Detection of Fish-Derived Ingredients in Animal Feeds by a TaqMan Real-Time PCR Assay

Animal species identification in food and feed has gained increasing interest in recent years due to public health, economic and legal concerns. In the present work, a TaqMan real-time PCR assay targeting the mitochondrial 12S rRNA gene was developed for detection of a specific DNA fragment (87 bp) common to the most economically important fish families in fish-derived ingredients used in feed formulations. Performance of the assay was evaluated against stringent acceptance criteria in terms of specificity and sensitivity. The TaqMan real-time fish-specific system proved to be highly sensitive, allowing the detection of 0.1 pg of fish DNA. The assay was successfully applied to the authentication of two types of real-world compound feeds: industrial farm animal feeds and commercial pet foods, allowing detection of small fish percentages in the samples. The quantitative potential of the real-time PCR assay was further performed, concluding that, although results were well reproducible under the experimental conditions tested, an accurate estimation of the target content in feeds is not feasible in practice. The reported methodology may be useful as a molecular analytical tool to support fisheries control and enforcement, as well as the verification of authenticity of fish and fish-derived products in the food and feed sectors.