Evol Ecol (2013) 27:769–782 DOI 10.1007/s10682-013-9635-1 ORIGINAL PAPER
Field but not lab paradigms support generalisation by predators of aposematic polymorphic prey: the Oophaga histrionica complex Adolfo Ame´zquita • Laura Castro • Mo´nica Arias Mabel Gonza´lez • Carolina Esquivel
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Received: 30 September 2012 / Accepted: 21 February 2013 Ó Springer Science+Business Media Dordrecht 2013
Abstract The persistence of novel aposematic forms, and thereby the evolution of aposematic polymorphism, remain intriguing. Novel and rare forms could be disproportionally attacked by predators that already learned to avoid a pre-existing and more common aposematic form. Alternatively, novel forms could be less frequently attacked if predators are reluctant to attack unknown potential prey (neophobia) or if previous learning allows them to generalise and recognise the novel form as toxic. We used colour variation in polymorphic poison frogs (Oophaga histrionica complex) to test whether predators familiar with one aposematic form do generalise their avoidance behaviour to other aposematic forms. To strengthen our inference, we combined a field test of attack rates to local and non-local models with a lab experiment of generalisation capabilities by newly born chicks. Field predators attacked a significantly lower proportion of 529 aposematic compared to 150 cryptic models. Predators co-occurring with the local aposematic form of O. histrionica equally avoided non-local forms, especially in areas where the species was abundant. Forty-two lab chicks learned to discriminate between an aposematic and a cryptic image, but failed to generalise to other aposematic images, even though we tried with six combinations of aposematic forms. To better mimic the situation in the field, we further tested whether chicks trained with a set of four simultaneous aposematic images would generalise better. They failed to learn the discrimination task. Our data contrast with previous field studies on other poison frogs, and support a role for generalisation, and arguably not neophobia, in predator avoidance of novel aposematic forms. Keywords
Aposematism Learning Poison frogs Polymorphism Predator cognition
Introduction Many predators avoid potential prey that signal their unprofitability by means of conspicuous coloration, odour or behaviour (Poulton 1890). Conspicuousness is thus A. Ame´zquita (&) L. Castro M. Arias M. Gonza´lez C. Esquivel Department of Biological Sciences, Universidad de los Andes, 4976 Bogota´, Colombia e-mail:
[email protected]
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considered an anti-predator defence in aposematic species. More often than not, conspicuous forms appear to be evolutionarily derived from inconspicuous forms (Guilford 1990). The very origin of aposematic variants appears perplexing because, the first conspicuous individuals in a population should increase the probability of being detected and attacked by ‘‘naı¨ve’’ predators. A few dispersed aposematic variants would then have few opportunities to spread and fixate their genes in the population. Predator avoidance of conspicuous prey is generally based upon learning and reinforcement, and thereby encounter rate (e.g. Skelhorn and Rowe 2006). Predators learn faster when prey are clumpled and look alike in their aposematic (e.g. coloration) signals (Mappes and Alatalo 1997; Greenwood et al. 1989). At the interspecific level, co-occurring aposematic species are predicted to evolve convergent coloration, because interspecific alikeness increases the apparent encounter rate with learning predators and therefore the protective value of aposematic coloration (Mu¨ller 1878, Lindstro¨m et al. 2006). In this context, the evolution of novel coloration forms in aposematic species remains paradoxical (Mallet and Joron 1999; Gray and McKinnon 2006). Although novel coloration may confer advantages in terms of mating success or thermoregulation (Poulton 1890; Maan and Cummings 2008), it would also reduce the probability of survival. Novel forms should be attacked more often than pre-existing and more common forms, already recognised by predators as toxic. At least three scenarios have been conceived to explain the persistence of novel coloration forms, and thus the evolution of aposematic polymorphism. (1) Novel forms in one species may not be rare for predators, for they resemble the coloration of other co-occurring aposematic species. This situation would explain the evolution of complex Mu¨llerian mimicry rings, where variation in one species can be explained from concomitant variation in co-occurring model species (Joron et al. 2001; Mallet and Joron 1999). (2) Novel forms may be rarely attacked by predators, if many of them experience neophobia or dietary conservatism (Marples et al. 1998; Lindstro¨m et al. 2001; Mappes et al. 2005), i.e. if they are reluctant to attack unknown potential prey that do not match their search images. And (3) novel forms may be protected by pre-existing aposematic forms, if trained predators extract certain aposematic cues (e.g. just red, circles, or bright coloration) and use them to avoid novel prey that share these cues with the preexisting ones (Gamberale-Stille and Tullberg 1999; Ruxton et al. 2008; Wollenberg et al. 2008). Predators would then generalise aposematic attributes and prey would be aposematically equivalent to them. The poison dendrobatid frogs have become nowadays a well established model for research on the evolution of aposematism. Most recent research has addressed the relative role of genetic structuring (Rudh et al. 2007; Wang and Shaffer 2008) and mate choice (Summers et al. 1999; Maan and Cummings 2008) in explaining the persistence of novel coloration forms, particularly in Oophaga pumilio. Investigations on the role of predators in the evolution of coloration are less common and deal mainly with the aposematic value of coloration (Saporito et al. 2007; Noonan and Comeault 2009; Chouteau and Angers 2011). The payasita (spanish word for clown) frogs, the O. histrionica complex, represent one of the most striking cases of polymorphism in an aposematic frog. Within a single species, dorsal coloration may be anything from red, yellow, orange, and blue to white, whereas the pattern may involve colour suffusions, vermiculations, few or many blotches, circles and transverse bands (Myers and Daly 1976; Mejı´a et al. 2013). In some populations, the range of intra-population variation in colour and pattern largely exceeds the range of interpopulation divergence. To better understand the persistence of novel coloration forms, and thereby the maintenance of aposematic polymorphism, we tested the general hypothesis
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that predators trained to avoid one aposematic form do generalise and avoid other aposematic forms. To strengthen our inferences we combined field experiments using wax aposematic models and the natural potential predators of O. histrionica (e.g. Noonan and Comeault 2009) with lab experiments using naı¨ve chicks as a paradigm for avian predators (Aronsson and Gamberale-Stille 2008). If the generalisation hypothesis holds, we predict that (1) field predators should avoid attacking wax models with aposematic coloration, including the local coloration pattern of O. histrionica and at least some of the other forms of the same species complex. And (2) naı¨ve chicks trained to avoid one coloration pattern of O. histrionica should later avoid other aposematic forms, i.e. they should generalise. In addition, because we obtained contrasting evidence of generalisation between field and lab experiments, we devised a third experiment aimed to explain the apparent contradiction. Because several aposematic albeit differently coloured prey may co-exist in the field with the same predators guild, we predicted that (3) chicks trained to simultaneously recognise several aposematic forms should generalise better to novel forms than chicks trained with a single aposematic form.
Materials and methods The O. histrionica complex includes four species of poison frogs occurring in the Pacific lowlands of Colombia and Ecuador: O. histrionica, O. lehmanni, O. occultator, and O. sylvatica. There is striking variation in coloration and pattern, mainly but not solely among populations of the widely distributed species: O. histrionica and O. sylvatica. There is also significant variation in the alkaloids cocktail secreted by their granular glands and at least part of this variation is correlated with distance between populations (Myers and Daly 1976). We are not aware of any report of predation on Payasita frogs but Rufous Motmots (Baryphthengus martii, Momotidae) are known to prey on frogs of the sister genus, Dendrobates (Master 1998). As other dendrobatid species, calling and foraging activity is diurnal in O. histrionica. Males are strongly territorial, call throughout most of day from elevated and exposed perches, and will readily react to nearby calling males by approaching and attacking them. Oviposition occurs outside water, females transport recently hatched tadpoles to phytotelmata, and return periodically to feed them with unfertilised eggs. Altogether, coloration, calling activity, and natural history render Payasita frogs a very conspicuous and easy to detect animal in the field. Field experiment To know whether predators in the field do generalise the local aposematic signal to other coloration forms we conducted a predation experiment. We built 800 paraffin wax models, similar in size to O. histrionica, and painted them with odourless paint according to the experimental treatment. The coloration pattern of O. histrionica at the study locality consists of red circles or blotches that contrast against a generally black background (Fig. 1, morph A). The morph is relatively widespread and the nearest known location with a different morph is at least 50 km away. Thus, the local predators are unlikely exposed to other colour patterns of the same species. We painted 200 models with the local aposematic pattern and 200 with brown, which simulates a cryptic frog and serves as control for eventual effects of the paint on the decision of predators. As generalisation tests, we painted 400 additional models with other coloration forms of the species complex that either shared the same general colour but differed in pattern (red transversal bands instead
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Fig. 1 Geographic location of the study site for the field experiment (morph A) and geographic origin of the O. histrionica (A–D) and O. lehmanni (E) morphs used in the field and lab experiments on generalisation skills in potential predators of poison frogs. Frog illustrations by C. Ocan˜a
of blotches, Fig. 1E), or shared the same general pattern but differed in colour (yellow instead of red blotches, Fig. 1D). Chicks and other birds are known to exhibit biased generalisation toward colour rather than pattern (Gamberale-Stille and Tullberg 1999). If predators do generalise, our prediction was that other aposematic forms should be as frequently attacked as the local form.
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At our study site, O. histrionica occurs in isolated aggregations with each ‘‘colony’’ separated from others by 1–3 km. To know whether local predators actually recognise the local morph of O. histrionica rather than react to colourful objects, we conducted the experiment in areas where O. histrionica was present (four transects) and where it was absent (two transects). The distance between two transects was never less than 1 km. We predicted that attacks on O. histrionica models bearing the local coloration should be lower in areas where the species was present because the co-occurring predators would already be trained. To set up the models in the field, we defined stations at 20 m steps along each transect. Between 4 and 12 randomly chosen models were then randomly distributed within 10 m around each station. To mimic the spatial distribution of calling males in the field, the distance between two models was never less than 1 m. The models remained in the field during 3 days and, when a model had been removed or had any evidence of attacks, it was replaced with a new model to initiate the next distribution round. All experimental rounds were conducted throughout 22 days, at the beginning of the rainy season. To test whether the Morph (three aposematic and one cryptic) or the Area (with or without O. histrionica) affected the probability of Attack (binary, attacked or not attacked), we built a Generalised Linear Model, with binomial distribution and logit link function. Because the models remained exposed in the field a variable number of days, we included the number of days as a weighting variable. Because the local model could be more frequently attacked than the other ones in areas where the species was present, we included the interaction term Morph x Area as an additional predictor in the model. As a post hoc test to compare the performance between pairs of colour morphs, we used the model profile plot; two colour morphs were considered to differ in the probability of being attacked when there was no overlap between their corresponding 95 % confidence intervals. Lab experiment We used domestic naı¨ve chicks Gallus gallus domesticus as a proxy for colour-vision predators. The species was chosen because (1) birds are reported predators of other poison frogs (Master 1998) or aposematic animals in general, (2) using chicks allows comparison with many other investigations on aposematism and mimicry (e.g. Gamberale-Stille and Tullberg 1999), and (3) avian (and in particular chicks) colour vision is relatively well understood at the physiological level (e.g. Osorio et al. 1999). We obtained from a shop 57 newly hatched (2–5 days old) chicks of unknown sex, that arrived in batches of 10–15 individuals. They were individually marked, housed in groups of four in metal cages (70 cm 9 50 cm 9 20 cm, length 9 width 9 height), and heated with incandescent 25 W bulbs. We fed them ad libitum with chick starter crumbs and water during the housing period, except during training and experimentation, when food deprivation was necessary. Training and testing took place in a rectangular arena (130 cm 9 60 cm 9 80 cm) with 10 circular wells, i.e. Petri dishes 4 cm in diameter, sunk about 1 cm into the floor, and spaced uniformly along the runway. We first trained the chicks to forage worms in the experimental arena, twice a day (morning and afternoon) and during 2 days. As experimental prey we used live Tenebrio molitor and Tribolium confusum larvae that were located in the middle of three wells. Chicks were deprived from food 1 h before each training session and moved in groups of 4–6. In the first session, the morning after arrival, chicks were released in groups of three to forage in the runway. The session ended when the birds passed the 10th well. We gradually reduced the number of companion chicks until the birds seemed to be not
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distressed by foraging alone, which happened around the fourth training session for most of them. To reduce stress effects, we only used for the main experiment the chicks that look not distressed during the initial training phase. The main experiment consisted of two parts: (1) a discrimination-learning test with seven trials, and (2) a generalisation test with one trial under extinction conditions. In the discrimination-learning experiment, we tested whether chicks learn to discriminate between a cryptic and an aposematic image. Two stimuli were presented simultaneously within each well. As unconditioned stimulus (US) we offered worms that were either untreated (palatable) or had been soaked for 2 h in a solution of 30 mL of water and 2 g of chloroquinine phosphate (distasteful). Each worm was placed centrally on top of the conditioned stimulus (CS), which was an oval 2.5 cm 9 1.5 cm photograph of a frog’s dorsal colour pattern printed in Kodak matte paper. As positive conditioned reinforcers we matched photographs of a brown and cryptic Hyloxalus frog to the palatable worms. And as negative conditioned reinforcers, we matched photographs of different aposematic frogs of the O. histrionica complex (Fig. 1, pattern A for 21 chicks and pattern C for 22 chicks) to the distasteful worms. Although we measured colour reflectance of the photographs, we did not calibrate or modify them according to the visual system of the chicks. Instead, we tested several combinations of learning and generalisation stimuli that mimicked the colour patterns of the live frogs. At the beginning of each session, we randomly assigned 10 positive and 10 negative reinforcers to the arena wells, but avoiding more than two consecutive reinforcers of the same type. We then placed the chick in any of several positions along the runway, waited until the chick passed all the 10 wells, and noted the type and number of attacked prey. To know whether trained chicks can generalise relevant aposematic cues from the discrimination-learning test to recognise other aposematic morphs, we conducted generalisation tests. Each of the two groups of chicks (trained with reinforcer A or C, Fig. 1) was split into three subgroups, that were presented respectively with the patterns E, B, and D (Fig. 1) in replacement of the original negative reinforcer. Because individual chicks may differ in their learning performance, we assigned chicks to the subgroups minimising among subgroups differences in the mean performance observed during the sixth learning session. All worms were palatable during the generalisation tests, to avoid the potential effect of any additional cue besides coloration pattern. Otherwise, the generalisation tests were conducted in the same way as the discrimination learning tests. The foraging decisions of chicks were quantified with an electivity score. In sum, each attack on a palatable/cryptic prey was given a value of 2 (right decision) whereas each attack on a distasteful/aposematic prey was given a 1. The order of attacks was weighted with a value of one applied to the first attack and the value each subsequent attack was reduced by a value 0.1. The electivity score for each session was defined as the weighted sum of attack scores divided by the total number of attacks. Our interpretation of the electivity score is in this way straightforward: if a chick avoids aposematic prey, then the electivity score should be high because its first attacks and most of the attacks will occur on cryptic prey. To test whether the chicks learned to discriminate between the aposematic and the cryptic images, we conducted a repeated measurement analyses on the electivity scores. As predictors we used the within-subjects factor Session (1–7), the between-subjects factor Morph (A or C in Fig. 1), and the interaction term Session x Morph. The identity of the chicks was treated as a random-effects factor nested within Morph. To test whether chicks were able to generalise the training aposematic stimulus to other aposematic stimuli, we followed a similar statistical approach, except that the factor Session had three levels (the
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first, the last, and the generalisation sessions) and the variable Morph pair had six levels, each describing two coloration forms, one used during the sessions 1–7 and the other during the generalisation session. A post hoc Student’s test was used to test for pairwise differences in the performance of chicks between Morph pairs. Generalisation after a complex task To test whether chicks do generalise when trained to discriminate a complex set of aposematic colorations against a cryptic one, we repeated the lab experiment with a few variants. Rather than using a single aposematic image, chicks were exposed to runaways where six of the wells contained the cryptic images but the other four contained dorsal photographs of O. histrionica (Fig. 1 morph A) and the three other aposematic frogs that co-occur at the field study site, namely Phyllobates aurotaenia, Andinobates fulguritus, and Dendrobates truncatus. The printouts mimicked the size of the original frogs. Moreover, we chose to conduct nine training sessions instead of seven due to the complexity of the task. And we used the morph C (Fig. 1) during the generalisation test.
Results Generalisation in the field After excluding the models with evidence of attacks by small arthropods (probably ants), we found direct evidence of attacks by potential predators in 150 (22 %) out of the 679 models that were set up in the field. The marks suggested beaks, claws and rodent-like teeth but, because we cannot ascertain the origin of each attack, we refrained ourselves from analysing the data with regard to the type of predator. In addition, 264 models (39 %) were removed from the ground. Thus, we decided to estimate attack rates in two ways: first, as a binary variable (0 = no evidence of attacks and 1 = removed or with evidence of attack); second, as an ordinal variable according to the strength of evidence of attack from predators with respective values 0,1,2 corresponding to no attack, removed and attacked. The probability of attack was significantly affected by our predictor variables (Generalised linear model, L-R Chi Square = 1496, P \ 0.0001, N = 679 models). All P values remained lower than 0.0001 and the statistical inferences were identical when we used the three-levels ordinal variable (intact, removed, with attack marks) to estimate the probability of attack. Therefore, we discuss below in detail only the first statistical model. Models were much less frequently attacked where O. histrionica was present than where it was absent (Area, L-R Chi Square = 909, P \ 0.0001) (Fig. 2). Brown models were more frequently attacked than any of the conspicuously coloured models (Morph, L-R Chi Square = 356, P \ 0.0001) and such difference was significantly accentuated in areas where O. histrionica was present (Area x Morph interaction, L-R Chi Square = 15, P = 0.0015) (Fig. 2). Considering pairwise contrasts between morphs, each aposematic morph was less frequently attacked than brown models (P \ 0.0001 in all cases). In areas where O. histrionica was present, all aposematic morphs were equally attacked (P [ 0.05 for all comparisons). However, in areas, where O. histrionica was absent, the local aposematic morph was slightly less frequently attacked than models with the same coloration but different pattern (P = 0.045) (Fig. 2).
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Fig. 2 Proportion of attacked models (centre of frog icons) as a function of the model coloration pattern (one aposematic local, two aposematic non-local, and one cryptic forms) and the area where the models were set up (with or without presence of O.histrionica frogs). Red (1) dots denote attacked models whereas green (0) denote non-attacked models. Dots were jittered and dot coloration was faded to improve visualisation of a large amount of data. N = 679 models
Generalisation in the lab In the first experiment, 66 % of variation in chicks’ electivity from session 1–7 was predictable from the input variables we tested (Repeated measurement analysis on Electivity score, R2 = 0.66, F = 9.1, P \ 0.0001, N = 294 sessions and 42 chicks). The performance of chicks improved along time (F ratio = 37.1, P \ 0.0001) and there were no detectable differences attributable to the aposematic model used to train them (Morph, F = 0.1, P = 0.7344; Morph 9 Session, F = 0.5, P = 0.7996) (Fig. 3). To test for generalisation, a second model compared the performance of chicks between the first (1) and the last (7) training sessions, and the generalisation test (G). The whole model was statistically significant (Repeated measurement analysis, R2 = 0.80, F = 5.4, P \ 0.0001, N = 126 sessions and 42 chicks), due to the effect of Session (F = 90.3, P \ 0.0001), but not the Morph combination (F = 1.5, P = 0.2095) or the interaction term (F = 1.1, P \ 0.3408) (Fig. 4). In detail, the electivity of chicks in the seventh session was 0.33 higher than in the first session (Student’s t, P \ 0.0001), but 0.25 lower in the
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Fig. 3 Performance along time of 42 chicks in a discrimination learning task. The electivity score is higher when chicks attack first and more often palatable worms associated with a cryptic frog image rather than distasteful worms associated with an aposematic frog image. The line passes through the average values and the vertical lines represent 95 % confidence intervals (CI). Non-overlapping CI denote statistically significant differences at P \ 0.05. Frog illustrations by C. Ocan˜a
generalisation test compared to the seventh session (P \ 0.0001). Comparing the generalisation test with the first session, the difference is just 0.09 (P = 0.0013), lower in the first session (Fig. 4). Generalisation after a complex task Chicks did not learn to discriminate four simultaneous aposematic images from a cryptic one (Session, F = 0.4, P = 0.5376) and thus the whole statistical model was just marginally significant (Generalised linear model, R2 = 0.21, F = 2.0, P = 0.0608, N = 72 sessions and 8 chicks). As expected from this result, there were no significant differences in the performance of chicks when comparing the first, the last (9) training session, and the generalisation test (R2 = 0.13, F = 0.2, P = 0.9917).
Discussion We tested whether predators familiar with one aposematic colour form do generalise their avoidance behaviour to other aposematic forms. To our knowledge, this is the first study to
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Fig. 4 Electivity of 42 chicks during the first and last (7) session of a discrimination learning experiment (see Fig. 3), and during a generalisation session. In the latter case, the aposematic frog image was replaced by another coloration form of the same (O. histrionica) or a closely related (O. lehmanni) species of poison frog. Frog illustrations by C. Ocan˜a
test this idea on a single prey system with both field and lab paradigms. We obtained contrasting results (Fig. 5). As predicted by the generalisation hypothesis, predators in the field equally avoided three colour forms of O. histrionica including the local one. Although we do not know which predators actually attacked the models, we believe they were familiar with O. histrionica, because the avoidance of aposematic models was significantly accentuated in areas where O. histrionica was present. Against the predictions of the generalisation hypothesis, chicks in the lab failed to avoid novel aposematic forms after being successfully trained to discriminate a single one. They did not generalise even though we tried six combinations of training and generalisation aposematic stimuli. The challenge is then to explain these contrasting results in the light of cognitive generalisation and alternative explanations. Predators (or most of them) may be neophobic. In the field, they could be reluctant to attack the local form because they learned it as distasteful and reluctant to attack the non-
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Fig. 5 Summary of the experimental results on the ability of field potential predators and lab chicks to generalise an aposematic stimulus. Continuous lines indicate positive and X-crossed lines lack of evidence of generalisation for a given pair of aposematic stimuli. Frog illustrations by C. Ocan˜a
local forms because they are new. If so, predators in the areas without O. histrionica should have rejected the non-local forms as often as predators in the area with O. histrionica. This prediction is clearly not met, as attack rates on non-local forms were much lower in frog vs. no-frog areas. In the lab experiment, chicks should have avoided the novel forms during the generalisation test (reviewed by Mappes et al. 2005); our data show that they attacked them as much as the cryptic stimuli that were much more familiar to them. Thus, neither field nor lab evidence supports neophobia as a plausible explanation for the patterns we observed. Alternatively, predators in the field may have successfully avoided the non-local forms because they are familiar with other local aposematic animals (e.g. insects) that look alike. Non-local forms would not be new to them, as the generalisation stimuli were to the lab
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chicks. We cannot discard unambiguously this explanation because we do not know the whole fauna of the megadiverse rain forest at the study site. Nonetheless, we have not seen any animal with similar coloration patterns (morphs D and E in Fig. 1) during more than 5 months of accumulated field work in the area, including about 6 years and different seasons. As a slight variation of the explanation above, field predators might perform better than lab chicks, because the former simultaneously learnt to avoid several aposematic animals. The complex task would prepare them better to generalise and avoid the novel models they encountered. We are aware of at least three other species of aposematic poison frogs that co-occur with O. histrionica at the study site (see Methods). To formally test this explanation, we designed the second lab experiment, where chicks were exposed to photographs of the four aposematic frog species and trained to discriminate them against the cryptic frog photograph. Chicks failed to learn the complex discrimination task in nine sessions. Either the task was too complex for the chicks or they needed additional sessions to learn it. Although our data do not allow us to reject the hypothesis that complex discrimination tasks allow better generalisation of aposematic stimuli, they support in any case that field predators did generalise. Last but not least, the contrasting results between the field and lab experiments may simply suggest that chicks are not appropriate proxies for the behaviour of predators in the field (e.g. Greenwood et al. 1981). The very idea sounds surprising and perhaps confusing given the considerable research based on the perceptual, learning, and behavioural abilities of chicks (e.g. Osorio et al. 1999; Gamberale-Stille and Tullberg 1999; Skelhorn and Rowe 2006). Most of those studies, however, used chicks as proxies for other bird predators in temperate field areas. Others used a different bird species (Parus major) and demonstrated their incapability to generalise aposematic stimuli (Sva´dova´ et al. 2009). We know too little on the natural predators of poison frogs as to make inferences about the validity of the chick predators paradigm. We know that poison frogs occur in the species-rich tropical rain forests where many species, reportedly birds, spiders, crabs, snakes and lizards, are their potential predators. We know that motmot birds (Master 1998) were reported to feed on Dendrobates auratus (Master 1998) and that most attacks on plasticine models mimicking O. pumilio were assignable to birds (Saporito et al. 2007); we have witnessed a snake and a crab eating O. histrionica, and we got beak-, teeth-, and claw-like marks on our experimental models. Perhaps, the whole guild of potential predators of poison frogs is too complex to be cognitively mimicked by chicks. If so, we should adopt new, more realistic paradigms to further understand the ecological consequences of polymorphism. Our field results strikingly contrast with similar field studies on other species of poison frogs. In two of them (Dendrobates tinctorius and Ranitomeya imitator), predators attacked novel aposematic forms more often than local forms (Noonan and Comeault 2009; Chouteau and Angers 2011, respectively); in another one (O. pumilio), the local aposematic form was much more often attacked than the novel forms (Hegna et al. 2012). In our study (O. histrionica), local and novel forms were equally attacked. The apparent contradiction strongly suggests geographic variation in the composition or behavioural reaction of potential predators. It also demands further field studies on the effect of interindividual, microgeographic and seasonal variation in toxicity, on predator reaction. Summing up, our data show that novel forms of O. histrionica would be protected (i.e. less attacked) in the field, probably because the guild of predators do generalise relevant cues from aposematic signals. Neophoby seems incompatible with our data and the question of whether complex discrimination tasks allow better generalisation remains open. If our patterns hold for other populations of Payasita frogs, then the ecological
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advantages of bearing novel colorations (e.g. in sexual selection) would more than counterbalance the relatively minor costs in terms of predation risk. We need more information on the identity, relative importance and cognitive abilities of various predators in shaping the evolutionary fate of polymorphic poison frogs. Acknowledgments The authors are very grateful to G. de Luna and C. Sarmiento for support during the initial phases of the experiment. C. Ocan˜a drew all the colour patterns of the study species complex. Financial support was provided by a grant (1204-452-21906) to A.A. by COLCIENCIAS and to C.E. by the Faculty of Sciences at the Universidad de los Andes. Research permits were provided by the Colombian Ministry of Environment (permits 004 for research and collection).
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