Caterpillars have evolved a multitude of biological defenses including defensive toxins, but the vast majority of these remain uncharacterised. We used proteotranscriptomics, recombinant expression and solid phase synthesis, and nociception assays to characterise defensive toxins produced by nettle caterpillars (Limacodidae), asp caterpillars (Megalopygidae), and processionary caterpillars (Thaumetopoeinae).
Pain-causing venom of the nettle caterpillar Doratifera vulnerans consists of more than 100 individual peptide toxins including many putative inhibitor cystine knots, with few proteins >10 kDa. The major pain-causing components are membrane-disrupting cationic peptides descended from the lepidopteran innate immunity peptide cecropin. In contrast, pain-inducing venom from the asp caterpillar Megalopyge opercularis was found to be radically different in composition, despite Limacodidae and Megalopygidae being closely related zygaenoid families. While also displaying membrane-disrupting activity, M. opercularis venom lacks inhibitor cystine knots or cecropins but instead features other classes of cystine-rich peptides as well as multiple aerolysin-like proteins. We show that these "megalysins" originate from a lateral gene transfer event from bacteria to an early lepidopteran ancestor, and hypothesise they are the primary agents responsible for membrane disruption and pain induction. Processionary caterpillars do not inject liquid venoms but instead possess millions of tiny urticating spines. Human contact with these spines causes strong allergic reactions rather than acute pain, while pregnant mares may suffer equine foetal loss syndrome. We show these defensive spines are loaded with multiple members of the secapin peptide family, which has also been recruited into some bee and wasp venoms. While hymenopteran venom secapins have been reported to induce inflammation and allodynia through modulation of the leukotriene and/or cyclooxygenase pathways, we are currently testing if processionary caterpillar spine secapins contribute to inflammation resulting from contact with their spines. These data highlight the diverse evolutionary trajectories taken by different lepidopteran taxa to arm their larvae with defensive toxins.