High-Resolution Melting Analysis of COI Sequences Distinguishes Pufferfish Species (Takifugu spp.) in China

Pufferfish is a traditional, delicious dish in Asia. However, eating wild or improperly processed pufferfish causes serious poisoning. This study aimed to exploit the high-resolution melting (HRM) method for authenticating four species of Takifugu pufferfish (Takifugu xanthopterus, T. fasciatus, T. flavidus, and T. rubripes). Candidate DNA barcodes, including the cytochrome c oxidase subunit I (COI), cytochrome oxidase b (Cytb), and the control region (D-loop), were analyzed, with COI selected as the optimal DNA barcode. An HRM method was developed to identify 57 commercial fish samples in China, including 33 commercial pufferfish products and 24 unlabeled fish products. The findings revealed that the pufferfish products were T. rubripes or T. fasciatus, and four T. xanthopterus samples were detected in unlabeled fish products. These results showed that DNA barcode coupled with HRM analysis was a rapid and efficient tool to identify pufferfish, which might aid in the prevention of consumer fraud or mislabeling of fish products.