Browsing by Person "Pokharel, Prayan"

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    Antioxidant availability trades off with warning signals and toxin sequestration in the large milkweed bug (Oncopeltus fasciatus)
    (2023) Heyworth, H. Cecilia; Pokharel, Prayan; Blount, Jonathan D.; Mitchell, Christopher; Petschenka, Georg; Rowland, Hannah M.
    In some aposematic species the conspicuousness of an individual's warning signal and the concentration of its chemical defense are positively correlated. Several mechanisms have been proposed to explain this phenomenon, including resource allocation trade‐offs where the same limiting resource is needed to produce both the warning signal and chemical defense. Here, the large milkweed bug (Oncopeltus fasciatus: Heteroptera, Lygaeinae) was used to test whether allocation of antioxidants, that can impart color, trade against their availability to prevent self‐damage caused by toxin sequestration. We investigated if (i) the sequestration of cardenolides is associated with costs in the form of changes in oxidative state; and (ii) oxidative state can affect the capacity of individuals to produce warning signals. We reared milkweed bugs on artificial diets with increasing quantities of cardenolides and examined how this affected signal quality (brightness and chroma) across different instars. We then related the expression of warning colors to the quantity of sequestered cardenolides and indicators of oxidative state—oxidative lipid damage (malondialdehyde), and two antioxidants: total superoxide dismutase and total glutathione. Bugs that sequestered more cardenolides had significantly lower levels of the antioxidant glutathione, and bugs with less total glutathione had less luminant orange warning signals and reduced chroma of their black patches compared to bugs with more glutathione. Bugs that sequestered more cardenolides also had reduced red–green chroma of their black patches that was unrelated to oxidative state. Our results give tentative support for a physiological cost of sequestration in milkweed bugs and a mechanistic link between antioxidant availability, sequestration, and warning signals.
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    Metabolization and sequestration of plant specialized metabolites in insect herbivores: Current and emerging approaches
    (2022) Jeckel, Adriana Moriguchi; Beran, Franziska; Züst, Tobias; Younkin, Gordon; Petschenka, Georg; Pokharel, Prayan; Dreisbach, Domenic; Ganal-Vonarburg, Stephanie Christine; Robert, Christelle Aurélie Maud
    Herbivorous insects encounter diverse plant specialized metabolites (PSMs) in their diet, that have deterrent, anti-nutritional, or toxic properties. Understanding how they cope with PSMs is crucial to understand their biology, population dynamics, and evolution. This review summarizes current and emerging cutting-edge methods that can be used to characterize the metabolic fate of PSMs, from ingestion to excretion or sequestration. It further emphasizes a workflow that enables not only to study PSM metabolism at different scales, but also to tackle and validate the genetic and biochemical mechanisms involved in PSM resistance by herbivores. This review thus aims at facilitating research on PSM-mediated plant-herbivore interactions.
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    Physiological and ecological implications of sequestered cardenolides in the milkweed bugs (Heteroptera: Lygaeinae)
    (2022) Pokharel, Prayan; Petschenka, Georg
    Milkweed bugs (Heteroptera: Lygaeinae) have a predilection for toxic plants, and possess a distinctive black and red coloration. Although many milkweed bugs are generalist seed predators, they commonly feed on plants in the family Apocynaceae (milkweed) which often contain toxic cardenolides. Cardenolides inhibit the ubiquitous Na+/K+-ATPase, an essential animal enzyme mediating essential physiological functions. Milkweed bugs possess pronounced insensitivity towards cardenolides due to a few amino acid substitutions in the Na+/K+-ATPase (i.e., target site insensitivity) and sequester cardenolides for protection against their predators. The overarching question remains whether chemical defenses, in aposematic individuals sequestering toxins, incur physiological costs, such as effects on growth or other fitness parameters like longevity and fecundity, production of color pigments, and handling oxidative stresses, and/or ecological costs, such as universality of toxin defense. Using an artificial diet, I raised larvae of four milkweed bug species (Oncopeltus fasciatus, Caenocoris nerii, Spilostethus pandurus and Arocatus longiceps) and a closely related pyrrhocorid bug species (Pyrrhocoris apterus) on three increasing dietary doses of cardenolides, and assessed the increase in growth by recording the mass until adult. Additionally, I investigated the life-history parameters only in O. fasciatus. To understand if milkweed bugs exhibit honest signaling, using same artificial diet treatment, the color intensity of O. fasciatus was measured by taking photographs in each larval stage until adulthood. To understand if toxin sequestration in milkweed bugs imposes oxidative stress, biomarkers of oxidative stress was measured through biochemical assays for lipid peroxidation (malondialdehyde, or MDA), superoxide dismutase (SOD), and total glutathione content (GSH). To understand why protection against certain predators is not observed in all bug species although they feed on seeds containing cardenolides, I tested if the outcome of predator- prey interaction was mediated by the structural variation within the same class of compound or by the insect species. For this purpose, I raised two milkweed bug species (Lygaeus equestris and Horvathiolus superbus) on the seeds of two phylogenetically unrelated host plants (Ranunculaceae: Adonis vernalis and Plantaginaceae: Digitalis purpurea) from which the bugs sequestered cardenolides, and carried out predation assays with lacewing larvae. The amount of toxins sequestered by the milkweed bugs was estimated using high performance liquid chromatography. My research revealed that dietary plant toxins increased growth in the sequestering specialists (O. fasciatus and C. nerii) but not in the sequestering generalist, S. pandurus, despite all possessing toxin-resistant Na+/K+-ATPases. Under exposure to the dietary toxins, the growth of A. longiceps nymphs (resistant and non-sequestering) was unaffected, while that of P. apterus (non-resistant and non-sequestering) was impaired. In addition, O. fasciatus nymphs developed to adults faster and lived longer as adults under toxin exposure when compared to individuals raised on the control diet, but produced significantly fewer offspring unless being transferred to a toxin-free diet after reaching adulthood. Furthermore, I showed that O. fasciatus raised on the high and medium levels of dietary cardenolides had significantly lower levels of GSH. Bugs with more GSH levels had brighter warning signals but these signals were not related to sequestration. Besides physiological aspects, the chance of milkweed bugs surviving a predator attack strongly depended on the structural differences of sequestered toxins. Overall, I found that cardenolide consumption exerts a positive effect on overall fitness in milkweed bugs, a conclusion in disagreement with current theory predicting costs of sequestration. Oxidative state may be a fundamental aspect where costs lie in aposematic individuals sequestering toxins, and the effect of plant-toxin sequestration on predators is affected by the structural variation of defensive compounds and therefore depends on the ecological context, i.e., host-plant use. My dissertation provides insight into the implications of physiology and ecology on sequestering aposematic insects, giving us a better understanding of plant-insect-predator interactions.