To investigate the pathologic differences in patients with pediatric-onset multiple sclerosis (POMS) and acute disseminated encephalomyelitis (ADEM) using diffusion tensor imaging (DTI) and diffusion basis spectrum imaging (DBSI). Fifteen children with POMS and eight children with ADEM underwent DTI and DBSI. The comparison of DTI and DBSI diffusivity measures of POMS (31 scans) and ADEM (17 scans) was performed as group comparison and association over time. In univariate analysis of average measures of DBSI and DTI over time, DBSI fractional anisotropy is lower in POMS than ADEM (P = 0.002), indicative of axonal injury of POMS. Higher DBSI fiber fraction (P = 0.046) and DBSI radial diffusivity (P = 0.016) but lower DBSI nonrestricted fraction (P = 0.005) in patients with POMS suggests higher axonal density, demyelination, and lower extra-axonal edema in POMS. However, there are no significant differences in DTI measures between POMS and ADEM over time. DBSI may be useful to monitor and quantitatively compare coexisting axonal injury, demyelination, and inflammation in central nervous system white matter tracts in children with POMS and ADEM, overcoming the disadvantages of DTI. Larger prospective longitudinal studies are required to confirm these results.