BACKGROUND: A pilot\'s workload induces autonomic nervous system modulations which could be related to a decrease of vigilance that could impair safety. Kinetics during flight and recovery are not well known. OBJECTIVE: The aim of this study was to assess linear and nonlinear heart rate variability (HRV) modulations and vigilance during a high mental workload induced by a complex flight and subsequent recovery. METHODS: There were 10 novice pilots (37.8 +/- 4.4 yr, 115.8 +/- 15.7 h flight experience) who performed a 3 h 30 min (09:30-13:00) multi-leg cross-country flight (Piper Pa28 airplane: 160 hp). We recorded electrocardiogram (ECG) during the flight and performed tests during the 24 h before and after the flight (13:30, 16:00, 18:30, 21:00, and 06:45). Tests included a stand test (10 min supine, 10 min standing), a Mackworth \'clock\' vigilance test, and a Karolinska Sleepiness Scale questionnaire. We assessed HRV components by time and frequency domains in parallel with the Poincare plot analysis. RESULTS: The flight induced a progressive decrease of RR intervals, standard deviation between normal-to-normal intervals (SDNN), Poincare SD1 and SD2 indices, and an increase of the low-frequency to high-frequency ratio (LF/HF). During recovery, vigilance remained depressed for 2 h 30 min after the flight. The decreased RR intervals and SD1 persisted for 5 h postflight both in supine and standing positions. LF/HF stayed elevated for 2 h 30 min after the flight. CONCLUSION: A multi-leg cross-country flight involves a vagal withdrawal and an increase of sympathetic activity lasting 5 h after landing. This delay could be recommended as a safety period.