Objective: To establish the relationship between HIV-induced cardiac diastolic dysfunction, immune responses, and virus replication in the heart using the simian immunodeficiency virus (SIV)/macaque model. Design: Cardiac diastolic dysfunction is common in HIV-infected individuals including asymptomatic patients and those treated with combination antiretroviral therapy. SIV-infected macaques develop cardiac dysfunction, serving as a useful model to establish mechanisms underlying HIV-induced cardiac dysfunction. To understand the relationship between functional cardiac impairment, viral replication in the heart, and associated host inflammatory responses, cardiac function was evaluated in SIV-infected macaques and functional decline was correlated with features of the host immune response and the extent of viral replication in both the myocardium and plasma. Methods: Cardiac function was evaluated longitudinally in 22 SIV-infected and eight uninfected macaques using mitral inflow and tissue Doppler echocardiography. Myocardial macrophage populations were evaluated by CD68 and CD163 immunostaining. SIV RNA levels in both myocardium and plasma were measured by qRT-PCR. Results: Echocardiographic abnormalities developed in SIV-infected macaques that closely resembled diastolic dysfunction reported in asymptomatic HIV-infected individuals. Although CD68 and CD163 were upregulated in the myocardium of SIV-infected animals, neither macrophage marker correlated with functional decline. SIV-induced diastolic dysfunction was strongly correlated with extent of SIV replication in the myocardium, implicating virus or viral proteins in the initiation and progression of cardiac dysfunction. Conclusion: This study demonstrated a strong correlation between cardiac functional impairment and extent of SIV replication in the myocardium, suggesting that persistent viral replication in myocardial macrophages induces cardiomyocyte damage manifest as diastolic dysfunction.