The Putative Involvement of Bacterial Symbionts in Cantharidin Biogenesis: An Explorative Study in Meloidae Insects

Insect-microbes holobionts integrate host and microbial functions, with symbionts supporting nutrition, immunity, and defence, while producing metabolites, including beetle-derived compounds with therapeutic potential. Cantharidin is a toxic terpene produced by blister beetles (Coleoptera: Meloidae), endowed with defensive and pharmacological properties. Male insects produce and contain cantharidin in large quantities and transfer it to females upon mating. This study is aimed to gain information about the involvement of insect-associated bacteria in cantharidin biogenesis. To support the possibility that bacteria participate in cantharidin biogenesis, cantharidin antibacterial activity was assessed against six reference strains of representative species of Bacillota and Pseudomonadota from publicly available culture collections. All bacterial strains tolerated concentrations up to 600 µg/ml cantharidin in a standard antibacterial susceptibility test. To identify candidate bacterial lineages, 16S rRNA metataxonomic profiling of the V5-V6 region was performed in males and females from different Meloidae subfamilies and tribes. Analysis of the insect-associated microbiomes of the five cantharidin-producing species (Lydus trimaculatus, Meloe proscarabaeus, Mylabris variabilis, Hycleus polymorphus, Zonitis flava) revealed communities dominated by Pseudomonadota, with secondary contributions from Actinomycetota in Z. flava and M. proscarabaeus and Cyanobacteriota in the other host insects. Although overall community structure and composition did not differ significantly between sexes, a few taxa displayed consistent male-associated patterns, with Staphylococcus, Cutibacterium and one Enterobacteriaceae ASV resulting more abundant in males across all species. The intrinsic bacterial resistance to cantharidin, with both quantitative and qualitative differences in microbiome structure between male and female insects, makes the hypothesis of a putative involvement of bacteria in cantharidin biogenesis still viable.