Abstract: To compare the otolith morphology of different Larimichthys polyactis populations, and to further clarify their population division, we applied the landmark method better at extracting morphological features for the first time, to explore the morphological differences of otoliths from L. polyactis populations in the Southern Yellow Sea and East China Sea. From March to December 2022, 30 individuals of 1-year old fish had been collected from each from Haizhou Bay (HZB), Lyusi (LS), Yangtze Estuary (YTE) and Zhoushan (ZS) fishing grounds. After extracting the left sagittal otoliths, we seclected even landmarks in otolith contour and four landmarks in sulcus acusticus for the landmark analysis. Results show that: 1) Among the 11 landmarks, those in sulcus acusticus (Landmark 8−11) had main contribution (91.88%) in explaining the main source of morphological variation, while those in otolith contour (Landmark 1−7) just had 8.12%, which reveals that the differences in otolith morphology among the four geographical groups mainly came from the auditory sulcus of otolith. 2) According to the scatter plot of principal component analysis, part of the Haizhou Bay population was significantly divided into one group, while the remaining Haizhou Bay fishing grounds and all other three fishing grounds had highly overlapped and clustered into another group, indicating that there might be two L. polyactis populations in the Southern Yellow Sea area, namely the Southern Yellow Sea-East China Sea group and the Northern Yellow Sea-Bohai Sea group. This also indicates that there were mixed L. polyactis populations in the Haizhou Bay fishing grounds. This may be explained by the long-term egg and larval dispersal process controlled by the current and active migratory routes during feeding and overwintering period. The results verified the potential of otolith morphology in delineating the populations of L. polyactis, providing valuable references for their spatiotemporal distribution.