Background: Although the mutual relationship between ambulation and Physical Activity (PA) in people with Multiple Sclerosis (pwMS) has been described in several studies, there is still a lack of detailed information about the way in which specific aspects of the gait cycle are associated with amount and intensity of PA. This study aimed to verify the existence of possible relationships among PA parameters and the spatio-temporal parameters of gait when both are instrumentally assessed.Methods: Thirty-one pwMS (17F, 14M, mean age 52.5, mean Expanded Disability Status Scale score 3.1) were requested to wear a tri-axial accelerometer 24h/day for 7 consecutive days and underwent an instrumental gait analysis, performed using an inertial sensor located on the low back, immediately before the PA assessment period. Main spatio-temporal parameters of gait (i.e. gait speed, stride length, cadence and duration of stance, swing and double support phase) were extracted by processing trunk accelerations. PA was quantified using average number of daily steps and percentage of time spent at different PA intensity, the latter calculated using cut-point sets previously validated for MS. The existence of possible relationships between PA and gait parameters was assessed using Spearman’s rank correlation coefficient rho.Results: Gait speed and stride length were the parameters with the highest number of significant correlations with PA features. In particular, they were found moderately to largely correlated with number of daily steps (rho 0.62, p<0.001), percentage of sedentary activity (rho = -0.44, p<0.001) and percentage of moderate-to-vigorous activity (rho = 0.48, p<0.001). Small to moderate significant correlations were observed between PA intensity and duration of stance, swing and double support phases.Conclusions: The data obtained suggest that the most relevant determinants associated with higher and more intense levels of physical activity in free-living conditions are gait speed and stride length.The simultaneous quantitative assessment of gait parameters and PA levels might represent a useful support for physical therapists in tailoring optimized rehabilitative and training interventions.