A conserved supergene locus controls colour pattern diversity in Heliconius butterflies

We studied whether similar developmental genetic mechanisms are involved in both convergent and divergent evolution. Mimetic insects are known for their diversity of patterns as well as their remarkable evolutionary convergence, and they have played an important role in controversies over the respective roles of selection and constraints in adaptive evolution. Here we contrast three butterfly species, all classic examples of Müllerian mimicry. We used a genetic linkage map to show that a locus, Yb, which controls the presence of a yellow band in geographic races of Heliconius melpomene, maps precisely to the same location as the locus Cr, which has very similar phenotypic effects in its co-mimic H. erato. Furthermore, the same genomic location acts as a "supergene", determining multiple sympatric morphs in a third species, H. numata. H. numata is a species with a very different phenotypic appearance, whose many forms mimic different unrelated ithomiine butterflies in the genus Melinaea. Other unlinked colour pattern loci map to a homologous linkage group in the co-mimics H. melpomene and H. erato, but they are not involved in mimetic polymorphism in H. numata. Hence, a single region from the multilocus colour pattern architecture of H. melpomene and H. erato appears to have gained control of the entire wing-pattern variability in H. numata, presumably as a result of selection for mimetic "supergene" polymorphism without intermediates. Although we cannot at this stage confirm the homology of the loci segregating in the three species, our results imply that a conserved yet relatively unconstrained mechanism underlying pattern switching can affect mimicry in radically different ways. We also show that adaptive evolution, both convergent and diversifying, can occur by the repeated involvement of the same genomic regions.     

Genetic differentiation without mimicry shift in a pair of hybridizing Heliconius species (Lepidoptera: Nymphalidae)

Butterflies in the genus Heliconius have undergone rapid adaptive radiation for warning patterns and mimicry, and are excellent models to study the mechanisms underlying diversification. In Heliconius, mimicry rings typically involve distantly related species, whereas closely related species often join different mimicry rings. Genetic and behavioural studies have n how reproductive isolation in many pairs of Heliconius taxa is largely mediated by natural and sexual selection on wing colour patterns. However, recent studies have uncovered new cases in which pairs of closely related species are near‐perfect mimics of each other. Here, we provide morphometric and genetic evidence for the coexistence of two closely related, hybridizing co‐mimetic species on the eastern slopes of the Andes, H. melpomene amaryllis and H. timareta ssp. nov. , which is described here as H. timareta thelxinoe . A joint analysis of multilocus genotyping and geometric morphometrics of wing shape shows a high level of differentiation between the two species, with only limited gene flow and mixing. Some degree of genetic mixing can be detected, but putative hybrids were rare, only one of 175 specimens being a clear hybrid. In contrast, we found phenotypic differentiation between populations of H. timareta thelxinoe , possibly indicative of strong selection for local mimicry in different communities. In this pair of species, the absence of breakdown of genetic isolation despite near‐identical wing patterns implies that factors other than wing patterns keep the two taxa apart, such as chemical or behavioural signals, or ecological adaptation along a strong altitudinal gradient. © 2013 The Linnean Society of London, Biological Journal of the Linnean Society, 2013, 109 , 830–847.     

Müllerian mimicry of a quantitative trait despite contrasting levels of genomic divergence and selection

Hybrid zones, where distinct populations meet and interbreed, give insight into how differences between populations are maintained despite gene flow. Studying clines in genetic loci and adaptive traits across hybrid zones is a powerful method for understanding how selection drives differentiation within a single species, but can also be used to compare parallel divergence in different species responding to a common selective pressure. Here, we study parallel divergence of wing colouration in the butterflies Heliconius erato and H. melpomene, which are distantly related Müllerian mimics which show parallel geographic variation in both discrete variation in pigmentation, and quantitative variation in structural colour. Using geographic cline analysis, we show that clines in these traits are positioned in roughly the same geographic region for both species, which is consistent with direct selection for mimicry. However, the width of the clines varies markedly between species. This difference is explained in part by variation in the strength of selection acting on colour traits within each species, but may also be influenced by differences in the dispersal rate and total strength of selection against hybrids between the species. Genotyping‐by‐sequencing also revealed weaker population structure in H. melpomene, suggesting the hybrid zones may have evolved differently in each species, which may also contribute to the patterns of phenotypic divergence in this system. Overall, we conclude that multiple factors are needed to explain patterns of clinal variation within and between these species, although mimicry has probably played a central role.     

Evolution of novel mimicry rings facilitated by adaptive introgression in tropical butterflies

Understanding the genetic basis of phenotypic variation and the mechanisms involved in the evolution of adaptive novelty, especially in adaptive radiations, is a major goal in evolutionary biology. Here, we used whole‐genome sequence data to investigate the origin of the yellow hindwing bar in the Heliconius cydno radiation. We found modular variation associated with hindwing phenotype in two narrow noncoding regions upstream and downstream of the cortex gene, which was recently identified as a pigmentation pattern controller in multiple species of Heliconius. Genetic variation at each of these modules suggests an independent control of the dorsal and ventral hindwing patterning, with the upstream module associated with the ventral phenotype and the downstream module with the dorsal one. Furthermore, we detected introgression between H. cydno and its closely related species Heliconius melpomene in these modules, likely allowing both species to participate in novel mimicry rings. In sum, our findings support the role of regulatory modularity coupled with adaptive introgression as an elegant mechanism by which novel phenotypic combinations can evolve and fuel an adaptive radiation.     

Balanced polymorphisms and their divergence in a Heliconius butterfly

The evolution of mimicry in similarly defended prey is well described by the Müllerian mimicry theory, which predicts the convergence of warning patterns in order to gain the most protection from predators. However, despite this prediction, we can find great diversity of color patterns among Müllerian mimics such as Heliconius butterflies in the neotropics. Furthermore, some species have evolved the ability to maintain multiple distinct warning patterns in single populations, a phenomenon known as polymorphic mimicry. The adaptive benefit of these polymorphisms is questionable since variation from the most common warning patterns is expected to be disadvantageous as novel signals are punished by predators naive to them. In this study, we use artificial butterfly models throughout Central and South America to characterize the selective pressures maintaining polymorphic mimicry in Heliconius doris. Our results highlight the complexity of positive frequency‐dependent selection, the principal selective pressure driving convergence among Müllerian mimics, and its impacts on interspecific variation of mimetic warning coloration. We further show how this selection regime can both limit and facilitate the diversification of mimetic traits.     

Development of microsatellite loci from a reference genome for the Neotropical butterfly Heliconius numata and its close relatives

The Neotropical butterfly Heliconius numata (Lepidoptera: Nymphalidae: Heliconiinae) is known for its striking diversity of wing color patterns driven by the Müllerian mimicry of multiple local models and controlled by a single supergene locus. Such fine‐scale variation of traits under strong selection offers a unique opportunity for the study of the ecology and genetics of adaptation. However, little is still known of the population processes driving geographical variation in wing‐pattern phenotypes. We report the characterization of 26 microsatellite markers for the butterfly H. numata, including six located inside the wing color‐pattern supergene region. All markers are polymorphic, with allele numbers ranging from 2 to 21 per locus, an observed heterozygosity of 0.111 to 0.848 and an expected heterozygosity of 0.126 to 0.942. A subset of 18 of these markers was tested on five closely related sympatric Heliconius species with an amplification success ranging from 88% to 94%. The obtained set of microsatellite markers provides a new and useful set of tools to investigate patterns of differentiation and selection in populations of mimetic Heliconius butterflies. Moreover, markers developed within the color‐pattern supergene will facilitate characterization of the association between the genetic architecture and the functional diversity of wing patterns. Finally, the cross‐species amplification success of the described markers extends their utility to also encompass comparative population genetic studies of closely related species within a clade of rapidly diversifying species.     

A single origin of Batesian mimicry among hybridizing populations of admiral butterflies (Limenitis arthemis) rejects an evolutionary reversion to the ancestral phenotype

Batesian mimicry is a fundamental example of adaptive phenotypic evolution driven by strong natural selection. Given the potentially dramatic impacts of selection on individual fitness, it is important to understand the conditions under which mimicry is maintained versus lost. Although much empirical and theoretical work has been devoted to the maintenance of Batesian mimicry, there are no conclusive examples of its loss in natural populations. Recently, it has been proposed that non-mimetic populations of the polytypic Limenitis arthemis species complex represent an evolutionary loss of Batesian mimicry, and a reversion to the ancestral phenotype. Here, we evaluate this conclusion using segregating amplified fragment length polymorphism markers to investigate the history and fate of mimicry among forms of the L. arthemis complex and closely related Nearctic Limenitis species. In contrast to the previous finding, our results support a single origin of mimicry within the L. arthemis complex and the retention of the ancestral white-banded form in non-mimetic populations. Our finding is based on a genome-wide sampling approach to phylogeny reconstruction that highlights the challenges associated with inferring the evolutionary relationships among recently diverged species or populations (i.e. incomplete lineage sorting, introgressive hybridization and/or selection).     

On the evolution of mimicry in avian nestlings

Batesian mimicry (BM), where a nontoxic species resembles a toxic species with aposematic coloring, has been recently described for a Neotropical species of the suboscine passerine (Laniocera hypopyrra). Understanding the order and series in which these characteristics evolved is unknown and requires character information from closely related taxa. Here, we trace the origin of mimetic traits and how they evolved by examining antipredator characteristics using images and other field‐collected trait data from nest and nestlings along with data available in the literature for the Laniisominae clade and closely related taxa. We found that morphological modifications of the downy feathers appeared first in the broader clade leading to the Laniisominae clade followed by further morphological and behavioral characteristics within the Laniisominae clade leading to the full BM. Images of nestlings in the Laniisominae and closely related clades demonstrated the extent of antipredator and camouflage characteristics. We found a complex set of behavioral and morphological traits in this clade for reducing predation from hiding to camouflage to mimicry. We further propose the evolution of two distinctive mimicry strategies in the Laniisominae clade: (1) Batesian Mimicry, as described above and (2) Masquerade, resemblance to inedible objects commonly found in their local environment. This complex set of antipredator traits shed light on the diversity of antipredator characteristics in avian nestlings, particularly in neotropical areas where the avian diversity is highest. Unfortunately, there are hundreds of species in the neotropics that lack basic natural history information on nesting traits, and therefore, we are likely missing critical information on the diversity of antipredator characteristics across avian nestlings.     

Study of Natural Genetic Variation in Early Fitness Traits Reveals Decoupling Between Larval and Pupal Developmental Time in <Emphasis Type="Italic">Drosophila melanogaster</Emphasis>

Characterizing the relationships between genotype and phenotype for developmental adaptive traits is essential to understand the evolutionary dynamics underlying biodiversity. In holometabolous insects, the time to reach the reproductive stage and pupation site preference are two such traits. Here we characterize aspects of the genetic architecture for Developmental Time (decomposed in Larval and Pupal components) and Pupation Height using lines derived from three natural populations of Drosophila melanogaster raised at two temperatures. For all traits, phenotypic differences and variation in plasticity between populations suggest adaptation to the original thermal regimes. However, high variability within populations shows that selection does not exhaust genetic variance for these traits. This could be partly explained by local adaptation, environmental heterogeneity and modifications in the genetic architecture of traits according to environment and ontogenetic stage. Indeed, our results show that the genetic factors affecting Developmental Time and Pupation Height are temperature-specific. Varying relationships between Larval and Pupal Developmental Time between and within populations also suggest stage-specific modifications of genetic architecture for this trait. This flexibility would allow for a somewhat independent evolution of adaptive traits at different environments and life stages, favoring the maintenance of genetic variability and thus sustaining the traits’ evolvabilities.     

Evolutionary rates for multivariate traits: the role of selection and genetic variation

A fundamental question in evolutionary biology is the relative importance of selection and genetic architecture in determining evolutionary rates. Adaptive evolution can be described by the multivariate breeders'' equation (), which predicts evolutionary change for a suite of phenotypic traits () as a product of directional selection acting on them (β) and the genetic variance–covariance matrix for those traits (G). Despite being empirically challenging to estimate, there are enough published estimates of G and β to allow for synthesis of general patterns across species. We use published estimates to test the hypotheses that there are systematic differences in the rate of evolution among trait types, and that these differences are, in part, due to genetic architecture. We find some evidence that sexually selected traits exhibit faster rates of evolution compared with life-history or morphological traits. This difference does not appear to be related to stronger selection on sexually selected traits. Using numerous proposed approaches to quantifying the shape, size and structure of G, we examine how these parameters relate to one another, and how they vary among taxonomic and trait groupings. Despite considerable variation, they do not explain the observed differences in evolutionary rates.     

Experimental Evolution of Alkaloid Tolerance in Sibling <Emphasis Type="Italic">Drosophila</Emphasis> Species with Different Degrees of Specialization

Drosophila buzzatii and Drosophila koepferae are sibling species with marked ecological differences related to their patterns of host exploitation. D. buzzatii is a polyphagous species with a sub-cosmopolitan distribution, while D. koepferae is endemic to the mountain plateaus of the Andes, where it exploits alkaloidiferous columnar cacti as primary hosts. We use experimental evolution to study the phenotypic response of these cactophilic Drosophila when confronting directional selection to cactus chemical defenses for 20 generations. Flies adapted to cactus diets also experienced higher viability on alkaloid-enriched media, suggesting the selection of adaptive genetic variation for chemical-stress tolerance. The more generalist species D. buzzatii showed a rapid adaptive response to moderate levels of secondary metabolites, whereas the columnar cacti specialist D. koepferae tended to maximize fitness under harder conditions. The evolutionary dynamic of fitness-related traits suggested the implication of metabolic efficiency as a key mediator in the adaptive response to chemical stress. Although we found no evidence of adaptation costs accompanying specialization, our results suggest the involvement of compensatory evolution. Overall, our study proposes that differential adaptation to secondary metabolites may contribute to varying degrees of host specialization, favoring niche partitioning among these closely related species.     

Evolutionary processes and its environmental correlates in the cranial morphology of western chipmunks (Tamias)

The importance of the environment in shaping phenotypic evolution lies at the core of evolutionary biology. Chipmunks of the genus Tamias (subgenus Neotamias) are part of a very recent radiation, occupying a wide range of environments with marked niche partitioning among species. One open question is if and how those differences in environments affected phenotypic evolution in this lineage. Herein we examine the relative importance of genetic drift versus natural selection in the origin of cranial diversity exhibited by clade members. We also explore the degree to which variation in potential selective agents (environmental variables) are correlated with the patterns of morphological variation presented. We found that genetic drift cannot explain morphological diversification in the group, thus supporting the potential role of natural selection as the predominant evolutionary force during Neotamias cranial diversification, although the strength of selection varied greatly among species. This morphological diversification, in turn, was correlated with environmental conditions, suggesting a possible causal relationship. These results underscore that extant Neotamias represent a radiation in which aspects of the environment might have acted as the selective force driving species’ divergence.     

Genetic basis of sex-specific color pattern variation in Drosophila malerkotliana

Pigmentation is a rapidly evolving trait that can play important roles in mimicry, sexual selection, thermoregulation, and other adaptive processes in many groups of animals. In Drosophila, pigmentation can differ dramatically among closely related taxa, presenting a good opportunity to dissect the genetic changes underlying species divergence. In this report, we investigate the genetic basis of color pattern variation between two allopatric subspecies of Drosophila malerkotliana, a widespread member of the ananassae species subgroup. In D. malerkotliana malerkotliana, the last three abdominal segments are darkly pigmented in males but not in females, while in D. malerkotliana pallens both sexes lack dark pigmentation. Composite interval mapping in F(2) hybrid progeny shows that this difference is largely controlled by three quantitative trait loci (QTL) located on the 2L chromosome arm, which is homologous to the 3R of D. melanogaster (Muller element E). Using highly recombinant introgression strains produced by repeated backcrossing and phenotypic selection, we show that these QTL do not correspond to any of the candidate genes known to be involved in pigment patterning and synthesis in Drosophila. These results, in combination with similar analyses in other Drosophila species, indicate that different genetic and molecular changes are responsible for the evolution of similar phenotypic traits in different lineages. This feature makes Drosophila color patterns a powerful model for investigating how the genetic basis of trait evolution is influenced by the intrinsic organization of regulatory pathways controlling the development of these traits.     

Model toxin level does not directly influence the evolution of mimicry in the salamander <Emphasis Type="Italic">Plethodon cinereus</Emphasis>

The resemblance between palatable mimics and unpalatable models in Batesian mimicry systems is tempered by many factors, including the toxicity of the model species. Model toxicity is thought to influence both the occurrence of mimicry and the evolution of mimetic phenotypes, such that mimicry is most likely to persist when models are particularly toxic. Additionally, model toxicity may influence the evolution of mimetic phenotype by allowing inaccurate mimicry to evolve through a mechanism termed ‘relaxed selection’. We tested these hypotheses in a salamander mimicry system between the model Notophthalmus viridescens and the mimic Plethodon cinereus, in which N. viridescens toxicity takes the form of tetrodotoxin. Surprisingly, though we discovered geographic variation in model toxin level, we found no support for the hypotheses that model toxicity directly influences either the occurrence of mimicry or the evolution of mimic phenotype. Instead, a link between N. viridescens size and toxicity may indirectly lead to relaxed selection in this mimicry system. Additionally, limitations of predator perception or variation in the rate of phenotypic evolution of models and mimics may account for the evolution of imperfect mimicry in this salamander species. Finally, variation in predator communities among localities or modern changes in environmental conditions may contribute to the patchy occurrence of mimicry in P. cinereus.     

Adaptive divergence in body size overrides the effects of plasticity across natural habitats in the brown trout

The evolution of life-history traits is characterized by trade-offs between different selection pressures, as well as plasticity across environmental conditions. Yet, studies on local adaptation are often performed under artificial conditions, leaving two issues unexplored: (i) how consistent are laboratory inferred local adaptations under natural conditions and (ii) how much phenotypic variation is attributed to phenotypic plasticity and to adaptive evolution, respectively, across environmental conditions? We reared fish from six locally adapted (domesticated and wild) populations of anadromous brown trout (Salmo trutta) in one semi-natural and three natural streams and recorded a key life-history trait (body size at the end of first growth season). We found that population-specific reaction norms were close to parallel across different streams and Q_ST was similar – and larger than F_ST – within all streams, indicating a consistency of local adaptation in body size across natural environments. The amount of variation explained by population origin exceeded the variation across stream environments, indicating that genetic effects derived from adaptive processes have a stronger effect on phenotypic variation than plasticity induced by environmental conditions. These results suggest that plasticity does not “swamp” the phenotypic variation, and that selection may thus be efficient in generating genetic change.     

Hybridization and transgressive exploration of colour pattern and wing morphology in Heliconius butterflies

Hybridization can generate novel phenotypes distinct from those of parental lineages, a phenomenon known as transgressive trait variation. Transgressive phenotypes might negatively or positively affect hybrid fitness, and increase available variation. Closely related species of Heliconius butterflies regularly produce hybrids in nature, and hybridization is thought to play a role in the diversification of novel wing colour patterns despite strong stabilizing selection due to interspecific mimicry. Here, we studied wing phenotypes in first‐ and second‐generation hybrids produced by controlled crosses between either two co‐mimetic species of Heliconius or between two nonmimetic species. We quantified wing size, shape and colour pattern variation and asked whether hybrids displayed transgressive wing phenotypes. Discrete traits underlain by major‐effect loci, such as the presence or absence of colour patches, generate novel phenotypes. For quantitative traits, such as wing shape or subtle colour pattern characters, hybrids only exceed the parental range in specific dimensions of the morphological space. Overall, our study addresses some of the challenges in defining and measuring phenotypic transgression for multivariate traits and our data suggest that the extent to which transgressive trait variation in hybrids contributes to phenotypic diversity depends on the complexity and the genetic architecture of the traits.     

Conserving marine biodiversity: insights from life-history trait candidate genes in Atlantic cod (Gadus morhua)

Recent technological developments have facilitated an increased focus on identifying genomic regions underlying adaptive trait variation in natural populations, and it has been advocated that this information should be important for designating population units for conservation. In marine fishes, phenotypic studies have suggested adaptation through divergence of life-history traits among natural populations, but the distribution of adaptive genetic variation in these species is still relatively poorly known. In this study, we extract information about the geographical distribution of genetic variation for 33 single nucleotide polymorphisms (SNPs) associated with life-history trait candidate genes, and compare this to variation in 70 putatively neutral SNPs in Atlantic cod (Gadus morhua). We analyse samples covering the major population complexes in the eastern Atlantic and find strong evidence for non-neutral levels and patterns of population structuring for several of the candidate gene-associated markers, including two SNPs in the growth hormone 1 gene. Thus, this study aligns with findings from phenotypic studies, providing molecular data strongly suggesting that these or closely linked genes are under selection in natural populations of Atlantic cod. Furthermore, we find that patterns of variation in outlier markers do not align with those observed at selectively neutral markers, and that outlier markers identify conservation units on finer geographical scales than those revealed when analysing only neutral markers. Accordingly, results also suggest that information about adaptive genetic variation will be useful for targeted conservation and management in this and other marine species.     

Genetic architecture of a feeding adaptation: garter snake (Thamnophis) resistance to tetrodotoxin bearing prey

Detailing the genetic basis of adaptive variation in natural populations is a first step towards understanding the process of adaptive evolution, yet few ecologically relevant traits have been characterized at the genetic level in wild populations. Traits that mediate coevolutionary interactions between species are ideal for studying adaptation because of the intensity of selection and the well-characterized ecological context. We have previously described the ecological context, evolutionary history and partial genetic basis of tetrodotoxin (TTX) resistance in garter snakes (Thamnophis). Derived mutations in a voltage-gated sodium channel gene (Na_v1.4) in three garter snake species are associated with resistance to TTX, the lethal neurotoxin found in their newt prey (Taricha). Here we evaluate the contribution of Na_v1.4 alleles to TTX resistance in two of those species from central coastal California. We measured the phenotypes (TTX resistance) and genotypes (Na_v1.4 and microsatellites) in a local sample of Thamnophis atratus and Thamnophis sirtalis. Allelic variation in Na_v1.4 explains 23 per cent of the variation in TTX resistance in T. atratus while variation in a haphazard sample of the genome (neutral microsatellite markers) shows no association with the phenotype. Similarly, allelic variation in Na_v1.4 correlates almost perfectly with TTX resistance in T. sirtalis, but neutral variation does not. These strong correlations suggest that Na_v1.4 is a major effect locus. The simple genetic architecture of TTX resistance in garter snakes may significantly impact the dynamics of phenotypic coevolution. Fixation of a few alleles of major effect in some garter snake populations may have led to the evolution of extreme phenotypes and an ‘escape’ from the arms race with newts.     

QUANTITATIVE GENETICS OF BRYOZOAN PHENOTYPIC EVOLUTION. I. RATE TESTS FOR RANDOM CHANGE VERSUS SELECTION IN DIFFERENTIATION OF LIVING SPECIES

The possible roles of random genetic change and natural selection in bryozoan speciation were analyzed using quantitative genetic methods on breeding data for traits of skeletal morphology in two closely related species of the cheilostome Stylopoma. The hypothesis that morphologic differences between the species are caused entirely by mutation and genetic drift could not be rejected for reasonable rates of mutation maintained for as few as 10³ to 10⁴ generations. Divergence times this short or shorter are consistent with the abrupt appearances of many invertebrate species in the fossil record, commonly followed by millions of years of morphologic stasis. To produce these differences over 10³ generations or fewer, directional selection acting alone would require unrealistically high levels of minimum selective mortality throughout divergence. Thus, selection is unnecessary to explain the divergence of these species, except as a means of accelerating the effects of random genetic change on shorter time scales (directional selection), or decelerating them over longer ones (stabilizing selection). These results are consistent with a variety of models of phenotypic evolution involving random shifts between multiple adaptive peaks. Similar results were obtained by substituting trait heritabilities and genetic covariances reconstructed by partitioning within- and among-colony phenotypic variance in place of the values based on breeding data. Quantitative genetic analysis of speciation in fossil bryozoan lineages is thus justified.