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In veterinary education, data from biomedical or natural sciences are mostly presented in the form of static or animated graphics with no or little amount of interactivity. These kinds of presentations are, however, often not sufficient to depict the complexity of the data or the presented topic. Interactive graphics, which allow to dynamically change data and related graphics, have rarely been considered as teaching tool in higher education of biomedical disciplines for veterinary education so far. In order to study the applicability and the usefulness of interactive graphics in biomedical disciplines for lecturers and students in veterinary education, three different courses from biomedical disciplines were exemplarily implemented as interactive graphics and evaluated in a pilot study by a survey amongst lecturers and students of our university. The interactive graphics were built using the Shiny environment, a web-based application framework for the statistic software R. The survey amongst lecturers and students was based on questionnaires covering questions on the handling and usefulness of the digital teaching tools. In total, n = 327 students and n = 5 lecturers participated in the evaluation study which revealed that the interactive graphics are easy to handle for lecturers and students, and that they can increase the motivation for either teaching or learning. In total, 71% of the students affirmed that interactive graphics led to an increased interest for the presented contents and 76% expressed the wish to get taught more topics with interactive graphics. We also provide a workflow that can be used as a guideline to develop interactive graphics.

Phenotypic adaptation in terms of background color matching to the local habitat is an important mechanism for survival in prey species. Thus, intraspecific variation in cryptic coloration is expected among localities with dissimilar habitat features (e.g., soil, vegetation). Yellow‐bellied toads (Bombina variegata) display a dark dorsal coloration that varies between populations, assumed to convey crypsis. In this study, we explored I) geographic variation in dorsal coloration and II) coloration plasticity in B. variegata from three localities differing in substrate coloration. Using avian visual modeling, we found that the brightness contrasts of the cryptic dorsa were significantly lower on the local substrates than substrates of other localities. In experiments, individuals from one population were able to quickly change the dorsal coloration to match a lighter substrate. We conclude that the environment mediates an adaptation in cryptic dorsal coloration. We suggest further studies to test the mechanisms by which the color change occurs and explore the adaptive potential of coloration plasticity on substrates of varying brightness in B. variegata and other species.

Revealing patterns of genetic diversity and barriers for gene flow are key points for successful conservation in endangered species. Methods based on molecular markers are also often used to delineate conservation units such as evolutionary significant units and management units. Here we combine phylo-geographic analyses (based onmtDNA) with population and landscape genetic analyses (based on microsatellites) for the endangered yellow-bellied toadBombina variegataover a wide distribution range in Germany. Our analyses show that two genetic clusters are present in the study area, a northern and a southern/central one, but that these clusters are not deeply divergent. The genetic data suggest high fragmentation among toad occurrences and consequently low genetic diversity. Genetic diversity and genetic connectivity showed a negative relationship with road densities and urban areas surrounding toad occurrences, indicating that these landscape features act as barriers to gene flow. To preserve a maximum of genetic diversity, we recommend considering both genetic clusters as management units, and to increase gene flow among toad occurrences with the aim of restoring and protecting functional meta-populations within each of the clusters. Several isolated populations with especially low genetic diversity and signs of inbreeding need particular short-term conservation attention to avoid extinction. We also recommend to allow natural gene flow between both clusters but not to use individuals from one cluster for translocation or reintroduction into the other. Our results underscore the utility of molecular tools for species conservation, highlight outcomes of habitat fragmentation onto the genetic structure of an endangered amphibian and reveal particularly threatened populations in need for urgent conservation efforts.

The chytrid fungusBatrachochytrium dendrobatidis(Bd) infects numerous amphibian species worldwide and is suggested to drive population declines and extinction events. We report a study ofBdinfection at the northernmost distribution of the European yellow-bellied toadBombina variegata. A total of 577 individuals from ponds in 16 study sites were sampled for DNA andBdthroughout the breeding season. Microsatellite genotyping revealed 3 genetic clusters for the hostB. variegatawith an overall low genetic diversity. One of the clusters displayed a low microsatellite heterozygosity, a high inbreeding coefficient as well as highBdinfection prevalence and intensities. Multi-model estimates identified site, time of sampling, and heterozygosity to be important predictors of an individual’sBdinfection status, and identified a strong effect of site on individualBdinfection intensity. The study site effects are suggestive of localized infection peaks, and the increase of individualBdinfection probabilities towards the end of the sampling period suggests cumulative infection during the breeding season. This study highlights the need for regular monitoring ofBdinfection variables at multiple localities and times to gain insights intoBddynamics. Due to the detected relationship between individualBdinfection status and heterozygosity, conservation measures should focus on the maintenance of high genetic diversity and connectivity within and among amphibian populations.

Amphibian populations world-wide are threatened by declines and extinctions mainly due to habitat loss and fragmentation. Habitat fragmentation threatens the yellow-bellied toad Bombina variegata in the northern and western regions of its distribution where it is strictly protected. We studied the genetic structure and diversity of populations at three geographical scales using microsatellite loci to detect potential threats for population persistence. At the local scale, we sampled four neighbouring localities at 1–2.6 km distance to detect effects of short-term (decades) fragmentation on connectivity. At the regional scale, five additional localities in the mountains of the Westerwald (Rhineland-Palatinate, Germany) were studied at up to 50.1 km distance to analyse genetic diversity and population structure. At the continental scale, we included data from regions in the northern distribution with fragmented populations (Hesse and Lower Saxony, Germany) and more continuous populations in the South (Alsace, France; Geneva, Switzerland; Trentino, Italy) to evaluate variation of genetic diversity. At the local scale, short-term fragmentation caused significant genetic differentiation between breeding assemblages only 1.4 km apart from each other. At the regional scale, we found notable genetic distance among localities. At the continental scale, we identified Alsace, Trentino and Geneva in the South as regions with low genetic structuring and high allelic richness, and the northern remaining regions in Germany as deeply structured with reduced allelic richness. We suggest that reduced genetic diversity and habitat fragmentation in northern regions makes these populations particularly vulnerable to decline. In conclusion, informed conservation management of B. variegata should focus on measures maintaining or improving connectivity among neighbouring populations.

Frogs in the genus Phyllobates are known for the presence of batrachotoxin, a highly toxic alkaloid, in their skin. Nevertheless, Phyllobates frogs from Costa Rica and Panama (P. lugubris and P. vittatus) are considered non-toxic, as they have been reported to harbor low concentrations of this alkaloid. However, the potential toxicity of Central American Phyllobates has not been assessed experimentally. Our goal was to determine the toxicity of the whole skin of P. vittatus, an endemic species from the Southeastern Pacific region of Costa Rica. We performed median lethal dose (LD50) tests in mice to determine general toxicity, and an irritant assay based on the behavioral responses of mice to subcutaneous injection, to determine differences in irritability, as a measure of toxicity, among three study localities. Using UPLC-ESI-QTOF, we obtained chemical profiles of the methanolic extract of frog skins. Due to the absence of mortality at the studied doses, we were unable to estimate LD50. However, we recorded a list of toxicity symptoms in mice that are consistent with cardiotoxic effects, and found that mice presented more symptoms at higher concentrations of skin extracts during the first hour of the LD50 assays, recovering completely at all doses by the end of the assay. On the other hand, we did not detect differences in irritability among studied localities. Additionally, we putatively identified three toxic alkaloids (Batrachotoxinin A, DHQ 251A and Lehm 275A). This study provides the first experimental data on the toxicity and associated symptoms in mice, as well as the chemical profile of the skin of P. vittatus. We suggest that the skin alkaloids of P. vitattus may confer a chemical defense towards predators.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Codivergence of mating traits and mate preferences can lead to behavioral isolation among lineages in early stages of speciation. However, mate preferences limit gene flow only when expressed as mate choice, and numerous factors might be more important than preferences in nature. In the extremely color polytypic strawberry poison frog (Oophaga pumilio), female mate preferences have codiverged with color in most allopatric populations tested. Whether these lab-assayed preferences predict mating (gene flow) in the wild remains unclear. We observed courting pairs in a natural contact zone between red and blue lineages until oviposition or courtship termination. We found color-assortative mating in a disturbed habitat with high population density but not in a secondary forest with lower density. Our results suggest color-assortative O. pumilio mate choice in the wild but also mating patterns that do not match those predicted by lab-assayed preferences.

Habitat occupancy by territorial animals is expected to depend on the distribution of critical resources. Knowledge on female territoriality is scarce, but it has been suggested as a mechanism to defend limited resources for reproduction. A previous study showed female intrasexual aggression to be associated with territorial behaviour in the strawberry poison frog Oophaga pumilio, a diurnal aposematic species with complex maternal care. Here, we investigate the link between spatial distribution of resources important for reproduction and female distribution and behaviour. We observed focal females in their natural habitat in Costa Rica, and recorded the distribution of ecological predictor variables in a grid system. We used the data for calculating home range and territory sizes and for connecting female habitat use to the distribution of potential resources by computing spatial habitat occupancy models. Even though we found females to occupy large home ranges, they were highly aggressive towards other females only inside a small part of their home range, here termed core area. Among the ecological factors, the sustained abundance of ants (main food item of the frogs), the presence of leaf litter and suitable rearing sites for tadpoles predicted female site occupancy patterns. The number of ants per grid was twice as high in the core areas compared to the rest of the female home ranges. Our results suggest that female spacing behaviour is principally driven by the spatial distribution of its main food resource, but that hiding places (leaf litter) and tadpole-rearing sites also play a role. The defence of areas with sustainably high abundance of ants could be relevant, as egg production and maternal care are energetically highly demanding in this prolonged-breeding species. Regarding the link between resource defence and maternal care, the reproductive strategy of female strawberry poison frogs resembles that of the females of small mammals comprising same-sex competition for food and high investment in producing and rearing young.

Experimental work in the early 20th century showed that background albedo experienced by larvae of the fire salamander (Salamandra salamandra) induce a durable morphological modification of the postmetamorphic color pattern, which needed confirmation due to the controversies regarding Paul Kammerer's experiments. Such a carry-over effect would be relevant as the black and yellow pattern of the alkaloid-containing adult fire salamanders has been suggested to serve as an aposematic signal. Hence, we hypothesized that (a) adult coloration is conspicuous to potential predators under light conditions at night, given the nocturnal activity of this species, and (b) a condition affecting the salamander's coloration pattern would also affect its toxicity to maintain a quantitatively honest aposematic signal. To test the first hypothesis, we used spectrometry to model the vision of potential avian and snake predators and confirmed that fire salamander's black-and-yellow pattern is contrasting enough against the forest leaf litter to be considered conspicuous at night. To test the second hypothesis, we first confirmed the background carry-over effect on black and yellow proportions in the dorsal skin of experimentally reared fire salamanders, using a rigorous experimental design. Then, we calculated the conspicuousness and determined the alkaloid profiles of these individuals. We did not find a correlation between conspicuousness and toxicity at the intrapopulation level. Moreover, there was no background carry-over effect on the alkaloid profile. We discuss our results in a physiological, ecological, evolutionary, and historical context.

Sexual signals are important for intraspecific communication and mate selection, but their evolution may be driven by both natural and sexual selection, and stochastic processes. Strawberry poison frogs (Oophaga pumilio) show strong color divergence among populations, but coloration also varies among individuals of the same population. The importance of coloration for female mate choice has been studied intensely, and sexual selection seems to affect color divergence in strawberry poison frogs. However, the effect of coloration on mating success under field conditions has received very little attention. Furthermore, few studies examined how phenotypic variation among individuals of the same color morph affects mate selection under natural conditions. We measured the spectral reflectance of courting and noncourting individuals and their background substrates in three geographically separated populations. In one population (Sarapiquí, Costa Rica), we found that naturally occurring courting pairs of males and females had significantly brighter dorsal coloration than individual males and females not engaged in courtship interactions. Our field observations suggest that, in the wild, females prefer brighter males while the reason for the higher courtship activity of brighter females remains unclear. Overall our results imply that brightness differences among individuals of the same color morph may actually affect reproductive success in some populations of strawberry poison frogs.

Evolutionary divergence in the coloration of toxic prey is expected when geographic variation in predator composition and behavior favours shifts in prey conspicuousness. A fundamental prediction of predator-driven colour divergence is that the local coloration should experience lower predation risk than novel prey phenotypes. The dorsal coloration of the granular poison frog varies gradually from populations of conspicuous bright red frogs to populations of dull green and relatively cryptic frogs. We conducted experiments with clay models in four populations to examine the geographic patterns of taxon-specific predation. Birds avoided the local phenotype while lizards consistently selected for decreased conspicuousness and crab predation did not depend on frog coloration. Importantly, birds and lizards favoured low conspicuousness in populations where relatively cryptic green morphs have evolved. This study provides evidence for the interplay among distinct selective pressures, from multiple-predator taxa, acting on the divergence in protective coloration of prey species.(c) 2014 The Linnean Society of London, Biological Journal of the Linnean Society, 2014, 113, 580-589.

The evolution of aposematism is linked to increased opportunities for conspicuous sexual displays since detection by potential predators is no longer disadvantageous. Therefore, phenotypic divergence in aposematic species leading to relatively cryptic forms is expected to constrain such opportunities, by restoring the trade-off between natural and sexual selection on the boldness of sexual displays. We asked if and how a derived phenotype of the poison-dart frog Oophaga granulifera that appears relatively cryptic to potential predators exhibits conspicuous sexual displays for potential mates. We used visual modeling of frog contrasts against their natural backgrounds to test if for conspecifics green frogs appear less conspicuous than red frogs as they do for birds. We conducted behavioral observations of focal red and green males to determine if green frogs adjust their display behavior to the availability of potential mates. Dorsal brightness is known to influence female preferences in at least one poison frog species. We found that, despite being less visible under some measures, green frogs may appear as bright as red frogs for conspecifics but not birds, when viewed on dark backgrounds. Additionally, green males called more intermittently than red males when advertising to distant females, but they exhibited a dramatic increase in calling activity in proximity of a female and were as active as red males in this context. Together, our results suggest that green frogs retain context-dependent conspicuousness to conspecifics despite the evolution of relative crypsis to potential predators.

Acoustic and visual signals like mating calls or colour patterns, which are involved in courtship displays, can be affected by the health status of an individual. For example, infection with parasites can result in sexual signals that are less intense or conspicuous than in healthy individuals; thus it is generally assumed that sexual signals indicate the quality of the holder. We use strawberry poison frogs from Costa Rica to find out (1) whether there is variation in the parasite load among individuals, (2) whether the parasite load varies over time, (3) whether the parasite load has an effect on acoustic and visual signals, which are supposed to be important for female mate choice and (4) whether the parasite load influences the reproductive behaviour. We found that 60 % of our focal males were infested with an intestinal nematode of the genus Cosmocerca while the parasite load changed considerably over time. We found slight associations between the intensity of acoustic signals (advertisement calls), visual signals (colour and brightness contrasts) and parasite load. Non-infected males produced significantly lower pulse rates than parasitised males. As pulse rate has been shown to decrease with age, we suggest that males without parasites become older than infected males. Moreover, the visual contrasts of the frogs varied across natural backgrounds. An interaction between parasite infection and the background of the calling place affected the ventral colour contrast. This finding suggests that at least some males without parasites defend calling places that render them more conspicuousness to conspecifics than other potential positions would. There were no differences in several elements of reproductive behaviour between parasitised and healthy males. Further studies are necessary to find out whether parasite infection varies among populations, and whether at higher levels it would have a stronger influence on sexual signals, behaviour and lifetime fitness.

Aim To investigate the intra-specific genetic divergence and phenotypic diversification in Oophaga granulifera, analysing phylogeographical patterns, population structure, coloration and traits potentially involved in reproductive isolation. Location Costa Rica. Methods Our phylogeographical analysis used the mitochondrial genes cytochrome b and 16S. We sequenced the two markers in 28 individuals belonging to 10 populations and 16 individuals from nine populations, respectively. The population structure was analysed based on seven polymorphic microsatellite loci in 7–15 individuals from 11 populations using maximum likelihood and Bayesian analyses. The colour phenotypes of 189 frogs and the reflectance spectra of 15 frogs from five populations were measured. In nine populations, we recorded the advertisement calls of 59 males and measured 10 morphological variables from 147 frogs. Results Our phylogeographical reconstruction supported the divergence into southern and northern lineages in O. granulifera. In a colour transition area between the main red and green morphs, we describe novel gradual intermediate phenotypes occurring in polymorphic populations. The colour distribution was asymmetrical compared to the genetic variation: the southern lineage included only monomorphic red populations while the northern lineage contained red, green and polymorphic populations. The two lineages differed significantly in advertisement calls and morphology. Microsatellites confirmed the clear inter-lineage divergence. In the northern lineage, the red morph differed from the intermediate and green morphs in the pulse rate of the advertisement call and in body size. In the colour transition area, there was no simple genetic or colour cline. Main conclusions Oophaga granulifera has evolved into two genetic lineages. The reduced gene flow and diversification in advertisement calls and morphology point to reproductive isolation between lineages. Within the northern lineage, the presence of gradual intermediate morphs and the population structure support the presence of gene flow across the colour transition area while the red versus intermediate–green divergence in both calls and morphology reveals the potential for assortative mating. Our data all congruently show that O. granulifera presents deep intra-specific divergence. The lack of genetic diversification among colour morphs and the genetic connectivity among the populations defining the colour transition area suggest that the colour diversification in the species is a recent phenomenon.

In many species male reproductive success is limited by access to females. Territoriality is one behavioural strategy which helps to acquire females. In the present study, we investigated the correlation between territory size and (1) female availability and (2) rate of intrusion by conspecific males in strawberry poison frogs, Oophaga pumilio. Males defended smaller territories in areas with a high female density and high rate of intrusion by conspecific males. Only males with high body condition values were able to establish territories in areas of high female density probably due to better fighting abilities. Moreover, dominant calling frequency was lower during agonistic interactions. Because only males with high body condition values were able to produce very low dominant frequencies, the acoustic properties appear to be an honest signal containing information about fighting abilities. Thus, the negative correlation between dominant frequency and mating success of males found in previous studies seems to be in part the result of intrasexual competition between males.

Females of the Strawberry Poison Frog (Oophaga pumilio) are known to be aggressive toward other females. However, the function of this behavior in females has not been identified. We hypothesized that females are territorial, occupying and defending specific areas in defense of food resources. To test this hypothesis, we calculated the position and size of core areas and home ranges for each female in the study area and the frequency of aggressive and feeding behavior exhibited by females in these areas. We provide evidence, for the first time, that females are territorial; they defend their core area against other intruding females. To determine the potential resource defended by females, we examined aggressive behavior associated with defense of mates, oviposition, and tadpole-rearing sites. We found no evidence that females defended areas with males, or that oviposition and tadpole-rearing sites were limited and defended by females. Instead, our observations suggest that female territoriality ...

The coexistence of both aposematic and cryptic morphs as different anti-predator strategies within a species seems to be an unusual phenomenon in nature. The strawberry poison frog, Oophaga pumilio, shows an astonishing colour diversity among populations in western Panama. In this study we selected a red and a green colour morph from two Panamanian islands (Isla Solarte and Isla Colon) for behavioural observations and measurements of conspicuousness. We found that red frogs were more visible to both conspecific frogs and potential predators than green frogs. Interestingly the difference in conspicuousness was most pronounced at the substrate that males used as principal calling places. Red males were more active and spent more time foraging than green males, which spent more time hidden. The association between conspicuousness of colouration and behaviour results in a more aposematic and a more cryptic anti-predator strategy. This is the first study which links differences in conspicuousness between animals on their natural backgrounds to differences in foraging as well as anti-predator behaviour and discusses the results in light of previous findings of toxicity analyses and potential costs and benefits of aposematism. To this end, our study adds a novel perspective for explaining extreme colour diversity between populations within an initially aposematic species.

Aim To analyse the phylogeographic structure of the strawberry poison frog, Oophaga pumilio (Dendrobatidae), across a large part of its range using a combination of mitochondrial and nuclear markers.

The tadpole of Ranitomeya variabilis is described based on preserved specimens from captive breed specimens. The tadpole of R. variabilis is similar to other dendrobatid tadpoles regarding tooth row formula, body shape, location of the spiracle, vent tube, and oral disc. It differs from other dendrobatid species, e.g., Allobates, Oophaga, and Phyllobates, in various aspects such as the absence of submarginal papillae, the shape of the jaw sheath, interorbital distance, and total length. Ranitomeya variabilis tadpoles undergo ontogenetic changes in the number of marginal papillae, the presence of jaw sheath serrations, jaw sheath keratinization, and tail color. The facultative cannibalistic feeding behavior of R. variabilis is discussed with respect to the species feeding ecology.

The present study investigates foraging and territorial behaviour of the strawberry poison frog (Oophaga pumilio) in dependence of the presence of formicine and myrmicine ants, which constitute the main food source of the frogs. Species of the formicine ant genera Brachymyrmex and Paratrechina contain highly toxic alkaloids (pumiliotoxins), which the frogs incorporate and accumulate in their skin what may serve for predator deterrence. Twelve male frogs of two populations (primary and secondary forest) in Hitoy Cerere, Costa Rica, were observed each for a full day. Calling time, feeding attempts and time spent inside and outside the core area of their territories were recorded. Furthermore, twelve males of both populations were observed during the main foraging time to determine, whether the frogs search for prey in specific patches of their territories. The ants inside the core areas of twenty four frog territories were collected and classified to genus. Ants of the genera Brachymyrmex and Paratrechina were classified to species or morphospecies, respectively. The presence of formicine and myrmicine ants in territorial areas was compared to non-territorial sites. We found that formicine ants (Brachymyrmex and Paratrechina) were more present inside the territorial core areas than outside. The higher presence of these ants in the core areas was associated with longer foraging times. We verified that toxic alkaloids of the pumiliotoxin group are present in the dendrobatid frogs of Hitoy Cerere. The results of this study suggest that toxic diet may be linked to territoriality in this frog species.

In this paper, we compare the advertisement calls of 207 neotropical strawberry poison frogs (Dendrobates pumilio) collected in 21 localities along a transect from northern Costa Rica to western Panama. Populations varied most in call duration and call rate, while pulse rate and duty cycle were less variable. Multivariate analyses showed that call variation followed a cline with higher call rates, shorter calls, lower duty cycles and higher pulse rates in the southeast. Body size decreased towards the southeast and explained most variation in dominant frequency, as well as some residual variation in call rate. We conclude that a combination of geography and morphology is largely responsible for call variation within this species. Two inferred bio-acoustic groups were roughly in accordance with two genetic groups, geographically separated in central Costa Rica. However, genetic distances among populations did not co-vary with call dissimilarity after correction for geographic distances. Thus, differences in calls between genetic groups are probably mainly a result of clinal variation. These findings agree with the general observation that bio-acoustic variation is often not (highly) associated with genetic divergence. Moreover, colour polymorphism observed among Panamanian populations was not reflected in a higher variability in call parameters relative to the monomorphic Costa Rican populations.

We use a combination of microsatellite marker analysis and mate-choice behavior experiments to assess patterns of reproductive isolation of the tungara 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 tungara 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 tungara 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 tungara 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.

One or both sexes exhibit territorial behavior in all dendrobatids that have been studied. Two types of territoriality are identified. In some Colosthethus species only females or both sexes defend small nonreproductive territories that offer access to water, food, and shelter. In other species, only males defend large reproductive territories against conspecific calling males, suggesting that females are the limited resource. However, in no case has the defended resource been unambiguously identified. I hypothesize that, in at least some species, males compete for calling sites in areas with high female density and with favorable sound transmission features. I found little support for the hypothesis that dendrobatids exhibit resource defense polygyny. More research is necessary to understand the function of territoriality and to identify limiting resources in poison frogs.

Both modelling and field data from three breeding seasons show that an environmental factor, clutch loss (CL), affects the operational sex ratio (OSR) and therefore male mating frequency in strawberry poison frogs. Clutch loss affects the length of reproductive cycles of both sexes: with increasing clutch losses, males spend proportionately more time than females in parental investment activities. Because of this, males spend relatively less time in the mating pool, i.e. exhibit proportionately more “time-out” than females in comparison to a situation with low or no clutch loss. Hence, clutch loss leads to a less male-biased OSR, coupled with a decrease in the opportunity for sexual selection. Furthermore, this study resolves an apparent paradox, the negative correlation between mating frequency and reproductive success (=number of produced tadpoles) of individual males in one breeding season. Clutch loss decouples the correlation between mating frequency and reproductive success because females re-enter the mating pool when they lose their offspring. However, clutch loss diminishes the reproductive output. Similar consequences of clutch loss on the OSR may be true for many species where both sexes reproduce frequently in one breeding season.

Calling behaviour and the characteristics of the males’ call are important for mate attraction, female mate choice and male mating success in anurans. In this study I investigated the variation in calling activity and the variation in one spectral (dominant frequency) and four temporal properties (call rate, call duration, duty cycle and pulse rate) of the advertisement call of strawberry poison frogs, Dendrobates pumilio, in two Costa Rican populations during two study periods. Regarding all call properties, no differences were found between populations or years, but between-male variation was significantly higher than within-recording and within-male variation. Dominant frequency was less variable within and among males than temporal call properties and I classified the former as static and the latter as intermediate properties of the call. No call property seemed to be strongly affected by body length or weight. Moreover, I used behavioural observations to relate male calling behaviour to mating success. Calling activity was more variable between males and accounted for more variability in mating success in multiple regression models than any call property. Some call properties (call rate, dominant frequency and pulse rate) were also related to male mating success in one or two study periods. These call properties are probably important for females during courtship to assess their prospective mate and/or for competitive male–male interactions. Furthermore, I compared call properties and mating success of some males for which data were available for both study periods. Pulse rate was lower in the second than in the first year, suggesting age-related changes in this call property. Moreover, male D. pumilio increased their relative mating success from the first to the second study year. Together with other studies, the data of this study suggest that, in anurans, endurance rivalry contributes more to variation in male mating success than does female choice based on male characteristics.

In this study I examined the relationship among abundance of reproductive resources, population density, and adult sex ratio in the strawberry dart-poison frog, Dendrobates pumilio, and how these variables in turn influence the mating system, male reproductive success, and sexual selection. I studied the mating behavior in two populations of D. pumilio living in a primary and secondary rainforest on the Caribbean slope of Costa Rica. The abundance of tadpole-rearing sites (reproductive resources for females) was approximately 10-fold higher in the secondary forest. Accordingly, the population density was higher and the adult sex ratio was strongly female biased in the secondary forest, whereas the adult sex ratio was even in the primary forest. The female-biased sex ratio was associated with a higher level of polygyny and higher male mating and reproductive success in the secondary forest. In contrast, the level of polyandry did not differ between habitats. As expected, the opportunity for sexual selection on male mating success was lower in the secondary forest, the habitat with high female density. In conclusion, my results suggest that ecological variables such as resource availability have a great impact on the mating system and sexual selection through their effect on population structure. Moreover, the results of this study give further evidence that the opportunity for sexual selection is influenced by the adult sex ratio and hence by the operational sex ratio in a population. Copyright 2002.

Sexual signals indicate species identity and mate quality, and their importance for mate attraction is largely recognized. Recently, research in animal communication has started to integrate multiple signal modalities and evaluate their interactions. However, mate choice experiments across animal taxa have been limited to laboratory conditions, and assessments of multiple sexual signals under field conditions are still lacking. We take advantage of the divergence in visual and acoustic signals among populations of the Neotropical poison frog Oophaga pumilio to evaluate the importance of male advertisement calls and color patterns in female mate selection. Previous mate choice experiments in this species suggested color-assortative female mate preferences across many populations. Nevertheless, acoustic signals are crucial for sexual selection in frogs, and males of O. pumilio use advertisement calls to attract females. We hypothesize that both advertisement calls and coloration affect female mate selection in O.pumilio. To test this hypothesis we tested 452 receptive females from six populations in Costa Rica and Panama in their natural home ranges for preferences regarding local vs. non-local advertisement calls and color patterns. Overall, the calls overrode the effect of coloration, whereby most females preferred local over non-local calls. We found a tendency to prefer brighter (but not necessarily local) males in two populations. Furthermore, the strength of preferences varied geographically, and thus might be involved in prezygotic isolation among populations. The stronger effect of calls on mate attraction is associated with acoustic divergence between genetic groups in the species, while color pattern diversity is mostly located within one genetic group, i.e., not linked to large-scale population structure. Finally our data highlights the importance to consider an array of signal modalities in multiple wild populations in studies of behavioral isolation. DFG/PR/626/4-1 DFG/PR/626/4-2

In many species with a resource-based mating system, males defend resources to increase their attractiveness to females. In the strawberry poison frog, Dendrobates pumilio, suitable tadpole-rearing sites appear to be a limited resource for females. Territorial males have been suggested to defend tadpole-rearing sites to increase their access to females. In this study we investigate the spatial association between tadpole-rearing sites and the sexes as well as the spatial association of males and females. If strawberry poison frogs have resource defense polygyny, we expect males and females to be associated with tadpole-rearing sites and that females will deposit their offspring in tadpole-rearing sites inside the territories of their mates. To test this hypothesis, home range and core area sizes were calculated for both sexes and the association patterns were compared in two areas that differed in their abundance of tadpole-rearing sites. Home ranges and core areas of females were much larger than male home ranges. Females showed a clumped distribution in the vicinity of tadpole-rearing sites. Males were not clumped and were less associated with tadpole-rearing sites. Females generally did not use tadpole-rearing sites in the territory of their mates and we therefore conclude that males did not defend tadpole-rearing sites for females. Our data are consistent with the general assumption that female distribution is influenced by resource distribution and that male distribution depends on female distribution. Nevertheless, the distribution of D. pumilio females was also influenced by male spacing patterns. Males probably initially establish their core areas where female density is high and then females move among territories to sample males. Males compete vigorously for places with high female density, the defense of which is likely important for enhancing their mating success. In general, the spacing patterns did not differ between populations but the sex ratio was strongly female biased in the habitat with more tadpole-rearing sites, reflecting the direct reliance of females on these resources.

We studied the effect of relative parental investment on potential reproductive rates (PRRs) to explain sex differences in selectivity and competition in the dart-poison frog Dendrobates pumilio. We recorded the reproductive behavior of this species in a Costa Rican lowland rainforest for almost 6 months. Females spent more time on parental care than males, and `time out' estimates suggest that PRRs of males are much higher than than those of females, rendering females the limiting sex in the mating process. Males defended territories that provide suitable calling sites, space for courtship and oviposition, and prevent interference by competitors. Male mating success was highly variable, from 0 to 12 matings, and was significantly correlated with calling activity and average perch height, but was independent of body size and weight. Estimates of opportunity for sexual selection and variation in male mating success are given. The mating system is polygamous: males and females mated several times with different mates. Females were more selective than males and may sample males between matings. The discrepancy in PRRs between the sexes due to differences in parental investment and the prolonged breeding season is sufficient to explain the observed mating pattern i.e., selective females, high variance in male mating success, and the considerable opportunity for sexual selection.

Aposematism and crypsis are often viewed as two extremes of a continuum of visual conspicuousness to predators. Theory predicts that behavioral and coloration conspicuousness should vary in tandem along the conspicuousness spectrum for antipredator strategies to be effective. Here we used visual modeling of contrast and behavioral observations to examine the conspicuousness of four populations of the granular poison frog, Oophaga granulifera, which exhibits almost continuous variation in dorsal color. The patterns of geographic variation in color, visual contrast, and behavior support a gradient of overall conspicuousness along the distribution of O. granulifera. Red and green populations, at the extremes of the color distribution, differ in all elements of color, contrast, and behavior, strongly reflecting aposematic and cryptic strategies. However, there is no smooth cline in any elements of behavior or coloration between the two extremes. Instead populations of intermediate colors attain intermediate conspicuousness by displaying different combinations of aposematic and cryptic traits. We argue that coloration divergence among populations may be linked to the evolution of a gradient of strategies to balance the costs of detection by predators and the benefits of learned aversion.

Aposematism and crypsis are often viewed as two extremes of a continuum of visual conspicuousness to predators. Theory predicts that behavioral and coloration conspicuousness should vary in tandem along the conspicuousness spectrum for antipredator strategies to be effective. Here we used visual modeling of contrast and behavioral observations to examine the conspicuousness of four populations of the granular poison frog, Oophaga granulifera, which exhibits almost continuous variation in dorsal color. The patterns of geographic variation in color, visual contrast, and behavior support a gradient of overall conspicuousness along the distribution of O. granulifera. Red and green populations, at the extremes of the color distribution, differ in all elements of color, contrast, and behavior, strongly reflecting aposematic and cryptic strategies. However, there is no smooth cline in any elements of behavior or coloration between the two extremes. Instead populations of intermediate colors attain intermediate conspicuousness by displaying different combinations of aposematic and cryptic traits. We argue that coloration divergence among populations may be linked to the evolution of a gradient of strategies to balance the costs of detection by predators and the benefits of learned aversion.

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.

Background: Females have often been shown to exhibit preferences for certain male traits. However, little is known about behavioural rules females use when searching for mates in their natural habitat. We investigated mate sampling tactics and related costs in the territorial strawberry poison frog (Oophaga pumilio) possessing a lek-like mating system, where both sequential and simultaneous sampling might occur. We continuously monitored the sampling pattern and behaviour of females during the complete period between two successive matings. Results: We found no evidence that females compared males by visiting them. Instead females mated with the closest calling male irrespective of his acoustic and physical traits, and territory size. Playback experiments in the natural home ranges of receptive females revealed that tested females preferred the nearest speaker and did not discriminate between low and high call rates or dominant frequencies. Conclusions: Our results suggest that females of O. pumilio prefer the closest calling male in the studied population. We hypothesize that the sampling tactic in this population is affected by 1) a strongly female biased sex ratio and 2) a low variance in traits of available males due to strong male-male competition, preventing low quality males from defending a territory and mating.

We describe primers and PCR conditions to amplify nine new tetranucleotide loci and one new dinucleotide locus isolated from the strawberry poison frog (Oophaga pumilio). In 21 individuals from Costa Rica, the number of alleles ranged from 4 to 16, observed heterozygosities from 40 to 100%, and polymorphic information content ranged from 0.60 to 0.90 per locus. Evidence for linkage disequilibrium was found only between two loci, but this pattern was not found in other populations tested. All primer pairs cross-amplified in Oophaga vicentei from Panama.

Background Uncovering how populations of a species differ genetically and ecologically is important for understanding evolutionary processes. Here we combine population genetic methods (microsatellites) with phylogenetic information (mtDNA) to define genetic population clusters of the wide-spread Neotropical túngara frog (Physalaemus pustulosus). We measure gene flow and migration within and between population clusters and compare genetic diversity between population clusters. By applying ecological niche modeling we determine whether the two most divergent genetic groups of the túngara frog (1) inhabit different habitats, and (2) are separated geographically by unsuitable habitat across a gap in the distribution. Results Most population structure is captured by dividing all sample localities into two allopatric genetic lineages. The Northern genetic lineage (NW Costa Rica) is genetically homogenous while the Southern lineage (SW Costa Rica and Panama) is sub-divided into three population clusters by both microsatellite and mtDNA analyses. Gene flow is higher within the Northern lineage than within the Southern lineage, perhaps due to increased landscape heterogeneity in the South. Niche modeling reveals differences in suitable habitat between the Northern and Southern lineages: the Northern lineage inhabits dry/pine-oak forests, while the Southern lineage is confined to tropical moist forests. Both lineages seem to have had little movement across the distribution gap, which persisted during the last glacial maximum. The lack of movement was more pronounced for the Southern lineage than for the Northern lineage. Conclusions This study confirms the finding of previous studies that túngara frogs diverged into two allopatric genetic lineages north and south of the gap in the distribution in central Costa Rica several million years ago. The allopatric distribution is attributed to unsuitable habitat and probably other unknown ecological factors present across the distribution gap. Niche conservatism possibly contributes to preventing movements across the gap and gene flow between both groups. Genetic and ecological data indicate that there is the potential for ecological divergence in allopatry between lineages. In this context we discuss whether the Northern and Southern lineages should be recognized as separate species, and we conclude that further studies of pre- and post-zygotic isolation are needed for a final assessment. Identified population clusters should motivate future behavioral and ecological research regarding within-species biodiversity and speciation mechanisms.