ObjectivesOur purpose was to investigate the immediate and prolonged effects of hamstrings fatigue on quadriceps neuromuscular function in individuals with anterior cruciate ligament reconstruction (ACLR) and matched uninjured controls. DesignCross-Sectional. SettingLaboratory. Participants16 participants with a history of ACLR and 16 uninjured controls. Main outcome measuresQuadriceps peak torque (PT), central activation ratio (CAR), early (RTD100) and late (RTD200) rate of torque development, vastus medialis and lateralis electromyographic (EMG) activity, and hamstrings-to-quadriceps co-activation assessed at baseline. Outcomes were evaluated pre-fatigue (PRE), immediately post-fatigue (POST), and 30min post-fatigue (POST30). The involved limbs of individuals with ACLR were assessed and control limbs were matched based on limb dominance. ResultsIndividuals with ACLR demonstrated lesser quadriceps PT (p = 0.004), CAR (p < 0.001), RTD100 (p = 0.042), RTD200 (p = 0.028), and vastus medialis EMG (p = 0.040) than controls, regardless of time. Quadriceps CAR (p < 0.001) and RTD200 (p < 0.001) decreased at POST and POST30, whereas RTD100 (p < 0.001) decreased at POST, regardless of group. ConclusionsThe observed reductions in quadriceps neuromuscular function may suggest involvement of central fatigue mechanisms, which should be explored prior to recommending hamstrings fatigue as a therapeutic intervention.