Organ-Specific Responses to Plant Secondary Metabolites Reveal Mechanisms of Adaptation.

Many insects feed on plants that produce toxic secondary metabolites. The Rattlebox plant (Crotalaria) produces Pyrrolizidine Alkaloids (PAs) that protect it from most herbivores, but some insects, including the Bella moth (Utetheisa ornatrix), can metabolize, sequester, and repurpose its chemical defenses. However, organ-specific responses to Crotalaria secondary metabolites remain poorly understood, and current knowledge is largely derived from a few model species and isolated organ studies. This study bridges this gap by evaluating organ-specific responses of the Bella moth to the Showy Rattlebox, C. spectabilis, through an integrative morphological, molecular, and evolutionary framework. Our results provide quantitative and qualitative characterizations of the fat body, gut, integument, Malpighian tubules, and testes of male Bella moth larvae and reveal two distinct midgut morphologies, suggesting regional specialization within the gut. Additionally, we used real-time quantitative PCR to demonstrate variation of detoxification-related gene expression among organs, with FMO2 highly expressed in Malpighian tubules and SNO predominantly expressed in testes. We found that the genes SOD2 and UGT exhibited significant diet-dependent expression differences between larvae fed with and without secondary metabolites. Our comparative analyses across Erebidae species known to feed on PA-containing host plants identified specific lineages and amino acid residues under positive selection in genes encoding N-oxidation enzymes, FMO2 and SNO. Collectively, these integrated morphological and molecular insights offer a framework of how the Bella moth larvae systematically manage C. spectabilis consumption. This work highlights the critical role of larval metabolic defense and opens broader research avenues into the mechanisms driving insect specialization on toxic host plants.