Detection of Alphitobius diaperinus by Real-Time Polymerase Chain Reaction With a Single-Copy Gene Target

Abstract

Use of edible insects as an alternative source of proteins in food and feed is increasing. These last years, numerous companies in Europe have started producing insects for food and feed purposes. In the European Union, the use of edible insects for human consumption falls within Regulation (EU) No. 2015/2283 on novel foods. For feed, Commission Regulation (EU) 2017/893 authorizes seven insect species as processed animal proteins for aquaculture. Methods of authentication are required to check the conformity of the products. In this study, we propose a real-time polymerase chain reaction (PCR) method for the specific detection of the lesser mealworm (Alphitobius diaperinus), one of the species included in the shortlist of authorized insects. The selected target is the cadherin gene with a single-copy (per haploid genome) illustrated by our experimental evidence. The PCR test amplified a 134-bp fragment of the cadherin gene. The qualitative method was assessed toward several performance criteria. Specificity was checked against 54 insect species next to other animal and plant species. The sensitivity, efficiency, robustness, and transferability of the PCR assay were also successfully tested. Finally, the applicability of the test was assessed on real-life processed samples (industrial meals) of A. diaperinus. The study also showed that there seems to be a huge confusion on the correct labeling of the marketed mealworms. We did not succeed to get Alphitobius laevigatus samples. They all appeared to belong to the A. diaperinus taxon.

A validated real-time PCR test for simultaneous detection of Gallus gallus and Meleagris gallopavo in feed instead of an impossible poultry test

“…The quantity of recovered plasmid DNA was converted into copy numbers as usual (Debode et al, 2010;Marien et al, 2018;Marien et al, 2022), with similar assumptions taken into consideration as previously (Marien et al, 2018): -1 unit of absorbance at 260 nm corresponds to a concentration of 50 µg . ml -1 ; -the mean molar weight of one base pair is set at 635 Da.…”

“…The efficiency of the PCR assay was calculated as previously (Marien et al, 2022) using a series of dilutions of plasmid material at target levels of 5000, 2500, 1000, 500 and 100 copies. Each dilution was analyzed in six replicates and on four runs.…”

Alphitobius diaperinus larvae (lesser mealworm) as human food – An approval of the European Commission – A critical review

“…The lesser mealworms are processed and sold in the form of freeze-dried snacks or processed foods such as pasta, burger patties or snack bars (Van Huis et al, 2013;Foodnavigator, 2018;Nutraingredients, 2018). The lesser mealworms are sometimes marketed as buffalo worms, which might be confusing due to the same term used for larvae of Alphitobius laevigatus (Fabricius, 1781) (Marien et al, 2022).…”

Mealworms for food and feed: are they only yellow?

“…The dark or mini mealworm, Tenebrio obscurus (F.), which has also recently been shown to be able to biodegrade polystyrene (Peng et al, 2019), or the black fungus beetle, Alphitobius laevigatus (F.), which is already used as feed for house pets, e.g. reptiles, amphibians (Marien et al, 2022;Żuk-Gołaszewska et al, 2022), could be notable mealworm candidates for future research and subsequent commercial exploitation as nutrient sources. Only the future will tell, but it appears that the use of mealworms as food and feed is only just beginning!…”