BACKGROUND Prolonged and strenuous exercise (PSE) induces transient left ventricular (LV) dysfunction. Although a consensus exists regarding the decrease in diastolic function, the existence of a decrease in systolic function by a PSE remains controversial, probably due to the transient tachycardia and changes in loading conditions observed upon the completion of exercise. Therefore, the objective was to evaluate LV systolic function before and after a PSE using two-dimensional speckle tracking echocardiography not only at rest but also during incremental tests to adjust heart rates (HR). RESULTS Sixteen healthy young men (23 ± 3 yr old) performed a 3-h period of intensity-controlled upright cycling. LV strain (S), systolic strain rate (SR), rotation, and systolic rotational rate were evaluated by two-dimensional speckle tracking echocardiography before and after a 3-h period of PSE at rest and during incremental tests. Posttest evaluation was performed once the HR had returned to the pretest value. Under resting conditions, parameters of systolic function were either unchanged or increased after the PSE. However, during the incremental test, all LV systolic SR and apical rotational rates were decreased after PSE (radial SR at workload 3 (W3): 2.21 ± 0.12.s(-1) vs 1.87 ± 0.10.s(-1), P < 0.01 and apical rotational rate at W3: 128 ± 28 deg.s(-1) vs 105 ± 26 deg.s(-1), P < 0.05). Regression analyses between LV systolic SR and HR showed lower y-intercepts without differences in slopes, suggesting a decrease of both global and regional systolic functions irrespective of HR after the PSE. CONCLUSIONS Our findings based on LV S and SR data during incremental tests demonstrate that the 3-h period of PSE induces LV systolic dysfunction.