Prevalence of human T. cruzi DTUs and Chagas disease manifestations in countries of North, Central and South America. CCC, Chronic Chagas Cardiomyopathy; DIG, Digestive form. Display omitted •Trypanosoma cruzi is partitioned into a minimum of seven discrete typing units (DTUs), TcI–TcVI and Tcbat.•DTUs have distinct, but not exclusive ecological and epidemiological associations.•Hybridization between parasite strains is frequent and challenges the paradigm of prevailing clonal evolution in T. cruzi.•The potential association between DTUs and Chagas disease manifestations, serology and chemotherapy is discussed.•Research priorities for these themes are pointed out and recommendations are issued. The genetic diversity of Trypanosoma cruzi, the protozoan agent of Chagas disease, is widely recognized. At present, T. cruzi is partitioned into seven discrete typing units (DTUs), TcI–TcVI and Tcbat. This article reviews the present knowledge on the parasite population structure, the evolutionary relationships among DTUs and their distinct, but not exclusive ecological and epidemiological associations. Different models for the origin of hybrid DTUs are examined, which agree that genetic exchange among T. cruzi populations is frequent and has contributed to the present parasite population structure. The geographic distribution of the prevalent DTUs in humans from the southern United States to Argentina is here presented and the circumstantial evidence of a possible association between T. cruzi genotype and Chagas disease manifestations is discussed. The available information suggests that parasite strains detected in patients, regardless of the clinical presentation, reflect the principal DTU circulating in the domestic transmission cycles of a particular region. In contrast, in several orally transmitted outbreaks, sylvatic strains are implicated. As a consequence of the genotypic and phenotypic differences of T. cruzi strains and the differential geographic distribution of DTUs in humans, regional variations in the sensitivity of the serological tests are verified. The natural resistance to benznidazole and nifurtimox, verified in vivo and in vitro for some parasite stocks, is not associated with any particular DTU, and does not explain the marked difference in the anti-parasitic efficacy of both drugs in the acute and chronic phases of Chagas disease. Throughout this review, it is emphasized that the interplay between parasite and host genetics should have an important role in the definition of Chagas disease pathogenesis, anti-T. cruzi immune response and chemotherapy outcome and should be considered in future investigations.