Short Amplicon-Length PCR Assay Targeting Mitochondrial Cytochrome b Gene for the Detection of Feline Meats in Burger Formulation

Consumption or mixing of feline ingredients in halal and kosher foods is forbidden, and various diseases such as SARS, anthrax, and hepatitis could be transmitted through feline meats. However, since feline species are abundant across the world without any market price and their meats are consumed in exotic foods, the chances of their adulteration in common meats are very high. Several recent reports appreciated short amplicon-length PCR assays for species authentication in processed foods assuming that shorter targets would be thermodynamically more stable than longer ones under natural decomposition and food processing treatments. However, scientific evidence to prove this hypothesis is rarely documented. For the first time, we developed here a PCR assay targeting only a 69-bp site of feline mitochondrial cytochrome b gene, and its authenticity was confirmed by AluI restriction enzyme followed by its separation and detection on a lab-on-a-chip-based automated electrophoretic system. The exceptional target stability was systematically proven over the previously documented shortest target (108 bp) under extreme autoclaving and microwaving treatments both in pure and mixed matrices. The assay specificity was tested against 14 terrestrial and aquatic species commonly consumed in foods, and no cross-species detection was observed. The limit of detection of the assay was 0.1 pg of feline DNA and 0.01 % (w/w) of feline meats in raw meats and cooked burgers, respectively.