Patterns of genetic variation and local adaptation of a native herbivore to a lethal invasive plant.

Understanding the evolutionary processes that influence fitness is critical to predicting species' responses to selection. Interactions among evolutionary processes including gene flow, drift and the strength of selection can lead to either local adaptation or maladaptation, especially in heterogenous landscapes. Populations experiencing novel environments or resources are ideal for understanding the mechanisms underlying adaptation or maladaptation, specifically in locally co-evolved interactions. We used the interaction between a native herbivore that oviposits on a patchily distributed introduced plant that in turn causes significant mortality to the larvae to test for signatures of local adaptation in areas where the two co-occurred. We used whole-genome sequencing to explore population structure, patterns of gene flow and signatures of local adaptation. We found signatures of local adaptation in response to the introduced plant in the absence of strong population structure with no genetic differentiation and low genetic variation. Additionally, we found localized allele frequency differences within a single population between habitats with and without the lethal plant, highlighting the effects of strong selection. Finally, we identified that selection was acting on larval ability to feed on the plant rather than on females' ability to avoid oviposition, thus uncovering the specific ontogenetic target of selection. Our work highlights the potential for adaptation to occur in a fine-grained landscape in the presence of gene flow and low genetic variation.

Current and lagged climate affects phenology across diverse taxonomic groups.

The timing of life events (phenology) can be influenced by climate. Studies from around the world tell us that climate cues and species' responses can vary greatly. If variation in climate effects on phenology is strong within a single ecosystem, climate change could lead to ecological disruption, but detailed data from diverse taxa within a single ecosystem are rare. We collated first sighting and median activity within a high-elevation environment for plants, insects, birds, mammals and an amphibian across 45 years (1975-2020). We related 10 812 phenological events to climate data to determine the relative importance of climate effects on species' phenologies. We demonstrate significant variation in climate-phenology linkage across taxa in a single ecosystem. Both current and prior climate predicted changes in phenology. Taxa responded to some cues similarly, such as snowmelt date and spring temperatures; other cues affected phenology differently. For example, prior summer precipitation had no effect on most plants, delayed first activity of some insects, but advanced activity of the amphibian, some mammals, and birds. Comparing phenological responses of taxa at a single location, we find that important cues often differ among taxa, suggesting that changes to climate may disrupt synchrony of timing among taxa.

Two sides of the same wing: ventral scales enhance dorsal wing color in the butterfly Speyeria mormonia.

Biological visual signals are often produced by complex interactions between light-absorbing and light-scattering structures, but for many signals, potential interactions between different light-interacting components have yet to be tested. Butterfly wings, for example, are thin enough that their two sides may not be optically isolated. We tested whether ventral wing scales of the Mormon fritillary, Speyeria mormonia, affect the appearance of dorsal orange patches, which are thought to be involved in sexual signaling. Using reflectance spectroscopy, we found that ventral scales, either silvered or non-silvered, make dorsal orange patches significantly brighter, with the silvered scales having the greater effect. Computational modeling indicates that both types of ventral scale enhance the chromatic perceptual signal of dorsal orange patches, with only the silvered scales also enhancing their achromatic perceptual signal. A lack of optical independence between the two sides of the wings of S. mormonia implies that the wing surfaces of butterflies have intertwined signaling functions and evolutionary histories.

Nutrient acquisition across a dietary shift: fruit feeding butterflies crave amino acids, nectivores seek salt.

Evolutionary dietary shifts have major ecological consequences. One likely consequence is a change in nutrient limitation-some nutrients become more abundant in the diet, others become more scarce. Individuals' behavior should change accordingly to match this new limitation regime: they should seek out nutrients that are deficient in the new diet. We investigated the relationship between diet and responses to nutrients using adult Costa Rican butterflies with contrasting feeding habits, testing the hypothesis that animals will respond more positively to nutrients that are scarcer in their diets. Via literature searches and our own data, we showed that nitrogen and sodium are both at lower concentration in nectar than in fruit. We therefore assessed butterflies' acceptance of sodium and four nitrogenous compounds that ranged in complexity from inorganic nitrogen (ammonium chloride) to protein (albumin). We captured wild butterflies, offered them aqueous solutions of each substance, and recorded whether they accepted (drank) or rejected each substance. Support for our hypothesis was mixed. Across the sexes, frugivores were four times more likely to accept amino acids (hydrolyzed casein) than nectivores, in opposition to expectation. In males, nectivores accepted sodium almost three times more frequently than frugivores, supporting expectations. Together, these results suggest that in butterflies, becoming frugivorous is associated with an increased receptivity to amino acids and decreased receptivity to sodium. Nectivory and frugivory are widespread feeding strategies in organisms as diverse as insects, birds, and bats; our results suggest that these feeding strategies may put different pressures on how animals fulfill their nutritional requirements.

From global change to a butterfly flapping: biophysics and behaviour affect tropical climate change impacts.

Difficulty in characterizing the relationship between climatic variability and climate change vulnerability arises when we consider the multiple scales at which this variation occurs, be it temporal (from minute to annual) or spatial (from centimetres to kilometres). We studied populations of a single widely distributed butterfly species, Chlosyne lacinia, to examine the physiological, morphological, thermoregulatory and biophysical underpinnings of adaptation to tropical and temperate climates. Microclimatic and morphological data along with a biophysical model documented the importance of solar radiation in predicting butterfly body temperature. We also integrated the biophysics with a physiologically based insect fitness model to quantify the influence of solar radiation, morphology and behaviour on warming impact projections. While warming is projected to have some detrimental impacts on tropical ectotherms, fitness impacts in this study are not as negative as models that assume body and air temperature equivalence would suggest. We additionally show that behavioural thermoregulation can diminish direct warming impacts, though indirect thermoregulatory consequences could further complicate predictions. With these results, at multiple spatial and temporal scales, we show the importance of biophysics and behaviour for studying biodiversity consequences of global climate change, and stress that tropical climate change impacts are likely to be context-dependent.

Genomic inference accurately predicts the timing and severity of a recent bottleneck in a nonmodel insect population.

The analysis of molecular data from natural populations has allowed researchers to answer diverse ecological questions that were previously intractable. In particular, ecologists are often interested in the demographic history of populations, information that is rarely available from historical records. Methods have been developed to infer demographic parameters from genomic data, but it is not well understood how inferred parameters compare to true population history or depend on aspects of experimental design. Here, we present and evaluate a method of SNP discovery using RNA sequencing and demographic inference using the program δaδi, which uses a diffusion approximation to the allele frequency spectrum to fit demographic models. We test these methods in a population of the checkerspot butterfly Euphydryas gillettii. This population was intentionally introduced to Gothic, Colorado in 1977 and has as experienced extreme fluctuations including bottlenecks of fewer than 25 adults, as documented by nearly annual field surveys. Using RNA sequencing of eight individuals from Colorado and eight individuals from a native population in Wyoming, we generate the first genomic resources for this system. While demographic inference is commonly used to examine ancient demography, our study demonstrates that our inexpensive, all-in-one approach to marker discovery and genotyping provides sufficient data to accurately infer the timing of a recent bottleneck. This demographic scenario is relevant for many species of conservation concern, few of which have sequenced genomes. Our results are remarkably insensitive to sample size or number of genomic markers, which has important implications for applying this method to other nonmodel systems.

Resource allocation as a driver of senescence: life history tradeoffs produce age patterns of mortality.

We investigate the effects of optimal time and resource allocation on age patterns of fertility and mortality for a model organism with (1) fixed maximum lifespan, (2) distinct juvenile and adult diets, and (3) reliance on nonrenewable resources for reproduction. We ask when it is optimal to tolerate starvation vs. conserve resources and then examine the effects of these decisions on adult mortality rates. We find that (1) age-related changes in tradeoffs partition the life cycle into as many as four discrete phases with different optimal behavior and mortality patterns, and (2) given a cost of reproduction, terminal investment can produce a signal of actuarial senescence. Also, given limitations imposed by non-replenishable resources, individuals beginning adult life with more replenishable resources do not necessarily live longer, since they can engage in capital breeding and need not defer reproduction to forage; low reproductive overheads and low costs of starvation also encourage capital breeding and may lead to earlier terminal investment and earlier senescence. We conclude that, even for species with qualitatively similar life histories, differences in physiological, behavioral and environmental tradeoffs or constraints may strongly influence optimal allocation schedules and produce variation in mortality patterns and life expectancy.

Fitness costs of butterfly oviposition on a lethal non-native plant in a mixed native and non-native plant community.

Non-native plants may be unpalatable or toxic, but have oviposition cues similar to native plants used by insects. The herbivore will then oviposit on the plant, but the offspring will be unable to develop. While such instances have been described previously, the fitness costs at the population level in the wild due to the presence of the lethal host have not been quantified, for this or other related systems. We quantified the fitness cost in the field for the native butterfly Pieris macdunnoughii in the presence of the non-native crucifer Thlaspi arvense, based on the spatial distributions of host plants, female butterflies and eggs in the habitat and the survival of the larvae in the wild. We found that 2.9% of eggs were laid on T. arvense on average, with a survival probability of 0, yielding a calculated fitness cost of 3.0% (95% confidence interval 1.7-3.6%) due to the presence of the non-native in the plant community. Survival probability to the pre-pupal stage for eggs laid on two native crucifers averaged 1.6% over 2 years. The magnitude of the fitness cost will vary temporally and spatially as a function of the relative abundance of the non-native plant. We propose that the fine-scale spatial structure of the plant community relative to the butterflies' dispersal ability, combined with the females' broad habitat use, contributes to the fitness costs associated with the non-native plant and the resulting evolutionary trap.

Antibody development to identify components of IIS and mTOR signaling pathways in lepidopteran species, a set of non-model insects.

Nutritional stress impacts many insect species that have differing reproductive strategies and life histories, yet it is unclear how nutrient-sensing signaling pathways mediate tissue-specific responses to changes in dietary input. In Drosophila melanogaster , insulin/insulin-like growth factor (IIS) and mTOR-mediated signaling within adipocytes regulates oogenesis. To facilitate comparative study of nutrient-sensing pathway activity in the fat body, we developed antibodies to assess IIS (anti-FOXO) and mTOR signaling (anti-TOR) across three nymphalid species (Lepidoptera). By optimizing whole-mount fat body immunostaining, we find FOXO nuclear enrichment in adult adipocytes, like that observed in Drosophila . Additionally, we show a previously uncharacterized TOR localization pattern in the fat body.

A single climate driver has direct and indirect effects on insect population dynamics.

Weather drives population dynamics directly, through effects on vital rates, or indirectly, through effects on the population's competitors, predators or prey and thence on vital rates. Indirect effects may include non-additive interactions with density dependence. Detection of climate drivers is critical to predicting climate change effects, but identification of potential drivers may depend on knowing the underlying mechanisms. For the butterfly Speyeria mormonia, one climate driver, snow melt date, has multiple effects on population growth. Snow melt date in year t has density-dependent indirect effects. Through frost effects, early snow melt decreases floral resources, thence per-capita nectar availability, which determines fecundity in the lab. Snow melt date in year t + 1 has density-independent direct effects. These effects explain 84% of the variation in population growth rate. One climate parameter thus has multiple effects on the dynamics of a species with non-overlapping generations, with one effect not detectable without understanding the underlying mechanism.

Novel host unmasks heritable variation in plant preference within an insect population.

Introductions of novel plant species can disturb the historical resource environment of herbivorous insects, resulting in strong selection to either adopt or exclude the novel host. However, an adaptive response depends on heritable genetic variation for preference or performance within the targeted herbivore population, and it is unclear how heritability of host-use preference may differ between novel and historical hosts. Pieris macdunnoughii butterflies in the Rocky Mountains lay eggs on the nonnative mustard Thlaspi arvense, which is lethal to their offspring. Heritability analyses revealed considerable sex-linked additive genetic variation in host preference within a population of this butterfly. This was contrary to general predictions about the genetic basis of preference variation, which are hypothesized to be sex linked between populations but autosomal within populations. Evidence of sex linkage disappeared when butterflies were tested on methanol-based chemical extracts, suggesting these chemicals in isolation may not be the primary driver of female choice among available host plants. Although unexpected, evidence for within-population sex-linked genetic variation in preference for T. arvense over native hosts indicates that persistent maladaptive oviposition on this lethal plant must be maintained by alternative evolutionary dynamics such as migration- or drift-selection balance or pleiotropic constraints.

The Genome of the Margined White Butterfly (Pieris macdunnoughii): Sex Chromosome Insights and the Power of Polishing with PoolSeq Data.

We report a chromosome-level assembly for Pieris macdunnoughii, a North American butterfly whose involvement in an evolutionary trap imposed by an invasive Eurasian mustard has made it an emerging model system for studying maladaptation in plant-insect interactions. Assembled using nearly 100× coverage of Oxford Nanopore long reads, the contig-level assembly comprised 106 contigs totaling 316,549,294 bases, with an N50 of 5.2 Mb. We polished the assembly with PoolSeq Illumina short-read data, demonstrating for the first time the comparable performance of individual and pooled short reads as polishing data sets. Extensive synteny between the reported contig-level assembly and a published, chromosome-level assembly of the European butterfly Pieris napi allowed us to generate a pseudochromosomal assembly of 47 contigs, placing 91.1% of our 317 Mb genome into a chromosomal framework. Additionally, we found support for a Z chromosome arrangement in P. napi, showing that the fusion event leading to this rearrangement predates the split between European and North American lineages of Pieris butterflies. This genome assembly and its functional annotation lay the groundwork for future research into the genetic basis of adaptive and maladaptive egg-laying behavior by P. macdunnoughii, contributing to our understanding of the susceptibility and responses of insects to evolutionary traps.

Unprecedented reorganization of holocentric chromosomes provides insights into the enigma of lepidopteran chromosome evolution.

Chromosome evolution presents an enigma in the mega-diverse Lepidoptera. Most species exhibit constrained chromosome evolution with nearly identical haploid chromosome counts and chromosome-level gene collinearity among species more than 140 million years divergent. However, a few species possess radically inflated chromosomal counts due to extensive fission and fusion events. To address this enigma of constraint in the face of an exceptional ability to change, we investigated an unprecedented reorganization of the standard lepidopteran chromosome structure in the green-veined white butterfly (Pieris napi). We find that gene content in P. napi has been extensively rearranged in large collinear blocks, which until now have been masked by a haploid chromosome number close to the lepidopteran average. We observe that ancient chromosome ends have been maintained and collinear blocks are enriched for functionally related genes suggesting both a mechanism and a possible role for selection in determining the boundaries of these genome-wide rearrangements.

Longevity can buffer plant and animal populations against changing climatic variability.

Both means and year-to-year variances of climate variables such as temperature and precipitation are predicted to change. However, the potential impact of changing climatic variability on the fate of populations has been largely unexamined. We analyzed multiyear demographic data for 36 plant and animal species with a broad range of life histories and types of environment to ask how sensitive their long-term stochastic population growth rates are likely to be to changes in the means and standard deviations of vital rates (survival, reproduction, growth) in response to changing climate. We quantified responsiveness using elasticities of the long-term population growth rate predicted by stochastic projection matrix models. Short-lived species (insects and annual plants and algae) are predicted to be more strongly (and negatively) affected by increasing vital rate variability relative to longer-lived species (perennial plants, birds, ungulates). Taxonomic affiliation has little power to explain sensitivity to increasing variability once longevity has been taken into account. Our results highlight the potential vulnerability of short-lived species to an increasingly variable climate, but also suggest that problems associated with short-lived undesirable species (agricultural pests, disease vectors, invasive weedy plants) may be exacerbated in regions where climate variability decreases.

The fingerprints of global climate change on insect populations.

Synthesizing papers from the last two years, I examined generalizations about the fingerprints of climate change on insects' population dynamics and phenology. Recent work shows that populations can differ in response to changes in climate means and variances. The part of the thermal niche occupied by an insect population, voltinism, plasticity and adaptation to weather perturbations, and interactions with other species can all exacerbate or mitigate responses to climate change. Likewise, land use change or agricultural practices can affect responses to climate change. Nonetheless, our knowledge of effects of climate change is still biased by organism and geographic region, and to some extent by scale of climate parameter.

The amino acids used in reproduction by butterflies: a comparative study of dietary sources using compound-specific stable isotope analysis.

It is a nutritional challenge for nectar-feeding insects to meet the amino acid requirements of oviposition. Here we investigate whether egg amino acids derive from larval diet or are synthesized from nectar sugar in four species of butterfly: Colias eurytheme, Speyeria mormonia, Euphydryas chalcedona, and Heliconius charitonia. These species exhibit a range of life history and differ in degree of shared phylogeny. We use 13C differences among plants to identify dietary sources of amino acid carbon, and we measure amino acid 13C using compound-specific stable isotope analysis. Egg essential amino acids derived solely from the larval diet, with no evidence for metabolic carbon remodeling. Carbon in nonessential amino acids from eggs derived primarily from nectar sugars, with consistent variation in amino acid turnover. There was no relationship between the nonessential amino acids of eggs and host plants, demonstrating extensive metabolic remodeling. Differences between species in carbon turnover were reflected at the molecular level, particularly by glutamate and aspartate. Essential amino acid 13C varied in a highly consistent pattern among larval host plants, reflecting a common isotopic "fingerprint" associated with plant biosynthesis. These data demonstrate conservative patterns of amino acid metabolism among Lepidoptera and the power of molecular stable isotope analyses for evaluating nutrient metabolism in situ.

Fine-Grained Distribution of a Non-Native Resource Can Alter the Population Dynamics of a Native Consumer.

New interactions with non-native species can alter selection pressures on native species. Here, we examined the effect of the spatial distribution of a non-native species, a factor that determines ecological and evolutionary outcomes but that is poorly understood, particularly on a fine scale. Specifically, we explored a native butterfly population and a non-native plant on which the butterfly oviposits despite the plant's toxicity to larvae. We developed an individual-based model to describe movement and oviposition behaviors of each butterfly, which were determined by plant distribution and the butterfly's host preference genotype. We estimated the parameter values of the model from rich field data. We simulated various patterns of plant distributions and compared the rates of butterfly population growth and changes in the allele frequency of oviposition preference. Neither the number nor mean area of patches of non-native species affected the butterfly population, whereas plant abundance, patch shape, and distance to the nearest native and non-native patches altered both the population dynamics and genetics. Furthermore, we found a dramatic decrease in population growth rates when we reduced the distance to the nearest native patch from 147 m to 136 m. Thus changes in the non-native resource distribution that are critical to the fate of the native herbivore could only be detected at a fine-grained scale that matched the scale of a female butterfly's movement. In addition, we found that the native butterfly population was unlikely to be rescued by the exclusion of the allele for acceptance of the non-native plant as a host. This study thus highlights the importance of including both ecological and evolutionary dynamics in analyses of the outcome of species interactions and provides insights into habitat management for non-native species.

Effects of Increased Flight on the Energetics and Life History of the Butterfly Speyeria mormonia.

Movement uses resources that may otherwise be allocated to somatic maintenance or reproduction. How does increased energy expenditure affect resource allocation? Using the butterfly Speyeria mormonia, we tested whether experimentally increased flight affects fecundity, lifespan or flight capacity. We measured body mass (storage), resting metabolic rate and lifespan (repair and maintenance), flight metabolic rate (flight capacity), egg number and composition (reproduction), and food intake across the adult lifespan. The flight treatment did not affect body mass or lifespan. Food intake increased sufficiently to offset the increased energy expenditure. Total egg number did not change, but flown females had higher early-life fecundity and higher egg dry mass than control females. Egg dry mass decreased with age in both treatments. Egg protein, triglyceride or glycogen content did not change with flight or age, but some components tracked egg dry mass. Flight elevated resting metabolic rate, indicating increased maintenance costs. Flight metabolism decreased with age, with a steeper slope for flown females. This may reflect accelerated metabolic senescence from detrimental effects of flight. These effects of a drawdown of nutrients via flight contrast with studies restricting adult nutrient input. There, fecundity was reduced, but flight capacity and lifespan were unchanged. The current study showed that when food resources were abundant, wing-monomorphic butterflies living in a continuous meadow landscape resisted flight-induced stress, exhibiting no evidence of a flight-fecundity or flight-longevity trade-off. Instead, flight changed the dynamics of energy use and reproduction as butterflies adopted a faster lifestyle in early life. High investment in early reproduction may have positive fitness effects in the wild, as long as food is available. Our results help to predict the effect of stressful conditions on the life history of insects living in a changing world.

The evolution of wing color in Colias butterflies: heritability, sex linkage, and population divergence.

We investigated the genetic background of intraspecific variation in wing color across an elevational gradient in the butterfly Colias philodice eriphyle. The degree of wing melanization was an accelerating function of elevation, and differences in wing melanization persisted in a common environment. Full-sibling analysis and parent-offspring regression yielded consistent, moderate to high heritabilities for the degree of wing melanization. The breeding experiments also demonstrated that wing melanization is strongly sex linked. Because traits that differentiate sister species also tend to be sex linked, our results suggest that the genetic mechanisms underlying intraspecific differences in wing melanization are not fundamentally different from those that have been shown to differentiate sister species.

The evolution of wing color: male mate choice opposes adaptive wing color divergence in Colias butterflies.

Correlated evolution of mate signals and mate preference may be constrained if selection pressures acting on mate preference differ from those acting on mate signals. In particular, opposing selection pressures may act on mate preference and signals when traits have sexual as well as nonsexual functions. In the butterfly Colias philodice eriphyle, divergent selection on wing color across an elevational gradient in response to the thermal environment has led to increasing wing melanization at higher elevations. Wing color is also a long-range signal used by males in mate searching. We conducted experiments to test whether sexual selection on wing melanization via male mate choice acts in the same direction as natural selection on mate signals due to the thermal environment. We performed controlled mate choice experiments in the field over an elevational range of 1500 meters using decoy butterflies with different melanization levels. Also, we obtained a more direct estimate of the relation between wing color and sexual selection by measuring mating success in wild-caught females. Both our experiments showed that wing melanization is an important determinant of female mating success in C. p. eriphyle. However, a lack of elevational variation in male mate preference prevents coevolution of mate signals and mate preference, as males at all elevations prefer less-melanized females. We suggest that this apparently maladaptive mate choice may be maintained by differences in detectability between the morphs or by preservation of species recognition.