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In the present study the population genetic structure of European catfish in the Anzali Lagoon and Aras Lake were examined using microsatellite markers. Sixty fin clip samples of Silurus glanis from two regions were collected and for genetic analysis 6 microsatellite loci were used to assess the population genetic structure of S. glanis. There were significant differences based on average number of alleles per locus and heterozygosity between two populations (p<0.01). The observed heterozygosity (Ho) for each population (He) per locus was from 0.297 to 0.733 in Aras lake and Anzali lagoon samples, respectively.The Analysis of molecular variance (AMOVA) indicated that the proportion of the genetic variation attributed to differences among populations of the S. glanis was highly significant for both FST and RST (FST = 0.165, RST = 0.38, p<0.001). Excess or lacks of heterozygosity was observed but most of used microsatellite loci in selected areas were at Hardy-Weinberg equilibrium. Our finding showed the two populations are genetically separated, therefore fisheries management and restocking program of this species especially in Anzali Lagoon is recommended.