Cardenolides, toxicity, and the costs of sequestration in the coevolutionary interaction between monarchs and milkweeds

Abstract

For highly specialized insect herbivores, plant chemical defenses are often co-opted as cues for oviposition and sequestration. In such interactions, can plants evolve novel defenses, pushing herbivores to trade off benefits of specialization with costs of coping with toxins? We tested how variation in milkweed toxins (cardenolides) impacted monarch butterfly (Danaus plexippus) growth, sequestration, and oviposition when consuming tropical milkweed (Asclepias curassavica), one of two critical host plants worldwide. The most abundant leaf toxin, highly apolar and thiazolidine ring–containing voruscharin, accounted for 40% of leaf cardenolides, negatively predicted caterpillar growth, and was not sequestered. Using whole plants and purified voruscharin, we show that monarch caterpillars convert voruscharin to calotropin and calactin in vivo, imposing a burden on growth. As shown by in vitro experiments, this conversion is facilitated by temperature and alkaline pH. We next employed toxin-target site experiments with isolated cardenolides and the monarch’s neural Na+/K+-ATPase, revealing that voruscharin is highly inhibitory compared with several standards and sequestered cardenolides. The monarch’s typical >50-fold enhanced resistance to cardenolides compared with sensitive animals was absent for voruscharin, suggesting highly specific plant defense. Finally, oviposition was greatest on intermediate cardenolide plants, supporting the notion of a trade-off between benefits and costs of sequestration for this highly specialized herbivore. There is apparently ample opportunity for continued coevolution between monarchs and milkweeds, although the diffuse nature of the interaction, due to migration and interaction with multiple milkweeds, may limit the ability of monarchs to counteradapt.

Impacts of larval host plant species on dispersal traits and free-flight energetics of adult butterflies

“…Disentangling the relative contribution of nutrition, defenses, flight muscle would require additional studies focusing on just this species. Work by Agrawal et al 32 recently examined such mechanisms for larval performance on tropical milkweed.…”

“…Although relationships between milkweeds, monarchs, and toxins have played a central role in our understanding of coevolution, plant defense, sequestration, and animal behavior 17 , understanding the mechanistic link between host plant chemicals, sequestration and adult flight is challenging. There is some recent evidence that cardenolides can be a burden for monarchs and that there are costs to sequestration 32 . In vitro work by Petschenka et al 75 demonstrated that certain cardenolides can be strong inhibitors of neural function in monarchs.…”

“…Thus, our work utilizing different milkweed species with unique cardenolide composition allows for potential costs to become apparent. There’s strong evidence suggesting monarchs selectively sequester more polar compounds, by either modification, detox, or transport 76 and may even reduce exposure of larvae to more toxic compounds via ovipositional preference 32 . Thus, monarch mitigation strategies to sequester these poisonous compounds suggests costs not only to growth, survival, lifespan 77 , and here we find support for costs of sequestration that affects energetics as well.…”

“…While monarch Na + /K + -ATPase is considered relatively insensitive to cardenolide action 31 , this notion is primarily based on larval work focused on the cardenolide oubain, although several other cardenolides are present in milkweed for which Na + /K + -ATPase sensitivity is less well-understood. Furthermore, recent work has shown that variation levels of specific cardenolides can negatively impact monarch growth 32 . Overall, these findings indicate that milkweed host plants differ in their suitability to monarch larvae and may differentially affect the physiology adult monarchs.…”

Evidence for tissue‐specific defence‐offence interactions between milkweed and its community of specialized herbivores

“…The most beneficial substitutions occur in the amino acid residues 111, 119 and 122, and additionally in 786 and 797 in specific clades, conferring enhanced tolerance to cardenolides (Dobler et al, 2012; Karageorgi et al, 2019). In response, defence evolution in plants has resulted in the production of specific cardenolide compounds with particularly high potency against resistant sodium pumps (Agrawal et al, 2021; Petschenka et al, 2018). Nonetheless, since not all milkweed insects show the same sodium pump insensitivity (i.e., different number of genetic substitutions), whether the degree of cardenolide tolerance in insects targeting different plant tissues matches tissue‐specific defence chemistry remains untested.…”