Abstract: We collected 254 individuals of nine wild Pelteobagrus fulvidraco populations from Yangtze River and Huaihe River in Anhui Province, then genotyped them by using ten microsatellite markers. The result shows a high level of genetic diversity. Altogether 245 alleles had been detected in ten loci with an average of effective allele number (N_e) of 9.75 per locus. In all P. fulvidraco populations, the allele number (N_a) was 5.20−14.80 and the N_e was 2.64−8.93; the observed heterozygosity (H_o) was 0.496−0.671 and the expected heterozygosity (H_e) was 0.557−0.818; the polymorphism information content (PIC) was 0.500−0.790. The molecular variance analysis (AMOVA) exhibited only 8.15% genetic variation among the populations and showed a low and middle level genetic differentiation (F_ST: 0.006−0.236). The UPGMA tree of P. fulvidraco based on Nei's genetic distance (0.079−0.640) and population genetic structure analysis based on Structure software both indicate that the nine populations belonged to four or five genetic lineages; ST, FN and MC populations were all independent groups which were genetically distantly related to the others. The results reveal that there is genetic differentiation among the nine P. fulvidraco populations and some populations have experienced bottleneck effects, but the wild resources still have high genetic diversity.