Aims Violacein (VIO), a bacterial pigment produced by Chromobacterium violaceum, was examined to evaluate the antichagasic activity and its action mechanism against Trypanosoma cruzi Y strain. Methods and Results Violacein was tested against the epimastigote, trypomastigote and amastigote forms of T. cruzi Y strain (benznidazole-resistant strain). VIO inhibited all T. cruzi developmental forms, including amastigotes, which is implicated in the burden of infection in the chronic phase of Chagas disease (CD). VIO induced cell death in T. cruzi through apoptosis, as determined by flow cytometry analyses with specific molecular probes and morphological alterations, such as involvement of reactive oxygen species and changes in mitochondrial membrane potential and cell shrinkage. Conclusion The results suggest antichagasic activity of VIO against T. cruzi Y strain with apoptotic involvement. Significance and Impact of the Study The treatment of CD has limited efficacy and side effects that restrict patient tolerability and compliance. The VIO molecule could be used as a model for therapeutic alternatives for this disease.