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Study Design: A retrospective, cross-sectional cohort study., Objective: This study aims to investigate the association between paraspinal muscle parameters and single-segment degenerative lumbar spondylolisthesis (DLS)., Summary of Background Data: The relationship between lumbar paraspinal muscles morphology and single-segment DLS remains unclear., Methods: A retrospective review was conducted on 115 patients with L4/5 single-segment DLS and 105 subjects without DLS. Two independent investigators assessed the relative cross-sectional area and fat infiltration rate of the multifidus, erector spinae, and psoas major at L3/4, L4/5, and L5/S1 levels, comparing these measurements between the two groups. Additionally, binary logistic regression analysis was performed with DLS as the dependent variable to analyze the relative cross-sectional area and fat infiltration rate of different paraspinal muscles. Within the DLS group, the correlation between paraspinal muscle characteristics and the anteroposterior diameter of the spinal canal was examined., Results: The fat infiltration rate of multifidus, erector spinae, and psoas major were higher in the DLS group than in the control group, while the relative cross-sectional area of multifidus and psoas major were lower in the DLS group. Binary logistic regression analysis revealed a significant correlation between fat infiltration rate of multifidus and psoas major and DLS. The relative cross-sectional area of multifidus and erector spinae was significantly smaller below the affected segment in the DLS group compared to the control group. A significant positive correlation was observed between the relative cross-sectional area of multifidus and erector spinae and the anteroposterior diameter of the spinal canal., Conclusion: There is a close association between paraspinal muscle degeneration and single-segment DLS, with increased relative cross-sectional area of the multifidus and psoas major possibly being risk factors for single-segment DLS. The restoration or enhancement of paraspinal muscle function could potentially serve as a pivotal target for the prevention and treatment of single-segment DLS., Level of Evidence: III., (Copyright © 2024 The Author(s). Published by Wolters Kluwer Health, Inc.)