Biology subjects

Merrill, R. M.

Publications and source records attributed to Merrill, R. M..

At least 19 recordsLinked to original sources

bioRxiv2026

Adaptation in the eye and brain contributes to species divergence in visual perception in Heliconius butterflies

Sensory systems mediate the interaction between organisms and their environment, but how complex sensory pathways evolve and relate to variation in perception and behavior across ecological contexts, remains poorly understood, especially for terrestrial taxa. Here, we investigate whole-visual-system adaptation in Heliconius erato butterflies. Using continent-wide sampling, we demonstrate that within H. erato, facet count significantly decreased with increasing elevation. Common-garden rearing of low-elevation H. erato populations from Ecuador and their high-elevation sister species, H. himera, showed that eye and brain morphology are heritable, and comparisons to genomic measures of divergence indicates that this variation is due to divergent selection. Parallel comparisons from Colombia involving H. chestertonii (high elevation) and H. erato venus (low elevation) further revealed that eye and brain morphology can evolve as independent, decoupled traits. For both locations, differences in visual acuity correlated with variation in facet count. We also observed parallel evolution of spectral sensitivity, with independent high-elevation populations having fewer red-reflecting lateral filtering pigments. To experimentally link visual system morphology to behavior, we assessed visual acuity in second-generation H. erato cyrbia-H. himera hybrids. Overall, acuity was influenced by facet count, and when analyzed together with brain morphology, by a positive interaction between facet count and optic lobe volume, demonstrating that structural investment in the eye and neural expansion combine to maximize visual perception. This work shows that visual adaptation is a multi-layered process whereby sensory traits can evolve independently under localized ecological pressures, but evolution across the visual pathway contributes to refinements in behavioral performance.

evolutionary biology↗

bioRxiv2026

Ecological divergence and post-eclosion brain development shape visual performance during Heliconius speciation

Sensory adaptation is increasingly recognized as a key driver of ecological speciation, but how visual system divergence is coordinated across development, and how this translates into behavioral differences, remains poorly understood. The butterfly Heliconius cydno, which inhabits closed-canopy forests, has larger eyes and greater investment in visual brain centers than its sympatric close-relative H. melpomene, which occupies more open forest-edge habitats, suggesting divergent ecological selection on the visual system. However, the behavioral consequences of these visual adaptations, their developmental trajectories, and whether they break down in hybrids is unknown. To address these questions, we combined ecological field data with behavioral assays and deep-learning-assisted segmentation of neuroanatomy. Visual acuity - the ability to resolve spatial detail - was higher in H. cydno, consistent with its greater ommatidia number, but also increased with age in both species despite no change in external eye morphology. These improvements coincided with the onset of male courtship and female oviposition, suggesting that early adult neurodevelopment shapes visual performance and may support the demands of reproduction. Brain morphology showed species-specific trajectories of post-eclosion optic lobe growth that broadly paralleled increases in acuity and were accompanied by ongoing neurogenesis in the adult optic lobes. While hybrids exhibited intermediate visual acuity, relationships among different components of the visual system were disrupted in hybrids. Together, these results show that ommatidia number alone cannot explain variation in visual acuity, and highlight how coordinated sensory evolution, and its breakdown in hybrids, may contribute to divergence during the early stages of speciation. Significance StatementAdaptation of sensory systems is increasingly recognized as a key driver of species formation, but it is unclear how these systems develop and shape behavioral differences. Using closely related Heliconius butterflies adapted to different light environments, we show that visual acuity improves during early adulthood through neural development, despite no change in eye structure. This maturation coincides with the onset of reproductive behaviors and is accompanied by growth and neurogenesis in visual brain regions. In hybrids, coordination among sensory traits breaks down, resulting in intermediate visual performance. These results show that sensory development is critical for behavioral adaptation and suggest that its integration contributes to the maintenance of species boundaries.

evolutionary biology↗

bioRxiv2025

Sexual dimorphism in pollen foraging and sensory traits in Heliconius butterflies

Sexual dimorphism in foraging behaviour is widespread in insects and may arise from differences in nutritional demands, sensory systems, or cognition. Heliconius pollen-feeding is an evolutionary innovation among butterflies that supports extended lifespans and sustained reproduction. However, how foraging behaviour varies between the sexes and how it relates to sexually dimorphic traits remains poorly understood. We investigated sex-specific foraging strategies in wild Heliconius himera, a highland specialist from southern Ecuador, using field surveys and DNA metabarcoding. Females carried more pollen than males, consistent with higher nutritional demands, yet we observed no differences in plant richness and composition. This indicates that sex differences reflect effort in foraging behaviour rather than shifts in plant choice. Gut samples revealed greater pollen diversity and a more consistent community profile than proboscis samples, suggesting they better capture cumulative foraging history. We also quantified sexually dimorphic sensory traits and found that males had larger eyes and more ommatidia, whereas females had larger mushroom bodies. While the functional significance of these differences remains unclear, these patterns are consistent with sexual dimorphism reported across Heliconius and suggest males and females may be under divergent selective pressures. Our findings highlight how sex-specific foraging differences can arise from differential effort on shared floral resources and co-occur with divergent sensory and neural investment, offering insights into the ecological basis of intraspecific variation in pollen use.

animal behavior and cognition↗

bioRxiv2025

Evolution of the olfactory system during the radiation of Heliconiini butterflies

Sensory system evolution plays a crucial role in shaping species interactions with their environment, yet the extent to which olfactory system diversity reflects ecological and evolutionary pressures at a macroevolutionary scale remains unclear. Here, we investigate the evolution of the olfactory system across the Heliconiini butterfly tribe, an ecologically diverse but closely related group. Using a comparative approach, we examined variation in antennal lobe morphology and its constituent structures, the glomeruli and antennal lobe hub, as well as olfactory receptor repertoires across species. We found that antennal lobe size variation is driven by independent shifts in glomerular and antennal lobe hub volumes, with species-specific differences occurring against a backdrop of broader phylogenetic stability. While no direct associations with ecological traits were observed, certain species showed large expansions in total glomerular volume and olfactory receptor numbers, warranting further investigation into unmeasured ecological or behavioural factors. Additionally, comparisons between wild-caught and insectary-reared individuals revealed a surprising pattern of developmental plasticity, with antennal lobe hub volumes increasing and glomeruli volumes decreasing in captivity, highlighting the influence of environmental conditions on neural development. These findings suggest that olfactory evolution in Heliconiini is shaped by both evolutionary divergence and developmental plasticity, emphasizing the need to integrate phylogenetic, ecological, and developmental perspectives to fully understand sensory system adaptation.

evolutionary biology↗

bioRxiv2024

Repeated evolution of reduced visual investment at the onset of ecological speciation in high-altitude Heliconius butterflies.

Colonisation of new habitats is typically followed by divergent selection acting on traits that are immediately important for fitness in the new habitat. For example, shifting sensory environments are often associated with variation in sensory traits critical for navigation and foraging. However, the extent to which the initial response to novel sensory conditions is mediated by phenotypic plasticity, and its contribution to early species divergence remains unclear. We took advantage of repeated cases of speciation in Heliconius butterflies with independent allopatric distributions in the west of the Colombian and Ecuadorian Andes. Using volumetric brain measurements, we analysed patterns of investment in sensory processing in brain components across different localities and habitats. We find that a higher-altitude species, H. chestertonii, differs in levels of investment in visual and olfactory brain centres compared to its lower altitude relative H. erato venus, mainly attributable to heritable variation as inferred from comparisons between wild and common-garden reared individuals. We compared these shifts with those reported for another high-altitude species, H. himera, and its parapatric lowland counterpart, H. erato cyrbia, and demonstrate parallel reductions in the size of specific optic lobe neuropils. Conversely, for the antennal lobe, we detected disparate trait shifts in H. himera and H. chestertonii in respect to their lowland erato neighbours. Overall, our findings add weight to the adaptive potential for neuroanatomical divergence related to sensory processing during early species formation. Lay summaryRepeated associations between trait variation and environmental shifts may indicate adaptation to local sources of natural selection. For instance, in fish, the presence of certain morphological traits in specific ecological conditions across independent populations is well documented, suggesting equivalent phenotypic responses to shared sources of natural selection. We compared independent cases of ecological divergence in Heliconius butterflies distributed along altitude gradients from sea level to mid mountain in the west of the Colombian and Ecuadorian Andes. Shifts in altitude involve repeated, abrupt transitions from wet, large-leaved, warm forests to higher dry, open, cold scrubs. We tested hypotheses about the role of these ecological shifts in driving adaptive evolution in neuroanatomical traits during early speciation. We showed that in Heliconius, independent changes in forest-type have been accompanied by heritable parallel patterns of divergence in sensory investment in visual processing in the brain. We propose these differences likely facilitate species divergence in the face of ongoing geneflow.

evolutionary biology↗

bioRxiv2024

Sensory weighting reflects changing patterns of visual investment during ecological divergence in Heliconius butterflies.

Integrating information across sensory modalities enables animals to orchestrate a wide range of complex behaviours. The relative importance placed on one sensory modality over another reflects the reliability of cues in a particular environment and corresponding differences in neural investment. As populations diverge across environmental gradients, the reliability of sensory cues may shift, favouring divergence in neural investment and the weight given to different sensory modalities. During their divergence across closed-forest and forest-edge habitats, closely related butterflies Heliconius cydno and H. melpomene evolved distinct brain morphologies, with the former investing more in vision. Quantitative genetic analyses suggest selection drove these changes, but their behavioural effects remain uncertain. We hypothesised that divergent neural investment may alter sensory weighting. We trained individuals in an associative learning experiment using multimodal colour and odour cues. When positively rewarded stimuli were presented in conflict pairing positively trained colour with negatively trained odour, and vice-versa, H. cydno favoured visual cues more strongly than H. melpomene. Hence, differences in sensory weighting may evolve early during divergence and are predicted by patterns of neural investment. These findings, alongside other examples, imply that differences in sensory weighting stem from divergent investment as adaptations to local sensory environments.

evolutionary biology↗

bioRxiv2023

Mating preferences act independently on individual elements of visual signals in Heliconius butterflies

Mating cues are often comprised of several elements, which can act independently, or in concert to attract a suitable partner. Individual elements may also function in other contexts, such as anti-predator defense or camouflage. In Heliconius butterflies, wing patterns comprise several individual color pattern elements, which advertise the butterflies toxicity to predators. These wing patterns are also mating cues, and males predominantly court females that possess the same wing pattern as their own. However, it is not known whether male preference is based on the full wing pattern or only individual pattern elements. We compared preferences of male H. erato lativitta between female models with the full wing pattern and those with some pattern elements removed. We found no differences in preference between the full wing pattern model and a model with pattern elements removed, indicating that the complete composition of all elements is not essential to the mating signal. Wing pattern preferences also contribute to pre-mating isolation between two other Heliconius taxa, H. erato cyrbia and H. himera, therefore, we next compared preferences for the same models in these species. H. erato cyrbia and H. himera strongly differed in preferences for the models, potentially providing a mechanism for how pre-mating isolation acts between these species. These findings suggest that contrasting levels of selective constraint act on elements across the wing pattern.

evolutionary biology↗

bioRxiv2023

Adaptive divergence in the eyes of Heliconius butterflies likely contributes to pre- and post mating isolation

When populations experience different sensory conditions, natural selection may favor whole sensory system divergence, from the peripheral structures to the brain. We characterized the outer eye morphology of sympatric Heliconius species from different forest types, and their first-generation reciprocal hybrids to test for adaptive visual system divergence and hybrid disruption. In Panama, Heliconius cydno occurs in closed forests, whereas Heliconius melpomene resides in more open areas. Previous work has shown that, among wild individuals, H. cydno has larger eyes than H. melpomene, and there are heritable, habitat-associated differences in the visual brain structures that exceed neutral divergence expectations. Notably, hybrids have intermediate neural phenotypes, suggesting disruption. To test for similar effects in the visual periphery, we reared both species and their hybrids in common garden conditions. We confirm that H. cydno has larger eyes and provide new evidence that this is driven by selection. Hybrid eye morphology is more H. melpomene-like despite body size being intermediate, contrasting with neural trait intermediacy. Thus, eye morphology differences between H. cydno and H. melpomene are consistent with adaptive divergence, and when combined with previous neuroanatomy data, suggest hybrid visual system disruption due to mismatched patterns of intermediacy and dominance in the visual pathway.

evolutionary biology↗

bioRxiv2023

Divergent warning patterns influence male and female mating behaviours in a tropical butterfly

Traits under divergent ecological selection that also function during mating can be important in maintaining species boundaries. Few studies have considered mutual mate choice, where both males and females base mating decisions on the same trait. We investigated the contribution of visual preference to assortative mating between an aposematic butterfly Heliconius cydno and its sympatric relative H. melpomene. Wing colouration in Heliconius butterflies evolved as a warning signal but also functions as a mating cue. We predicted that both sexes of H. cydno contributed to assortative mating by exhibiting visual preference towards conspecific wing colouration. We analysed published and new data from preference experiments, in which males were presented with conspecific and H. melpomene females. We also recorded female responses and mating outcomes in choice experiments, involving conspecific males with either the original white or artificially painted red forewing bands. Both sexes of H. cydno responded more positively towards the conspecific colouration, yet male colouration did not predict mating outcomes in female choice experiments. As courtships are initiated by males in butterflies, our findings suggest that female visual preference might be of secondary importance in preventing gene flow between H. cydno and H. melpomene.

animal behavior and cognition↗

bioRxiv2023

Adaptive introgression of a visual preference gene

Visual preferences are important drivers of mate choice and sexual selection, but little is known of how they evolve at the genetic level. Here we take advantage of the diversity of bright warning patterns displayed by Heliconius butterflies, which are also used during mate choice. We show that two Heliconius species have evolved the same visual mating preferences for females with red patterns by exchanging genetic material through hybridization. Extensive behavioral experiments reveal that male preferences are associated with a genomic region of increased admixture between these two species. Variation in neural expression of regucalcin1, located within this introgressed region, correlates with visual preference across populations, and disruption of regucalcin1 with CRISPR/Cas9 impairs courtship towards conspecific females, proving a direct link between gene and behavior. Our results support a role for hybridization during behavioral evolution, and show how visually-guided behaviors contributing to adaptation and speciation are encoded within the genome.

evolutionary biology↗

bioRxiv2023

Quantifying visual acuity in Heliconius butterflies

Heliconius butterflies are well-known for their colourful wing patterns, which advertise distastefulness to potential predators and are used during mate choice. However, the relative importance of different aspects of these signals will depend on the visual abilities of Heliconius and their predators. Previous studies have investigated colour sensitivity and neural anatomy, but visual acuity (the ability to perceive detail) has not been studied in these butterflies. Here, we provide the first estimate of visual acuity in Heliconius: from a behavioural optomotor assay, we found that mean visual acuity = 0.49 cycles-per-degree (cpd), with higher acuity in males than females. We also estimated visual acuity from eye morphology and reported slightly lower values (mean visual acuity = 0.38 cpd), but acuity was still higher in males. Finally, we estimated how visual acuity affects Heliconius visual perception compared to a potential avian predator. Whereas the bird predator maintained high resolving power, Heliconius lost the ability to resolve detail at greater distances, though colours may remain salient. These results will inform future studies of Heliconius wing pattern evolution, as well as other aspects in these highly visual butterflies, which have emerged as an important system in studies of adaptation and speciation.

evolutionary biology↗

bioRxiv2023

Parallel shifts in flight-height associated with altitude across incipient Heliconius species

Vertical gradients in microclimate, resource availability and interspecific interactions are thought to underly stratification patterns in tropical insect communities. However, only a few studies have explored the adaptive significance of vertical space use during early population divergence. We analysed flight-height variation across speciation events in Heliconius butterflies representing parallel instances of divergence between low and high-altitude populations. We measured flight-height in wild H. erato venus and H. chestertonii, lowland and mountain specialists respectively, and found that H. chestertonii consistently flies at a lower height. We compared these data with previously published results for H. e. cyrbia and H. himera, the latter of which flies lower and, like H. chestertonii, recently colonised high-altitude, dry forests. We show that these repeated trends largely result from shared patterns of selection across equivalent environments, producing parallel trait-shifts in H. himera and H. chestertonii. Although our results imply a signature of local adaptation, we did not find an association between resource distribution and flight-height in H. e. venus and H. chestertonii. We discuss how this pattern may be explained by variation in forest structure and microclimate. Overall, our findings underscore the importance of behavioural adjustments during early divergence mediated by altitude-shifts.

evolutionary biology↗

bioRxiv2023

Oviposition behaviour is not affected by ultraviolet light in a butterfly with sexually-dimorphic expression of a UV-sensitive opsin

Animal vision is important for mediating multiple complex behaviours. In Heliconius butterflies, vision guides fundamental behaviours such as oviposition, foraging and mate choice. Colour vision in Heliconius involves ultraviolet (UV), blue and long-wavelength sensitive photoreceptors (opsins). Additionally, Heliconius possess a duplicated UV opsin, and its expression varies widely within the genus. In Heliconius erato, opsin expression is sexually dimorphic; only females express both UV-sensitive opsins, enabling UV wavelength discrimination. However, the ecological pressures that have driven these sex-specific differences in visual perception remain unresolved. Heliconius females invest heavily in finding hostplants to lay their eggs, a behaviour heavily reliant on visual cues. We tested whether UV vision is used for oviposition in H. erato and Heliconius himera females by manipulating the availability of UV in behavioural experiments under naturalistic conditions. We found that UV did not influence the number of oviposition attempts or the number of eggs laid. In addition, their hostplant, Passiflora punctata, does not reflect UV wavelengths, and models of H. erato female vision suggest only minimal stimulation of the UV opsins. Overall, these findings suggest that UV wavelengths do not directly affect the ability of Heliconius females to find suitable oviposition sites. Alternatively, UV discrimination could be used in the context of foraging or mate choice, but this remains to be tested.

evolutionary biology↗

bioRxiv2022

Does sexual conflict contribute to the evolution of novel warning patterns?

Why warning patterns are so diverse is an enduring evolutionary puzzle. Because predators associate particular patterns with unpleasant experiences, an individuals predation risk should decrease as the local density of its warning pattern increases, promoting pattern monomorphism. Distasteful Heliconius butterflies are known for their diversity of warning patterns. Here, we explore whether interlocus sexual conflict can contribute to their diversification. Male Heliconius use warning patterns as mating cues, but mated females may suffer costs if this leads to harassment, favoring novel patterns. Using simulations, we show that drift alone is unlikely to cause pattern diversification, but that sexual conflict can assist such process. We also find that genetic architecture influences the evolution of male preferences, which track changes in warning pattern due to sexual selection. When male attraction imposes costs on females, this affects the speed at which novel pattern alleles increase. In two experiments, females laid fewer eggs with males present. However, although males in one experiment showed less interest in females with manipulated patterns, we found no evidence that female coloration mitigates sex-specific costs. Overall, male attraction to conspecific warning patterns may impose an unrecognized cost on Heliconius females, but further work is required to determine this experimentally.

evolutionary biology↗

bioRxiv2020

Light environment influences mating behaviours during the early stages of divergence in tropical butterflies

Species divergence is facilitated when traits under divergent selection also act as mating cues. Fluctuations in sensory conditions can alter signal perception independently of adaptation to contrasting sensory environments, but how this fine scale variation affects behavioural isolation has received less attention, especially in terrestrial organisms. The warning patterns of Heliconius butterflies are under selection for aposematism and act as mating cues. Using computer vision, we extracted behavioural data from 1481 hours of video footage for 387 individuals. We show that the putative hybrid species H. heurippa and its close relative H. timareta linaresi differ in their response to divergent warning patterns, and that these differences are strengthened with increased local illuminance. Trials with live individuals reveal low-level assortative mating that are sufficiently explained by differences in visual attraction. Finally, results from hybrid butterflies are consistent with linkage between a major warning pattern gene and the corresponding behaviour, though the differences in behaviour we observe are unlikely to cause rapid reproductive isolation as predicted under a model of hybrid trait speciation. Overall, our results highlight that the role of ecological mating cues for behavioural isolation may depend on the immediate sensory conditions during which they are displayed to conspecifics.

evolutionary biology↗

bioRxiv2020

Neural divergence and hybrid disruption between ecologically isolated Heliconius butterflies

The importance of behavioural evolution during speciation is well established, but we know little about how this is manifest in sensory and neural systems. Although a handful of studies have linked specific neural changes to divergence in host or mate preferences associated with speciation, how brains respond to broad environmental transitions, and whether this contributes to reproductive isolation, remains unknown. Here, we examine divergence in brain morphology and neural gene expression between closely related, but ecologically distinct, Heliconius butterflies. Despite on-going gene flow, sympatric species pairs within the melpomene-cydno complex are consistently separated across a gradient of open to closed forest and decreasing light intensity. By generating quantitative neuroanatomical data for 107 butterflies, we show that H. melpomene and H. cydno have substantial shifts in brain morphology across their geographic range, with divergent structures clustered in the visual system. These neuroanatomical differences are mirrored by extensive divergence in neural gene expression. Differences in both morphology and gene expression are heritable, exceed expected rates of neutral divergence, and result in intermediate traits in first generation hybrid offspring. This likely disrupts neural system function, leading to a mismatch between the environment and the behavioral response of hybrids. Our results suggest that disruptive selection on both neural function and external morphology result in coincident barriers to gene flow, thereby facilitating speciation.

evolutionary biology↗

bioRxiv2020

Clustering of loci controlling species differences in chemical bouquets of sympatric Heliconius butterflies

The degree to which loci promoting reproductive isolation cluster in the genome - i.e. the genetic architecture of reproductive isolation - can influence the tempo and mode of speciation. Tight linkage between these loci can facilitate speciation in the face of gene flow. Pheromones play a role in reproductive isolation in many Lepidoptera species, and the role of endogenously-produced compounds as secondary metabolites decreases the likelihood of pleiotropy associated with many barrier loci. Heliconius butterflies use male sex pheromones to both court females (aphrodisiac wing pheromones) and ward off male courtship (male-transferred anti-aphrodisiac genital pheromones), and it is likely that these compounds play a role in reproductive isolation between Heliconius species. Using a set of backcross hybrids between H. melpomene and H. cydno, we investigated the genetic architecture of putative male pheromone compound production. We found a set of 40 significant quantitative trait loci (QTL) representing 33 potential pheromone compounds. QTL clustered significantly on two chromosomes, chromosome 8 for genital compounds and chromosome 20 for wing compounds, and chromosome 20 was enriched for potential pheromone biosynthesis genes. There was minimal overlap between pheromone QTL and known QTL for mate choice and color pattern. Nonetheless, we did detect linkage between a QTL for wing androconial area and optix, a color pattern locus known to play a role in reproductive isolation in these species. This tight clustering of putative pheromone loci might contribute to coincident reproductive isolating barriers, facilitating speciation despite ongoing gene flow.

evolutionary biology↗

bioRxiv2020

Neural processing genes are linked to divergence in visual mate preference in sympatric Heliconius butterflies

Many animal species remain separate not because they fail to produce viable hybrids, but because their individuals choose not to mate. However, we still know very little of the genetic mechanisms underlying changes in these mate preference behaviours. Heliconius butterflies display bright warning patterns, which they also use to recognize conspecifics. Here, we couple QTL for divergence in visual preference behaviours with population genomic and gene expression analyses of neural tissue (central brain, optic lobes and ommatidia) across development in two sympatric Heliconius species. Within a region containing 200 genes, we identify five genes that are strongly associated with divergent visual preferences. Three of these have previously been implicated in key components of neural signalling (specifically an ionotropic glutamate receptor and two regucalcins), and overall our candidates suggest shifts in behaviour involve changes in visual integration or processing. This would allow preference evolution without altering perception of the wider environment.

evolutionary biology↗