Monitoring Yellow Mealworm (Tenebrio molitor) as a Potential Novel Allergenic Food: Effect of Food Processing and Matrix

Abstract

The consumption of insects has increased in western countries, raising concerns about their potential to induce food allergic reactions in sensitized/allergic individuals. This work intended to develop a real-time PCR approach for the detection/quantification of yellow mealworm (Tenebrio molitor) as a potential allergenic food in complex matrices. For this purpose, reference mixtures simulating the production of pork sausages and wheat biscuits containing known amounts of mealworm were used. Real-time PCR with TaqMan probe targeting the cytochrome b gene of T. molitor was able to detect up to 2 fg of insect DNA, and 1.0 and 0.1 mg/kg of mealworm flour in autoclaved sausages and baked biscuits, respectively. Generally, the method showed acceptable analytical performance parameters, confirming its suitability/applicability for a wide range of foods. However, real-time PCR data showed significant differences among food matrix and processing, highlighting the importance of using appropriate calibration models for quantitative analysis. Finally, the real-time PCR approach was successfully validated with blind mixtures and applied to commercial samples, demonstrating its efficacy and reliability in the quantification of mealworm in processed foodstuffs.

Keywords: Tenebrio molitor; food matrix; heat processing; insects; novel allergens; real-time PCR.

Figure 1

Amplification curves (a) and respective calibration curve (b) obtained by real-time PCR with TaqMan probe targeting the cytochrome b gene and using serially diluted (1/10) DNA extracts of T. molitor from 20 ng to 2 fg (n = 4 replicates). Cq—cycle of quantification.

Figure 2

Calibration curves obtained by real-time PCR with TaqMan probe targeting cytochrome b gene using sausages (a) and biscuits (b) model mixtures before and after thermal treatment, containing 100,000 mg/kg; 10,000 mg/kg; 1000 mg/kg; 100 mg/kg; 10 mg/kg; 1 mg/kg; and 0.1 mg/kg (w/w) of mealworm (n = 8 replicates). For the same spiking level, * means statistically significant differences between raw and processed mixtures (p < 0.05), following t-test analysis.

Figure 3

Comparison of the dynamic range of the calibration curves obtained by real-time PCR targeting cytochrome b gene according to the type of thermal treatment and food matrix (mean ± SD of n = 8 replicates from 2 independent runs). Different letters for the same spiking level mean statistically significant differences between matrices (p < 0.05) following ordinary one-way ANOVA or ANOVA analysis.