Plants are important ecological factors and food resources, which can significantly affect the occurrence and distribution of insects. The metabolites in host plants can affect the feeding, spawning, and avoidance behaviors of herbivorous insects. Galeruca daurica (Joannis) is a phytophagous pest that has seriously occurred in the desert steppe of Inner Mongolia in recent years, only infesting the leaves of Allium plants. In order to clarify the effects of plant metabolites on the gene expression in G. daurica larvae at the transcriptome level, we fed the larvae of G. daurica with Allium tuberosum leaves soaked in 10% DMSO solutions containing d-galactose, β-d-glucopyranose, l-rhamnose, isoquercitrin, isoflavone, and rutin, respectively, used the larvae fed on A. tuberosum leaves soaked in a 10% DMSO solution as the control, and screened out the differentially expressed genes (DEGs) by performing high-throughput transcriptome sequencing. The results showed that a total of 291 DEGs were identified compared to the solvent control (DMSO), including 130, 34, 29, 21, 72, and 97 in the isoquercitrin, isoflavone, rutin, d-galactose, β-d-glucopyranose, and l-rhamnose treatment groups, respectively. GO and KEGG enrichment analysis showed that most DEGs were enriched in various metabolic pathways, implying that these six main primary and secondary metabolites in Allium plants may affect various metabolic processes in the larvae of G. daurica.

Metamorphosis in insects is regulated by hormonal signals and metabolic pathways. In this study, we characterized a 20E-mediated miR-2788/Treh1 regulatory axis involved in the larval-to-pupal transition in the leaf beetle Galeruca daurica. Dual-luciferase assays and expression profiling suggest that miR-2788 directly targets and suppresses trehalase 1 (Treh1), a key enzyme for trehalose hydrolysis. We found that exogenous 20E treatment significantly downregulated miR-2788, leading to the derepression of Treh1 and increased expression of chitin biosynthetic genes (CHI, CHS, and GPI). Conversely, miR-2788 overexpression or Treh1 silencing via RNAi resulted in massive trehalose accumulation, reduced chitin synthesis, and lethal developmental arrest during the metamorphic transition. Furthermore, pharmacological inhibition of Treh1 with Validamycin not only recapitulated these chitin-deficiency phenotypes but was also associated with an upregulation of miR-2788. These results describe a signaling cascade where 20E modulates the conversion of carbohydrate to chitin through the miR-2788/Treh1 axis. This study advances our understanding of hormonal–miRNA–metabolic crosstalk and identifies miR-2788 as a potential target for RNAi-based biopesticide development.

Lipophorin receptors play a crucial role in the reproductive development of insects. However, their regulatory function in insect reproductive diapause remains poorly understood. In this study, a lipophorin receptor gene (GdLpR) was cloned from Galeruca daurica using RT-PCR. The open reading frame (ORF) of GdLpR is 2589 bp in length, encoding 862 amino acids, and possesses typical structural characteristics of the lipophorin receptor family. RT-qPCR analysis indicated that the expression of GdLpR is up-regulated in G. daurica adults before diapause, exhibits an expression pattern of initial down-regulation, subsequent up-regulation, and then further down-regulation during diapause, and is up-regulated again after diapause termination. After RNA interference of GdLpR, the expression levels of diapause-related genes ecdysone receptor (EcR), nuclear hormone receptor (HR3), and vitellogenin (Vg) were downregulated, while the expression level of fatty acid synthase (FAS) and total lipid content were upregulated, leading to premature diapause in adult G. daurica. These results indicate that GdLpR is involved in the regulation of reproductive diapause in G. daurica.

Galeruca daurica (Joannis) (Coleoptera: Chrysomelidae) is an oligophagous pest in which both adults and larvae prefer to feed on Allium forage grasses of the Liliaceae family. In this study, we identified gustatory receptor (GR) genes based on the transcriptome data of G. daurica; analyzed the expression profiles of these GR genes across different larval instars and various tissues of male and female adults using quantitative real-time PCR (qRT-PCR); detected the electrophysiological responses of the mouthparts of male and female G. daurica adults to flavonoids and carbohydrates using single sensillum recording (SSR); and recorded the changes in food consumption of G. daurica adults after feeding on six host plant-derived metabolites. A total of 26 GR genes were identified from the transcriptome data of adult and larval of G. daurica. Phylogenetic analysis was performed to screen candidate functional gustatory receptor genes, including four sugar receptors (GdauGR7, GdauGR10, GdauGR14 and GdauGR28), seven bitter receptors (GdauGR11, GdauGR16~17, GdauGR22, GdauGR25~26 and GdauGR30), and two CO₂ receptors (GdauGR15 and GdauGR20). Larval expression profiling of GdauGRs in G. daurica revealed that the relative expression levels of 17 genes exhibited dynamic changes during larval growth and development. GdauGRs were expressed to varying degrees in the antennae, mouthparts, brain, gut, and forelegs of adult G. daurica, with sex-specific differences. Notably, the expression levels of GdauGR4, GdauGR9 and GdauGR16 in the gut were extremely significantly higher than those in other tissues. In the SSR test, the six tested flavonoids and one carbohydrate were able to induce robust electrophysiological responses in the gustatory sensilla on the antennae and mouthparts of adult G. daurica at specific concentrations. In addition, the supplementation of several host-derived metabolites altered the food consumption of adult G. daurica. These findings lay a solid foundation for elucidating the molecular mechanisms underlying gustatory recognition and host adaptation in G. daurica.