Much of the research using diffusion tensor imaging (DTI) in stroke focuses on characterizing the microstructural status of corticospinal tracts and its utility as a prognostic biomarker. However, the ischemic event in the lesioned cortex also triggers structural and functional alterations in its contralateral homolog through the corpus callosum (CC), known as transcallosal diaschisis. The few studies that have characterized the microstructural status of the CC using DTI only examine its relationship with paretic limb performance. Given the well-established role of the CC for bimanual coordination, especially fibers connecting the larger sensorimotor networks such as prefrontal, premotor and supplementary motor regions, we examine the relationship between the microstructural status of the CC and bimanual performance in chronic stroke survivors (n = 41). We used movement times for two self-initiated and self-paced bimanual tasks to capture bimanual performance. Using publicly available control datasets (n = 52), matched closely for acquisition parameters, including sequence, diffusion gradient strength and number of directions, we also explored the effect of age and stroke on callosal microstructure. We found that callosal microstructure was significantly associated with bimanual performance in chronic stroke survivors such that those with lower callosal FA were slower at completing the bimanual task. Notably, while the primary sensorimotor regions (CC3) showed the strongest relationship with bimanual performance, this was closely followed by the premotor/supplementary motor (CC2) and the prefrontal (CC1) regions. We used multiple mixed regression to systematically account for loss of callosal axons (i.e., normalized callosal volume) as well as differences in lesion size and other metrics of structural damage. Chronic stroke survivors presented with significantly greater loss of callosal fiber orientation (lower mean FA) compared to neurologically intact, age-similar controls, who in turn presented with lower callosal FA compared to younger controls. The effect of age and stroke were observed for all regions of the CC except the splenium. These preliminary findings suggest that in chronic stroke survivors with relatively localized lesions, callosal microstructure can be expected to change beyond the primary sensorimotor regions and might impact coordinated performance of self-initiated and cooperative bimanual tasks.