Objective: To examine the effect of induced fatigue on spatiotemporal gait parameters in persons with multiple sclerosis (PwMS) by using 6‐min walk test (6MWT). Methods: A cross‐sectional study with a control group (25 healthy persons) was performed. Fifty‐six PwMS (37 female) were divided into three groups according to their level of disability, as measured by the expanded disability status scale (EDSS): mild (n = 23, EDSS = 1.0–3.5), moderate (n = 19, EDSS = 4.0–5.5), and severe (n = 12, EDSS = 6.0–6.5). Spatiotemporal gait parameters were measured by the GAITRite electronic walkway before and after 6MWT performance. In addition, to determine the level of fatigue in PwMS, the participants completed the questionnaire modified fatigue impact scale (MFIS) before performing the 6MWT. Statistical analyses were performed to compare intragroup and intergroup differences. Results: Fatigue level was lower in the mild (37.6 ± 20.5) versus moderate (54.3 ± 17.2) versus severe (53.6 ± 12.9) groups (p < 0.05). Significant differences were found among all the groups in terms of the distance walked during the 6MWT (p = 0.001) and of the spatiotemporal gait parameters: gait velocity (cm/s), cadence (steps/min), and step length (cm) decreased and, on the contrary, step time (sec), stance, and double support time (% gait cycle) increased when EDSS scores were higher (p < 0.05). The percentage of change (%) in the spatiotemporal gait parameters, after 6MWT performance, was statistically significant in the moderate and severe groups: gait velocity (−8.8%, −25.9%) and step length (−6.5%, −13.4%) decreased, whereas step time (3.0%, 15.0%), double support time (8.8%, 19.1%), step time asymmetry (32.1%, 64.0%), and single support asymmetry (60.0%, 74.7%) increased (p < 0.05). Conclusions: Gait pattern worsen after performing a walking‐induced fatigue test, such as 6MWT, in moderate–severe PwMS (EDSS ≥ 4.0). Identifying these gait alterations will allow physiotherapists to determine specific therapeutic objectives. [ABSTRACT FROM AUTHOR]