Development of a Taqman-based real time PCR method for the detection of the banded cricket (Gryllodes sigillatus) in feed

The demand for sustainable protein sources has led to a growing interest in the use of insects in animal feed. This study focuses on the detection of banded crickets ( Gryllodes sigillatus ) in feed using a real time PCR method. The research aims to address the challenges associated with identifying insects in feed, in light of European regulations governing the use of processed animal proteins. The methodology involved the development of a specific probe, to be combined with a previously developed primer pair. Method’s specificity, efficiency and sensitivity were internally validated. The technical detection limit was 4 pg of DNA/reaction. The method was able to successfully detect target species G. sigillatus at a concentration of 0.1% in various model feed samples. The results indicate that the developed real time PCR method reliably detects G. sigillatus in feed, providing a means to ensure compliance with regulatory requirements. Judging from similar cases, the method appears to be transferable to food analysis, subject to verification of practical sensitivity, especially in ultra-processed foods. This research contributes to ongoing efforts to promote the use of insects as a sustainable source, emphasizing the importance of accurate species identification in the context of food safety and regulatory compliance. • Development of a real time PCR protocol to detect Gryllodes sigillatus in feed • Reliable, robust and selective identification in mixed feed compounds • Increased method specificity and versatility