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Human brain connectivity is extremely complex and variable across subjects. While long association and projection bundles are stable and have been deeply studied, short association bundles present higher intersubject variability, and few studies have been carried out to adequately describe the structure, shape, and reproducibility of these bundles. However, their analysis is crucial to understand brain function and better characterize the human connectome. In this study, we propose an automatic method to identify reproducible short association bundles of the superficial white matter, based on intersubject hierarchical clustering. The method is applied to the whole brain and finds representative clusters of similar fibers belonging to a group of subjects, according to a distance metric between fibers. We experimented with both affine and non-linear registrations and, due to better reproducibility, chose the results obtained from non-linear registration. Once the clusters are calculated, our method performs automatic labeling of the most stable connections based on individual cortical parcellations. We compare results between two independent groups of subjects from a HARDI database to generate reproducible connections for the creation of an atlas. To perform a better validation of the results, we used a bagging strategy that uses pairs of groups of 27 subjects from a database of 74 subjects. The result is an atlas with 44 bundles in the left hemisphere and 49 in the right hemisphere, of which 33 bundles are found in both hemispheres. Finally, we use the atlas to automatically segment 78 new subjects from a different HARDI database and to analyze stability and lateralization results.

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White matter bundle segmentation using diffusion MRI fiber tractography has become the method of choice to identify white matter fiber pathways in vivo in human brains. However, like other analyses of complex data, there is considerable variability in segmentation protocols and techniques. This can result in different reconstructions of the same intended white matter pathways, which directly affects tractography results, quantification, and interpretation. In this study, we aim to evaluate and quantify the variability that arises from different protocols for bundle segmentation. Through an open call to users of fiber tractography, including anatomists, clinicians, and algorithm developers, 42 independent teams were given processed sets of human wholebrain streamlines and asked to segment 14 white matter fascicles on six subjects. In total, we received 57 different bundle segmentation protocols, which enabled detailed volume-based and streamline-based analyses of agreement and disagreement among protocols for each fiber pathway. Results show that even when given the exact same sets of underlying streamlines, the variability across protocols for bundle segmentation is greater than all other sources of variability in the virtual dissection process, including variability within protocols and variability across subjects. In order to foster the use of tractography bundle dissection in routine clinical settings, and as a fundamental analytical tool, future endeavors must aim to resolve and reduce this heterogeneity. Although external validation is needed to verify the anatomical accuracy of bundle dissections, reducing heterogeneity is a step towards reproducible research and may be achieved through the use of standard nomenclature and definitions of white matter bundles and well-chosen constraints and decisions in the dissection process.