Abstract: Background The Manila clam (Ruditapes philippinarum) is one of the important naturally distributed bivalve species in East Asia and is widely cultured in China. Objective This study aims to comparatively analyze the genetic diversity between cultured and wild populations of R. philippinarum. Methods A total of 162 samples from two cultured populations (Fujian, China) and three wild populations (Laizhou, China; Nampo and Haeju, North Korea) were analyzed by amplifying and sequencing a partial segment of the mitochondrial COⅠ gene. Results Within the 646 bp COⅠ gene fragment, a total of 63 polymorphic sites were detected across five populations, defining 59 haplotypes. The haplotype diversity indices (Hd) ranged from 0.880 to 0.938, and nucleotide diversity indices (π) varied between 0.001 and 0.010 among the five populations. Wild populations exhibited haplotype diversity indices of 0.905~0.938 and nucleotide diversity values of 0.005~0.010, with 41 haplotypes identified, accounting for 69.5% of the total haplotypes. Cultured populations showed haplotype diversity of 0.880~0.902 and nucleotide diversity of 0.001~0.006, defining 22 haplotypes, which represented 37.3% of the total haplotypes. AMOVA analysis revealed a low genetic differentiation index (F_st=0.23) between cultured and wild populations, accounting for only 8.13% of the total variation, which was not statistically significant (P>0.01). Most genetic variation (80.34%, P<0.001) originated from differences among individuals within populations. Pairwise F_st and genetic distance analyses indicated significant divergence (P<0.01) between North Korean and Chinese populations, while no significant differentiation was observed among other populations (P>0.01). Phylogenetic trees and network analysis showed that the five populations formed two distinct genetic clusters, primarily influenced by geographic isolation. Conclusion The findings suggest that aquaculture practices have not significantly affected the genetic diversity of R. philippinarum. However, wild populations exhibited higher genetic diversity and more unique haplotypes compared to cultured populations. This implies that the reduced genetic diversity in cultured populations may result from smaller effective population sizes and inbreeding within closed-cycle farming systems. The study provides valuable insights for the sustainable management and genetic breeding of R. philippinarum in future aquaculture.