Predicting spring migration of two European amphibian species with plant phenology using citizen science data.

Habitat fragmentation is one of the drivers for amphibian population declines globally. Especially in industrialized countries roads disrupt the seasonal migration of amphibians between hibernation and reproduction sites, often ending in roadkills. Thus, a timely installing of temporary mitigation measures is important for amphibian conservation. We wanted to find out if plant phenology can be a proxy in advance to determine the start of amphibian migration, since both phenomena are triggered by temperature. We analysed data of 3751 amphibian and 7818 plant phenology observations from citizen science projects in Austria between 2000 and 2018. Using robust regression modelling we compared the migration of common toads (Bufo bufo) and common frogs (Rana temporaria) with the phenology of five tree, one shrub, and one herb species. Results showed close associations between the migration of common frogs and phenological phases of European larch, goat willow and apricot. Models based on goat willow predict migration of common frog to occur 21 days after flowering, when flowering was observed on 60th day of year; apricot based models predict migration to occur 1 day after flowering, observed on the 75th day of year. Common toads showed weaker associations with plant phenology than common frogs. Our findings suggest that plant phenology can be used to determine the onset of temporary mitigation measures for certain amphibian species to prevent roadkills.

Adaptation to elevation but limited local adaptation in an amphibian.

We performed a reciprocal transplant experiment to estimate "parallel" adaptation to elevation and "unique" adaptation to local sites at the same elevation, using the frog Rana temporaria in the Swiss Alps. It is important to distinguish these two processes because they have different implications for population structure and ecological specialization. Larvae were reared from hatching to metamorphosis within enclosures installed in their pond of origin, in three foreign ponds at the same elevation, and in four ponds at different elevation (1500-2000 m higher or lower). There were two source populations from each elevation, and adults were held in a common environment for 1 year before they were crossed to produce offspring for the experiment. Fitness was a measure that integrated larval survival, development rate, and body size. Parallel adaptation to elevation was indicated by an advantage at the home elevation (11.5% fitness difference at low elevation and 47% at high elevation). This effect was stronger than that observed in most other studies, according to a survey of previous transplant experiments across elevation (N = 8 animal species and 71 plants). Unique local adaptation within elevational zones was only 0.3-0.7 times as strong as parallel adaptation, probably because gene flow is comparatively high among nearby wetlands at the same elevation. The home-elevation advantage may reduce gene flow across the elevational gradient and enable the evolution of habitat races specialized on elevation.

Altered thyroid hormone levels affect the capacity for temperature-induced developmental plasticity in larvae of Rana temporaria and Xenopus laevis.

Anuran larvae show phenotypic plasticity in age and size at metamorphosis as a response to temperature variation. The capacity for temperature-induced developmental plasticity is determined by the thermal adaptation of a population. Multiple factors such as physiological responses to changing environmental conditions, however, might influence this capacity as well. In anuran larvae, thyroid hormone (TH) levels control growth and developmental rate and changes in TH status are a well-known stress response to sub-optimal environmental conditions. We investigated how chemically altered TH levels affect the capacity to exhibit temperature-induced developmental plasticity in larvae of the African clawed frog (Xenopus laevis) and the common frog (Rana temporaria). In both species, TH level influenced growth and developmental rate and modified the capacity for temperature-induced developmental plasticity. High TH levels reduced thermal sensitivity of metamorphic traits up to 57% (R. temporaria) and 36% (X. laevis). Rates of growth and development were more plastic in response to temperature in X. laevis (+30%) than in R. temporaria (+6%). Plasticity in rates of growth and development is beneficial to larvae in heterogeneous habitats as it allows a more rapid transition into the juvenile stage where rates of mortality are lower. Therefore, environmental stressors that increase endogenous TH levels and reduce temperature-dependent plasticity may increase risks and the vulnerability of anuran larvae. As TH status also influences metabolism, future studies should investigate whether reductions in physiological plasticity also increases the vulnerability of tadpoles to global change.

Endocrine Disruption Alters Developmental Energy Allocation and Performance in Rana temporaria.

Environmental change exposes wildlife to a wide array of environmental stressors that arise from both anthropogenic and natural sources. Many environmental stressors with the ability to alter endocrine function are known as endocrine disruptors, which may impair the hypothalamus-pituitary-thyroid axis resulting in physiological consequences to wildlife. In this study, we investigated how the alteration of thyroid hormone (TH) levels due to exposure to the environmentally relevant endocrine disruptor sodium perchlorate (SP; inhibitory) and exogenous L-thyroxin (T4; stimulatory) affects metabolic costs and energy allocation during and after metamorphosis in a common amphibian (Rana temporaria). We further tested for possible carry-over effects of endocrine disruption during larval stage on juvenile performance. Energy allocated to development was negatively related to metabolic rate and thus, tadpoles exposed to T4 could allocate 24% less energy to development during metamorphic climax than control animals. Therefore, the energy available for metamorphosis was reduced in tadpoles with increased TH level by exposure to T4. We suggest that differences in metabolic rate caused by altered TH levels during metamorphic climax and energy allocation to maintenance costs might have contributed to a reduced energetic efficiency in tadpoles with high TH levels. Differences in size and energetics persisted beyond the metamorphic boundary and impacted on juvenile performance. Performance differences are mainly related to strong size-effects, as altered TH levels by exposure to T4 and SP significantly affected growth and developmental rate. Nevertheless, we assume that juvenile performance is influenced by a size-independent effect of achieved TH. Energetic efficiency varied between treatments due to differences in size allocation of internal macronutrient stores. Altered TH levels as caused by several environmental stressors lead to persisting effects on metamorphic traits and energetics and, thus, caused carry-over effects on performance of froglets. We demonstrate the mechanisms through which alterations in abiotic and biotic environmental factors can alter phenotypes at metamorphosis and reduce lifetime fitness in these and likely other amphibians.

Hindered and constrained: limited potential for thermal adaptation in post-metamorphic and adult Rana temporaria along elevational gradients.

Adaptation to warming climates could counteract the effects of global warming. Thus, understanding how species cope with contrasting climates may inform us about their potential for thermal adaptation and which processes may hamper that ability (e.g. evolutionary trade-offs, phenology or behavioural thermoregulation). In addition to temperature, time constraints may also exert important selective pressures. Here, we compare the thermal sensitivity of locomotion of metamorphic and adult European common frogs (Rana temporaria) originating from populations along an elevational gradient. We employed the template mode of variation (TMV) analysis to decompose the thermal sensitivity of locomotion and explore the existence of trade-offs ('hotter is better' and 'specialist-generalist') and the degree of local adaptation. To that end, we studied the relationship between TMV parameters and local environmental conditions. Further, we compared preferred temperatures to assess whether behavioural thermoregulation could dampen the effects of thermal variation, reducing the intensity of selection and limiting thermal adaptation (i.e. 'Bogert effect'). We suggest that behavioural thermoregulation has promoted the conservatism of thermal sensitivity in R. temporaria. Yet, we observed a trend towards narrower thermal niches shifted towards warmer temperature in populations with severe temporal constraints, conforming to the 'generalist-specialist' trade-off. Apparently, this enables time-constrained populations - especially in the case of metamorphs - to effectively exploit resources during the warmest periods. The limited potential of R. temporaria for thermal adaptation suggests that forecasts of global warming should incorporate thermoregulation and explore its potential to buffer species from rising temperatures.

European common frog Rana temporaria (Anura: Ranidae) larvae show subcellular responses under field-relevant Bacillus thuringiensis var. israelensis (Bti) exposure levels.

Bacillus thuringiensis var. israelensis (Bti) is presumed to be an environmental friendly agent for the use in either health-related mosquito control or the reduction of nuisance associated with mosquitoes from seasonal wetlands. Amphibians inhabiting these valuable wetlands may be exposed to Bti products several times during their breeding season. Up until now, information regarding effects on the non-targeted group of amphibians has to be considered rather inconsistent. On this account, we evaluated how three repeated exposures to frequently used Bti formulations (VectoBac®12AS, VectoBac®WG) in field-relevant rates affect European common frog (Rana temporaria) larvae. In a laboratory approach, we assessed potential effects with regard to enzymatic biomarkers (glutathione-S-transferase (GST), glutathione reductase (GR), acetylcholine esterase (AChE)), development, body condition and survival until the end of metamorphosis. Although survival and time to metamorphosis were not significantly affected, larval development tended to be shortened in the Bti treated water phase. Furthermore, exposure to Bti induced significant increases of GST (37-550%), GR (5-140%) and AChE (38-137%) irrespectively of the applied formulation, indicating detoxification, antioxidant responses as well as an alteration of neuronal activity. GST activity increased twice as much after two repeatedly executed Bti applications within a time period of 6 days. The examination of several biochemical markers is needed to fully evaluate the ecotoxicological risk of Bti for amphibian populations, especially in the context of worldwide amphibian declines. Nevertheless, following the precautionary principle, it may be advisable to implement certain thresholds for application numbers and intervals in order to ensure environmentally friendly mosquito control programs, especially in areas designated for nature conservation.

Electrophysiological differences in cholinergic signaling between the hearts of summer and winter frogs (Rana temporaria).

The striking seasonal difference in sensitivity of frog cardiac muscle to acetylcholine or stimulation of parasympathetic nervous fibers has been noted almost a century ago, although its electrophysiological basis has never been revealed. The present study compares the effects of the muscarinic agonist carbamylcholine chloride (CCh 10-8-10-5 M) on electrical activity in isolated atrial and ventricular preparations from dormant frogs (Rana temporaria) caught in January (winter-acclimatized, WA) and from active frogs caught in July (summer-acclimatized, SA). Seasonal differences in the density of potassium acetylcholine-dependent current (IKACh) were also studied in atrial and ventricular myocytes from both summer and winter groups. In atrial myocardium, CCh produced concentration-dependent shortening of action potentials (APs). CCh concentration producing a 50% reduction of AP duration was lower in WA (1.03 × 10-7 M) than SA atria (2.7 × 10-7 M). 10-6 M CCh induced drastic reduction of AP amplitude rendering the tissue unexcitable in both WA or SA atrial preparations. Ventricular preparations showed greater seasonal difference in CCh sensitivity. While 10-6 M induced inexcitability in 50% of tested WA preparations, in SA preparations even 10-5 M CCh was without effect. This striking difference between cholinergic effects in SA and WA frog ventricle could be explained by seasonal changes in the IKACh density. The density of IKACh induced by 10-5 M CCh and measured at 0 mV was 14.4 ± 3.45 pA/pF in WA, but only 1.5 ± 0.4 pA/pF in SA atrial cells. In ventricular cells, the respective values were 2.61 ± 0.56 and 0.71 ± 0.09 pA/pF. Thus, hibernating winter frog has a much greater electrophysiological cholinergic response than active summer frog due to up-regulation of IKACh.

The phenotypic variability in Rana temporaria decreases in response to drying habitats.

In this study, we evaluated the diversity of skin coloration as a proxy for phenotypic diversity. The European common frog (Rana temporaria) populations from the Southern slope of central Pyrenees lie at the limit of the species distribution in latitude and altitude. We analysed the relationship of skin color typology with different environmental variables and found a large decrease in skin type variety in frogs developing in temporary water bodies when compared to those developing in permanent water bodies. Our results show that our method can be used as a non-invasive way to study phenotypic diversity and suggest that adaptation to an early metamorphosis in a rapidly-drying habitat can have negative effects on adult phenotypic diversity. In light of these results, we argue that access to permanent water bodies is important to prevent loss of diversity in anuran populations and reduce their vulnerability to environmental impacts as well as pathogens.

Differing long term trends for two common amphibian species (Bufo bufo and Rana temporaria) in alpine landscapes of Salzburg, Austria.

This study focuses on the population trends of two widespread European anuran species: the common toad (Bufo bufo) and the common frog (Rana temporaria). The basis of this study is data gathered over two decades of amphibian fencing alongside roads in the Austrian state of Salzburg. Different statistical approaches were used to analyse the data. Overall average increase or decrease of each species was estimated by calculating a simple average locality index. In addition the statistical software TRIM was used to verify these trends as well as to categorize the data based on the geographic location of each migration site. The results show differing overall trends for the two species: the common toad being stable and the common frog showing a substantial decline over the last two decades. Further analyses based on geographic categorization reveal the strongest decrease in the alpine range of the species. Drainage and agricultural intensification are still ongoing problems within alpine areas, not only in Salzburg. Particularly in respect to micro-climate and the availability of spawning places these changes appear to have a greater impact on the habitats of the common frog than the common toad. Therefore we consider habitat destruction to be the main potential reason behind this dramatic decline. We also conclude that the substantial loss of biomass of a widespread species such as the common frog must have a severe, and often overlooked, ecological impact.

Physiological mechanisms of adaptive developmental plasticity in Rana temporaria island populations.

BACKGROUND: Adaptive plasticity is essential for many species to cope with environmental heterogeneity. In particular, developmental plasticity allows organisms with complex life cycles to adaptively adjust the timing of ontogenetic switch points. Size at and time to metamorphosis are reliable fitness indicators in organisms with complex cycles. The physiological machinery of developmental plasticity commonly involves the activation of alternative neuroendocrine pathways, causing metabolic alterations. Nevertheless, we have still incomplete knowledge about how these mechanisms evolve under environments that select for differences in adaptive plasticity. In this study, we investigate the physiological mechanisms underlying divergent degrees of developmental plasticity across Rana temporaria island populations inhabiting different types of pools in northern Sweden. METHODS: In a laboratory experiment we estimated developmental plasticity of amphibian larvae from six populations coming from three different island habitats: islands with only permanent pools, islands with only ephemeral pools, and islands with a mixture of both types of pools. We exposed larvae of each population to either constant water level or simulated pool drying, and estimated their physiological responses in terms of corticosterone levels, oxidative stress, and telomere length. RESULTS: We found that populations from islands with only temporary pools had a higher degree of developmental plasticity than those from the other two types of habitats. All populations increased their corticosterone levels to a similar extent when subjected to simulated pool drying, and therefore variation in secretion of this hormone does not explain the observed differences among populations. However, tadpoles from islands with temporary pools showed lower constitutive activities of catalase and glutathione reductase, and also showed overall shorter telomeres. CONCLUSIONS: The observed differences are indicative of physiological costs of increased developmental plasticity, suggesting that the potential for plasticity is constrained by its costs. Thus, high levels of responsiveness in the developmental rate of tadpoles have evolved in islands with pools at high but variable risk of desiccation. Moreover, the physiological alterations observed may have important consequences for both short-term odds of survival and long term effects on lifespan.

Comparative High-Density Linkage Mapping Reveals Conserved Genome Structure but Variation in Levels of Heterochiasmy and Location of Recombination Cold Spots in the Common Frog.

By combining 7077 SNPs and 61 microsatellites, we present the first linkage map for some of the early diverged lineages of the common frog, Rana temporaria, and the densest linkage map to date for this species. We found high homology with the published linkage maps of the Eastern and Western lineages but with differences in the order of some markers. Homology was also strong with the genome of the Tibetan frog Nanorana parkeri and we found high synteny with the clawed frog Xenopus tropicalis We confirmed marked heterochiasmy between sexes and detected nonrecombining regions in several groups of the male linkage map. Contrary to the expectations set by the male heterogamety of the common frog, we did not find male heterozygosity excess in the chromosome previously shown to be linked to sex determination. Finally, we found blocks of loci showing strong transmission ratio distortion. These distorted genomic regions might be related to genetic incompatibilities between the parental populations, and are promising candidates for further investigation into the genetic basis of speciation and adaptation in the common frog.

Volunteer Conservation Action Data Reveals Large-Scale and Long-Term Negative Population Trends of a Widespread Amphibian, the Common Toad (Bufo bufo).

Rare and threatened species are the most frequent focus of conservation science and action. With the ongoing shift from single-species conservation towards the preservation of ecosystem services, there is a greater need to understand abundance trends of common species because declines in common species can disproportionately impact ecosystems function. We used volunteer-collected data in two European countries, the United Kingdom (UK) and Switzerland, since the 1970s to assess national and regional trends for one of Europe's most abundant amphibian species, the common toad (Bufo bufo). Millions of toads were moved by volunteers across roads during this period in an effort to protect them from road traffic. For Switzerland, we additionally estimated trends for the common frog (Rana temporaria), a similarly widespread and common amphibian species. We used state-space models to account for variability in detection and effort and included only populations with at least 5 years of data; 153 populations for the UK and 141 for Switzerland. Common toads declined continuously in each decade in both countries since the 1980s. Given the declines, this common species almost qualifies for International Union for the Conservation of Nature (IUCN) red-listing over this period despite volunteer conservation efforts. Reasons for the declines and wider impacts remain unknown. By contrast, common frog populations were stable or increasing in Switzerland, although there was evidence of declines after 2003. "Toads on Roads" schemes are vital citizen conservation action projects, and the data from such projects can be used for large scale trend estimations of widespread amphibians. We highlight the need for increased research into the status of common amphibian species in addition to conservation efforts focusing on rare and threatened species.

The relative importance of prey-borne and predator-borne chemical cues for inducible antipredator responses in tadpoles.

Chemical cues that evoke anti-predator developmental changes have received considerable attention, but it is not known to what extent prey use information from the smell of predators and from cues released through digestion. We conducted an experiment to determine the importance of various types of cues for the adjustment of anti-predator defences. We exposed tadpoles (common frog, Rana temporaria) to water originating from predators (caged dragonfly larvae, Aeshna cyanea) that were fed different types and quantities of prey outside of tadpole-rearing containers. Variation among treatments in the magnitude of morphological and behavioural responses was highly consistent. Our results demonstrate that tadpoles can assess the threat posed by predators through digestion-released, prey-borne cues and continually released predator-borne cues. These cues may play an important role in the fine-tuning of anti-predator responses and significantly affect the outcome of interactions between predators and prey in aquatic ecosystems. There has been much confusion regards terminology used in the literature, and therefore we also propose a more precise and consistent binomial nomenclature based on the timing of chemical cue release (stress-, attack-, capture-, digestion- or continually released cues) and the origin of cues (prey-borne or predator-borne cues). We hope that this new nomenclature will improve comparisons among studies on this topic.

Behavioural adaptations of Rana temporaria to cold climates.

Environmental conditions at the edge of a species' ecological optimum can exert great ecological or evolutionary pressure at local populations. For ectotherms like amphibians temperature is one of the most important abiotic factors of their environment as it influences directly their metabolism and sets limits to their distribution. Amphibians have evolved three ways to cope with sub-zero temperatures: freeze tolerance, freeze protection, freeze avoidance. The aim of this study was to assess which strategy common frogs at mid and high elevation use to survive and thrive in cold climates. In particular we (1) tested for the presence of physiological freeze protection, (2) evaluated autumnal activity and overwintering behaviour with respect to freeze avoidance and (3) assessed the importance of different high-elevation microhabitats for behavioural thermoregulation. Common frogs did not exhibit any signs of freeze protection when experiencing temperatures around 0 °C. Instead they retreated to open water for protection and overwintering. High elevation common frogs remained active for around the same period of time than their conspecifics at lower elevation. Our results suggest that at mid and high elevation common frogs use freeze avoidance alone to survive temperatures below 0 °C. The availability of warm microhabitats, such as rock or pasture, provides high elevation frogs with the opportunity of behavioural thermoregulation and thus allows them to remain active at temperatures at which common frogs at lower elevation cease activity.

Reproductive interference between Rana dalmatina and Rana temporaria affects reproductive success in natural populations.

Experimental evidence suggests that reproductive interference between heterospecifics can seriously affect individual fitness; support from field studies for such an effect has, however, remained scarce. We studied reproductive interference in 25 natural breeding ponds in an area where two ranid frogs, Rana dalmatina and Rana temporaria, co-occur. The breeding seasons of the two species usually overlap and males of both species are often found in amplexus with heterospecific females, even though matings between heterospecifics produce no viable offspring. We estimated species abundance ratios based on the number of clutches laid and evaluated fertilization success. In ponds with low spatial complexity and a species abundance ratio biased towards R. temporaria, the average fertilization success of R. dalmatina eggs decreased, while this relationship was not detectable in spatially more complex ponds. Fertilization success of R. temporaria did not decrease with increasing relative numbers of heterospecifics. This asymmetry in fitness effects of reproductive interference may be attributed to R. temporaria males being more competitive in scramble competition for females than R. dalmatina males. Our study is among the first to demonstrate that in natural breeding populations of vertebrates interference among heterospecifics has the potential to substantially lower reproductive success at the population level, which may in turn affect population dynamics.

Why do green rods of frog and toad retinas look green?

Amphibian "green" rods express a blue-sensitive cone visual pigment, and should look yellow. However,when observing them axially under microscope one sees them as green. We used single-cell microspectrophotometry (MSP) to reveal the basis of the perceived color of these photoreceptors. Conventional side-on MSP recording of the proximal cell segments reveals no selective longwave absorbing pigment explaining the green color. End-on MSP recording shows, in addition to the green rod visual pigment, an extra 2- to 4-fold attenuation being almost flat throughout the visible spectrum. This attenuation is absent in red (rhodopsin) rods, and vanishes in green rods when the retina is bathed in high-refractive media, and at wide illumination aperture. The same treatments change the color from green to yellow. It seems that the non-visual pigment attenuation is a result of slender green rod myoids operating as non-selective light guides. We hypothesize that narrow myoids, combined with photomechanical movements of melanin granules, allow a wide range of sensitivity regulation supporting the operation of green rods as blue receptors at mesopic-to low-photopic illumination levels.End-on transmittance spectrum of green rods looks similar to the reflectance spectrum of khaki military uniforms. So their greenness is the combined result of optics and human color vision.

Behavioural and physiological adaptations to low-temperature environments in the common frog, Rana temporaria.

BACKGROUND: Extreme environments can impose strong ecological and evolutionary pressures at a local level. Ectotherms are particularly sensitive to low-temperature environments, which can result in a reduced activity period, slowed physiological processes and increased exposure to sub-zero temperatures. The aim of this study was to assess the behavioural and physiological responses that facilitate survival in low-temperature environments. In particular, we asked: 1) do high-altitude common frog (Rana temporaria) adults extend the time available for larval growth by breeding at lower temperatures than low-altitude individuals?; and 2) do tadpoles sampled from high-altitude sites differ physiologically from those from low-altitude sites, in terms of routine metabolic rate (RMR) and freeze tolerance? Breeding date was assessed as the first day of spawn observation and local temperature recorded for five, paired high- and low-altitude R. temporaria breeding sites in Scotland. Spawn was collected and tadpoles raised in a common laboratory environment, where RMR was measured as oxygen consumed using a closed respiratory tube system. Freeze tolerance was measured as survival following slow cooling to the point when all container water had frozen. RESULTS: We found that breeding did not occur below 5°C at any site and there was no significant relationship between breeding temperature and altitude, leading to a delay in spawning of five days for every 100 m increase in altitude. The relationship between altitude and RMR varied by mountain but was lower for individuals sampled from high- than low-altitude sites within the three mountains with the highest high-altitude sites (≥900 m). In contrast, individuals sampled from low-altitudes survived freezing significantly better than those from high-altitudes, across all mountains. CONCLUSIONS: Our results suggest that adults at high-altitude do not show behavioural adaptations in terms of breeding at lower temperatures. However, tadpoles appear to have the potential to adapt physiologically to surviving at high-altitude via reduced RMR but without an increase in freeze tolerance. Therefore, survival at high-altitude may be facilitated by physiological mechanisms that permit faster growth rates, allowing completion of larval development within a shorter time period, alleviating the need for adaptations that extend the time available for larval growth.

Phenotypic plasticity in the hepatic transcriptome of the European common frog (Rana temporaria): the interplay between environmental induction and geographical lineage on developmental response.

Phenotypic plasticity might facilitate adaptation to new environmental conditions through the enhancement of initial survival of organisms. Once a population is established, further adaptation and diversification may occur through adaptive trait evolution. While several studies have found evidence for this mechanism using phenotypic traits, much less is known at the level of gene expression. Here, we use an islands system of frog populations that show local adaptation and phenotypic plasticity to pool drying conditions in development time until metamorphoses. We examined gene expression differences in Rana temporaria tadpole livers with respect to pool drying at the source population and in response to simulated pool drying in the laboratory. Using a MAGEX cDNA microarray and quantitative real-time polymerase chain reaction (qPCR), we identified an increase in several gene transcripts in response to artificial pool drying including thyroid hormone receptor alpha and beta, carbamoyl phosphate synthetase 1, ornithine transcarbamylase and catalase. In addition, these gene transcripts also showed greater abundance in island populations that developed faster. Hence, the gene transcripts were related to both constitutive response (higher levels in island populations that developed faster) and plastic response (increased abundance under decreasing water levels). This pattern is in accordance with genetic accommodation, which predicts similarities between plastic gene expression and constitutive expression in locally adapted populations.

Hot tadpoles from cold environments need more nutrients--life history and stoichiometry reflects latitudinal adaptation.

1. High-latitude species (and populations within species) are adapted to short and cold summers. They often have high growth and development rates to fully use the short growing season and mature before the onset of winter. 2. Within the context of ecological stoichiometry theory, this study combines ecology with evolution by relating latitudinal life-history adaptations to their molecular consequences in body nutrient composition in Rana temporaria tadpoles. 3. Temperature and food quality were manipulated during the development of tadpoles from Arctic and Boreal origins. We determined tadpole growth rate, development rate, body size and nutrient content, to test whether (i) Arctic tadpoles could realize higher growth rates and development rates with the help of higher-quality food even when food quantity was unchanged, (ii) Arctic and Boreal tadpoles differed in their stoichiometric (and life history) response to temperature changes, (iii) higher growth rates lead to higher tadpole P content (growth rate hypothesis) and (iv) allometric scaling affects tadpole nutrient allocation. 4. We found that especially Arctic tadpoles grew and developed faster with the help of higher-quality food and that tadpoles differed in their stoichiometric (and life history) response to temperature changes depending on region of origin (probably due to different temperature optima). There was no evidence that higher growth rates mediated the positive effect of temperature on tadpole P content. On the contrary, the covariate growth rate was negatively connected with tadpole P content (refuting the growth rate hypothesis). Lastly, tadpole P content was not related to body size, but tadpole C content was higher in larger tadpoles, probably due to increased fat storage. 5. We conclude that temperature had a strong effect on tadpole life history, nutrient demand and stoichiometry and that this effect depended on the evolved life history.

Using genetic variation to infer associations with climate in the common frog, Rana temporaria.

Recent and historical species' associations with climate can be inferred using molecular markers. This knowledge of population and species-level responses to climatic variables can then be used to predict the potential consequences of ongoing climate change. The aim of this study was to predict responses of Rana temporaria to environmental change in Scotland by inferring historical and contemporary patterns of gene flow in relation to current variation in local thermal conditions. We first inferred colonization patterns within Europe following the last glacial maximum by combining new and previously published mitochondrial DNA sequences. We found that sequences from our Scottish samples were identical to (92%), or clustered with, the common haplotype previously identified from Western Europe. This clade showed very low mitochondrial variation, which did not allow inference of historical colonization routes but did allow interpretation of patterns of current fine-scale population structure without consideration of confounding historical variation. Second, we assessed fine-scale microsatellite-based patterns of genetic variation in relation to current altitudinal temperature gradients. No population structure was found within altitudinal gradients (average FST=0.02), despite a mean annual temperature difference of 4.5 °C between low- and high-altitude sites. Levels of genetic diversity were considerable and did not vary between sites. The panmictic population structure observed, even along temperature gradients, is a potentially positive sign for R. temporaria persistence in Scotland in the face of a changing climate. This study demonstrates that within taxonomic groups, thought to be at high risk from environmental change, levels of vulnerability can vary, even within species.