Make it a standard? The creation and variability assessment of a consensus standard protocol for Tenebrio molitor larvae feeding trials

Abstract Interest in the nutrition of the yellow mealworm ( Tenebrio molitor L.) larvae is on the rise, leading to an increase in publications on this topic. The absence of a standard protocol and resulting differences in experimental designs reduces comparability among studies and impedes research on mealworm nutrition. To address this, a consensus standardised protocol was developed specifically for the evaluation of mealworm larval growth and performance in feeding trials. The efficacy of this protocol was evaluated through an international ring test involving seven partners using two wheat brans as dry feed (a standard bran and a local bran) at 27 °C and 60% relative humidity. As experimental units, plastic crates filled with 2.1 kg of bran and 10,000 4-week-old larvae were used with six replicates. Agar gel was provided as wet feed ad libitum . The mean individual larval weight and the number of larvae per crate were determined weekly until either three or more replicates ran out of feed or pupation exceeded 10%. At harvest, the total larval fresh biomass and amount of frass was determined. Larval samples were taken for chemical analysis. To assess the protocol, the within (repeatability) and between (reproducibility) laboratory variability was calculated for each parameter. The repeatability was good (limit at 12% (standard) and 14% (local)). The reproducibility was poorer with a limit 2.7 times higher for the standard feed (36%) and 3.8 times higher for the local feed (55%). For both feeds, the total larval fresh harvest, amount of frass and the larval protein concentration were the most consistent both within and among laboratories. The highest variability was observed at the early life stages and for the larvae ash content. The detailed consensus standard protocol and repeatability/reproducibility estimates can be used as basis for future mealworm feeding trials, comparing results and future improvements.