Given the evolutionary similarities between the bacterial and mitochondrial protein synthesis machinery, we tested the hypothesis that the widely used antibiotic doxycycline reduces mitochondrial function, and results in cardiac contractile dysfunction. Indeed, doxycycline exposure resulted in a dose-dependent reduction in maximal uncoupled mitochondrial respiration in cultured H9C2 cells. Maximal mitochondrial respiration was also reduced in mice treated with doxycycline. MRI of treated mice revealed contractile dysfunction as evidenced by marked diastolic and a mild systolic dysfunction. Moreover, doxycycline exacerbated mitochondrial and contractile dysfunction in animals with type 2 diabetes mellitus.