Silkworms (Bombyx mori) reared on artificial diets during all instars have the advantages of simplicity and efficiency, year-round production, and reduced risk of poisoning. However, low silk yield remains a challenge, limiting its industrial application. To address this issue, the spinning behavior, nutrient absorption, and transcriptomics of silkworms were investigated. Compared with silkworms reared on mulberry leaves during all instars, those fed with artificial diets showed significantly lower cocoon weight, cocoon shell weight, cocoon shell rate, and silk gland tissue somatic index at the end of the fifth instar (P < 0.01). The spinning duration and crawling distance of silkworms reared on artificial diets were also significantly lower than those reared on mulberry leaves (P < 0.01). Regarding nutrient absorption, the dietary efficiency indexes of silkworms fed with artificial diets were significantly lower than those fed with mulberry leaves, except for the efficiency conversion of digesta to cocoon (P < 0.01). Further RNA-Seq analysis revealed 386 differentially transcribed genes between the 2 groups, with 242 upregulated and 144 downregulated genes. GO enrichment analysis showed that differential transcriptional genes were mainly enriched in organic acid metabolism, oxidation–reduction, and drug catabolism. KEGG enrichment analysis showed that differential transcriptional genes were mainly enriched in genetic information processing and metabolism pathways. Our findings provide new insights into the silk secretion and can serve as a reference for future research and application of silkworms fed with artificial diets.

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