BACKGROUND: We aimed to estimate the prevalence of and explore risk factors for Mycobacterium tuberculosis infection among adolescents in a high tuberculosis (TB) and human immunodeficiency virus (HIV) prevalence setting. METHODS: A cross-sectional study of adolescents (10-19 years) randomly selected from a demographic surveillance area (DSA) in rural KwaZulu-Natal, South Africa. We determined M tuberculosis infection status using the QuantiFERON-TB Gold-plus assay. We used HIV data from the DSA to estimate community-level adult HIV prevalence and random-effects logistic regression to identify risk factors for TB infection. RESULTS: We enrolled 1094 adolescents (548 [50.1%] female); M tuberculosis infection prevalence (weighted for nonresponse by age, sex, and urban/rural residence) was 23.0% (95% confidence interval [CI], 20.6-25.6%). Mycobacterium tuberculosis infection was associated with older age (adjusted odds ratio [aOR], 1.37; 95% CI, 1.10-1.71, for increasing age-group [12-14, 15-17, and 18-19 vs 10-11 years]), ever (vs never) having a household TB contact (aOR, 2.13; 95% CI, 1.25-3.64), and increasing community-level HIV prevalence (aOR, 1.43 and 95% CI, 1.07-1.92, for increasing HIV prevalence category [25%-34.9%, 35%-44.9%, ≥45% vs <25%]). CONCLUSIONS: Our data support prioritizing TB prevention and care activities in TB-affected households and high HIV prevalence communities.