Little evidence exists to support the presence of differences in spatiotemporal gait parameters and ambulation ability between those individuals with traumatic and nontraumatic lower-limb amputation (LLA). We conducted an exploratory study of 81 male Veterans with unilateral amputation to quantify differences in spatiotemporal gait parameters and ambulatory mobility between Veterans with traumatic and nontraumatic LLA. Furthermore, we identified variables that significantly contributed to the explanation of variability in modified 2-min walk test distance. All participants completed the modified 2-min walk test and a spatiotemporal gait analysis using an instrumented walkway during a routine physical therapy visit. Veterans with nontraumatic LLA walked significantly shorter mean distances during a modified 2-min walk test than Veterans with traumatic LLA. Variables identified as significant contributors to modified 2-min walk test variability were amputated limb stance time, amputated limb step length, and percentage of the gait cycle spent in double support. These findings demonstrate that differences in spatiotemporal gait parameters and ambulatory mobility exist between Veterans with traumatic and nontraumatic LLA and identify important spatiotemporal parameters of gait contributing to this decline. These parameters should be considered as targets for intervention and future investigation.