Direct Sequencing Method for Species Identification of Canned Sardine and Sardine-Type Products

A direct sequencing method based on a 103 bp diagnostic sequence derived from a species-specific mitochondrial DNA cytochrome b sequence of 150 bp obtained by Polymerase Chain Reaction was tested for the identification of 47 commercial canned sardine and sardine-type products from various countries. Multiple alignment of 14 analyzed reference samples belonging to Clupeomorpha species was performed versus the canned samples. Low intraspecific variability was observed for canned sardine (</=0.03), whereas mean interspecific variability was 0.23. A phylogenetic tree was constructed, and the calculated bootstrap values (BP, 88-99%) were used as indicators of the correct assignment of unknown canned samples to reference species. According to this methodology, the 26 commercial canned sardines analyzed were grouped in the same clade as the Sardina pilchardus reference and identified unequivocally. These assignments were confirmed by the high BP value of 99%.