Abstract: 【Aim】Existing in vitro artificial diets for honey bees are formulated based on the nutritional requirements of Apis mellifera worker bees; however, their suitability for drone in vitro development remains unclear. This study focused on the effects of artificial diets on the development of A. m. ligustica drones through the investigation of body weight and hemolymph proteome to provide a theoretical basis for studies on drone in vitro artificial diet rearing. 【Methods】 The A. m. ligustica drones were reared under in vitro rearing condition (treatment group) and natural condition (control group), then the body weight at the larval, pupal and newly emerged adult stages was measured, and the corresponding hemolymph proteins were collected and identified by liquid chromatography-tandem mass spectrometer (LC-MS/MS) and analyzed by bioinformatics approaches, such as GO and KEGG enrichment analyses, to reveal the differences in the functional classification and biological pathways of hemolymph proteins between the two rearing methods. 【Results】 The body weight of A. m. ligustica drones at various developmental stages in the treatment group was significantly lower than that in the control group. Qualitative analysis of the hemolymph proteome indicated that the treatment group had a total of 1 097 expressed proteins, while the control group had 1 074 expressed proteins. Furthermore, quantitative analysis of the hemolymph proteome revealed that the expression of 78 proteins was down-regulated and that of 10 proteins was up-regulated during the larval stage of A. m. ligustica in the treatment group as compared to that in the control group. The elevated expression of hemdymph proteins activated the Toll and Imd signaling pathways, harvested energy through the degradation of complex sugars and made lysosomal functions to be delayed in launching. Compared to the control group, treatment group had 122 down-regulated and 15 up-regulated proteins during the pupal stage, indicating that artificial diet rearing activates the Hippo signaling pathway to reduce the developmental rate to cope with the undernutrition. Compared to the control group, the treatment group had 37 up-regulated and 168 down-regulated proteins during the newly emerged adult stage, indicating that artificial diet rearing synthesizes a large number of proteins through triggering the ribosome pathway. 【Conclusion】 The findings demonstrate that in vitro reared drones are undernourished in the supporting of artificial diets. To establish a scientific and efficient drone-rearing approach, it is imperative to conduct further investigation on the nutritional requirements of drone development, and optimize the nutritional composition of artificial diets.

Key words: Apis mellifera ligustica, drone, artificial diet, hemolymph, proteomics; in vitro rearing