[Aim]  Analyzing the genetic diversity and genetic structure characteristics of the pine sawyer beetle Monochamus alternatus in the Guangxi Zhuang Autonomous Region, to deepen the understanding of population differentiation and ecological adaptation patterns of the M. alternatus. [Methods]  Variation in the sequence of the mitochondrial cytochrome coxidase subunit Ⅰ (COⅠ) gene between specimens from 11 geographical populations in Guangxi was measured and compared. Bioedit and DnaSP 5.0 software were used to analyze the genetic diversity of M. alternatus, and Arlequin 3.5 software to perform a neutrality test and analyze genetic differentiation. Popart 11 software was used to draw a haplotype network diagram, and MEGA 6 to draw a phylogenetic tree including other domestic and foreign specimens. [Results]  In the analysis of 635 bp COⅠ sequences from 167 individuals of M. alternatus, 29 variable sites were detected, accounting for 4.57% of all sites. The overall haplotype diversity was 0.858, and nucleotide diversity was 0.004 52. The genetic distances among the 11 geographic populations ranged from 0.001 to 0.010, and the Mantel test results indicated no significant correlation between genetic distance and geographic distance (P > 0.05). AMOVA analysis showed that within-population genetic variation was higher than between-population variation (60.09% > 39.91%), with Fst values ranging from -0.047 to 0.889, indicating varying degrees of genetic differentiation among different populations. The haplotype network diagram divided 32 haplotypes into 3 evolutionary centers, with Hap8 being the ancestral haplotype, and shared haplotypes accounting for 34.38% of all haplotypes. [Conclusion]  The M. alternatus in Guangxi exhibits high genetic diversity, strong potential for genetic variation, and environmental adaptability. Factors such as human activities, environmental characteristics, and social features influence the genetic structure of M. alternatus populations in Guangxi, enabling gene flow over long distances.