Production, composition, and mode of action of the painful defensive venom produced by a limacodid caterpillar, Doratifera vulnerans

Abstract

Venoms have evolved independently several times in Lepidoptera. Limacodidae is a family with worldwide distribution, many of which are venomous in the larval stage, but the composition and mode of action of their venom is unknown. Here, we use imaging technologies, transcriptomics, proteomics, and functional assays to provide a holistic picture of the venom system of a limacodid caterpillar, Doratifera vulnerans. Contrary to dogma that defensive venoms are simple in composition, D. vulnerans produces a complex venom containing 151 proteinaceous toxins spanning 59 families, most of which are peptides <10 kDa. Three of the most abundant families of venom peptides (vulnericins) are 1) analogs of the adipokinetic hormone/corazonin-related neuropeptide, some of which are picomolar agonists of the endogenous insect receptor; 2) linear cationic peptides derived from cecropin, an insect innate immune peptide that kills bacteria and parasites by disrupting cell membranes; and 3) disulfide-rich knottins similar to those that dominate spider venoms. Using venom fractionation and a suite of synthetic venom peptides, we demonstrate that the cecropin-like peptides are responsible for the dominant pain effect observed in mammalian in vitro and in vivo nociception assays and therefore are likely to cause pain after natural envenomations by D. vulnerans. Our data reveal convergent molecular evolution between limacodids, hymenopterans, and arachnids and demonstrate that lepidopteran venoms are an untapped source of novel bioactive peptides.

Venomics of the Central European Myrmicine Ants Myrmica rubra and Myrmica ruginodis

“…Although the acid-based defense system is better known to the general public, the number of species that deploy this mechanism is limited, whereas ~75% of all ant species inject venom [ 22 , 54 ]. Across the animal kingdom, defensive venoms are characterized by their ability to induce pain [ 55 , 56 , 57 ].…”

Multipurpose peptides: The venoms of Amazonian stinging ants contain anthelmintic ponericins with diverse predatory and defensive activities

“…The synthetic ponericins Na1b, Nc1a and Nc3a were found to inhibit H. contortus development with an IC50 of 2.8-5.6 µM, which is only 4-8-fold lower potency than the commercial anthelmintic levamisole but considerably less active than macrocyclic lactone anthelmintics [43,44]. These ponericins have similar efficacy against H. contortus to previously reported anthelmintic venom peptides, including the heterodimeric peptide ∆myrtoxin-Mp1a (Mp1a) from the Australian jack jumper ant Myrmecia pilosula (IC50 6.8 µM) [13] and the disulfide-rich inhibitor cystine knot (ICK) peptide U-LCTX-Dv33 (Dv33) from venom of the Australian caterpillar Doratifera vulnerans (IC50 2.6 µM) [45]. The disulfiderich cyclotide kalata B1, derived from the flowering plant Oldenlandia affinis, has similar potency against H. contortus (IC50 2.3 µM) [46].…”

“…Nc2a and Nc3b had moderate anthelmintic activity (IC50 23-38 µM), similar to the double-ICK peptide Hi1a from venom of the Australian funnel-web spider Hadroncyhe infensa (IC50 22.9 µM) [27], and cecropin-like venom peptides from D. vulnerans (IC50 24.5-30.5 µM) [45]. These cecropin-like peptides are similar to the ant-derived ponericins in that they are also short, lack disulfide-bridges, and disrupt membranes, conferring broad-spectrum activity against mammalian cells, helminths and microbes, and inducing nocifensive behaviour in mice [45]. Kalata B1 and Mp1a also exert anthelmintic activity via membrane disruption, and Mp1a induces spontaneous pain behaviour in mice [36,46].…”

“…The disulfiderich cyclotide kalata B1, derived from the flowering plant Oldenlandia affinis, has similar potency against H. contortus (IC50 2.3 µM) [46]. Nc2a and Nc3b had moderate anthelmintic activity (IC50 23-38 µM), similar to the double-ICK peptide Hi1a from venom of the Australian funnel-web spider Hadroncyhe infensa (IC50 22.9 µM) [27], and cecropin-like venom peptides from D. vulnerans (IC50 24.5-30.5 µM) [45]. These cecropin-like peptides are similar to the ant-derived ponericins in that they are also short, lack disulfide-bridges, and disrupt membranes, conferring broad-spectrum activity against mammalian cells, helminths and microbes, and inducing nocifensive behaviour in mice [45].…”

Evolution, Expression Patterns, and Distribution of Novel Ribbon Worm Predatory and Defensive Toxins

“…However, the processes underlying toxin and venom evolution remain poorly understood, in particular for many invertebrate groups. Nevertheless, the advancement of sequencing and analytical techniques has expanded research to traditionally neglected taxa, shedding light into venom biology of species that were previously challenging to work with, like mammals ( Casewell et al 2019 ; Nekaris et al 2020 ) or small invertebrates ( Gorson et al 2015 ; Robinson et al 2018 ; Jenner et al 2019 ; von Reumont et al 2020 ; Walker et al 2021 ). This increase in the taxonomic coverage of the venomous taxa being investigated is revealing a high genetic and functional diversity of venom compounds across taxa ( Madio et al 2018 ; Robinson et al 2018 ; Giorgianni et al 2020 ), as well as novel mechanisms of venom evolution ( Sachkova et al 2020 ; Undheim and Jenner 2021 ), challenging traditional views in venom research.…”