Number of CTD biosynthesis genes in six insects.

Cantharidin (CTD) is a well-established defensive toxin synthesized by blister beetles, displaying both therapeutic potential and toxicity. Among these beetles, Hycleus cichorii and Hycleus phaleratus are the two most commercially significant species due to their capacity to produce CTD in males. In this investigation, we conducted a gene expression profiling analysis of male and female individuals of these two species, utilizing the Illumina Hiseq4000 platform. We identified 7,983 expressed genes, including 2,823 differentially expressed genes (DEGs) shared by both male and female blister beetles. Nineteen genes related to CTD biosynthesis in the terpenoid backbone biosynthesis pathway were identified, including hydroxymethylglutaryl-CoA reductase (HMGR; EC:1.1.1.34), which demonstrated a significant correlation with CTD content. Furthermore, hydroxymethylglutaryl-CoA synthase (HMGS; EC:2.3.3.10) and isopentenyl-diphosphate Delta-isomerase (IDI; EC:5.3.3.2) were also found to be significantly up-regulated in males. Comparative analysis revealed that NADP+-dependent farnesol dehydrogenase (FOHSDR; EC:1.1.1.216) and farnesyl diphosphate synthase (FDPS; EC:2.5.1.1) had the highest copy number in these beetles, significantly higher than the copy number of the other four non-Meloidae insects. The analysis of the protein-protein interaction network of genes related to CTD biosynthesis revealed that the acetyl-CoA C-acetyltransferase (ACAT; EC:2.3.1.9) gene was the central gene, exhibiting greater expression in male blister beetles than in females. This study offers novel insights into the mechanisms of CTD biosynthesis in blister beetles and enhances our comprehensions of the association between particular genes and CTD content.