The protozoan parasite Trypanosoma cruzi is the causative agent of the Chagas disease, which is endemic in southeastern Mexico and is transmitted by the vector Triatoma dimidiata (triatomide). T. cruzi infect a great variety of domestic and wild mammals; rodents are considered one of the most important reservoirs of the parasite in the transmission cycles of T. cruzi. The objective of this study was to determine the frequency of T. cruzi infection and to determine the parasitic load in synanthropic and wild rodents from the rural community of southern Mexico. A total of 41 blood samples and 68 heart tissue samples were collected from various species of synanthropic (n= 48 in 2 species) and wild rodents (n= 35 in 5 species). DNA was extracted from samples to detect the presence of T. cruzi through quantitative PCR (qPCR). T. cruzi DNA was detected in the 9.75% of the blood samples of the synanthropic species (4/41) (14.28%) for Rattus rattus samples and 25% for Ototylomys phyllotis samples, with an average of parasitic load of 4.80 ± 1.17 parasites/μL. In the case of heart tissue samples, 10.29% were positive for T. cruzi (7/68) (8.7% for Rattus rattus, 40% for Peromyscus yucatanicus, and 42.8% for Ototylomys phyllotis) with an average parasite load of 3.15 ± 1.98 eq-parasites/mg. The active and chronic infection of T. cruzi in synanthropic or wild rodents of the rural community of southern Mexico evidences the natural infection in these reservoirs which contribute to maintaining the agent in the wild and domestic environments and can represent a risk of infection for the human population when the vector is present.