Nineteen Years of Consistently Positive and Strong Female Mate Preferences despite Individual Variation.

Sexual selection driven by mate choice has generated some of the most astounding diversity in nature, suggesting that population-level preferences should be strong and consistent over many generations. On the other hand, mating preferences are among the least repeatable components of an individual animal's phenotype, suggesting that consistency should be low across an animal's lifetime. Despite decades of intensive study of sexual selection, there is almost no information about the strength and consistency of preferences across many years. In this study, we present the results of more than 5,000 mate choice tests with a species of wild frog conducted over 19 consecutive years. Results show that preferences are positive and strong and vary little across years. This consistency occurs despite the fact that there are substantial differences among females in their strength of preference. We also suggest that mate preferences in populations that are primarily the result of sensory exploitation might be more stable over time than preferences that are primarily involved in assessing male quality.

Female túngara frogs do not experience the continuity illusion.

In humans and some nonhuman vertebrates, a sound containing brief silent gaps can be rendered perceptually continuous by inserting noise into the gaps. This so-called "continuity illusion" arises from a phenomenon known as "auditory induction" and results in the perception of complete auditory objects despite fragmentary or incomplete acoustic information. Previous studies of auditory induction in gray treefrogs (Hyla versicolor and H. chrysoscelis) have demonstrated an absence of this phenomenon. These treefrog species produce pulsatile (noncontinuous) vocalizations, whereas studies of auditory induction in other taxa, including humans, often present continuous sounds (e.g., frequency-modulated sweeps). This study investigated the continuity illusion in a frog (Physalaemus pustulosus) with an advertisement vocalization that is naturally continuous and thus similar to the tonal sweeps used in human psychophysical studies of auditory induction. In a series of playback experiments, female subjects were presented with sets of stimuli that included complete calls, calls with silent gaps, and calls with silent gaps filled with noise. The results failed to provide evidence of auditory induction. Current evidence, therefore, suggests that mammals and birds experience auditory induction, but frogs may not. This emerging pattern of taxonomic differences is considered in light of potential methodological, neurophysiological, and functional explanations.

Harmonic calls and indifferent females: no preference for human consonance in an anuran.

The human music faculty might have evolved from rudimentary components that occur in non-human animals. The evolutionary history of these rudimentary perceptual features is not well understood and rarely extends beyond a consideration of vertebrates that possess a cochlea. One such antecedent is a preferential response to what humans perceive as consonant harmonic sounds, which are common in many animal vocal repertoires. We tested the phonotactic response of female túngara frogs (Physalaemus pustulosus) to variations in the frequency ratios of their harmonically structured mating call to determine whether frequency ratio influences attraction to acoustic stimuli in this vertebrate that lacks a cochlea. We found that the ratio of frequencies present in acoustic stimuli did not influence female response. Instead, the amount of inner ear stimulation predicted female preference behaviour. We conclude that the harmonic relationships that characterize the vocalizations of these frogs did not evolve in response to a preference for frequency intervals with low-integer ratios. Instead, the presence of harmonics in their mating call, and perhaps in the vocalizations of many other animals, is more likely due to the biomechanics of sound production rather than any preference for 'more musical' sounds.

Treatment with arginine vasotocin alters mating calls and decreases call attractiveness in male túngara frogs.

The peptide hormone arginine vasotocin (AVT) and its mammalian homolog arginine vasopressin modulate a variety of social behaviors in vertebrates. In anurans, AVT influences the production of advertisement calls, the acoustic signals that males use to attract females and repel rival males. In this study, we investigate the effects of AVT on call characteristics in the túngara frog (Physalaemus pustulosus). Túngara frogs produce a "whine" that is important for species recognition; they may also produce a second, attractive call component, the "chuck". We used a field playback experiment to determine changes in male calling behavior following treatment with AVT. A previous study showed that AVT alters call rate and the production of chucks; in the current analysis, we focus on changes in the whine. Males produce shorter whines with higher initial frequencies following treatment with AVT. Call changes do not vary with a social stimulus. We also used female phonotaxis experiments to investigate the effects of call changes on female mate choice. Females disfavor the calls produced by males treated with exogenous AVT. We suggest that AVT influences motivation to call and the motor control of call production, but that over-stimulation of the vocal system limited the production of attractive calls in this experimental context.

Task differences confound sex differences in receiver permissiveness in túngara frogs.

In many mating systems, both sexes respond to the same sexual signal. In frogs, males typically call in response to advertisement calls, while females approach male calls in choosing a mate. The costs of signal detection errors are expected to differ between the sexes. Missed opportunities are costly for males because ignoring a signal results in failing to compete with rivals for mates, while their cost for misidentification is lower (time and energy displaying to the incorrect target). By contrast, for females, the cost of misidentification is high (mating with incorrect species or low-quality partner), while their cost for missed opportunity is lower because the operational sex ratio puts females at a premium. Consequently, females should be more selective in their response to signal variation than males. We report that presumed sexual differences in selectivity in túngara frogs (Physalaemus pustulosus) are task-specific rather than sex-specific. As predicted, male túngara frogs are less selective in their vocal responses than are females in their phonotactic responses. Males exhibiting phonotaxis to the same calls, however, are as selective as females, and are significantly more selective than when they respond vocally to the same calls. Our study shows that apparent differences between the sexes emerge from differences in the behaviours themselves and are not intrinsic to each sex. Analogous behavioural differences might confound sex differences in other systems; thus, we suggest consideration of the behavioural plasticity of sex as well as its stereotypy.

Visual sensitivity to a conspicuous male cue varies by reproductive state in Physalaemus pustulosus females.

The vocal sac is a visually conspicuous attribute of most male frogs, but its role in visual communication has only been demonstrated recently in diurnally displaying frogs. Here we characterized the spectral properties of the inflated vocal sac of male túngara frogs (Physalaemus pustulosus), a nocturnal species, and túngara visual sensitivity to this cue across reproductive state and sex. We measured the spectral and total reflectance of different male body regions, including inflated and non-inflated vocal sacs, along with samples of the visual background against which males are perceived. Inflated vocal sacs were the most reflective of all body parts, being one log unit more reflective than background materials. We utilized an optomotor drum with black stripes and stripes that mimicked the spectral reflectance of the inflated vocal sacs with various nocturnal light intensities to measure the visual sensitivity thresholds of males, non-reproductive females and reproductive females. All three groups exhibited visual sensitivities corresponding to intensities below moonless conditions in open habitats or at the edge of secondary tropical forests. Reproductive females exhibited the greatest visual sensitivity of all groups, and were significantly more sensitive than non-reproductive females. Though the mechanism for this physiological difference between reproductive and non-reproductive females is unknown, it is consistent with previously observed patterns of light-dependent phonotaxic behavior in túngaras. We suggest that the visual ecology of the vocal sac, especially in nocturnal frogs, offers a rich source for investigations of visual ecology and physiological regulation of vision.

Cues for eavesdroppers: do frog calls indicate prey density and quality?

Predators and parasites that eavesdrop on the mating signals of their prey often preferentially select individuals within a prey/host species that produce specific cues. Mechanisms driving such signal preferences are poorly understood. In the tungara frog Physalaemus pustulosus, conspecific females, frog-eating bats, and blood-sucking flies all prefer complex to simple mating calls. In this study we assess the natural signal variation in choruses in the wild and test two hypotheses for why eavesdroppers prefer complex calls: (1) prey quality: complex calls indicate better quality of prey/host, and (2) prey density: complex calls indicate higher prey/host density. Call complexity is not correlated with frog length, mass, or body condition, but it does signal higher abundance of prey/host. Thus, increased effectiveness of attack may have played a role favoring the preference for complex calls in eavesdropping heterospecifics.

Geographic variation of genetic and behavioral traits in northern and southern tüngara frogs.

We use a combination of microsatellite marker analysis and mate-choice behavior experiments to assess patterns of reproductive isolation of the túngara frog Physalaemus pustulosus along a 550-km transect of 25 populations in Costa Rica and Panama. Earlier studies using allozymes and mitochondrial DNA defined two genetic groups of túngara frogs, one ranging from Mexico to northern Costa Rica (northern group), the second ranging from Panama to northern South America (southern group). Our more fine-scale survey also shows that the northern and southern túngara frogs are genetically different and geographically separated by a gap in the distribution in central Pacific Costa Rica. Genetic differences among populations are highly correlated with geographic distances. Temporal call parameters differed among populations as well as between genetic groups. Differences in calls were explained better by geographic distance than by genetic distance. Phonotaxis experiments showed that females preferred calls of males from their own populations over calls of males from other populations in about two-thirds to three-fourths of the contrasts tested. In mating experiments, females and males from the same group and females from the north with males from the south produced nests and tadpoles. In contrast, females from the south did not produce nests or tadpoles with males from the north. Thus, northern and southern túngara frogs have diverged both genetically and bioacoustically. There is evidence for some prezygotic isolation due to differences in mate recognition and fertilization success, but such isolation is hardly complete. Our results support the general observation that significant differences in sexual signals are often not correlated with strong genetic differentiation.

The vocal sac increases call rate in the Tungara frog Physalaemus pustulosus.

In most anurans, the production of advertisement calls is accompanied by the inflation of a vocal sac. Current functions of the vocal sac, however, are not fully understood, although several hypotheses have been proposed. One hypothesis suggests that the vocal sac decreases the intercall interval (i.e., increases call rate) by reinflating the lungs more rapidly than is possible with the buccal pump. We investigate this hypothesis by analyzing audio and video recordings of calling tungara frogs. We compare the first two call bouts emitted by an originally uninflated male. The first call bout requires lung inflation via buccal pumping, but in the second, the male is already inflated because of capture of air and reinflation of the lungs by the vocal sac. Lung inflation to typical field levels requires 26-51 buccal pumps, which takes at least 4.4 s. This estimate is more than 2.5 times the typical intercall interval with lung reinflation via a vocal sac (ca. 1.7 s). Evidence from phonotaxis tests demonstrates that these differences in intercall intervals are salient to females and that female Physalaemus pustulosus prefer the shorter intercall interval/higher call rate. Acoustic analyses demonstrate that the first call of bout 1, which requires buccal pumping, is usually shorter, of lower amplitude, and spans a smaller frequency range than the first call of bout 2, which does not require buccal pumping. Because females prefer longer, more intense calls, these results suggest that the vocal sac not only increases call rate but also allows males to produce more calls of increased attractiveness to females.

A cognitive framework for mate choice and species recognition.

Mating decisions contribute to both the fitness of individuals and the emergence of evolutionary diversity, yet little is known about their cognitive architecture. We propose a simple model that describes how preferences are translated into decisions and how seemingly disparate patterns of preference can emerge from a single perceptual process. The model proposes that females use error-prone estimates of attractiveness to select mates based on a simple decision rule: choose the most attractive available male that exceeds some minimal criterion. We test the model in the tungara frog, a well-characterized species with an apparent dissociation between mechanisms of mate choice and species recognition. As suggested by our model results, we find that a mate attraction feature alters assessments of species status. Next, we compare female preferences in one-choice and two-choice tests, contexts thought to emphasize species recognition and mate choice, respectively. To do so, we use the model to generate maximum-likelihood estimators of preference strengths from empirical data. We find that a single representation of preferences is sufficient to explain response probabilities in both contexts across a wide range of stimuli. In this species, mate choice and species recognition are accurately and simply summarized by our model. While the findings resolve long-standing anomalies, they also illustrate how models of choice can bridge theoretical and empirical treatments of animal decisions. The data demonstrate a remarkable congruity of perceptual processes across contexts, tasks, and taxa.

Biogeography of the túngara frog, Physalaemus pustulosus: a molecular perspective.

Physalaemus pustulosus, a small leptodactylid frog with South American affinities, ranges across northern South America through Middle America to southern Mexico. To investigate its geographic variation and evolutionary origins, we analysed the presumptive gene products of 14 allozyme loci and sequenced a portion of the mitochondrial COI gene from individuals sampled throughout the distribution. Generally, allozyme dissimilarities and sequence divergences are correlated with each other and with geographic proximity. The greatest discontinuity in genetic variation was found between populations in Middle America vs. South America + Panama. Based on two Bayesian MCMC (Markov chain Monte Carlo) divergence time estimates involving two independent temporal constraints, the timing of the separation of northern and southern túngara frog lineages is significantly older than the time since completion of the current Panama land bridge. P. pustulosus first invaded Middle America from South America about 6-10 million years ago giving rise to the northern lineage. The southern lineage then invaded Panama independently after land bridge completion. Despite millions of years of independent evolution, the multilocus allozyme data revealed that western Panama populations represent a contact zone containing individuals with alleles from both groups present.

The effects of time, space and spectrum on auditory grouping in túngara frogs.

Male túngara frogs (Physalaemus pustulosus) produce complex calls consisting of two components, a approximately 350 ms FM sweep called the "whine" followed by up to seven approximately 40 ms harmonic bursts called "chucks". In order to choose and locate a calling male, females attending to choruses must group call components into auditory streams to correctly assign calls to their sources. Previously we showed that spatial cues play a limited role in grouping: calls with normal spectra and temporal structure are grouped over wide angular separations (< or =135 degrees ). In this study we again use phonotaxis to first test whether an alternative cue, the sequence of call components, plays a role in auditory grouping and second, whether grouping is mediated by peripheral or central mechanisms. We found that while grouping is not limited to the natural call sequence, it does vary with the relative onset times of the two calls. To test whether overlapping stimulation in the periphery is required for grouping, the whine and chuck were filtered to restrict their spectra to the sensitivity ranges of the amphibian and basilar papillae, respectively. For these dichotic-like stimuli, grouping still occurred (albeit only to 45 degrees separation), suggesting that stream formation is mediated by central mechanisms.

Functional mapping of the auditory midbrain during mate call reception.

We examined patterns of neural activity as assayed by changes in gene expression to localize representation of acoustic mating signals in the auditory midbrain of frogs. We exposed wild-caught male Physalaemus pustulosus to conspecific mating calls that vary in their behavioral salience, nonsalient mating calls, or no sound. We measured expression of the immediate early gene egr-1 (also called ZENK, zif268, NGFI-A, and krox-24) throughout the torus semicircularis, the auditory midbrain homolog of the inferior colliculus. Differential egr-1 induction in response to the acoustic stimuli occurred in the laminar, midline, and principal nuclei of the torus semicircularis, whereas the ventral region did not show significant effects of stimulus. The laminar nucleus differentially responded to conspecific mating calls compared with nonsalient mating calls, whereas the midline and principal nuclei responded preferentially to one of two conspecific calls. These responses were not explained by simple acoustic properties of the stimuli, and they demonstrate a functional heterogeneity of auditory processing of complex biological signals within the frog midbrain. Moreover, using analyses that assess the ability of the torus semicircularis as a whole to discriminate among acoustic stimuli, we found that activity patterns in the four regions together provide more information about biologically relevant acoustic stimuli than activity in any single region.

Sexual selection in female perceptual space: how female túngara frogs perceive and respond to complex population variation in acoustic mating signals.

Female preferences for male mating signals are often evaluated on single parameters in isolation or small suites of characters. Most signals, however, are composites of many individual parameters. In this study we quantified multivariate traits in the advertisement call of the túngara frog, Physalaemus pustulosus. We represented the calls in multidimensional scaling space and chose nine test calls to represent the range of population variation. We then tested females for phonotactic preference between calls in each pair of the nine test calls. We used statistics developed for paired comparisons in such "round robin" competitions to evaluate the null hypothesis of equal attractiveness, and to examine the degree to which females responded to calls as being different from or similar to one another in attractiveness. We then examined the attractiveness of each test call relative to all other test calls as a function of their location in multivariate acoustic space (the acoustic landscape) to visualize sexual selection on calls. Finally, we used methods from cognitive psychology to illustrate the females' perception of call attractiveness in multivariate space, and compared this perceptual landscape to the acoustic landscape of quantitative call variation. We show that correlations between individual call characters are not strong and thus there are few biomechanical constraints on their independent evolution. Most call variables differed among males, and there was high repeatability of call characters within males. Females often discriminated between pairs of calls from the population, and there were significant differences among calls in their attractiveness. Female preferences for calls were not stabilizing. The region of the acoustic landscape that was most attractive to females included the mean call but was not centered around it. The females' perceptual or preference landscape did not correlate with the call's acoustic landscape, and female perception of calls decreased rather than enhanced call differences.

Fine-scale genetic pattern and evidence for sex-biased dispersal in the túngara frog, Physalaemus pustulosus.

Túngara frogs (Physalaemus pustulosus) are a model system for sexual selection and communication. Population dynamics and gene flow are of major interest in this species because they influence speciation processes and microevolution, and could consequently provide a deeper understanding of the evolutionary processes involved in mate recognition. Although earlier studies have documented genetic variation across the species' range, attempts to investigate dispersal on a local level have been limited to mark-recapture studies. These behavioural studies indicated high mobility at a scale of several hundred metres. In this study we used seven highly polymorphic microsatellite loci to investigate fine-scaled genetic variation in the túngara frog. We analysed the influence of geographical distance on observed genetic patterns, examined the influence of a river on gene flow, and tested for sex-biased dispersal. Data for 668 individuals from 17 populations ranging in distance from 0.26 to 11.8 km revealed significant levels of genetic differentiation among populations. Genetic differentiation was significantly correlated with geographic distance. A river acted as an efficient barrier to gene flow. Several tests of sex-biased dispersal were conducted. Most of them showed no difference between the sexes, but variance of Assignment Indices exhibited a statistically significant male bias in dispersal.

Generalization in response to mate recognition signals.

Females usually exhibit strong and unequivocal recognition of conspecific mating signals and reject those of other sympatric heterospecifics. However, most species are allopatric with one another, and the degree to which females recognize mating signals of allopatric species is more varied. Such mating signals are often rejected but are sometimes falsely recognized as conspecific. We studied the dynamics of mate recognition in female túngara frogs (Physalaemus pustulosus) in response to a series of calls that were intermediate between the conspecific and each of five allopatric-heterospecific calls: two that elicited recognition from females in previous studies and three that did not. This study shows that females perceive variation in allopatric mating signals in acontinuous manner with no evidence of perceptual category formation. The strength of recognition is predicted by how different the target stimulus is from the conspecific call within a series of calls. But the differences in recognition responses among call series are not predicted by the similarity of the call series to the conspecific call. The latter result suggests that the strength of recognition of allopatric signals might be influenced by processes of stimulus generalization and past evolutionary history.

The effects of spatially separated call components on phonotaxis in túngara frogs: evidence for auditory grouping.

Numerous animals across disparate taxa must identify and locate complex acoustic signals imbedded in multiple overlapping signals and ambient noise. A requirement of this task is the ability to group sounds into auditory streams in which sounds are perceived as emanating from the same source. Although numerous studies over the past 50 years have examined aspects of auditory grouping in humans, surprisingly few assays have demonstrated auditory stream formation or the assignment of multicomponent signals to a single source in non-human animals. In our study, we present evidence for auditory grouping in female túngara frogs. In contrast to humans, in which auditory grouping may be facilitated by the cues produced when sounds arrive from the same location, we show that spatial cues play a limited role in grouping, as females group discrete components of the species' complex call over wide angular separations. Furthermore, we show that once grouped the separate call components are weighted differently in recognizing and locating the call, so called 'what' and 'where' decisions, respectively.

An ecosystem report on the Panama Canal: monitoring the status of the forest communities and the watershed.

In 1996, the Smithsonian Tropical Research Institute and the Republic of Panama's Environmental Authority, with support from the United States Agency for International Development, undertook a comprehensive program to monitor the ecosystem of the Panama Canal watershed. The goals were to establish baseline indicators for the integrity of forest communities and rivers. Based on satellite image classification and ground surveys, the 2790 km2 watershed had 1570 km2 of forest in 1997, 1080 km2 of which was in national parks and nature monuments. Most of the 490 km2 of forest not currently in protected areas lies along the west bank of the Canal, and its management status after the year 2000 turnover of the Canal from the U.S. to Panama remains uncertain. In forest plots designed to monitor forest diversity and change, a total of 963 woody plant species were identified and mapped. We estimate there are a total of 850-1000 woody species in forests of the Canal corridor. Forests of the wetter upper reaches of the watershed are distinct in species composition from the Canal corridor, and have considerably higher diversity and many unknown species. These remote areas are extensively forested, poorly explored, and harbor an estimated 1400-2200 woody species. Vertebrate monitoring programs were also initiated, focusing on species threatened by hunting and forest fragmentation. Large mammals are heavily hunted in most forests of Canal corridor, and there was clear evidence that mammal density is greatly reduced in hunted areas and that this affects seed predation and dispersal. The human population of the watershed was 113 000 in 1990, and grew by nearly 4% per year from 1980 to 1990. Much of this growth was in a small region of the watershed on the outskirts of Panama City, but even rural areas, including villages near and within national parks, grew by 2% per year. There is no sewage treatment in the watershed, and many towns have no trash collection, thus streams near large towns are heavily polluted. Analyses of sediment loads in rivers throughout the watershed did not indicate that erosion has been increasing as a result of deforestation, rather, erosion seems to be driven largely by total rainfall and heavy rainfall events that cause landslides. Still, models suggest that large-scale deforestation would increase landslide frequency, and failure to detect increases in erosion could be due to the gradual deforestation rate and the short time period over which data are available. A study of runoff showed deforestation increased the amount of water from rainfall that passed directly into streams. As a result, dry season flow was reduced in a deforested catchment relative to a forested one. Currently, the Panama Canal watershed has extensive forest areas and streams relatively unaffected by humans. But impacts of hunting and pollution near towns are clear, and the burgeoning population will exacerbate these impacts in the next few decades. Changes in policies regarding forest protection and pollution control are necessary.

ALLOZYME AND ADVERTISEMENT CALL VARIATION IN THE TÚNGARA FROG, PHYSALAEMUS PUSTULOSUS.

We analyzed variation in advertisement calls and allozymes in 30 populations along a 5000-km transect throughout most of the range of the túngara frog, Physalaemus pustulosus. All 12 call variables measured show significant differences among populations despite the importance of the advertisement call in species recognition. Some call variables exhibited clinal variation, whereas most others differed between the two major allozyme groups that have invaded Panama at different times, perhaps 4-4.5 million yr apart. Call variables that primarily affect discrimination among conspecifics tended to exhibit greater variation than call variables that are crucial for species recognition. The proximate mechanism of production underlying a call variable, however, is a better predictor of its variation. Contrary to predictions of some sexual selection models, call variation exhibits predictable patterns of geographical variation, although a substantial portion of variation among populations is not explained by geographic position. Although allozymes, calls, and geography usually covary, closer populations can have more similar calls independent of allozyme similarity.

SPECIES RECOGNITION AND SEXUAL SELECTION AS A UNITARY PROBLEM IN ANIMAL COMMUNICATION.

We investigated patterns of mating call preference and mating call recognition by examining phonotaxis of female túngara frogs, Physalaemus pustulosus, in response to conspecific and heterospecific calls. There are four results: females always prefer conspecific calls; most heterospecific calls do not elicit phonotaxis; some heterospecific calls do elicit phonotaxis and thus are effective mate recognition signals; and females prefer conspecific calls to which a component of a heterospecific call has been added to a normal conspecific call. We use these data to illustrate how concepts of species recognition and sexual selection can be understood in a unitary framework by comparing the distribution of signal traits to female preference functions.