To visualize the course of the tympanic segment of chorda tympani nerve (CTN) using ultra-high-resolution computed tomography (U-HRCT).A hundred and fourteen ears with no evident otologic pathologies were included. The tympanic segment of CTN was divided into four portions: periannular, posteromalleal, malleal, and anteromalleal. The length of the periannular portion running along the tympanic annulus was recorded. Four points of interest (the beginning and end of the posteromalleal and anteromalleal portions) were selected to perform distance measurements relative to the tip of the malleus manubrium. Differences in lengths and distances were compared in terms of ear sides and sexes.The length of the periannular portion was 2.49 ± 1.16 mm. The beginning of the posteromalleal portion was located more laterally on the right side than on the left side (mean: 4.09 mm vs. 3.92 mm, P = 0.016). The end of the posteromalleal portion was located more inferiorly on the right (mean: 2.11 mm vs. 2.26 mm, P = 0.018). The beginning of the anteromalleal portion on the right was located more laterally than that on the left (mean: 2.60 mm vs. 2.45 mm, P = 0.027). The start and end of the anteromalleal portion were more posteriorly located in women than in men (both Ps < 0.001).The course of the tympanic segment of normal CTN was comprehensively visualized by U-HRCT. Preoperative evaluation of the tympanic segment of CTN might be helpful in avoiding iatrogenic injury during middle ear surgery.Copyright © 2022 The Author(s). Published by Elsevier Inc. All rights reserved.