BACKGROUND: Greater gait variability is associated with falls in aging and multiple sclerosis. However, whether older adults with MS (OAMS), show higher gait variability relative to healthy older adults (HOA), under single and dual-task walking conditions, has not been reported. Furthermore, it is unclear whether practice may improve gait variability in both groups. RESEARCH QUESTION: Is gait variability higher in OAMS relative to HOA, particularly in DTW compared to STW? Furthermore, does practice result in decreased gait variability in both groups, notably under DTW compared to STW? METHODS: We examined the effect of within-session practice on gait variability during single (STW) and dual (DTW) task gait conditions. OAMS (n = 97, mean±SD age: 65 ± 5 years, 66 females) and HOA (n = 113, mean±SD age: 68 ± 7 years, 73 females) were recruited. Practice effects on gait variability were evaluated over three repeated counterbalanced STW and DTW trials. Gait variability measures included Coefficient of Variation (CV) in stride velocity, stride length, and swing time. RESULTS: OAMS demonstrated higher gait variability, on all measures, relative to HOA during both STW and DTW (P < 0.001). Gait variability on all measures was higher in DTW compared to STW, (P < 0.05). Practice resulted in decreased gait variability (P < 0.01) on all measures in both OAMS and HOA. Furthermore, practice resulted in decreased temporal gait variability, as measured by swing time CV, under DTW in particular (P < 0.05). SIGNIFICANCE: In conclusion, OAMS exhibited greater gait variability than HOA, yet both groups demonstrated decreases in temporal and spatial gait variability after within-session practice, notably under DTW, which in turn may reduce fall risk.