The prevalence of Latent Tuberculosis Infection (LTBI) and Annual Risk of Tuberculosis Infection (ARTI), key epidemiologic indicators used to estimate the transmission of tuberculosis in the community, have not been evaluated in Romania. The aim of the study is to estimate the prevalence of LTBI and ARTI in children and adolescents from two areas with increased TB endemics: Constanta and Timis. Material and methods: We performed an epidemiological observational study to estimate the prevalence of LTBI, using tuberculin skin test (TST), in a representative sample size of the population aged 0-16 years in both areas, regardless of the presence or absence of BCG post vaccination scarring. Results: 1,928 children and adolescents were evaluated, 1,088 from Constanta (CT) and 840 from Timis (TM). The LTBI prevalence for the studied group was 12.81% (CT-11.86% and TM - 14.05%), for boys 12.46% (CT-11.34% and TM-14.11%) and girls 13.21% (CT-12.53% and TM-13.98%). The prevalence of LTBI was 8.2% in the urban area (CT-9.19%, TM-7.02%) and 4.5% in the rural area (CT-2.67% TM -7.02%). ARTI for the study cohort was 1.64% (1.58% CT and 2.19% TM). In 2016, the TB global incidence (GI) in the two analyzed areas (CT = 73.7% per 100,000, and TM = 70 per 100,000) was higher than the national one (GI = 64.8 per 100,000). Conclusions The values of LTBI and ARTI prevalence in the two analyzed territories were comparable, but significantly higher than in other developed countries, and similar to those with high TB global incidence.