In Mexico City, the incidence of childhood acute lymphoblastic leukemia (ALL) is one of the highest in the world; epidemiologic evidence suggests that infectious agents could be involved in the genesis of this disease. Early transmitted oncogenic retroviruses infecting lymphocytes are important candidates. PCR-based assays were used to screen viral genomic sequences of human T-cell lymphotrophic virus, type 1 (HTLV1) and mouse mammary tumor virus (MMTV)-like virus (MMTV-LV) in leukemic cells from 67 pediatric patients with ALL. Viral genomic sequences were not detected in any sample by neither standard nor nested PCR. Because of the methodologic strictness and high statistical power of the study, these results suggest that HTLV1 and MMTV-LV are not involved in the genesis of childhood ALL in Mexican children. To our knowledge, this is the first work exploring the direct participation of HTLV1 and MMTV-LV retroviruses in childhood ALL development. ©2013 AACR.