Different molecular markers have been employed for typing Trypanosoma cruzi strains from endemic areas of Chagas disease. The low-stringency single specific primer–polymerase chain reaction (LSSP–PCR) has been a sensitive and informative technique that uses the variable region of kinetoplast DNA minicircles as a genetic marker, allowing detection of DNA sequence variation. In the present study, we analyzed the intra-lineage genetic variability of the T. cruzi strains obtained from triatomine feces collected on filter paper FTA card by LSSP–PCR. The hybridization of the PCR products with a probe for the subgenus Schizotrypanum and a clone-specific probe from Dm28c confirmed the subgenus as T. (S.) cruzi and respective lineages as T. cruzi I. Phenetic analysis showed the presence of three clusters that diverged by different coefficients of similarity. Thirteen T. cruzi I genotypes were observed circulating among Triatoma pseudomaculata and Rhodnius nasutus from peridomiciliary and natural environments in five peri-urban and urban localities of Jaguaruana, Ceará, Brazil. These data indicate the importance of the circulation of T. cruzi I genotypes among T. pseudomaculata and R. nasutus in different environments and the possible risk of Chagas disease domestic transmission.