Insects represent a sustainable, protein-rich food source widely consumed in Asia, Africa, and South America. Eating Clanis bilineata tsingtauica Mell is common in the eastern part of China. A comparative characterization of nutrients in the meat and epidermis of C. bilineata tsingtauica was performed in this study. The results showed this insect to be high in nutrients, particularly in the epidermis where protein total was 71.82%. Sixteen different amino acids were quantified in C. bilineata tsingtauica, and the ratio of essential to nonessential amino acids in the epidermis and meat was 68.14% and 59.27%, respectively. The amino acid composition of C. bilineata tsingtauica is balanced, representing a high-quality protein source. Eight minerals were quantified in C. bilineata tsingtauica, including four macro and four trace elements. Fe in the epidermis and Zn in the meat were abundant at 163.82 and 299.31 μg/g DW, respectively. The presence of phytic acid impacted the absorption of mineral elements in food. We also detected phytic acid in C. bilineata tsingtauica. The molar ratio of phytic acid to zinc (PA/Zn) in C. bilineata tsingtauica was very low (3.28) compared to Glycine max and Cryptotympana atrata, which indicated that mineral utilization was high. In conclusion, this study confirms that C. bilineata tsingtauica is a highly nutritious food source for human consumption, and the results provide a basis for further consumption and industrialization of this edible insect.

Clanis bilineata tsingtauica Mell, 1922 (Lepidoptera, Sphingidae), also known as “Doudan” in China, is an important pest in legume crops. As an edible insect, it is most commonly consumed in Jiangsu, Shandong, and Henan Provinces. Mass rearing requires access to large numbers of eggs. This stage, however, is of short duration and supplies are frequently not sufficient for insect production. Therefore, we identified the cold storage conditions for C. bilineata tsingtauica that can effectively prolong the storage time of the eggs, to make supplies more readily available. We found that when stored at 4 °C, only 7.5% of the eggs hatched after 7 days, while at 10 °C the hatch rate was 78.3%. At 15 °C, the egg hatch rate remained at this same level (77.8% even after 14–20 days). Considering various combinations, we found that optimal egg hatch occurred if eggs were stored at 15 °C for 11 days, and then held at 15–20 °C under dark conditions. Stored as described above, the egg hatch rate was not significantly different from the control group (at 28 °C). These conditions allow for easier mass rearing of C. bilineata tsingtauica by providing a stable supply of eggs.

Quantitative real-time PCR (qRT-PCR) is a high-reliability, -sensitivity, and -operability technique to quantify gene expression. It is necessary to select stable reference genes for normalization. Water plays important roles in the metabolism, physiology, distribution, and so on, in insects. In this study, the suitability of various reference genes for qRT-PCR analysis was evaluated in different developmental stages of Chilo suppressalis exposed to desiccation or rehydration stress. The ∆Ct method, geNorm, NormFinder, and BestKeeper were used to evaluate the suitability of nine reference genes for normalizing gene expression in the third instar larvae, the fifth instar larvae, male pupae, female pupae, male adults, and female adults under different humidities. The results indicated that 18S rRNA was the most stable reference gene for monitoring gene expression in the third instar larvae, while ACTIN, TUB, UBI, UBI, and EF1 were the optimal genes for the fifth instar larvae, male pupae, female pupae, male adults, and female adults, respectively. The optimal number of reference genes recommended by geNorm analysis indicated that two candidate reference genes were sufficient for data normalization under all experimental conditions tested. To validate these recommendations, the expression profile of the gene encoding heat shock protein 60 (Hsp60) was investigated. Hsp60 transcript levels showed significant differences when normalized to the most stable single reference gene, or combined reference genes, compared with the least stable reference gene. The reference genes identified in the present study will enhance the reliability of gene expression data for C. suppressalis under humidity stress.