013: Rapid and Reliable Silkworm Identification in Food Products Using a CRISPR/Cas12a Coupled LAMP Assay

Introduction

Edible insects are gaining attention as future foods due to their high protein content, low breeding costs per unit area, and low greenhouse gas emissions. As the edible insect market expands, concerns such as adulteration and allergies are increasingly coming into focus. Silkworms, in particular, have been traditionally consumed, and their low cost coupled with reported allergy incidents necessitating careful management. This study developed a detection method for silkworms to ensure food safety and protect individuals from potential allergy incidents.

Methods

The CRISPR/Cas12a coupled LAMP assay was developed for the rapid on-site detection of silkworm. Silkworm-specific LAMP primers and guide RNA were designed based on the cytochrome c oxidase subunit I (COI) of mitochondrial DNA. Ten ng of genomic DNA from twenty-two samples, including ten insect species, nine seafoods, and three meats, were used to test specificity. Sensitivity was determined by serially diluting silkworm DNA from 1 ng to 0.0001 ng and silkworm-wheat mixed sample with varying amount of silkworm, ranging from 1% to 0.005%. The applicability of developed methods was evaluated using forty-five commercial processed edible insect products.

Results

The CRISPR/Cas12a coupled LAMP assay was developed for detecting silkworm, with its specificity validated using genomic DNA from various insects, seafood, and meats. The limit of detection was about 0.0001 ng of silkworm DNA and 0.01% of silkworm-wheat mixed sample. Furthermore, signal intensity was observed in only twenty-one out of the forty-five commercial processed edible insect products that contained silkworms. These results demonstrate that the developed assay provides a reliable and practical solution for food authentication and food safety.

Significance

The CRISPR/Cas12 coupled LAMP assay developed in this study is both specific and sensitive for detecting silkworms in food. It also presents benefits such as being rapid, simple, and suitable for on-site applications, unlike conventional methods that require a thermal cycler. This method is anticipated to be used in the future to prevent mislabeling, adulteration, and allergic incidents.

Authors: Seung-Man Suh, Sumin Kim, Hae-Yeong Kim