DNA Based Tools to Deter Fraudulent Incidents in Fishery Sector: An Update on the Bulgarian Market Survey

Species substitution represents a major issue in fishery resulting in economic and public health consequences. Therefore, European Union set a regulatory framework for seafood traceability encouraging the use of DNA-based methods for deterring fraudulent incidents. A first DNA barcoding survey on labelling compliance of seafood sold on the Bulgarian market, highlighted a mislabelling rate of 17.7% plausibly due to both food business operators improper training on species identification and labeling and few major deliberate substitutions of high-value with less valuable white fish species. This considered, a second survey on labelling compliance was performed between 2019 and 2020 and 147 products (white fish N=100; cephalopods N=28, molluscs bivalves N=16 and gastropods N=3) were collected. A COI gene fragment was amplified and sequenced using universal primers. In case of failure of species allocation by COI, a 16SrRNA gene fragment was applied as alternative target. The sequences were queried against reference databases. Molecular species identity was univocally assigned for identity percentage rates > 98% for COI gene and =100% for the 16SrRNA gene. Finally, a RFLP analysis on the non-repetitive region of the nuclear Polyphenolic Adhesive Foot Protein (PAP) was included in the study for the identification of 13 mollusk bivalve products belonging to Mytilus sp. Within fish and cephalopods products were generally allocated at species level confirming COI effectiveness in species discrimination. As fish, an overall mislabelling rate of 14% was found due to substitutions of Gadus chalcogrammus, Gadus morhua or Merluccius productus with less valuable Gadiformes species. Within cephalopods category, a mislabelling rate of 10.7% was highlighted due to the replacement of Ommastrephes bartramii and Todarodes pacificus with Dosidicus gigas known as a frequent substituent species. As regards molluscs bivalves, the use of a multi-target approach and RFLP analysis allowed the univocal species identification of all the products highlighting a final mislabelling rate of 18%. No mislabelling incidents were highlighted for gastropods. The overall mislabelling rate of 13.6% (20/147) emphasize the relevance of the application of molecular tools for verifying the products’ identity to protect both seafood supply chain and consumers’ rights.