Accumulating studies have exploited the qPCR method for the qualitative and quantitative detection of fish adulteration, including the rapid identification of Atlantic salmon, Atlantic cod ( Gadus morhua), and European plaice ( Pleuronectes platessa), as well as the accurate quantification of two closely related tuna species ( Thunnus obesus and Thunnus albacares) in a binary mix in tuna cans. Of these, real-time quantitative polymerase chain reaction (qPCR) testing is the most outstanding representative. To date, DNA-based methods are the most widely applied for identifying and quantifying fish adulteration. Spectral and chromatographic techniques have been developed and applied to the detection of salmon adulteration, but these suffer from suboptimal levels of specificity and sensitivity. Traditionally, the identification of rainbow trout and Atlantic salmon mainly relies on their morphological features however, this is a difficult task since Atlantic salmon is often sold in processed forms in the market, as fillets or blocks, dried or grilled, and canned or smoked. Therefore, efficient and accurate methods for the detection of Atlantic salmon adulterated with rainbow trout are urgently needed. These incidents of fraud not only harm consumers’ economic interests, but also increase health and safety risks, as rainbow trout cultured in freshwater may have a high risk of parasite infections. Driven by high economic profits, the substitution of low-cost rainbow trout for high-priced Atlantic salmon as well as its adulteration frequently occur at every stage of the food supply chain. Due to its rapid growth rate and strong environmental tolerance, rainbow trout is becoming one of the most common cultured fish in China, and hence, its price is much lower than that of Atlantic salmon in the Chinese market. Rainbow trout ( Oncorhynchus mykiss) is a freshwater fish belonging to the Salmonidae family, which is visually similar to Atlantic salmon in terms of meat color and texture. Due to the huge demand for salmon in the Chinese market, foreign imports of Atlantic salmon have rapidly increased in China in recent years. Overall, the ddPCR-based quantitative method exhibited high levels of accuracy, stability, sensitivity, and practicability, suitable for applications in the routine surveillance and quality assurance of salmon products.Ītlantic salmon ( Salmo salar) is highly appreciated worldwide for its unique taste and rich nutrient content, which consists of significant amounts of polyunsaturated fatty acids and essential trace elements crucial for human health. In comparison to real-time quantitative PCR (qPCR) testing, the established ddPCR method exhibited a higher level of stability and accuracy. The method was successfully applied to detect rainbow trout adulteration in commercial raw and processed Atlantic salmon products. Thermal and freezing treatments, as well as adding food additives within the recommended dosage limits, had no significant effect on the quantification accuracy. The quantification method demonstrated an acceptable level of repeatability and reproducibility, as the values of the relative standard deviation (RSD) for the tested meat mixtures with the known fractions were all less than 5%. The dynamic range of the established ddPCR method was from 1% to 90%, with a limit of detection (LOD) of 0.2% and a limit of quantification (LOQ) of 0.8% for rainbow trout in Atlantic salmon, respectively. A quantitative formula for calculating the mass fraction of adulterated Atlantic salmon with rainbow trout was established based on a one-step conversion strategy, in which the DNA copy number ratios were directly transformed to meat mass fractions by introducing a fixed constant (the transfer coefficient). Species-specific primers and probes were designed targeting the single-copy nuclear gene myoglobin of two salmonids. In this study, a precise method for identifying and quantifying adulterated Atlantic salmon with rainbow trout based on droplet digital PCR (ddPCR) testing was developed. Hence, reliable techniques are urgently needed to detect Atlantic salmon adulteration. Low-cost fish species are often used to adulterate or substitute for Atlantic salmon products, posing a serious threat to market order and public health.
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