GC–MS‐based metabonomic analysis of silkworm haemolymph reveals four‐stage metabolic responses to nucleopolyhedrovirus infection

Abstract Silkworm, Bombyx mori , an economically significant insect, plays a crucial role in silk production. However, silkworm breeding is highly susceptible to various pathogens, particularly the Bombyx mori nucleopolyhedrovirus (BmNPV), which poses a serious threat. Recent metabonomic studies have provided insights into the metabolic changes associated with BmNPV infection. BmNPV infection has obvious temporal characteristics. However, few studies have investigated the silkworms infected in different periods. This study employed gas chromatography–mass spectrometry (GC–MS) to perform a comprehensive analysis of haemolymph metabolites in silkworms at 48, 72, 96 and 120 h post‐infection (h.p.i.). Through the integration of time‐course analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment, the study revealed distinct four‐stage metabolic characteristics in the silkworm's response to BmNPV infection. At Stage 1 (48 h.p.i.), silkworms activate antioxidant defence mechanisms, with significant enrichment in metabolic pathways involving key antioxidants such as glutathione, to mitigate oxidative stress induced by viral invasion. By Stage 2 (72 h.p.i.), pathways related to amino acid metabolism and protein synthesis become active, indicating an increase in protein synthesis. In Stage 3 (96 h.p.i.), energy metabolism and substance transport pathways are significantly upregulated to support the rapid viral replication and the enhanced locomotor behaviour of silkworm. Finally, at Stage 4 (120 h.p.i.), there is a further enhancement of pathways related to energy metabolism, nucleic acid synthesis, and substance transport, which align with peak viral assembly and release. These findings contribute to an in‐depth understanding of the biochemical basis of silkworm resistance to NPV.