*2.6. Tractography Based on Navigated Transcranial Magnetic Stimulation*

Deterministic fiber tractography was based on nTMS language mapping data without additional anatomical seeding (Brainlab Elements, version 3.1.0; Brainlab AG, Munich, Germany). Both export datasets were first transferred to an external server and then auto-fused with the MRI sequences of the respective patient, using manual correction in case of registration misalignments and eddy current correction for DTI data. The datasets containing language-positive or language-negative nTMS points were separately defined as three-dimensional objects, followed by generation of regions of interest (ROIs) out of these objects by adding a rim of 5 mm to each stimulation point [14,15,17–19].

An individual fractional anisotropy (FA) value, the fractional anisotropy threshold (FAT), was defined separately in each patient for nTMS-based DTI FT using the ROI constituted of language-positive or language-negative nTMS points, which was determined by setting angulation to 90◦ , the minimum fiber length (FL) to 30 mm, and increasing the FA stepwise until no fibers were displayed, followed by decreasing the FA by 0.01, thus visualizing a minimum fiber course. The corresponding FA value was defined as 100% FAT. A similar approach using a higher minimum FL for FAT determination has been used previously for nTMS-based DTI FT [17,19,47].

After FAT definition, nTMS-based DTI FT was carried out separately with the ROIs of language-positive and language-negative nTMS points using the respective FA values for 100% FAT, 75% FAT, 50% FAT, and 25% FAT, and a minimum FL of 30 mm as well as 3 mm, respectively. The values of 30 and 3 mm were considered for the minimum FL due to the characteristic length of U-fibers that have been described to be typically in this range [48].
