A novel screening approach based on six real-time PCR systems for the detection of crustacean species in food

•A real time PCR based screening assay was developed for the detection of crustaceans in food.•Economical relevant shrimps, lobster and prawn species are detected in one analytical step.•The Chinese mitten crab and the Common shrimp are captured by the assay.•Pre-spotted primer and probes on 96-well PCR plates makes the assay easy-to-use and flexible.•Practical applicability was proven with market samples.

Analysis of crustacean species plays a role in authenticity issues as well as allergen detection. A real-time PCR-based screening assay was developed for the detection of crustaceans in food. In order to cover most relevant species in one analytical step, PCR systems were newly developed for the detection of shrimps (Penaeidae), lobster (Homarus sp.), Common shrimp (Crangon crangon), river prawns (Macrobrachium sp.) and Chinese mitten crab (Eriocheir sinensis). In addition a published system targeting Northern prawn (Pandalus borealis) was selected. All PCR-systems are based on mitochondrial 16S rRNA gene sequences and were optimized to be run with a standard program at a universal annealing temperature of 60 °C. Validation experiments confirmed a sensitivity of the PCR systems of 0.01-0.1 genome copies or 0.04-2.5 pg DNA, respectively. Specificity was demonstrated with 25,000 copies of pure genomic DNA from about thirty plant and animal species relevant in food and the performance in food matrix was evaluated. Finally primer and probes were pre-spotted steadily on 96-well PCR plates and the practical applicability of the assay was proven with selected food products. The assay enables detection of multiple species of market relevance.