Chagasic cardiomyopathy is caused by Trypanosoma cruzi infection, and it affects ~7 million individuals on the American continent. Poly(ADP-ribose) polymerase 1 (PARP1) is a nuclear DNA repair enzyme. In this study, we demonstrate that PARP1 tends to be localized to mitochondria in chagasic myocardium. The mtPARP1 was bound to mitochondrial DNA polymerase gamma (Pol ?) that is essential for mtDNA synthesis and repair. Genetic or chemical inhibition of PARP1 was beneficial in improving the mtDNA content, and mitochondrial and left ventricular (LV) function in Chagas disease. Our results suggest that mitochondrial transport of PARP1 adversely impacts the mtDNA maintenance by Pol ? replisome, and exacerbates the mitochondrial dysfunction, oxidative stress, and cardiac remodeling in Chagas disease. We propose that small molecules that prevent PARP1 transport to mitochondria or that arrest PARP1 effects on Pol ? activity will be beneficial in preserving the mitochondrial health and LV function in chronic cardiomyopathy of chagasic (and potentially other) etiologies.