A multiplex PCR forensic protocol for the molecular certification of sea catfishes (Ariidae - Siluriformes) from coastal Amazon, Brazil

The marine catfishes (family Ariidae) are important fisheries resources, particularly along the Amazon coast in northern Brazil. Whole fresh specimens and fish fillets are commercialized in both national and international markets, including several records of species substitution and frauds in the trade of processed products. In addition, the life traits of marine catfish species make them particularly vulnerable to overexploitation. Therefore, we developed a molecular certification system for the authentication of these species based on DNA markers. Specific and universal primers were used for the amplification of the mitochondrial Cytochrome b (Cytb) gene and combined with primers for the 16S ribosomal RNA gene (control marker) to provide the first forensic molecular protocol based on multiplex PCR for marine catfishes from Brazilian Amazon coast. Accordingly, distinct band profiles were obtained for each of the most exploited species along this region: Bagre bagre, Notarius grandicassis, Sciades couma, Sciades herzbergii, Sciades parkeri, Sciades passany, and Sciades proops. To amplify the band pattern required for species identification, a concentration of 20 ng/ul was required. In the cross-amplification assay, other fish species were discriminated from the Ariidae bands. The multiplex PCR was about 35x cheaper and saved 13 h of labor to identify the seven species of Ariidae when compared to DNA sequencing. Thus, this protocol represents a viable and efficient tool to the certification of processed products by inspection agencies and to assure the consumer's rights, eventually leading to an improved management in fisheries. The fishing industry might benefit from the multiplex protocol for the authentication of commercial products, adding market value to them based on DNA certification labels.