Despite the growing importance of the regulatory function of B cells in many infectious diseases, their immunosuppressive role remains elusive in chronic Chagas disease (CCD). Here, we studied the proportion of different B cell subsets and their capacity to secrete IL-10 ex vivo in peripheral blood from patients with or without CCD cardiomyopathy. First, we immunophenotyped peripheral blood mononuclear cells from patients according to the expression of markers CD19, CD24, CD38 and CD27 and we showed an expansion of total B cell and transitional CD24highCD38high B cell subsets in CCD patients with cardiac involvement compared to non-infected donors. Although no differences were observed in the frequency of total IL-10 producing B cells (B10) among the groups, CCD patients with cardiac involvement showed an increased proportion of naïve B10 cells and a tendency to a higher frequency of transitional B10 cells compared to non-infected donors. Our research demonstrates that transitional B cells are greatly expanded in patients with the cardiac form of CCD and these cells retain the ability to secrete IL-10. These findings provide insight into the phenotypic distribution of regulatory B cells in CCD, an important step towards new strategies to prevent cardiomyopathy associated with T. cruzi infection.