Monarch Larvae Research Articles

Low-Level Dietary Clothianidin Exposure Preferentially Causes Prepupal Mortality of Monarch Butterflies (Danaus plexippus).

Data from prior research indicate the prepupal stage of the monarch butterfly life cycle is more sensitive to clothianidin exposure than the larval stage. A set of experiments was conducted to determine if the dietary clothianidin exposures that cause prepupal mortality are environmentally relevant. Monarch larvae were raised from egg to pupae on clothianidin-contaminated swamp milkweed plants (Asclepias incarnata). Larval growth as well as larval and prepupal survival were monitored throughout the experiments, in which the exposures ranged from 1.4 to 2793.1 ng/g leaf. Exposures of 5.4 to 46.9 ng/g leaf resulted primarily in prepupal mortality, whereas higher exposures of 1042.4 to 2793.1 ng/g leaf resulted exclusively in larval mortality, indicating the prepupal stage is more sensitive to clothianidin exposure than the larval stage. A median lethal concentration and a 10% lethal concentration of 37 and 6 ng/g leaf, respectively, were estimated for prepupal mortality. Both effect concentrations are within the range of clothianidin concentrations reported in leaves collected from wild milkweed plants, indicating prepupal mortality is an environmentally relevant effect. Environ Toxicol Chem 2024;43:2039-2044. © 2024 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.

Exploring the interplay between host plant structural defences and origin on behavioural and life‐history traits of two Neotropical danaines

AbstractFactors that mediate insect herbivore sabotaging behaviour of host plant latex defences remain under debate. The most prominent hypothesis suggests that the anatomy of the host plant's laticifers determines trenching or vein‐cutting behaviour, but there are inconsistencies in the literature. In addition to latex, other plant defences have been shown to affect plant natural enemies, but experimental studies comparing the effects of multiple defences on herbivore performance and behaviour are scarce. In this study, we investigated the anatomy of the laticifers of Calotropis procera (Aiton) W.T. Aiton (Apocynaceae), an exotic milkweed of southeastern Brazil, as well as the sabotaging behaviour of larvae of southern monarch, Danaus erippus (Cramer), and queen butterfly, Danaus gilippus (Cramer) (Lepidoptera: Nymphalidae) on this host plant. We also reviewed the sabotaging behaviour of larvae reared on the native milkweed Asclepias curassavica L. (Apocynaceae) from previous studies, as well as the description of the anatomy of its laticifers. As we found no correspondence between sabotaging behaviour of southern monarchs and queens and the anatomy of the host plants' laticifers, we tested whether structural defences of C. procera leaves (latex, waxes and trichomes) affected sabotaging behaviour and larval performance of southern monarch larvae. The structural barriers of this well‐defended host affected larvae in different ways: Southern monarchs developed more slowly on control leaves than on leaves from which wax had been removed, and only latex removal reduced the frequency of sabotaging behaviour. Regardless of the defence removal treatment, frequencies of trenching behaviour decreased and vein‐cutting behaviour increased as larvae developed. Overall, our results show that several factors in addition to the anatomy of laticifers affect sabotaging behaviour of southern monarchs and queens, including larval ontogeny, danaine species and latex outflow. This evidence suggests a behavioural plasticity in the sabotaging behaviour of larvae of both southern monarchs and queens.

Host plants and landscape predict use of roadside habitat by breeding monarchs

Abstract Vegetation along roadside edges, while posing some risks, can also provide breeding habitat for monarch butterflies. We investigated how landscape and site‐specific factors affect the quality of roadsides as monarch breeding habitat. We surveyed more than 250 sites along roadsides throughout Minnesota for milkweeds, floral resources and monarch eggs and larvae. We found that milkweed distribution was affected by adjacent land use and geography, with milkweeds more likely to be present in roadsides adjacent to crop and grassland and the south and west of the state. The presence and density of monarch eggs and larvae were primarily driven by milkweed density, with the likelihood of monarch presence at each site increasing with milkweed density. The density of monarchs per plant increased in sites with lower densities of milkweed, suggesting that monarch eggs are concentrated more densely on fewer plants where milkweed is scarce and spread more thinly across plants where milkweed is common. Monarch density per milkweed increased as sites were farther from core grasslands, to the south, and on narrower roadsides, also suggesting use may be higher in areas with a scarcity of habitat. The density of monarch eggs and larvae was higher in mowed than unmowed milkweed. Our study suggests that roadside habitats effectively provide breeding locations for monarchs, regardless of adjacent land use type and even when distant to high‐quality natural areas. Our findings assist conservation practitioners in planning and managing roadside habitats for monarchs and other pollinators.

Do Invasive Jorō Spiders (Trichonephila clavata) from Asia Avoid Eating Unpalatable Monarch Butterflies (Danaus plexippus) in North America?

An invasive spider from East Asia has established in the U.S. southeast (the "jorō spider," Trichonephila clavata) and is rapidly expanding its range. Studies assessing the impact of this species are needed, including how expansive its diet is. An open question is whether monarch butterflies, Danaus plexippus, are a potential prey item for this spider, given that jorō spiders do not coexist with monarchs in their native range. Since monarch larvae feed on milkweed, they sequester cardiac glycosides into their adult tissues, rendering them unpalatable to many predators. At sites within northeast Georgia, we staged a series of trials (n = 61) where we tossed monarchs into jorō spider webs and, for comparison, performed similar trials with another aposematic species, gulf fritillary (Agraulis vanilla), and a palatable species, tiger swallowtail (Papilio glaucus). We recorded the outcome of the trials, which included whether the spider attacked or did not attack the prey. We also conducted a visual survey during the same fall season to look for evidence of jorō spiders consuming monarchs naturally. Our findings revealed that jorō spiders avoided eating monarchs; spiders only attacked monarchs 20% of the time, which was significantly less than the attack rates of similarly sized or larger butterflies: 86% for gulf fritillaries and 58% for tiger swallowtails. Some jorō spiders even removed monarchs from their webs. From our visual surveys of the surrounding area, we found no evidence of natural monarch consumption and, in general, butterflies made up only a fraction of the jorō spider diet. We conclude that jorō spiders appear to recognize monarch butterflies as being unpalatable, even without having a prior history with the species. This invokes questions about how these spiders can immediately recognize their unpalatability without touching the butterflies.

Revisiting geographic variation in melanism of monarch butterfly larvae in North America using iNaturalist photos

Monarch butterflies in North America have an exceptionally large breeding distribution, occupying regions west of the Rocky Mountains and throughout the eastern seaboard. An experimental study conducted 17 years ago and published in this journal appeared to show that western monarch larvae tend to have smaller black stripes than those from eastern parents, which at the time was thought to be an adaptation to higher solar exposure in California. Here, we revisit this question by measuring melanism of eastern and western larvae from online photographs submitted to iNaturalist by members of the public. We downloaded over 500 photos of larvae from 10 different states (4 in the west, 6 in the east) and used image analysis to quantify the size of each larva's black stripes (% of surface covered by black). We compared average melanism level between east and west, while also accounting for temperature (at the photo location), seasonal effects, and latitude. Results showed larvae tended to be darker with lower development temperatures, and later in the season, though there was no significant difference between eastern and western larvae in their degree of melanism. It is not yet clear why findings here were different from the prior experimental study. Also, the wild larvae appeared to be notably lighter in color than the captive-reared larvae of the original study, suggesting that the captive environment does not truly replicate the developmental experience of wild larvae, at least in terms of cuticular development. Thus, if there truly are innate tendencies for western larvae to differ (morphologically) from eastern, they are not observable in nature.

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

Animals derive resources from their diet and allocate them to organismal functions such as growth, maintenance, reproduction, and dispersal. How variation in diet quality can affect resource allocation to life-history traits, in particular those important to locomotion and dispersal, is poorly understood. We hypothesize that, particularly for specialist herbivore insects that are in co-evolutionary arms races with host plants, changes in host plant will impact performance. From their coevolutionary arms-race with plants, to a complex migratory life history, Monarch butterflies are among the most iconic insect species worldwide. Population declines initiated international conservation efforts involving the replanting of a variety of milkweed species. However, this practice was implemented with little regard for how diverse defensive chemistry of milkweeds experienced by monarch larvae may affect adult fitness traits. We report that adult flight muscle investment, flight energetics, and maintenance costs depend on the host plant species of larvae, and correlate with concentration of milkweed-derived cardenolides sequestered by adults. Our findings indicate host plant species can impact monarchs by affecting fuel requirements for flight.

Grappling with uncertainty in ecological projections: a case study using the migratory monarch butterfly

AbstractProjecting species’ responses to future climate conditions is critical for anticipating conservation challenges and informing proactive policy and management decisions. However, best practices for choosing climate models for projection ensembles are currently in flux. We compared including a maximum number of models against trimming ensembles based on model validation. This was done within the emerging practice of ensemble building using an increasingly larger number of global climate models (GCMs) for future projections. We used recently reported estimates on primary drivers of population fluctuations for the migratory monarch butterfly (Danaus plexippus) to examine how multiple sources of uncertainty impact population forecasts for a well‐studied species. We compared mean spring temperature and precipitation observed in Texas from 1980 to 2005 with predictions from 16 GCMs to determine which of the models performed best. We then built tailored climate projections accumulating both temperature (in the form of growing degree days) and rainfall using both “complete” (all 16) and “trimmed” (best‐performing) ensembles based on three emission scenarios. We built the tailored projections of spring growing conditions to assess the range of possible climate outcomes and their potential impacts on monarch development. Results were similar when mean predictions were compared between trimmed and complete ensembles. However, when daily projections and uncertainty were accumulated over the entire spring season, we showed substantial differences between ensembles in terms of possible ecological outcomes. Ensembles that used all 16 GCMs included so much uncertainty that projections for future spring conditions ranged from being too cold to too hot for monarch development. GCMs based on best‐performing metrics provided much more useful information, projecting higher spring temperatures for developing monarch larvae in the future which could lead to larger summer populations but also suggesting risk from excessive heat. When there is a strong basis for identifying mechanistic drivers of population dynamics, our results support using a smaller subset of validated GCMs to bracket a range of the most defensible future environmental conditions tailored to the species of interest. Yet understating uncertainty remains a risk, and we recommend clearly articulating the rationale and consequences of selecting GCMs for long‐term projections.

Does the Presence of the Biological Control Agent, Hypena opulenta (Lepidoptera: Erebidae) on Swallow-Worts Deter Monarch Oviposition?

Invasive black and pale swallow-worts (Vincetoxicum nigrum (L.) Moench, and Vincetoxicum rossicum Kelopow), which are related to milkweeds, can act as ecological traps for monarch butterflies (Danaus plexippus L. (Lepidoptera: Nymphalidae)) as they lay eggs on them that fail to develop. A recently approved biological control agent against swallow-worts, Hypena opulenta Christoph, occupies the same feeding guild on swallow-worts as monarch larvae and could be perceived as a competitor to monarchs. We tested how the presence of this defoliating moth on swallow-worts may influence monarch host selection. In a two-year field experiment, we placed pale swallow-wort plants that were either infested with H. opulenta or noninfested as well as common milkweed (Asclepias syriaca L.), into monarch habitats to assess oviposition rates. In the laboratory, monarchs were either given a choice or not between milkweeds and black swallow-worts with or without H. opulenta. While monarchs strongly preferred common milkweed in the field, up to 25% of the eggs we observed were laid on pale swallow-wort, without preference for swallow-wort with (10.7%) or without (14.3%) H. opulenta. In laboratory choice and no-choice tests, monarchs did not lay any eggs on black swallow-wort, likely because of the long-term laboratory rearing on common milkweeds. Our results confirm that pale swallow-wort may act as an oviposition sink to monarchs in Michigan as well. Since the biological control program is still in its infancy, the nature of interactions between monarchs and H. opulenta may change as the biocontrol agent becomes more widespread.

Quantifying neonicotinoid insecticide residues in milkweed and other forbs sampled from prairie strips established in maize and soybean fields

Adding habitat within agricultural ecosystems is necessary to reverse declines in pollinator abundance and diversity. Understanding pesticide exposure to pollinator habitat near crop fields is necessary to support risk assessments. Neonicotinoids applied to maize and soybean seeds can be transported from crop fields to adjacent habitat through dust drift during planting and/or through overland runoff or subsurface flow following planting. Pollinators, especially bees (e.g. Apidae) and monarch butterflies (Danaus plexippus), could be exposed to neonicotinoids through ingestion of contaminated plant material (e.g. milkweed leaves). Neonicotinoids have been detected in pollinator-attractive habitats near crop fields; however, the magnitude and seasonal variation of the concentration of these insecticides has not been determined. We quantified concentrations of clothianidin, thiamethoxam, and imidacloprid in soil and forb leaf tissue, including milkweed (Asclepias spp.) located within reconstructed prairies (3–4 years post-establishment) within or adjacent to maize or soybean fields. Samples taken from April through August in 2017 and 2018 were analyzed with liquid chromatography-tandem mass spectrometry; 100% of soil, 80% of vegetation from blooming forbs, and 80% of milkweed leaf tissue samples had at least one neonicotinoid present above the method detection limit (0.07–0.9 ng g-1). The maximum concentrations detected in 2017 or 2018 of clothianidin, thiamethoxam, and imidacloprid in milkweed leaf tissue samples were 6.6, 12.9, and 2.8 ng g-1, respectively. These values are 10 to 130-fold lower than the chronic LC10 values for monarch larvae, indicating it is unlikely that this route of neonicotinoid exposure will cause adverse effects to monarch larvae.

Landscape- and local-level variables affect monarchs in Midwest grasslands

ContextIt is estimated that over one billion milkweed stems need to be restored to sustain the eastern North American migratory population of monarch butterflies; where and in what context the stems should be placed on the landscape is key to addressing habitat deficits.ObjectivesWe assessed how the amount of appropriate habitat surrounding a particular patch of monarch habitat affects monarch presence and reproduction. To ensure that habitat restoration efforts are targeted towards areas that maximize monarch population growth, it is important to understand the effects of landscape heterogeneity on monarch occurrence in habitat patches (i.e. grasslands with milkweeds) across the landscape.MethodsOver two summers (2018–2019), we surveyed monarch adults, larvae, and eggs at sixty grassland sites in Wisconsin that varied in patch size and landscape context (proportion grassland, forest edge density, and road density). We also estimated milkweed density and floral richness to characterize local patch quality.ResultsAdult monarch abundance was highest at patches with the lowest proportion of surrounding grassland and lowest road density, and was heavily influenced by patch quality variables. Egg and larva density in a patch increased with milkweed density and floral richness within a patch. Patch size was unrelated to monarch abundance.ConclusionsThese results suggest that optimal sites for monarch habitat restoration are within landscapes which contain little habitat and that high milkweed density and floral richness and abundance should be conservation goals.

Disentangling the direct, indirect, and combined effects of experimental warming on a plant–insect herbivore interaction

AbstractThere is increasing evidence that climate warming will have both direct and indirect effects on species. Whereas the direct effects of climate warming represent the proximate physiological consequences of changing abiotic conditions, the indirect effects of climate change reflect changes mediated by at least one other interacting species. The relative importance of these two kinds of effects has been unclear, limiting our ability to generalize the response of different species to climate change. Here, we used a series of experiments to disentangle some of the key direct and indirect effects of warming on the growth of monarch butterfly caterpillars (Danaus plexippus) and showy milkweed plants (Asclepias speciosa) during a window of rapid growth for both species. The effects of warming differed between direct, indirect, and combined effect experiments. Warming from 26°C to 30°C directly increased the growth of both monarch larvae and milkweeds, with monarch and milkweed growth rates showing similar sensitivity to warming. However, in a subsequent experiment, we did not observe significantly increased growth when comparing caterpillars and plants reared at 27°C and 31°C, suggesting that small differences can change the direct effects of warming. When caterpillars that were maintained at laboratory temperatures were fed leaves from host plants that were exposed to warmer temperatures, warming had a negative indirect effect on larval growth rates likely mediated by decreases in milkweed leaf quality. In experiments combining direct and indirect effects, we observed a net positive effect of warming on larval growth rates. Warming had no combined effects on milkweed growth, potentially due to opposing positive direct and negative indirect effects on growth mediated via increased monarch herbivory. These results show how variability among the direct, indirect, and combined effects of even relatively simple, short‐term climatic perturbations can present challenges for predicting the broader effects of climatic warming in multispecies communities.

More milkweed in farmlands containing small, annual crop fields and many hedgerows

Milkweed has declined substantially, with over 80% declines in some agricultural regions. This threatens monarch butterfly (Danaus plexippus) persistence, because monarch larvae feed solely on milkweed. Thus conservation actions are needed to enhance the availability of milkweed, particularly in agricultural landscapes. Conservation actions to date have largely focused on reducing intensive agricultural practices, mainly use of herbicides. However, research suggests that landscape-scale alteration of the cropped portion of an agricultural landscape (the "farmland"), for example, to reduce crop field sizes, can benefit herbaceous plants such as milkweed. Here we collected data on milkweed occurrence and cover in agricultural landscapes in Ontario, Canada, capturing variability in milkweed from field edge to interior by sampling in the interior and along the edges of 68 crop fields. We used these data to evaluate the relative effects of farming practices within the sampled field (e.g. herbicide, fertilizer use) on milkweed versus the effects of mean field size, crop diversity, hedgerow cover, and the proportion of farmland in annual crops in the surrounding landscape. Additionally, we evaluated the effects of these variables on the cover of other herbaceous plants, to identify which—if any—could benefit milkweed without increasing overall weed cover. We found more milkweed at sites surrounded by landscapes with smaller crop fields, lower crop diversity, and higher cover of annual crops. Milkweed was more likely to occur at sites surrounded by landscapes with more hedgerows. These landscape-scale effects on milkweed were often larger than those of within-field farming practices. Importantly, we found that most variables had opposite effects on milkweed relative to other plants. Thus, altering the landscape to benefit milkweed does not imply an increase in weed cover.

Monarch Butterfly (Danaus plexippus) Life-Stage Risks from Foliar and Seed-Treatment Insecticides.

Conservation of North America's eastern monarch butterfly (Danaus plexippus) population would require establishment of milkweed (Asclepias spp.) and nectar plants in the agricultural landscapes of the north central United States. A variety of seed-treatment and foliar insecticides are used to manage early- and late-season pests in these landscapes. Thus, there is a need to assess risks of these insecticides to monarch butterfly life stages to inform habitat conservation practices. Chronic and acute dietary toxicity studies were undertaken with larvae and adults, and acute topical bioassays were conducted with eggs, pupae, and adults using 6 representative insecticides: beta-cyfluthrin (pyrethroid), chlorantraniliprole (anthranilic diamide), chlorpyrifos (organophosphate), imidacloprid, clothianidin, and thiamethoxam (neonicotinoids). Chronic dietary median lethal concentration values for monarch larvae ranged from 1.6 × 10-3 (chlorantraniliprole) to 5.3 (chlorpyrifos) μg/g milkweed leaf, with the neonicotinoids producing high rates of arrested pupal ecdysis. Chlorantraniliprole and beta-cyfluthrin were generally the most toxic insecticides to all life stages, and thiamethoxam and chlorpyrifos were generally the least toxic. The toxicity results were compared to insecticide exposure estimates derived from a spray drift model and/or milkweed residue data reported in the literature. Aerial applications of foliar insecticides are expected to cause high downwind mortality in larvae and eggs, with lower mortality predicted for adults and pupae. Neonicotinoid seed treatments are expected to cause little to no downslope mortality and/or sublethal effects in larvae and adults. Given the vagile behavior of nonmigratory monarchs, considering these results within a landscape-scale context suggests that adult recruitment will not be negatively impacted if new habitat is established in close proximity of maize and soybean fields in the agricultural landscapes of the north central United States. Environ Toxicol Chem 2021;40:1761-1777. © 2021 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.

Range-wide variations in common milkweed traits and their effect on monarch larvae.

Leaf economic spectrum (LES) theory has historically been employed to inform vegetation models of ecosystem processes, but largely neglects intraspecific variation and biotic interactions. We attempt to integrate across environment-plant trait-herbivore interactions within a species at a range-wide scale. We measured traits in 53 populations spanning the range of common milkweed (Asclepias syriaca) and used a common garden to determine the role of environment in driving patterns of intraspecific variation. We used a feeding trial to determine the role of plant traits in monarch (Danaus plexippus) larval development. Trait-trait relationships largely followed interspecific patterns in LES theory and persisted in a common garden when individual traits change. Common milkweed showed intraspecific variation and biogeographic clines in traits. Clines did not persist in a common garden. Larvae ate more and grew larger when fed plants with more nitrogen. A longitudinal environmental gradient in precipitation corresponded to a resource gradient in plant nitrogen, which produces a gradient in larval performance. Biogeographic patterns in common milkweed traits can sometimes be predicted from LES, are largely driven by environmental conditions, and have consequences for monarch larval performance. Changes to nutrient dynamics of landscapes with common milkweed could potentially influence monarch population dynamics. We show how biogeographic trends in intraspecific variation can influence key ecological interactions, especially in common species with large distributions.

Thermal tolerance and environmental persistence of a protozoan parasite in monarch butterflies

Many parasites have external transmission stages that persist in the environment prior to infecting a new host. Understanding how long these stages can persist, and how abiotic conditions such as temperature affect parasite persistence, is important for predicting infection dynamics and parasite responses to future environmental change. In this study, we explored environmental persistence and thermal tolerance of a debilitating protozoan parasite that infects monarch butterflies. Parasite transmission occurs when dormant spores, shed by adult butterflies onto host plants and other surfaces, are later consumed by caterpillars. We exposed parasite spores to a gradient of ecologically-relevant temperatures for 2, 35, or 93 weeks. We tested spore viability by feeding controlled spore doses to susceptible monarch larvae, and examined relationships between temperature, time, and resulting infection metrics. We also examined whether distinct parasite genotypes derived from replicate migratory and resident monarch populations differed in their thermal tolerance. Finally, we examined evidence for a trade-off between short-term within-host replication and long-term persistence ability. Parasite viability decreased in response to warmer temperatures over moderate-to-long time scales. Individual parasite genotypes showed high heterogeneity in viability, but differences did not cluster by migratory vs. resident monarch populations. We found no support for a negative relationship between environmental persistence and within-host replication, as might be expected if parasites invest in short-term reproduction at the cost of longer-term survival. Findings here indicate that dormant spores can survive for many months under cooler conditions, and that heat dramatically shortens the window of transmission for this widespread and virulent butterfly parasite.

Assessing risk of Lymantria dispar L. invasion management to monarch butterflies (Danaus plexippus)

AbstractConflicting conservation goals that lead to management tradeoffs are not uncommon in conservation practice. In this study, we explore a potential conflict between the management of a destructive invasive insect pest, Lymantria dispar, and conservation of a widely declining iconic species, the monarch butterfly (Danaus plexippus). Specifically, we assess the risk of exposure of susceptible monarch larvae to aerial applications of Btk, a Lepidoptera‐specific larvicide used in L. dispar suppression and eradication. Our findings indicate minimal conflict between L. dispar management and monarch conservation, as spatial overlap between Btk aerial applications and monarch larvae was found to be marginal. Furthermore, the results of our study indicated specific actions management can take to further minimize potential conflict between these conservation goals. Our study could serve as a guide to other efforts to evaluate potential tradeoffs between the management of invasive species and the conservation of vulnerable species.

Constant Light and Frequent Schedule Changes Do Not Impact Resistance to Parasites in Monarch Butterflies.

Organisms have evolved internal biological clocks to regulate their activities based on external environmental cues, such as light, temperature, and food. Environmental disruption of these rhythms, such as caused by constant light or frequent light schedule changes, has been shown to impair development, reduce survival, and increase infection susceptibility and disease progression in numerous organisms. However, the precise role of the biological clock in host-parasite interactions is understudied and has focused on unnatural host-parasite combinations in lab-adapted inbred models. Here, we use the natural interaction between monarch butterflies (Danaus plexippus) and their virulent protozoan parasite, Ophryocystis elektroscirrha, to investigate the effects of constant light and frequent light schedule changes on development, survival, and parasite susceptibility. We show that constant light exposure slows the monarchs' rate of development but does not increase susceptibility to parasitic infection. Furthermore, frequent schedule changes decrease parasite growth, but have no effect on egg-to-adult survival of infected monarchs. Interestingly, these conditions are usually disruptive to the biological clock, but do not significantly impact the clock of monarch larvae. These unexpected findings show that constant light and frequent schedule changes can uncouple host and parasite performance and highlight how natural relationships are needed to expand our understanding of clocks in host-parasite interactions.

Monarch caterpillars are robust to combined exposure to the roadside micronutrients sodium and zinc.

Human activities are increasing the environmental availability of micronutrients, including sodium and some essential metals. Micronutrients are often limiting in animal diets but may have negative effects when consumed in excess. Though prior research has documented how elevated exposure to individual micronutrients can impact organismal development and fitness, we know less about combined effects of multiple micronutrients. In the wild, monarch butterfly larvae (Danaus plexippus) commonly consume plants in roadside habitats that contain elevated levels of sodium (from road salt) and zinc (from vehicle wear-and-tear). We reared monarch caterpillars to adulthood to test individual and combined effects of dietary sodium and zinc on components of fitness, sodium-linked phenotypes (proxies for neural and flight muscle development) and concentrations of sodium and zinc in adult butterflies. Monarch survival was not impacted by elevated sodium or zinc individually or in combination. Yet, monarchs feeding on sodium-treated milkweed developed relatively larger eyes, consistent with a positive effect of sodium on neural development. Measurements of element concentrations in butterfly and plant tissue indicated that monarchs had higher zinc levels than those present in zinc-treated milkweed but lower sodium levels than those present in sodium-treated milkweed. Monarchs developing on sodium-treated milkweed also had prolonged development time, which might be a cost associated with developing extra neural tissue or investing in mechanisms to excrete excess dietary sodium during the larval stage. Our results indicate that sodium, more than zinc, is likely influencing phenotypic development and performance of insect pollinators in roadside habitats. Yet, in contrast to previous work, our experiment suggests that the highest levels of sodium found along roads are not always harmful for developing monarchs. Future work could consider how potentially stressful effects of micronutrients could be mitigated by increased macronutrient availability or how developmental factors such as migratory status might increase micronutrient requirements.

Elevated atmospheric concentrations of CO2 increase endogenous immune function in a specialist herbivore.

Animals rely on a balance of endogenous and exogenous sources of immunity to mitigate parasite attack. Understanding how environmental context affects that balance is increasingly urgent under rapid environmental change. In herbivores, immunity is determined, in part, by phytochemistry which is plastic in response to environmental conditions. Monarch butterflies Danaus plexippus, consistently experience infection by a virulent parasite Ophryocystis elektroscirrha, and some medicinal milkweed (Asclepias) species, with high concentrations of toxic steroids (cardenolides), provide a potent source of exogenous immunity. We investigated plant-mediated influences of elevated CO2 (eCO2 ) on endogenous immune responses of monarch larvae to infection by O. elektroscirrha. Recently, transcriptomics have revealed that infection by O. elektroscirrha does not alter monarch immune gene regulation in larvae, corroborating that monarchs rely more on exogenous than endogenous immunity. However, monarchs feeding on medicinal milkweed grown under eCO2 lose tolerance to the parasite, associated with changes in phytochemistry. Whether changes in milkweed phytochemistry induced by eCO2 alter the balance between exogenous and endogenous sources of immunity remains unknown. We fed monarchs two species of milkweed; A. curassavica (medicinal) and A. incarnata (non-medicinal) grown under ambient CO2 (aCO2 ) or eCO2 . We then measured endogenous immune responses (phenoloxidase activity, haemocyte concentration and melanization strength), along with foliar chemistry, to assess mechanisms of monarch immunity under future atmospheric conditions. The melanization response of late-instar larvae was reduced on medicinal milkweed in comparison to non-medicinal milkweed. Moreover, the endogenous immune responses of early-instar larvae to infection by O. elektroscirrha were generally lower in larvae reared on foliage from aCO2 plants and higher in larvae reared on foliage from eCO2 plants. When grown under eCO2 , milkweed plants exhibited lower cardenolide concentrations, lower phytochemical diversity and lower nutritional quality (higher C:N ratios). Together, these results suggest that the loss of exogenous immunity from foliage under eCO2 results in increased endogenous immune function. Animal populations face multiple threats induced by anthropogenic environmental change. Our results suggest that shifts in the balance between exogenous and endogenous sources of immunity to parasite attack may represent an underappreciated consequence of environmental change.

Suitability of native milkweed (Asclepias) species versus cultivars for supporting monarch butterflies and bees in urban gardens.

Public interest in ecological landscaping and gardening is fueling a robust market for native plants. Most plants available to consumers through the horticulture trade are cultivated forms that have been selected for modified flowers or foliage, compactness, or other ornamental characteristics. Depending on their traits, some native plant cultivars seem to support pollinators, specialist insect folivores, and insect-based vertebrate food webs as effectively as native plant species, whereas others do not. There is particular need for information on whether native cultivars can be as effective as true or “wild-type” native species for supporting specialist native insects of conservation concern. Herein we compared the suitability of native milkweed species and their cultivars for attracting and supporting one such insect, the iconic monarch butterfly (Danaus plexippus L.), as well as native bees in urban pollinator gardens. Wild-type Asclepias incarnata L. (swamp milkweed) and Asclepias tuberosa L. (butterfly milkweed) and three additional cultivars of each that vary in stature, floral display, and foliage color were grown in a replicated common garden experiment at a public arboretum. We monitored the plants for colonization by wild monarchs, assessed their suitability for supporting monarch larvae in greenhouse trials, measured their defensive characteristics (leaf trichome density, latex, and cardenolide levels), and compared the proportionate abundance and diversity of bee families and genera visiting their blooms. Significantly more monarch eggs and larvae were found on A. incarnata than A. tuberosa in both years, but within each milkweed group, cultivars were colonized to the same extent as wild types. Despite some differences in defense allocation, all cultivars were as suitable as wild-type milkweeds in supporting monarch larval growth. Five bee families and 17 genera were represented amongst the 2,436 total bees sampled from blooms of wild-type milkweeds and their cultivars in the replicated gardens. Bee assemblages of A. incarnata were dominated by Apidae (Bombus, Xylocopa spp., and Apis mellifera), whereas A. tuberosa attracted relatively more Halictidae (especially Lasioglossum spp.) and Megachilidae. Proportionate abundance of bee families and genera was generally similar for cultivars and their respective wild types. This study suggests that, at least in small urban gardens, milkweed cultivars can be as suitable as their parental species for supporting monarch butterflies and native bees.