The aim of this randomized counterbalanced, 2 × 2 cross-over study was to investigate the effects of mental fatigue on cognitive and aerobic performance in adolescent active endurance athletes. Ten active male endurance athletes (age = 16 ± 1.05 years, height = 1.62 ± 0.04 m, body mass = 55.5 ± 4.2 kg) were familiarized to all experimental procedures on day 1. On days 2 and 3, participants provided a rating of mental fatigue before and after completing a 30 min Stroop test that measures selective attention capacity and skills and their processing speed ability (mentally fatigued condition), or a 30 min control condition in a randomized counterbalanced order. They then performed d2 test and a 20 m multistage fitness test (MSFT), which was used to measure selective and sustained attention and visual scanning speed (i.e., concentration performance (CP) and total number of errors (E)) and aerobic fitness (i.e., maximum oxygen uptake (VO2max) and velocity at which VO2max occurs (vVO2max)), respectively. Rating of perceived exertion (RPE) was assessed after a MSFT. Subjective ratings of mental fatigue were higher after the Stroop task (p < 0.001). CP (p = 0.0.1), E (p < 0.001), vVO2max (p = 0.020), and estimated VO2max (p = 0.021) values were negatively affected by mental fatigue. RPE were significantly higher in the mentally fatigued than in the control conditions (p = 0.02) post-MSFT. Mental fatigue impairs aerobic and cognitive performance in active male endurance athletes.