Cryptic female Strawberry poison frogs experience elevated predation risk when associating with an aposematic partner

Population divergence in sexual signals may lead to speciation through prezygotic isolation. Sexual signals can change solely due to variation in the level of natural selection acting against conspicuousness. However, directional mate choice (i.e., favoring conspicuousness) across different environments may lead to gene flow between populations, thereby delaying or even preventing the evolution of reproductive barriers and speciation. In this study, we test whether natural selection through predation upon mate‐choosing females can favor corresponding changes in mate preferences. Our study system, Oophaga pumilio, is an extremely color polymorphic neotropical frog with two distinctive antipredator strategies: aposematism and crypsis. The conspicuous coloration and calling behavior of aposematic males may attract both cryptic and aposematic females, but predation may select against cryptic females choosing aposematic males. We used an experimental approach where domestic fowl were encouraged to find digitized images of cryptic frogs at different distances from aposematic partners. We found that the estimated survival time of a cryptic frog was reduced when associating with an aposematic partner. Hence, predation may act as a direct selective force on female choice, favoring evolution of color assortative mating that, in turn, may strengthen the divergence in coloration that natural selection has generated.     

No evidence for differential survival or predation between sympatric color morphs of an aposematic poison frog

Because variation in warning signals slows down the predator education process, aposematic theory predicts that animal warning signals should be monomorphic. Yet, warning color polytypisms are not uncommon in aposematic species. In cases where warning signal variants are separated geographically, adaptation to local predators could explain this variation. However, this cannot explain the persistence of sympatric polymorphisms in aposematic taxa. The strawberry poison frog (Oophaga pumilio) exhibits both allopatric and sympatric warning color variation in and around the Bocas del Toro archipelago of Panama. One explanation that has been proposed for the rapid diversification of O. pumilio coloration in this archipelago is low predation; if island populations have few predators, stabilizing selection would be relaxed opening the door for diversification via selection or genetic drift. Using a combination of mark-recapture and clay model studies, we tested for differences in survival and predation among sympatric red and yellow color morphs of O. pumilio from Bastimentos Island. We found no evidence for differential survival or predation in this population, despite the fact that one morph (red) is more common and widely distributed than the other (yellow). Even in an area of the island where the yellow morph is not found, predator attack rates were similar among morphs. Visual modeling suggests that yellow and red morphs are distinguishable and conspicuous against a variety of backgrounds and by viewers with different visual systems. Our results suggest that general avoidance by predators of red and yellow, both of which are typical warning colors used throughout the animal kingdom, may be contributing to the apparent stability of this polymorphism.     

Spatial variation in the fitness of divergent aposematic phenotypes of the poison frog, Dendrobates tinctorius

Aposematic species use brightly coloured signals to warn potential predators of their unpalatability. The function of these signals is largely believed to be frequency-dependent. All else being equal, stabilizing selection is expected to constrain the evolution of novel signals. However, despite the expected frequency-dependent function of aposematic signals, interpopulation variation in aposematic signals is ubiquitous in nature. Here, we used clay models of the poison frog Dendrobates tinctorius to test the nature of selection in regions containing varying frequencies of frogs possessing the local aposematic signal. Our findings support a role for stabilizing selection in maintaining the local signal type in a region of high signal frequency; however, we observe a lack of stabilizing selection at one site coincident with a decrease in the density of frogs possessing the local signal. Spatial variation in local aposematic signal frequencies may facilitate the evolution of novel signal types by altering the adaptive landscape for divergent aposematic phenotypes. Our results provide evidence for spatial variation in the selective regime acting on aposematic signals within an established aposematic system and highlight the need for further study of the nature of selection acting across different spatial scales in diverse aposematic systems.     

Combination of local selection pressures drives diversity in aposematic signals

The diversity of aposematic signals is one of the most difficult phenomena for understanding the evolution of such signals because aposematic animals are most effectively protected when they are common. Theoretical and experimental studies predict that a combination of local selection pressures could maintain variation in aposematic signals. However, the application of this hypothesis to large-scale geographic variation in aposematic signals, other than mimicry systems, is yet to be tested empirically. I investigated geographic variation in morphological and behavioural aposematic signals of the newts, Cynops pyrrhogaster, and in predation pressures on them in populations ranging over 800 km of latitude. Field experiments demonstrated that local differences in predation pressures explain well the island-mainland variation in the aposematic colouration and behaviour of newts. Furthermore, I found a latitudinal gradient in aposematic colouration but not in behaviour, independent of predation pressures. The results suggested that island-mainland variation in aposematic signals resulting from local differences in predation pressures might also be shaped by several factors, such as temperature, body size variation, and genetic differences, and such factors might act on each aposematic trait differently.     

Diversification and convergence of aposematic phenotypes: truncated receptors and cellular arrangements mediate rapid evolution of coloration in harlequin poison frogs

Aposematic signals represent one of the classical systems to study evolution and, as such, they have received considerable empirical and theoretical investigation. Despite the extensive literature on aposematic coloration, much uncertainty remains about genetic changes responsible for the repeated evolution of similar signals in multiple lineages. Here, we study the diversification and convergence of coloration among lineages of aposematic harlequin poison frogs (Oophaga histrionica complex). Our results suggest that different background phenotypes, showing different color and/or luminance contrast, have evolved independently at least twice in this group. We suggest that cellular arrangements are behind the striking diversity of color and patterns in this group and propose that differences in dorsal background color may be related to either or both, the presence/absence of xanthophores and the dispersion of melanosomes. Our genetic analyses support a role for the melanocortin receptor MC1R in melanosome aggregation, and we show evidence that two different mutations (?433 and C432A) are responsible for the darker phenotypes that may display a more detectable, easier to learn, aposematic signal.     

Assortative mating in poison-dart frogs based on an ecologically important trait

The origin of new species can be influenced by both deterministic and stochastic factors. Mate choice and natural selection may be important deterministic causes of speciation (as opposed to the essentially stochastic factors of geographic isolation and genetic drift). Theoretical models predict that speciation is more likely when mate choice depends on an ecologically important trait that is subject to divergent natural selection, although many authors have considered such mating/ecology pleiotropy, or "magic-traits" to be unlikely. However, phenotypic signals are important in both mate choice and ecological processes such as avoiding predation. In chemically defended species, it may be that the phenotypic characteristics influencing mate choice are the same signals being used to transmit a warning to potential predators, although few studies have demonstrated this in wild populations. We tested for assortative mating between two color morphs of the Strawberry Poison-Dart Frog, Dendrobates pumilio, a group with striking geographic variation in aposematic color patterns. We found that females significantly prefer individuals of their own morph under two different light treatments, indicating strong assortative mating based on multiple coloration cues that are also important ecological signals. This study provides a rare example of one phenotypic trait affecting both ecological viability and nonrandom mating, indicating that mating/ecology pleiotropy is plausible in wild populations, particularly for organisms that are aposematically colored and visually orienting.     

Fisherian and Wrightian theories of speciation

Fisher's theory of sexual selection, Wright's shifting-balance theory, and recent models based on them are reviewed as mechanisms of animal speciation. The joint evolution of mating preferences and secondary sexual characters can cause rapid nonadaptive phenotypic divergence and premating isolation between geographically separated populations, or along a cline. Extensive comparative data on Drosophila species support the suggestion of R. A. Fisher and T. Dobzhansky that the evolution of mating preferences can reinforce partial postmating isolation between sympatric populations. The interaction of natural selection and random genetic drift in local populations with a small effective size can produce a rapid transition between relatively stable phenotypes separated by an adaptive valley, or between chromosomal rearrangements with a heterozygote disadvantage. Large demographic fluctuations, such as frequent random local extinction and colonization, are required for the rapid spread of new adaptations (or karyotypes) when intermediate phenotypes (or rearrangement heterozygotes) are selected against.     

Warning signals are seductive: Relative contributions of color and pattern to predator avoidance and mate attraction in Heliconius butterflies

Visual signaling in animals can serve many uses, including predator deterrence and mate attraction. In many cases, signals used to advertise unprofitability to predators are also used for intraspecific communication. Although aposematism and mate choice are significant forces driving the evolution of many animal phenotypes, the interplay between relevant visual signals remains little explored. Here, we address this question in the aposematic passion‐vine butterfly Heliconius erato by using color‐ and pattern‐manipulated models to test the contributions of different visual features to both mate choice and warning coloration. We found that the relative effectiveness of a model at escaping predation was correlated with its effectiveness at inducing mating behavior, and in both cases wing color was more predictive of presumptive fitness benefits than wing pattern. Overall, however, a combination of the natural (local) color and pattern was most successful for both predator deterrence and mate attraction. By exploring the relative contributions of color versus pattern composition in predation and mate preference studies, we have shown how both natural and sexual selection may work in parallel to drive the evolution of specific animal color patterns.     

Aposematism increases acoustic diversification and speciation in poison frogs

Multimodal signals facilitate communication with conspecifics during courtship, but they can also alert eavesdropper predators. Hence, signallers face two pressures: enticing partners to mate and avoiding detection by enemies. Undefended organisms with limited escape abilities are expected to minimize predator recognition over mate attraction by limiting or modifying their signalling. Alternatively, organisms with anti-predator mechanisms such as aposematism (i.e. unprofitability signalled by warning cues) might elaborate mating signals as a consequence of reduced predation. We hypothesize that calls diversified in association with aposematism. To test this, we assembled a large acoustic signal database for a diurnal lineage of aposematic and cryptic/non-defended taxa, the poison frogs. First, we showed that aposematic and non-aposematic species share similar extinction rates, and aposematic lineages diversify more and rarely revert to the non-aposematic phenotype. We then characterized mating calls based on morphological (spectral), behavioural/physiological (temporal) and environmental traits. Of these, only spectral and temporal features were associated with aposematism. We propose that with the evolution of anti-predator defences, reduced predation facilitated the diversification of vocal signals, which then became elaborated or showy via sexual selection.     

The role of predators in maintaining the geographic organization of aposematic signals

Selective predation of aposematic signals is expected to promote phenotypic uniformity. But while these signals may be uniform within a population, numerous species display impressive variations in warning signals among adjacent populations. Predators from different localities who learn to avoid distinct signals while performing intense selection on others are thus expected to maintain such a geographic organization. We tested this assumption by placing clay frog models, representing distinct color morphs of the Peruvian poison dart frog Ranitomeya imitator and a nonconspicuous frog, reciprocally between adjacent localities. In each locality, avian predators were able to discriminate between warning signals; the adjacent exotic morph experienced up to four times more attacks than the local one and two times more than the nonconspicuous phenotype. Moreover, predation attempts on the exotic morph quickly decreased to almost nil, suggesting rapid learning. This experiment offers direct evidence for the existence of different predator communities performing localized homogenizing selection on distinct aposematic signals.     

Conditions for the spread of conspicuous warning signals: a numerical model with novel insights

The initial evolution of conspicuous warning signals presents an evolutionary problem because selection against rare conspicuous signals is presumed to be strong, and new signals are rare when they first arise. Several possible solutions have been offered to solve this apparent evolutionary paradox, but disagreement persists over the plausibility of some of the proposed mechanisms. In this paper, we construct a deterministic numerical simulation model that allows us to derive the strength of selection on novel warning signals in a wide range of biologically relevant situations. We study the effects of predator psychology (learning, rate of mistaken attacks, and neophobia) on selection. We also study the how prey escape, predation intensity, number of predators, and abundance of different prey types affects selection. The model provides several important results. Selection on novel warning signals is number rather than frequency dependent. In most cases, there exists a threshold number of aposematic individuals below which aposematism is selected against and above which aposematism is selected for. Signal conspicuousness (which increases detection rate) and distinctiveness (which allows predator to distinguish defended from nondefended prey) have opposing effects on evolution of warning signals. A more conspicuous warning signal cannot evolve unless it makes the prey more distinctive from palatable prey, reducing mistaken attacks by predators. A novel warning signal that is learned quickly can spread from lower abundance more easily than a signal that is learned more slowly. However, the relative rate at which the resident signal and the novel signal are learned is irrelevant for the spread of the novel signal. Long-lasting neophobia can facilitate the spread of novel warning signals. Individual selection via the ability of defended prey to escape from predator is not likely to facilitate evolution of conspicuous warning signals if both the resident (cryptic) morph and the novel morph have the same escape probability. Predation intensity (defined as the proportion of palatable prey eaten by the predator) has a strong effect on selection. More intense predation results in strong selection against rare signals, but also strong selective advantage to common signals. The threshold number of aposematic individuals is lower when predation is intense. Thus, the evolution of warning signals may be more likely in environments where predation is intense. The effect of numbers of predators depends on whether predation intensity also changes. When predation intensity is constant, increasing numbers of predators raises the threshold number of aposematic individuals, and thus makes evolution of aposematism more difficult. If predation intensity increases in parallel with number of predators, the threshold number of aposematic individuals does not change much, but selection becomes more intense on both sides of the threshold.     

Rapid population divergence linked with co-variation between coloration and sexual display in strawberry poison frogs

The likelihood of speciation is assumed to increase when sexually selected traits diverge together with ecologically important traits. According to sexual selection theory, the evolution of exaggerated display behavior is driven by increased mating success, but limited by natural selection, for example, through predation. However, the evolution of aposematic coloration (i.e., an ecologically important trait) could relieve the evolution of exaggerated display behavior from the bound of predation, resulting in joint divergence in aposematic coloration and sexual display behavior between populations. We tested this idea by examining conspicuousness, using color contrasts between individuals and their native backgrounds, and sexual display of 118 males from genetically diverged populations of the Strawberry poison frog, Dendrobates pumilio. Our results show that the level of conspicuousness of the population predicts the sexual display behavior of males. Males from conspicuous populations used more exposed calling sites. We argue that changes in aposematic coloration may rapidly cause not only postmating isolation due to poorly adapted hybrids, but also premating isolation through shifts in mating behaviors.     

Sexual selection and conflict as engines of ecological diversification

Ecological diversification presents an enduring puzzle: how do novel ecological strategies evolve in organisms that are already adapted to their ecological niche? Most attempts to answer this question posit a primary role for genetic drift, which could carry populations through or around fitness "valleys" representing maladaptive intermediate phenotypes between alternative niches. Sexual selection and conflict are thought to play an ancillary role by initiating reproductive isolation and thereby facilitating divergence in ecological traits through genetic drift or local adaptation. Here, I synthesize theory and evidence suggesting that sexual selection and conflict could play a more central role in the evolution and diversification of ecological strategies through the co-optation of sexual traits for viability-related functions. This hypothesis rests on three main premises, all of which are supported by theory and consistent with the available evidence. First, sexual selection and conflict often act at cross-purposes to viability selection, thereby displacing populations from the local viability optimum. Second, sexual traits can serve as preadaptations for novel viability-related functions. Third, ancestrally sex-limited sexual traits can be transferred between sexes. Consequently, by allowing populations to explore a broad phenotypic space around the current viability optimum, sexual selection and conflict could act as powerful drivers of ecological adaptation and diversification.     

Does spatial variation in predation pressure modulate selection for aposematism?

It is widely believed that aposematic signals should be conspicuous, but in nature, they vary from highly conspicuous to near cryptic. Current theory, including the honest signal or trade‐off hypotheses of the toxicity–conspicuousness relationship, cannot explain why adequately toxic species vary substantially in their conspicuousness. Through a study of similarly toxic Danainae (Nymphalidae) butterflies and their mimics that vary remarkably in their conspicuousness, we show that the benefits of conspicuousness vary along a gradient of predation pressure. Highly conspicuous butterflies experienced lower avian attack rates when background predation pressure was low, but attack rates increased rapidly as background predation pressure increased. Conversely, the least conspicuous butterflies experienced higher attack rates at low predation pressures, but at high predation pressures, they appeared to benefit from crypsis. Attack rates of intermediately conspicuous butterflies remained moderate and constant along the predation pressure gradient. Mimics had a similar pattern but higher attack rates than their models and mimics tended to imitate the signal of less attacked model species along the predation pressure gradient. Predation pressure modulated signal fitness provides a possible mechanism for the maintenance of variation in conspicuousness of aposematic signals, as well as the initial survival of conspicuous signals in cryptic populations in the process of aposematic signal evolution, and an alternative explanation for the evolutionary gain and loss of mimicry.     

Radiation of the polymorphic Little Devil poison frog (Oophaga sylvatica) in Ecuador

Some South American poison frogs (Dendrobatidae) are chemically defended and use bright aposematic colors to warn potential predators of their unpalatability. Aposematic signals are often frequency‐dependent where individuals deviating from a local model are at a higher risk of predation. However, extreme diversity in the aposematic signal has been documented in poison frogs, especially in Oophaga. Here, we explore the phylogeographic pattern among color‐divergent populations of the Little Devil poison frog Oophaga sylvatica by analyzing population structure and genetic differentiation to evaluate which processes could account for color diversity within and among populations. With a combination of PCR amplicons (three mitochondrial and three nuclear markers) and genome‐wide markers from a double‐digested RAD (ddRAD) approach, we characterized the phylogenetic and genetic structure of 199 individuals from 13 populations (12 monomorphic and 1 polymorphic) across the O. sylvatica distribution. Individuals segregated into two main lineages by their northern or southern latitudinal distribution. A high level of genetic and phenotypic polymorphism within the northern lineage suggests ongoing gene flow. In contrast, low levels of genetic differentiation were detected among the southern lineage populations and support recent range expansions from populations in the northern lineage. We propose that a combination of climatic gradients and structured landscapes might be promoting gene flow and phylogenetic diversification. Alternatively, we cannot rule out that the observed phenotypic and genomic variations are the result of genetic drift on near or neutral alleles in a small number of genes.     

Diversity in warning coloration: selective paradox or the norm?

Aposematic theory has historically predicted that predators should select for warning signals to converge on a single form, as a result of frequency‐dependent learning. However, widespread variation in warning signals is observed across closely related species, populations and, most problematically for evolutionary biologists, among individuals in the same population. Recent research has yielded an increased awareness of this diversity, challenging the paradigm of signal monomorphy in aposematic animals. Here we provide a comprehensive synthesis of these disparate lines of investigation, identifying within them three broad classes of explanation for variation in aposematic warning signals: genetic mechanisms, differences among predators and predator behaviour, and alternative selection pressures upon the signal. The mechanisms producing warning coloration are also important. Detailed studies of the genetic basis of warning signals in some species, most notably Heliconius butterflies, are beginning to shed light on the genetic architecture facilitating or limiting key processes such as the evolution and maintenance of polymorphisms, hybridisation, and speciation. Work on predator behaviour is changing our perception of the predator community as a single homogenous selective agent, emphasising the dynamic nature of predator–prey interactions. Predator variability in a range of factors (e.g. perceptual abilities, tolerance to chemical defences, and individual motivation), suggests that the role of predators is more complicated than previously appreciated. With complex selection regimes at work, polytypisms and polymorphisms may even occur in Müllerian mimicry systems. Meanwhile, phenotypes are often multifunctional, and thus subject to additional biotic and abiotic selection pressures. Some of these selective pressures, primarily sexual selection and thermoregulation, have received considerable attention, while others, such as disease risk and parental effects, offer promising avenues to explore. As well as reviewing the existing evidence from both empirical studies and theoretical modelling, we highlight hypotheses that could benefit from further investigation in aposematic species. Finally by collating known instances of variation in warning signals, we provide a valuable resource for understanding the taxonomic spread of diversity in aposematic signalling and with which to direct future research. A greater appreciation of the extent of variation in aposematic species, and of the selective pressures and constraints which contribute to this once‐paradoxical phenomenon, yields a new perspective for the field of aposematic signalling.     

Population structure and evolutionary progress

Wright's shifting-balance theory is discussed as an example of a process that can cause species to evolve combinations of characters that could not evolve under natural selection alone. A review of the existing theory of peak shifts indicates that the conditions of extreme isolation that are necessary to permit genetic drift to alter the outcome of natural selection in local populations would make gene flow too weak to spread a new combination of genes to other populations in a reasonable time. Instead, it seems likely that major demographic changes must occur in a species for the shifting-balance process to work. A discussion of direct and indirect studies of gene flow in natural populations suggests that the current genetic structure of many species is likely to reflect past demographic events rather than ongoing gene flow. It is possible then that demographic processes could be responsible for spreading new traits in a species, but that would be true whether those new traits evolved only owing to natural selection or owing in addition to genetic drift and other forces.     

Patterns of variation within and between Greek populations of Ceratitis capitata suggest extensive gene flow and latitudinal clines

Four natural Greek populations of Mediterranean fruit fly, Ceratitis capitata (Wiedemann), was studied for genetic variability at 25 enzyme loci. The comparison of polymorphism within and between populations shows two loci with high between-population heterozygosity (HT) and very high fixation index (F(ST)) values, suggesting the presence of balancing selection. The gradual decline of common allele frequency of the polymorphic loci tested indicated that latitudinal clines are present in Greece. Indirect estimates of gene flow based both on Wright's method (Nm*) and on the Slatkin's method (Nm*), which depends on the frequencies of rare alleles found in only one population, revealed a substantial amount of gene flow (Nm = 3.493 and Nm* = 3.197). These estimates of gene flow may well explain why the "introduced" Greek populations of C. capitata, in spite of their low genetic variability, display the same polymorphic loci. Gene flow in combination with natural selection and genetic drift may have played an important role to genetic differentiation in this species in Greece.     

Female preferences for aposematic signal components in a polymorphic poison frog

Aposematic signals may be subject to conflicting selective pressures from predators and conspecifics. We studied female preferences for different components of aposematic coloration in the polymorphic poison frog Oophaga pumilio across several phenotypically distinct populations. This frog shows striking diversity in color and pattern between geographically isolated populations in western Panama. Results indicate that male dorsal color is the most important determiner of female preferences. We did not find consistent evidence for effects of other signal components, such as spotting pattern or ventral color. Females in two populations showed assortative preferences mediated by male dorsal coloration. In a third population we found incomplete color-assortative preference behavior, with females exhibiting strong discrimination toward one novel color but not another. These results hint at a possible interaction between sexual and natural selection: female tolerance of unfamiliar coloration patterns could facilitate the establishment of novel phenotypes that are favored by other selective pressures (e.g., predator biases). Furthermore, our study suggests that specific components of the aposematic signal (i.e., dorsal color, ventral color, and spotting pattern) are affected differently by natural and sexual selection.     

Did natural selection or genetic drift produce the cranial diversification of neotropical monkeys?

A central controversy among biologists is the relative importance of natural selection and genetic drift as creative forces shaping biological diversification (Fisher 1930; Wright 1931). Historically, this controversy has been an effective engine powering several evolutionary research programs during the last century (Provine 1989). While all biologists agree that both processes operate in nature to produce evolutionary change, there is a diversity of opinion about which process dominates at any particular organizational level (from DNA and proteins to complex morphologies). To address this last level, we did a broadscale analysis of cranial diversification among all living New World monkeys. Quantitative genetic models yield specific predictions about the relationship between variation patterns within and between populations that may be used to test the hypothesis that genetic drift is a sufficient explanation for morphological diversification. Diversity at several levels in a hierarchy of taxonomic/phylogenetics relationship was examined from species within genera to families within superfamilies. The major conclusion is that genetic drift can be ruled out as the primary source of evolutionary diversification in cranial morphology among taxa at the level of the genus and above as well as for diversification of most genera. However, drift may account for diversification among species within some Neotropical primate genera, implying that morphological diversification associated with speciation need not be adaptive in some radiations.