conducted in urban centers. Rural clinics and residents may face unique barriers to care and treatment. METHODS: A prospective cohort study of HIV-infected children was conducted between September 2007 and September 2010 at the rural HIV clinic in Macha, Zambia. HIV-infected children younger than 16 years of age at study enrollment who received antiretroviral therapy (ART) during the study were eligible. Treatment outcomes during the first two years of ART, including mortality, immunologic status, and virologic suppression, were assessed and risk factors for mortality and virologic suppression were evaluated. RESULTS: A total of 69 children entered the study receiving ART and 198 initiated ART after study enrollment. The cumulative probabilities of death among children starting ART after study enrollment were 9.0% and 14.4% at 6 and 24 months after ART initiation. Younger age, higher viral load, lower CD4+ T-cell percentage and lower weight-for-age z-scores at ART initiation were associated with higher risk of mortality. The mean CD4(+) T-cell percentage increased from 16.3% at treatment initiation to 29.3% and 35.0% at 6 and 24 months. The proportion of children with undetectable viral load increased to 88.5% and 77.8% at 6 and 24 months. Children with longer travel times (≥ 5 hours) and those taking nevirapine at ART initiation, as well as children who were non-adherent, were less likely to achieve virologic suppression after 6 months of ART. CONCLUSIONS: HIV-infected children receiving treatment in a rural clinic experienced sustained immunologic and virologic improvements. Children with longer travel times were less likely to achieve virologic suppression, supporting the need for decentralized models of ART delivery.