COIBar-RFLP Molecular Strategy Discriminates Species and Unveils Commercial Frauds in Fishery Products

The DNA analysis is the best approach to authenticate species in seafood products and to unveil frauds based on species substitution. In this study, a molecular strategy coupling Cytochrome Oxidase I (COI) DNA barcoding with the consolidated methodology of Restriction Fragment Length Polymorphisms (RFLPs), named COIBar-RFLP, was applied for searching pattern of restriction enzyme digestion, useful to discriminate seven different fish species (juveniles of <i>Engraulis encrasicolus</i> and <i>Sardina pilchardus</i> sold in Italy as "bianchetto" and <i>Aphia minuta</i> sold as "rossetto"; icefish <i>Neosalanx tangkahkeii</i>; European perch, <i>Perca fluviatilis</i> and the Nile Perch, <i>Lates niloticus;</i> striped catfish, <i>Pangasianodon hypophthalmus</i>). A total of 30 fresh and frozen samples were processed for DNA barcoding, analyzed against a barcode library of COI sequences retrieved from GenBank, and validated for COIBar-RFLP analysis. Cases of misdescription were detected: 3 samples labeled as "bianchetto" were substituted by <i>N. tangkahkeii</i> (2 samples) and <i>A. minuta</i> (1 sample); 3 samples labeled as "persico reale" (<i>P. fluviatilis</i>) were substituted by <i>L. niloticus</i> and <i>P. hypophthalmus.</i> All species were simultaneously discriminated through the restriction pattern obtained with <i>Msp</i>I enzyme. The results highlighted that the COIBar-RFLP could be an effective tool to authenticate fish in seafood products by responding to the emerging interest in molecular identification technologies.