Abstract Background: The diagnostic importance of three-dimensional (3D) speckle-tracking strain-imaging echocardiography in patients with acute myocarditis has not yet been assessed. The aim of this study was to test the sensitivity and specificity of 3D-speckle-tracking echocardiography as compared to CMR (cardiovascular magnetic resonance imaging) for the diagnosis of acute myocarditis. Methods and Results: 45 patients that were admitted to the Medical Clinic II of the University Hospital Bonn with clinically suspected myocarditis were enrolled in our study (71% male, mean age: 43.9±16.3 years, peak troponin level: 1.38±3.51 ng/ml). 3D full-volume echocardiographic images were obtained and offline speckle-tracking analysis of regional and global LV deformation was performed. All patients received CMR scans and myocarditis was diagnosed in 29 subjects. The 16 patients, in whom myocarditis was excluded by CMR, served as controls. Regional changes in myocardial texture and tissue edema (diagnosed by CMR) were significantly associated with regional impairment of circumferential, longitudinal, and radial strain, as well as regional 3D displacement and total 3D strain. The 3D diastolic strain index was not associated with pathological findings in the CMR. However, the 3D global longitudinal strain (GLS) outperformed well-known 2D parameters associated with myocarditis, such as LVEF and LVEDV. Conclusions: This is the first study examining the use of 3D-speckle-tracking echocardiography in patients with acute myocarditis. Global longitudinal strain was significantly associated with and impaired in patients with myocarditis. Therefore, 3D echocardiography could become a useful diagnostic tool in the primary diagnosis of myocarditis.