A multifunctional role for riboflavin in the yellow nectar of Capsicum baccatum and Capsicum pubescens.

A few Capsicum (pepper) species produce yellow-colored floral nectar, but the chemical identity and biological function of the yellow pigment are unknown. A combination of analytical biochemistry techniques was used to identify the pigment that gives Capsicum baccatum and Capsicum pubescens nectars their yellow color. Microbial growth assays, visual modeling, and honey bee preference tests for artificial nectars containing riboflavin were used to assess potential biological roles for the nectar pigment. High concentrations of riboflavin (vitamin B2) give the nectars their intense yellow color. Nectars containing riboflavin generate reactive oxygen species when exposed to light and reduce microbial growth. Visual modeling also indicates that the yellow color is highly conspicuous to bees within the context of the flower. Lastly, field experiments demonstrate that honey bees prefer artificial nectars containing riboflavin. Some Capsicum nectars contain a yellow-colored vitamin that appears to play roles in (1) limiting microbial growth, (2) the visual attraction of bees, and (3) as a reward to nectar-feeding flower visitors (potential pollinators), which is especially interesting since riboflavin is an essential nutrient for brood rearing in insects. These results cumulatively suggest that the riboflavin found in some Capsicum nectars has several functions.

Metabolic stress as a driver of life-history plasticity: flight promotes longevity and antioxidant production in monarch butterflies.

Life-history theory predicts that increased investment in traits related to reproduction will be associated with a reduced ability to invest in survival or longevity. One mechanistic explanation for this trade-off is that metabolic stress generated from current fitness activities (e.g. reproduction or locomotion) will increase somatic damage, leading to reduced longevity. Yet, there has been limited support for this damage-based hypothesis. A possible explanation is that individuals can respond to increases in metabolic stress by plastically inducing cellular maintenance responses, which may increase, rather than decrease, longevity. We tested this possibility by experimentally manipulating investment in flight activity (a metabolic stressor) in the migratory monarch butterfly (Danaus plexippus), a species whose reproductive fitness is dependent on survival through a period of metabolically intensive migratory flight. Consistent with the idea that metabolic stress stimulated investment in self-maintenance, increased flight activity enhanced monarch butterfly longevity and somatic tissue antioxidant capacity, likely at a cost to reproductive investment. Our study implicates a role for metabolic stress as a driver of life-history plasticity and supports a model where current engagement in metabolically stressful activities promotes somatic survival by stimulating investment in self-maintenance processes.

Rearing the Cabbage White Butterfly (Pieris rapae) in Controlled Conditions: A Case Study with Heavy Metal Tolerance.

The cabbage white butterfly (Pieris rapae) is an important system for applied pest control research and basic research in behavioral and nutritional ecology. Cabbage whites can be easily reared in controlled conditions on an artificial diet, making them a model organism of the butterfly world. In this paper, a manipulation of heavy metal exposure is used to illustrate basic methods for rearing this species. The general protocol illustrates how butterflies can be caught in the field, induced to lay eggs in greenhouse cages, and transferred as larvae to artificial diets. The methods show how butterflies can be marked, measured, and studied for a variety of research questions. The representative results give an idea of how artificial diets that vary in components can be used to assess butterfly performance relative to a control diet. More specifically, butterflies were most tolerant to nickel and least tolerant to copper, with a tolerance of zinc somewhere in the middle. Possible explanations for these results are discussed, including nickel hyper-accumulation in some mustard host plants and recent evidence in insects that copper may be more toxic than previously appreciated. Finally, the discussion first reviews variations to the protocol and directions for troubleshooting these methods, before considering how future research might further optimize the artificial diet used in this study. Overall, by providing a detailed video overview of the rearing and measurement of cabbage whites on artificial diets, this protocol provides a resource for using this system across a wide range of studies.

Biology for biomimetics I: function as an interdisciplinary bridge in bio-inspired design.

In bio-inspired design, the concept of 'function' allows engineers and designers to move between biological models and human applications. Abstracting a problem to general functions allows designers to look to traits that perform analogous functions in biological organisms. However, the idea of function can mean different things across fields, presenting challenges for interdisciplinary research. Here we review core ideas in biology that relate to the concept of 'function,' including adaptation, tradeoffs, and fitness, as a companion to bio-inspired design approaches. We align these ideas with a top-down approach in biomimetics, where engineers or designers start with a problem of interest and look to biology for ideas. We review how one can explore a range of biological analogies for a given function by considering function across different parts of an organism's life, such as acquiring nutrients or avoiding disease. Engineers may also draw inspiration from biological traits or systems that exhibit a particular function, but did not necessarily evolve to do so. Such an evolutionary perspective is important to how biodesigners search biological space for ideas. A consideration of the evolution of trait function can also clarify potential trade-offs and biological models that may be more promising for an application. This core set of concepts from evolutionary and organismal biology can aid engineers and designers in their search for biological inspiration.

Developing the genotype-to-phenotype relationship in evolutionary theory: A primer of developmental features.

For decades, there have been repeated calls for more integration across evolutionary and developmental biology. However, critiques in the literature and recent funding initiatives suggest this integration remains incomplete. We suggest one way forward is to consider how we elaborate the most basic concept of development, the relationship between genotype and phenotype, in traditional models of evolutionary processes. For some questions, when more complex features of development are accounted for, predictions of evolutionary processes shift. We present a primer on concepts of development to clarify confusion in the literature and fuel new questions and approaches. The basic features of development involve expanding a base model of genotype-to-phenotype to include the genome, space, and time. A layer of complexity is added by incorporating developmental systems, including signal-response systems and networks of interactions. The developmental emergence of function, which captures developmental feedbacks and phenotypic performance, offers further model elaborations that explicitly link fitness with developmental systems. Finally, developmental features such as plasticity and developmental niche construction conceptualize the link between a developing phenotype and the external environment, allowing for a fuller inclusion of ecology in evolutionary models. Incorporating aspects of developmental complexity into evolutionary models also accommodates a more pluralistic focus on the causal importance of developmental systems, individual organisms, or agents in generating evolutionary patterns. Thus, by laying out existing concepts of development, and considering how they are used across different fields, we can gain clarity in existing debates around the extended evolutionary synthesis and pursue new directions in evolutionary developmental biology. Finally, we consider how nesting developmental features in traditional models of evolution can highlight areas of evolutionary biology that need more theoretical attention.

Post-secretory synthesis of a natural analog of iron-gall ink in the black nectar of Melianthus spp.

The black nectar produced by Melianthus flowers is thought to serve as a visual attractant to bird pollinators, but the chemical identity and synthesis of the black pigment are unknown. A combination of analytical biochemistry, transcriptomics, proteomics, and enzyme assays was used to identify the pigment that gives Melianthus nectar its black color and how it is synthesized. Visual modeling of pollinators was also used to infer a potential function of the black coloration. High concentrations of ellagic acid and iron give the nectar its dark black color, which can be recapitulated through synthetic solutions containing only ellagic acid and iron(iii). The nectar also contains a peroxidase that oxidizes gallic acid to form ellagic acid. In vitro reactions containing the nectar peroxidase, gallic acid, hydrogen peroxide, and iron(iii) fully recreate the black color of the nectar. Visual modeling indicates that the black color is highly conspicuous to avian pollinators within the context of the flower. Melianthus nectar contains a natural analog of iron-gall ink, which humans have used since at least medieval times. This pigment is derived from an ellagic acid-Fe complex synthesized in the nectar and is likely involved in the attraction of passerine pollinators endemic to southern Africa.

Anthropogenic sodium influences butterfly responses to nitrogen-enriched resources: implications for the nitrogen limitation hypothesis.

Humans are increasing the environmental availability of historically limited nutrients, which may significantly influence organismal performance and behavior. Beneficial or stimulatory responses to increases in nitrogen availability (i.e., nitrogen limitation) are generally observed in plants but less consistently in animals. One possible explanation is that animal responses to nitrogen enrichment depend on how nitrogen intake is balanced with sodium, a micronutrient crucial for animals but not plants. We tested this idea in the cabbage white butterfly (Pieris rapae), a species that frequently inhabits nutrient-enriched plants in agricultural settings and roadside verges. We asked (1) whether anthropogenic increases in sodium influence how nitrogen enrichment affects butterfly performance and (2) whether individuals can adaptively adjust their foraging behavior to such effects. Larval nitrogen enrichment enhanced growth of cabbage white larvae under conditions of low but not high sodium availability. In contrast, larval nitrogen enrichment increased egg production of adult females only when individuals developed with high sodium availability. Ovipositing females preferred nitrogen-enriched leaves regardless of sodium availability, while larvae avoided feeding on nitrogen-enriched leaves elevated in sodium. Our results show that anthropogenic increases in sodium influence whether individuals benefit from and forage on nitrogen-enriched resources. Yet, different nitrogen-to-sodium ratios are required to optimize larval and adult performance. Whether increases in sodium catalyze or inhibit benefits of nitrogen enrichment may depend on how evolved nutrient requirements vary across stages of animal development.

Broadening the Taxonomic Breadth of Organisms in the Bio-Inspired Design Process.

(1) Generating a range of biological analogies is a key part of the bio-inspired design process. In this research, we drew on the creativity literature to test methods for increasing the diversity of these ideas. We considered the role of the problem type, the role of individual expertise (versus learning from others), and the effect of two interventions designed to increase creativity-going outside and exploring different evolutionary and ecological "idea spaces" using online tools. (2) We tested these ideas with problem-based brainstorming assignments from a 180-person online course in animal behavior. (3) Student brainstorming was generally drawn to mammals, and the breadth of ideas was affected more by the assigned problem than by practice over time. Individual biological expertise had a small but significant effect on the taxonomic breadth of ideas, but interactions with team members did not. When students were directed to consider other ecosystems and branches of the tree of life, they increased the taxonomic diversity of biological models. In contrast, going outside resulted in a significant decrease in the diversity of ideas. (4) We offer a range of recommendations to increase the breadth of biological models generated in the bio-inspired design process.

Developmental plasticity in multimodal signals: light environment produces novel signalling phenotypes in a butterfly.

Developmental plasticity can alter the expression of sexual signals in novel environments and is therefore thought to play an important role in promoting divergence. Sexual signals, however, are often multimodal and mate choice multivariate. Hence, to understand how developmental plasticity can facilitate divergence, we must assess plasticity across signal components and its cumulative impact on signalling. Here, we examine how developmental plasticity influences different components of cabbage white butterfly Pieris rapae multimodal signals, its effects on their signalling phenotypes and its implications for divergence. To do this, we reared P. rapae caterpillars under two different light environments (low-light and high-light) to simulate conditions experienced by P. rapae colonizing a novel light habitat. We then examined plasticity in both visual (wing coloration) and olfactory (pheromone abundance) components of male sexual signals. We found light environments influenced expression of both visual and olfactory components and resulted in a trade-off between signal modalities. The 'low-light' phenotype had duller wing colours but higher abundance of the pheromone, indole, whereas the 'high-light' phenotype had comparatively brighter wings but lower abundance of indole. These results show that by simultaneously altering expression of different signal components, developmental plasticity can produce multiple signalling phenotypes, which may catalyse divergence.

Anthropogenic Zinc Exposure Increases Mortality and Antioxidant Gene Expression in Monarch Butterflies with Low Access to Dietary Macronutrients.

Biologists seek to understand why organisms vary in their abilities to tolerate anthropogenic contaminants, such as heavy metals. However, few studies have considered how tolerance may be affected by condition-moderating factors such as dietary resource availability. For instance, the availability of crucial limiting macronutrients, such as nitrogen and phosphorous, can vary across space and time either naturally or due to anthropogenic nutrient inputs (e.g., agricultural fertilizers or vehicle emissions). Organisms developing in more macronutrient-rich environments should be of higher overall condition, displaying a greater ability to tolerate metal contaminants. In monarch butterflies (Danaus plexippus), we factorially manipulated dietary macronutrient availability and exposure to zinc, a common metal contaminant in urban habitats that can be toxic but also has nutritional properties. We tested whether (1) the ability to survive zinc exposure depends on dietary macronutrient availability and (2) whether individuals exposed to elevated zinc levels display higher expression of antioxidant genes, given the roles of antioxidants in combatting metal-induced oxidative stress. Exposure to elevated zinc reduced survival only for monarchs developing on a low-macronutrient diet. However, for monarchs developing on a high-macronutrient diet, elevated zinc exposure tended to increase survival. In addition, monarchs exposed to elevated zinc displayed higher expression of antioxidant genes when developing on the low-macronutrient diet but lower expression when developing on the high-macronutrient diet. Altogether, our study shows that organismal survival and oxidative stress responses to anthropogenic zinc contamination depend on the availability of macronutrient resources in the developmental environment. In addition, our results suggest the hypothesis that whether zinc acts as a toxicant or a nutrient may depend on macronutrient supply. Environ Toxicol Chem 2022;41:1286-1296. © 2022 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.

Convergent evolution of a blood-red nectar pigment in vertebrate-pollinated flowers.

Nearly 90% of flowering plants depend on animals for reproduction. One of the main rewards plants offer to pollinators for visitation is nectar. Nesocodon mauritianus (Campanulaceae) produces a blood-red nectar that has been proposed to serve as a visual attractant for pollinator visitation. Here, we show that the nectar's red color is derived from a previously undescribed alkaloid termed nesocodin. The first nectar produced is acidic and pale yellow in color, but slowly becomes alkaline before taking on its characteristic red color. Three enzymes secreted into the nectar are either necessary or sufficient for pigment production, including a carbonic anhydrase that increases nectar pH, an aryl-alcohol oxidase that produces a pigment precursor, and a ferritin-like catalase that protects the pigment from degradation by hydrogen peroxide. Our findings demonstrate how these three enzymatic activities allow for the condensation of sinapaldehyde and proline to form a pigment with a stable imine bond. We subsequently verified that synthetic nesocodin is indeed attractive to Phelsuma geckos, the most likely pollinators of Nesocodon We also identify nesocodin in the red nectar of the distantly related and hummingbird-visited Jaltomata herrerae and provide molecular evidence for convergent evolution of this trait. This work cumulatively identifies a convergently evolved trait in two vertebrate-pollinated species, suggesting that the red pigment is selectively favored and that only a limited number of compounds are likely to underlie this type of adaptation.

Bioinspiration as a method of problem-based STEM education: A case study with a class structured around the COVID-19 crisis.

Bioinspiration is a promising lens for biology instruction as it allows the instructor to focus on current issues, such as the COVID-19 pandemic. From social distancing to oxygen stress, organisms have been tackling pandemic-related problems for millions of years. What can we learn from such diverse adaptations in our own applications? This review uses a seminar course on the COVID-19 crisis to illustrate bioinspiration as an approach to teaching biology content. At the start of the class, students mind-mapped the entire problem; this range of subproblems was used to structure the biology content throughout the entire class. Students came to individual classes with a brainstormed list of biological systems that could serve as inspiration for a particular problem (e.g., absorptive leaves in response to the problem of toilet paper shortages). After exploration of relevant biology content, discussion returned to the focal problem. Students dug deeper into the literature in a group project on mask design and biological systems relevant to filtration and transparency. This class structure was an engaging way for students to learn principles from ecology, evolution, behavior, and physiology. Challenges with this course design revolved around the interdisciplinary and creative nature of the structure; for instance, the knowledge of the participants was often stretched by engineering details. While the present class was focused on the COVID-19 crisis, a course structured through a bioinspired approach can be applied to other focal problems, or subject areas, giving instructors a powerful method to deliver interdisciplinary content in an integrated and inquiry-driven way.

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.

Evaluating costs of heavy metal tolerance in a widely distributed, invasive butterfly.

Organismal tolerance to environmental pollution is thought to be constrained by fitness costs, where variants with higher survival in polluted environments have lower performance in nonpolluted environments. Yet, costs are not always detected in empirical studies. One hypothesis suggests that whether tolerance costs emerge depends on the degree of heterogeneity populations experience with respect to pollution exposure. For instance, in populations confined to local environments where pollution is persistent, selection may favour alleles that enhance pollution tolerance but reduce performance in nonpolluted environments (costs). However, in broadly distributed populations that undergo selection in both polluted and nonpolluted patches, costs should be eroded. Understanding tolerance costs in broadly distributed populations is relevant to management of invasive species, which are highly dispersive, wide ranging, and often colonize disturbed or polluted patches such as agricultural monocultures. Therefore, we conducted a case study quantifying costs of tolerance to zinc pollution (a common heavy metal pollutant) in wild cabbage white butterflies (Pieris rapae). This wide ranging, highly dispersive and invasive pest periodically encounters metal pollution by consuming plants in urban and agricultural settings. In contrast to expected costs of tolerance, we found that cabbage white families with greater zinc tolerance also produced more eggs and had higher reproductive effort under nonpolluted conditions. These results contribute to a more general hypothesis of why costs of pollution tolerance vary across studies: patchy selection with pollutants should erode costs and may favour genotypes that perform well under both polluted and nonpolluted conditions. This might partly explain why widely distributed invasive species are able to thrive in diverse, polluted and nonpolluted habitats.

Lessons from movement ecology for the return to work: Modeling contacts and the spread of COVID-19.

Human behavior (movement, social contacts) plays a central role in the spread of pathogens like SARS-CoV-2. The rapid spread of SARS-CoV-2 was driven by global human movement, and initial lockdown measures aimed to localize movement and contact in order to slow spread. Thus, movement and contact patterns need to be explicitly considered when making reopening decisions, especially regarding return to work. Here, as a case study, we consider the initial stages of resuming research at a large research university, using approaches from movement ecology and contact network epidemiology. First, we develop a dynamical pathogen model describing movement between home and work; we show that limiting social contact, via reduced people or reduced time in the workplace are fairly equivalent strategies to slow pathogen spread. Second, we develop a model based on spatial contact patterns within a specific office and lab building on campus; we show that restricting on-campus activities to labs (rather than labs and offices) could dramatically alter (modularize) contact network structure and thus, potentially reduce pathogen spread by providing a workplace mechanism to reduce contact. Here we argue that explicitly accounting for human movement and contact behavior in the workplace can provide additional strategies to slow pathogen spread that can be used in conjunction with ongoing public health efforts.

Asymmetric interspecific competition drives shifts in signalling traits in fan-throated lizards.

Interspecific competition can occur when species are unable to distinguish between conspecific and heterospecific mates or competitors when they occur in sympatry. Selection in response to interspecific competition can lead to shifts in signalling traits-a process called agonistic character displacement. In two fan-throated lizard species-Sitana laticeps and Sarada darwini-females are morphologically indistinguishable and male agonistic signalling behaviour is similar. Consequently, in areas where these species overlap, males engage in interspecific aggressive interactions. To test whether interspecific male aggression between Si. laticeps and Sa. darwini results in agonistic character displacement, we quantified species recognition and signalling behaviour using staged encounter assays with both conspecifics and heterospecifics across sympatric and allopatric populations of both species. We found an asymmetric pattern, wherein males of Si. laticeps but not Sa. darwini showed differences in competitor recognition and agonistic signalling traits (morphology and behaviour) in sympatry compared with allopatry. This asymmetric shift in traits is probably due to differences in competitive abilities between species and can minimize competitive interactions in zones of sympatry. Overall, our results support agonistic character displacement, and highlight the role of asymmetric interspecific competition in driving shifts in social signals.

Genetic Variation Influences Tolerance to a Neonicotinoid Insecticide in 3 Butterfly Species.

Neonicotinoid pesticides harm nontarget insects, but their sublethal effects on butterflies are understudied. We exposed larvae of 3 butterfly species (Pieris rapae, Colias philodice, and Danaus plexippus) to low levels of the neonicotinoid imidacloprid in their host plants and followed individuals to adulthood. Imidacloprid altered adult body size, especially in female monarchs, but its effects varied across maternal families, highlighting the importance of considering genetic variation in ecotoxicological testing. Environ Toxicol Chem 2020;39:2228-2236. © 2020 SETAC.

Nutritional constraints on brain evolution: Sodium and nitrogen limit brain size.

Nutrition has been hypothesized as an important constraint on brain evolution. However, it is unclear whether the availability of specific nutrients or the difficulty of locating high-quality diets limits brain evolution, especially over long periods of time. We found that dietary nutrient content predicted brain size across 42 species of butterflies. Brain size, relative to body size, was associated with the sodium and nitrogen content of a species' diet. There was no evidence that host plant apparency (measured by plant height) was related to brain evolution. The timing of diet shifts across species varied from 3.5 to 90 million years ago, but nutritional constraints did not lessen over time as species adapted to a diet. Although nutrition was linked to overall brain volume, there was no evidence that nutrition was related to the relative size of individual brain regions. Laboratory rearing experiments confirmed the underlying assumption of most comparative studies that the majority of interspecific trait variation stems from genetically based differences across species rather than developmental plasticity. This study highlights a novel role of sodium and nitrogen in brain evolution, which is additionally interesting given current anthropogenic change in the availability of these nutrients.

Traffic influences nutritional quality of roadside plants for monarch caterpillars.

Roadside habitats are increasingly being targeted for restoration and conservation. Roadside habitats often exhibit altered soil and plant chemistry due to pollution from maintenance (e.g. de-icing salt), car deterioration, and exhaust. Roadside plants may attract animals due to elevated levels of sodium or nitrogen, but high concentrations of heavy metals and sodium can be toxic, potentially setting an ecological trap. In this study, we determine how roads influence the chemistry of common milkweed (Asclepias syriaca) as it is the primary roadside host plant for the declining monarch butterfly (Danaus plexippus) in the eastern United States. Even though road salt is applied during the winter, we detect enhanced sodium along roads the following growing season. Road salts increase soil sodium, which in turn elevates host-plant foliar sodium (occasionally to toxic levels in <10% of plants) and sodium content in monarch caterpillars feeding on these plants. Sodium levels of milkweed leaves are highest close to the edge of busy roads. Some heavy metals (lead, zinc) are also elevated in roadside soils or plants. Nitrogen content was affected by adjacent agricultural use, but not traffic volume or proximity to a road. Other potential road pollutants (e.g. nickel) were not elevated in soil or plants. Despite a clear signature of road pollution in the chemistry of milkweed, most plants are likely still suitable for developing monarchs. Nonetheless, restoration investments in snowy regions should prioritize sites with lower-traffic density that are further from the road edge to minimize toxic impacts of high sodium. To extend this research to other insects of conservation concern, future work should characterize the nutritional quality of nectar, pollen, and other species of host-plants in roadside habitats.

Tolerance of Novel Toxins through Generalized Mechanisms: Simulating Gradual Host Shifts of Butterflies.

Organisms encounter a wide range of toxic compounds in their environments, from chemicals that serve anticonsumption or anticompetition functions to pollutants and pesticides. Although we understand many detoxification mechanisms that allow organisms to consume toxins typical of their diet, we know little about why organisms vary in their ability to tolerate entirely novel toxins. We tested whether variation in generalized stress responses, such as antioxidant pathways, may underlie variation in reactions to novel toxins and, if so, their associated costs. We used an artificial diet to present cabbage white butterfly caterpillars (Pieris rapae) with plant material containing toxins not experienced in their evolutionary history. Families that maintained high performance (e.g., high survival, fast development time, large body size) on diets containing one novel toxic plant also performed well when exposed to two other novel toxic plants, consistent with a generalized response. Variation in constitutive (but not induced) expression of genes involved in oxidative stress responses was positively related to performance on the novel diets. While we did not detect reproductive trade-offs of this generalized response, there was a tendency to have less melanin investment in the wings, consistent with the role of melanin in oxidative stress responses. Taken together, our results support the hypothesis that variation in generalized stress responses, such as genes involved in oxidative stress responses, may explain the variation in tolerance to entirely novel toxins and may facilitate colonization of novel hosts and environments.