We investigated the thymic and peripheral T-lymphocyte subsets in BALB/c mice undergoing acute or chronic Trypanosoma cruzi infection, in terms of expression of particular Vb rearrangements of the T-cell receptor. We first confirmed the severe depletion of CD4+ CD8+ thymocytes following acute T. cruzi infection. By contrast, the numbers of CD4+ CD8+ cells in subcutaneous lymph nodes increased up to 16 times. In subcutaneous lymph nodes, we found CD4+ CD8+ cells that expressed prohibited segments TCRVb5 and TCRVb12 (which are physiologically deleted in the thymus of BALB/c mice), as did some mature single-positive cells (CD4+ CD8– and CD4– CD8+ ). In the thymus of infected animals, although higher numbers of immature cells bearing such Vb segments were seen, they were no longer detected in the mature single-positive stage, suggesting that negative selection occurs normally. We also found increased numbers of cells bearing the potentially autoreactive phenotype TCRVb5+ and TCRVb12+ in T-lymphocyte subsets from subcutaneous lymph nodes of T. cruzi chronically infected mice. In conclusion, our data indicate that immature T lymphocytes bearing prohibited TCRVb segments leave the thymus and gain the lymph nodes, where they further differentiate into mature CD4+ or CD8+ cells. Conjointly, these findings show changes in the shaping of the central and peripheral T-cell repertoire in both acute and chronic phases of murine T. cruzi infection. The release of potentially autoreactive T cells in the periphery of the immune system may contribute to the autoimmune process found in both murine and human Chagas’ disease.