Understanding the genetic diversity and population structure of fish species is crucial for the sustainable use and protection of fish germplasm resources. Hemibagrus guttatus (Bagridae, Siluriformes) is widely distributed in the large subtropical Pearl River (China) and is a commercially important fish. H. guttatus populations have been declining due to over-fishing, water pollution, and hydraulic constructions, it is now a wild animals under second class national protection. However, the genetic diversity of wild H. guttatus is not clear, despite its important ecological significance and its significance for conservation. In this paper, genes mitochondrial cytochrome c oxidase subunit I (COI) and cytochrome b (Cyt b) were used to analyze the genetic structure and found that there is degradation in germplasm resources of H. guttatus that could destabilize the sustainable use of this species.