BACKGROUND: Post-exercise recovery phase is associated with clustering of various cardiovascular events and, therefore, monitoring of cardiac autonomic control via heart rate variability (HRV) during this phase may allow identification of autonomic alterations that are not evident under resting conditions in type 2 diabetes mellitus (T2DM) patients. PURPOSE: To investigate and compare the diagnostic performance of resting and post-exercise HRV for detecting cardiac autonomic neuropathy (CAN) in T2DM patients. METHODS: Forty-two T2DM patients were categorized as CAN-positive and CAN-negative based on standard cardiovascular autonomic reflex tests (CARTs). Short-term resting and post-exercise HRV after a graded exercise test were evaluated for each participant. Diagnostic performance of both resting and post-exercise HRV measures was computed using standard statistical procedures. RESULTS: Diagnostic testing yielded superior diagnostic performance of post-exercise HRV than resting HRV measures. Root mean square of successive differences (RMSSD) between adjacent R-R intervals (p鈥?鈥?.01), percentage of consecutive N-N intervals that vary by >50鈥痬s (pNN50) (p鈥?鈥?.03) and total power (TP) (p鈥?鈥?.01) were significantly better diagnostic indicators of CAN under post-exercise conditions than at rest. Predictive ability of these post-exercise HRV measures for CAN was maintained after adjusting various clinical confounders to cardiac autonomic function. CONCLUSION: Post-exercise HRV measures such as TP, RMSSD and pNN50 were found to be more accurate diagnostic tests for detecting CAN than resting HRV. Hence, monitoring of the HRV measures proposed here during exercise testing protocols may provide important diagnostic information regarding CAN in T2DM.