Phylogenomics of Tetraopes longhorn beetles unravels their evolutionary history and biogeographic origins.

Tetraopes longhorn beetles are known for their resistance to milkweed plant toxins and their coevolutionary dynamics with milkweed plants (Asclepias). This association is considered a textbook example of coevolution, in which each species of Tetraopes is specialized to feed on one or a few species of Asclepias. A major challenge to investigating coevolutionary hypotheses and conducting molecular ecology studies lies in the limited understanding of the evolutionary history and biogeographical patterns of Tetraopes. By integrating genomic, morphological, paleontological, and geographical data, we present a robust phylogeny of Tetraopes and their relatives, using three inference methods with varying subsets of data, encompassing 2-12 thousand UCE loci. We elucidate the diversification patterns of Tetraopes species across major biogeographical regions and their colonization of the American continent. Our findings suggest that the genus originated in Central America approximately 21 million years ago during the Miocene and diversified from the Mid-Miocene to the Pleistocene. These events coincided with intense geological activity in Central America. Additionally, independent colonization events in North America occurred from the Late Miocene to the early Pleistocene, potentially contributing to the early diversification of the group. Our data suggest that a common ancestor of Tetraopini migrated into North America, likely facilitated by North Atlantic land bridges, while closely related tribes diverged in Asia and Europe during the Paleocene. Establishing a robust and densely sampled phylogeny of Tetraopes beetles provides a foundation for investigating micro- and macroevolutionary phenomena, including clinal variation, coevolution, and detoxification mechanisms in this ecologically important group.

Pyrethroid insecticide and milkweed cardenolide interactions on detoxification enzyme activity and expression in monarch caterpillars.

Declines of the monarch butterfly population have prompted large-scale plantings of milkweed to restore the population. In North America, there are >73 species of milkweed to choose from for these nationwide plantings. However, it is unclear how different milkweed species affect monarch caterpillar physiology, particularly detoxification enzyme activity and gene expression, given the highly variable cardenolide composition across milkweed species. Here, we investigate the effects of a high cardenolide, tropical milkweed species and a low cardenolide, swamp milkweed species on pyrethroid sensitivity as well as detoxification enzyme activity and expression in monarch caterpillars. Caterpillars fed on each species through the fifth-instar stage and were topically treated with bifenthrin after reaching this final-instar stage. Esterase, glutathione S-transferase, and cytochrome P450 monooxygenase activities were quantified as well as the expression of selected esterase, glutathione S-transferase, ABC transporter, and cytochrome P450 monooxygenase transcripts. There were no significant differences in survival 24 h after treatment with bifenthrin. However, bifenthrin significantly increased glutathione S-transferase activity in caterpillars feeding on tropical milkweed and significantly decreased esterase activity in caterpillars feeding on tropical and swamp milkweed. Significant differential expression of ABC transporter, glutathione S-transferase, and esterase genes was observed for caterpillars feeding on tropical and swamp milkweed and not receiving bifenthrin treatment. Furthermore, significant differential expression of glutathione S-transferase and esterase genes was observed for bifenthrin-treated and -untreated caterpillars feeding on tropical milkweed relative to swamp milkweed. These results suggest that feeding on different milkweed species can affect detoxification and development mechanisms with which monarch caterpillars rely on to cope with their environment.

Cardenolide, Potassium, and Pyrethroid Insecticide Combinations Reduce Growth and Survival of Monarch Butterfly Caterpillars (Lepidoptera: Nymphalidae).

The monarch butterfly, Danaus plexippus L., has evolved to be insensitive to milkweed cardenolides via genetic modifications of Na+/K+-ATPase. There is concern for insecticide exposures near agriculture, with little information on monarch caterpillar toxicology. It is unclear how cardenolide insensitivity may affect the sensitivity of monarch caterpillars to pyrethroid insecticides. Additionally, potassium fertilizers may affect monarch caterpillar physiology and cardenolide sequestration. Here, we investigated the growth, survival, and development of caterpillars exposed to the cardenolide ouabain, bifenthrin, and potassium chloride (KCl) alone and in combination. Caterpillars were either exposed to 1) ouabain from third- to fifth-instar stage, 2) KCl at fifth-instar stage, 3) KCl and bifenthrin at fifth-instar stage, or 4) combinations of ouabain at third-instar stage + KCl + bifenthrin at fifth-instar stage. Caterpillar weight, diet consumption, frass, and survival were recorded for the duration of the experiments. It was observed that 1-3 mg ouabain/g diet increased body weight and diet consumption, whereas 50 mg KCl/g diet decreased body weight and diet consumption. Caterpillars feeding on KCl and treated with 0.2 µg/µl bifenthrin consumed significantly less diet compared to individuals provided untreated diet. However, there was no effect on survival or body weight. Combinations of KCl + ouabain did not significantly affect caterpillar survival or body weight following treatment with 0.1 µg/µl bifenthrin. At the concentrations tested, there were no effects observed for bifenthrin sensitivity with increasing cardenolide or KCl concentrations. Further studies are warranted to understand how milkweed-specific cardenolides, at increasing concentrations, and agrochemical inputs can affect monarch caterpillar physiology near agricultural landscapes.

Pyrethroid Exposure Reduces Growth and Development of Monarch Butterfly (Lepidoptera: Nymphalidae) Caterpillars.

Insecticide exposure has been identified as a contributing stressor to the decline in the North American monarch butterfly Danaus plexippus L. (Lepidoptera: Nymphalidae) population. Monarch toxicity data are currently limited and available data focuses on lethal endpoints. This study examined the 72-h toxicity of two pyrethroid insecticides, bifenthrin and ß-cyfluthrin, and their effects on growth and diet consumption. The toxicity of bifenthrin to caterpillars was lower than ß-cyfluthrin after 72 h. Survival was the most sensitive endpoint for bifenthrin, but diet consumption and caterpillar growth were significantly reduced at sublethal levels of ß-cyfluthrin. Using AgDRIFT spray drift assessment, the aerial application of bifenthrin or ß-cyfluthrin is predicted to pose the greatest risk to fifth-instar caterpillars, with lethal insecticide deposition up to 28 m for bifenthrin and up to 23 m for ß-cyfluthrin from treated edges of fields. Low boom ground applications are predicted to reduce distances of lethal insecticide exposure to 2 m from the treated field edge for bifenthrin and ß-cyfluthrin. Growth and survival of fifth-instar monarch caterpillars developing within the margins of a treated field may be significantly impacted following foliar applications of bifenthrin or ß-cyfluthrin. These findings provide evidence that pyrethroid insecticides commonly used for soybean pest control are a potential risk to monarch caterpillars in agricultural landscapes.

Abundance and Diversity of Dung Beetles (Coleoptera: Scarabaeoidea) as Affected by Grazing Management in the Nebraska Sandhills Ecosystem.

Dung beetles (Coleoptera: Scarabaeoidea) serve a significant role in regulating ecosystem services on rangelands. However, the influence of grazing management on dung beetle communities remains largely unknown. The purpose of this study was to investigate dung beetle abundance and diversity throughout the grazing season in the Nebraska Sandhills Ecoregion. Grazing treatments included: continuous grazing (CONT), low-stocking rotational grazing (LSR), high-stocking rotational grazing (HSR), and no grazing (NG). The abundance and diversity of dung beetles were measured in the 2014 and 2015 grazing seasons using dung-baited pitfall traps. Dung beetle abundance for each grazing treatment was characterized through four indices: peak abundance, species richness, Simpson's diversity index, and Simpson's evenness. A total of 4,192 dung beetles were collected through both years of trapping in this study. Peak abundance and species richness were greater in grazed treatments when compared to NG in both years. Peak abundance in the HSR was 200% (2014) and 120% (2015) higher than in the LSR. Species richness in the HSR was 70% (2014) and 61% (2015) higher than in the LSR, and 89% (2014) and 133% (2015) higher than in CONT. Simpson's diversity index was lower in the NG and CONT treatments when compared to the LSR or HSR treatments for both years. We conclude that rotational grazing, regardless of stocking density, promoted dung beetle abundance and diversity within the Nebraska Sandhills Ecoregion.