Background Little is known about factors contributing to the altered movement patterns observed in many individuals with anterior cruciate ligament (ACL) injury. We addressed whether altered muscular activity is such a factor. Methods 16 participants with unilateral, non-reconstructed ACL rupture were scored for altered movement patterns according to Test for Substitution Patterns (TSP), which includes the single-leg squat (SLS). Surface electromyography (SEMG), was recorded in the lower extremities at initiation of weight-transfer from double-leg to single-leg stance (eyes closed), simulating the initiation of an SLS. Normalised SEMG amplitudes 200-300 ms after weight-transfer initiation were compared between injured and non-injured sides, and correlated to the TSP scores for the SLS. Peak absolute SEMG amplitudes during 5 TSP test movements were also compared between sides. Results At weight-transfer initiation, muscle activity was lower in the tibialis anterior, gastrocnemius and peroneus longus muscles on the injured side. Low muscle activity correlated moderately to worse TSP scores for the SLS for the gluteus medius (r s =-0.56, p=0.03), and gastrocnemius muscles (r s =-0.56, p=0.02). Median peak absolute amplitude during TSP movements was lower in the quadriceps, gastrocnemius and peroneus longus muscles on the injured side. Conclusions The altered patterns of muscle activity at weight-transfer initiation, correlations between lower activity at movement initiation and altered movement patterns during SLS and the altered peak amplitudes during TSP movements together indicate alterations in sensorimotor control that may contribute to the observed altered movement patterns. Future studies will determine if exercises targeting muscle activity initiation should complement customary ACL injury rehabilitation.