Triatoma dimidiata is one of the major Chagas disease vectors, with an extensive diversity in its morphology, habitat, and level of domiciliation. Molecular studies based on the internal transcribed spacer 2 (ITS-2) have subdivided this species into four potential taxonomic groups. Using both ITS-2 and cytochrome B markers, we confirmed the sibling species status of ITS-2 Group 3 and detected an apparent sympatry of ITS-2 Groups 2 and 3 in the Yucatan peninsula, Mexico. Here we examine the geographic distribution of T. dimidiata ITS-2 genotypes in the region and compare their egg production and Trypanosoma cruzi infection rates, as indicators of biological differences between groups. PCR genotyping of large natural populations showed an extensive sympatry of Groups 2 and 3 in most of the peninsula, often within the same house. We also detected a large proportion of individuals displaying ITS-2 sequences from both Groups 2 and 3, suggesting hybridization. Analysis of ITS-2 genotype frequencies indicated a strong departure from Hardy-Weinberg equilibrium in female hybrids, but not in males, due to a large heterozygote deficit. These results suggest random mating between ITS-2 Groups 2 and 3 combined with reduced viability and/or survival in female hybrids. This and other factors may allow for the maintenance of distinct ITS-2 Groups 2 and 3 populations despite high hybrid frequencies. Importantly, T. cruzi infection was much higher in hybrids compared to ITS-2 Groups 2 and 3 individuals, but all three genotypes appeared to seasonally infest houses in a similar manner in the region. These findings warrant further studies on T. dimidiata taxonomy and its epidemiologic implications.