Abstract Body: Introduction: Chagas Cardiomyopathy (CC) develops in around 30% of Trypanosoma cruzi-infected individuals decades after initial infection and was recently considered hypoendemic in the US by CDC. Research Questions: The mechanisms determining why only a subset of infected individuals develops CC, and why the disease manifests decades after infection in the apparent absence of parasite in the heart, remain critical unanswered questions. Goals: Elucidate disease mechanisms and identify potential therapeutic targets for CC treatment by applying single nuclei RNA sequencing (snRNAseq) to evaluate cell composition and molecular pathways altered in CC. Methods: snRNAseq was performed on explanted hearts from 6 CC individuals from different cardiac regions (11 LV, 1 RV), and compared to controls, dilated cardiomyopathies (DCM) and arrhythmogenic cardiomyopathy (ACM). B and T cell receptor sequences were analyzed using TRUSt4. Results: CC showed massive cardiomyocyte depletion versus all conditions (CC=10.05±6.84% vs ctrl=45.67±14.76%, p=7.6e-08) and profound lymphocyte infiltration (CC=30.35±14.26% vs ctrl=1.59±2.17%, p=2.74e-10). Despite 20–70% of the tissue area be covered by fibrosis (higher than other cardiomyopathies), fibroblast proportions were not different of other cardiomyopathies, same observed to myeloid cells. Remaining cardiomyocytes showed upregulated interferon-γ and -α pathway genes, suggesting ongoing inflammatory signaling in surviving cells. Lymphocytes were predominantly memory B cells and plasma cells (70%), with equal CD4 naïve/helper and CD8 cytotoxic T cells comprising the remainder. B cell clonality analysis revealed high clonal expansion (>100 cells/clone), with subject-specific clones present in both memory B and plasma cell states, distributed across LV and RV from the same individual. T cell clonal expansion was less pronounced but similarly distributed across tissue regions and cell states. Conclusion: CC is characterized by profound cardiomyocyte loss and massive tissue infiltration of clonally expanded, antibody-secreting plasma and memory B cells, suggesting an antigen-driven autoimmune mechanism in CC pathogenesis and pointing to novel immunotherapy targets.