The microbiome at the interface between environmental stress and animal health: an example from the most threatened vertebrate group.

Nitrate pollution and global warming are ubiquitous stressors likely to interact and affect the health and survival of wildlife, particularly aquatic ectotherms. Animal health is largely influenced by its microbiome (commensal/symbiotic microorganisms), which responds to such stressors. We used a crossed experimental design including three nitrate levels and five temperature regimes to investigate their interactive and individual effects on an aquatic ectotherm, the European common frog. We associated health biomarkers in larvae with changes in gut bacteria diversity and composition. Larvae experienced higher stress levels and lower body condition under high temperatures and nitrate exposure. Developmental rate increased with temperature but decreased with nitrate pollution. Alterations in bacteria composition but not diversity are likely to correlate with the observed outcomes in larvae health. Leucine degradation decreased at higher temperatures corroborating accelerated development, nitrate degradation increased with nitrate level corroborating reduced body condition and an increase in lysine biosynthesis may have helped larvae deal with the combined effects of both stressors. These results reinforce the importance of associating traditional health biomarkers with underlying microbiome changes. Therefore, we urge studies to investigate the effects of environmental stressors on microbiome composition and consequences for host health in a world threatened by biodiversity loss.

Mapping of quantitative trait loci for life history traits segregating within common frog populations.

The evolution of complex traits is often shaped by adaptive divergence. However, very little is known about the number, effect size, and location of the genomic regions influencing the variation of these traits in natural populations. Based on a dense linkage map of the common frog, Rana temporaria, we have localized, for the first time in amphibians, three significant and nine suggestive quantitative trait loci (QTLs) for metabolic rate, growth rate, development time, and weight at metamorphosis, explaining 5.6-18.9% of the overall phenotypic variation in each trait. We also found a potential pleiotropic QTL between development time and size at metamorphosis that, if confirmed, might underlie the previously reported genetic correlation between these traits. Furthermore, we demonstrate that the genetic variation linked to fitness-related larval traits segregates within Rana temporaria populations. This study provides the first insight into the genomic regions that affect larval life history traits in anurans, providing a valuable resource to delve further into the genomic basis of evolutionary change in amphibians.

Does Bti (Bacillus thuringiensis var. israelensis) affect Rana temporaria tadpoles?

Biocides based on toxins of Bacillus thuringiensis var. israelensis (Bti) are established as alternatives to conventional chemical insecticides for mosquito control all across the globe since they are regarded ecologically compatible and harmless to non-target species. Since recent studies on amphibian larvae have called this opinion into question, we exposed Rana temporaria tadpoles to single (1 mg/L), tenfold (10 mg/L) and hundredfold (100 mg/L) field concentrations of VectoBac® WG (a water dispersible granule Bti formulation) in the laboratory for eleven days to investigate whether larvae were adversely affected by Bti and its endotoxin proteins. In addition to a negative (water) control, a positive control based on organic rice protein (50 mg/L) was run to check for the nutritional relevance of Bti proteins. There was no Bti-related mortality and a histopathological analysis of tadpole intestines revealed no adverse effects. Analyses of biomarkers for proteotoxicity (stress protein family, Hsp70) and neurotoxicity or metabolic action (b-esterases acetylcholine esterase (AChE) and carboxylesterases) revealed no significant differences between Bti treatments and the negative control. The responses of tadpoles in the protein-supplemented positive control differed from those of the negative control and the Bti treatments. Tadpoles in the positive control had reduced body mass and elevated AChE activity.

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.

Cool tadpoles from Arctic environments waste fewer nutrients - high gross growth efficiencies lead to low consumer-mediated nutrient recycling in the North.

Endothermic organisms can adapt to short growing seasons, low temperatures and nutrient limitation by developing high growth rates and high gross growth efficiencies (GGEs). Animals with high GGEs are better at assimilating limiting nutrients and thus should recycle (or lose) fewer nutrients. Longer guts in relation to body mass may facilitate higher GGE under resource limitation. Within the context of ecological stoichiometry theory, this study combines ecology with evolution by relating latitudinal life-history adaptations in GGE, mediated by gut length, to its ecosystem consequences, such as consumer-mediated nutrient recycling. In common garden experiments, we raised Rana temporaria tadpoles from two regions (Arctic/Boreal) under two temperature regimes (18/23 °C) crossed with two food quality treatments (high/low-nitrogen content). We measured tadpole GGEs, total nutrient loss (excretion + egestion) rates and gut length during ontogeny. In order to maintain their elemental balance, tadpoles fed low-nitrogen (N) food had lower N excretion rates and higher total phosphorous (P) loss rates than tadpoles fed high-quality food. In accordance with expectations, Arctic tadpoles had higher GGEs and lower N loss rates than their low-latitude conspecifics, especially when fed low-N food, but only in ambient temperature treatments. Arctic tadpoles also had relatively longer guts than Boreal tadpoles during early development. That temperature and food quality interacted with tadpole region of origin in affecting tadpole GGEs, nutrient loss rates and relative gut length, suggests evolved adaptation to temperature and resource differences. With future climate change, mean annual temperatures will increase. Additionally, species and genotypes will migrate north. This will change the functioning of Boreal and Arctic ecosystems by affecting consumer-mediated nutrient recycling and thus affect nutrient dynamics in general. Our study shows that evolved latitudinal adaption can change key ecosystem functions.

Isocline analysis of competition predicts stable coexistence of two amphibians.

We investigated the interaction between larvae of two anuran amphibian species (Rana temporaria and Bufo bufo) to test models of two-species competition. The study had a response surface experimental design with four replicates, each consisting of 24 density combinations. Larval performance-and, by assumption, change in population size-was defined by a linear combination of survival, growth, and development. We fit six competition models from the literature and discovered that density dependence was strongly non-linear, with the highest support for the Hassel-Comins model. Rana temporaria was competitively superior to B. bufo; the impacts of both species on growth and development were about five- to tenfold greater than those on survival. Isocline analysis predicted a stable configuration, which agrees with the observation that these two species are syntopic in nature. The results of this study confirm competition theory by identifying a model structure that agrees with data and making predictions that are broadly supportive of the observations.

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.

Compensatory growth strategies are affected by the strength of environmental time constraints in anuran larvae.

Organisms normally grow at a sub-maximal rate. After experiencing a period of arrested growth, individuals often show compensatory growth responses by modifying their life-history, behaviour and physiology. However, the strength of compensatory responses may vary across broad geographic scales as populations differ in their exposition to varying time constraints. We examined differences in compensatory growth strategies in common frog (Rana temporaria) populations from southern and northern Sweden. Tadpoles from four populations were reared in the laboratory and exposed to low temperature to evaluate the patterns and mechanisms of compensatory growth responses. We determined tadpoles' growth rate, food intake and growth efficiency during the compensation period. In the absence of arrested growth conditions, tadpoles from all the populations showed similar (size-corrected) growth rates, food intake and growth efficiency. After being exposed to low temperature for 1 week, only larvae from the northern populations increased growth rates by increasing both food intake and growth efficiency. These geographic differences in compensatory growth mechanisms suggest that the strategies for recovering after a period of growth deprivation may depend on the strength of time constraints faced by the populations. Due to the costs of fast growth, only populations exposed to the strong time constraints are prone to develop fast recovering strategies in order to metamorphose before conditions deteriorate. Understanding how organisms balance the cost and benefits of growth strategies may help in forecasting the impact of fluctuating environmental conditions on life-history strategies of populations likely to be exposed to increasing environmental variation in the future.

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.

Replication of an experiment on extremely diluted thyroxine and highland amphibians.

INTRODUCTION: The purpose of this study was to reproduce an experiment with diluted thyroxine and amphibians. A detailed account of the difficulties of this line of research has been published by the initial team (Endler and Scherer-Pongratz). One experiment which has been reported reproducible by the initial team and independent researchers is the effect of extremely diluted agitated thyroxine (T30x) versus analogously prepared water (W30x) in amphibians from biotopes above the tree line (highland amphibians). METHODS: (A) The author replicated the experiment. Rana temporaria were taken from an alpine biotope and the methods given in the original protocols were followed. Animals were treated from the 2-legged stage on. (B), the author reanalyzed the results reported by the initial team and by independent researchers (van Wijk, Lassnig, Zausner-Lukitsch, Bach, Harrer). RESULTS: (A) In the author's own experiment, there was a clear trend of T30x animals developing more slowly (i.e. up to 6 h within 3 days) than W30x animals. This is in line with the previous experiments. Due to small numbers of animals, the differences in the frequency of larvae reaching the 4-legged stage and the stage with reduced tail were not statistically significant (p > 0.05). The effect size was large (d > 0.08). (B) In the analysis of all available data with regard to the 4-legged stage, pooled T30x values from the initial team were 10.1% smaller than W30x values (100%) and pooled T30x values from the 5 independent researchers were 12.4% smaller (p < 0.01 and d > 0.08). Analogously, the number of animals entering the juvenile stage with reduced tail was smaller for T30x than for W30x.

Geographic variation in corticosterone response to chronic predator stress in tadpoles.

Chronic stress often affects growth and development negatively, and these effects are often mediated via glucocorticoid hormones, which elevate during stress. We investigated latitudinal variation in corticosterone (CORT) response to chronic predator stress in Rana temporaria tadpoles along a 1500-km latitudinal cline in Sweden tadpoles, in a laboratory experiment. We hypothesized that more time-constrained high-latitude populations have evolved a lower CORT response to chronic stress to maintain higher growth under stressful conditions. Southern tadpoles had higher CORT content in response to predators after 1 day of exposure, whereas there was no increase in CORT in the northern populations. Two weeks later, there were no predator-induced CORT elevations. Artificially elevated CORT levels strongly decreased growth, development and survival in both northern and southern tadpoles. We suggest that the lower CORT response in high-latitude populations can be connected with avoidance of CORT-mediated reduction in growth and development, but also discuss other possible explanations.

Alteration of mitochondrial efficiency affects oxidative balance, development and growth in frog (Rana temporaria) tadpoles.

Mitochondria are known to play a central role in life history processes, being the main source of reactive oxygen species (ROS), which promote oxidative constraint. Surprisingly, although the main role of the mitochondria is to produce ATP, the plasticity of mitochondrial ATP generation has received little attention in life history studies. Yet, mitochondrial energy transduction represents the physiological link between environmental resources and energy allocated to animal performance. Studying both facets of mitochondrial functioning (ATP and ROS production) would allow better understanding of the proximate mechanisms underlying life history. We have experimentally modulated the mitochondrial capacity to generate ROS and ATP during larval development of Rana temporaria tadpoles, via chronic exposure (34 days) to a mitochondrial uncoupler (2,4-dinitrophenol, dNP). The aim was to better understand the impact of mitochondrial uncoupling on both responses in terms of oxidative balance, energy input (oxygen and feeding consumption) and energy output (growth and development of the tadpole). Exposure to 2,4-dNP reduced mitochondrial ROS generation, total antioxidant defences and oxidative damage in treated tadpoles compared with controls. Despite the beneficial effect of dNP on oxidative status, development and growth rates of treated tadpoles were lower than those in the control group. Treatment of tadpoles with 2,4-dNP promoted a mild mitochondrial uncoupling and enhanced metabolic rate. These tadpoles did not increase their food consumption, and thus failed to compensate for the energy loss elicited by the decrease in the efficiency of ATP production. These data suggest that the cost of ATP production, rather than the oxidative balance, is the parameter that constrains growth/development of tadpoles, highlighting the central role of energy transduction in larval performance.

Time constraints and flexibility of growth strategies: geographic variation in catch-up growth responses in amphibian larvae.

1. As size is tightly associated with fitness, compensatory strategies for growth loss can be vital for restoring individual fitness. However, immediate and delayed costs of compensatory responses may prevent their generalization, and the optimal strategy may depend on environmental conditions. Compensatory responses may be particularly important in high-latitude habitats with short growing seasons, and thus, high-latitude organisms might be more efficient at compensating after periods of unfavourable growth conditions than low-latitude organisms. 2. We investigated geographical differences in catch-up growth strategies of populations of the common frog (Rana temporaria) from southern and northern Sweden in two factorial common garden experiments involving predation risk and two different causes of growth arrest (nutritional stress and low temperatures) to evaluate how the compensatory strategies can be affected by context-dependent costs of compensation. Larval and metamorphic traits, and post-metamorphic performance were used as response variables. 3. Only northern tadpoles exposed to low food completely caught up in terms of metamorphic size, mainly by extending the larval period. Low food decreased survival and post-metamorphic jumping performance in southern, but not in northern tadpoles, suggesting that northern tadpoles have a better ability to compensate after periods of restricted food. 4. Both northern and southern tadpoles were able to metamorphose at the same size as control tadpoles after being exposed to low temperatures, indicating that consequences of variation in temperature and food availability differed for tadpoles. However, the combination of low temperatures and predation risk reduced survival in both southern and northern tadpoles. Also, predation risk decreased energy storage in both experiments. 5. Our results highlight the influence of climatic variation and the type of stressor as selective factors shaping compensatory strategies.

Seasonality determines patterns of growth and age structure over a geographic gradient in an ectothermic vertebrate.

Environmental variation connected with seasonality is likely to affect the evolution of life-history strategies in ectotherms, but there is no consensus as to how important life-history traits like body size are influenced by environmental variation along seasonal gradients. We compared adult body size, skeletal growth, mean age, age at first reproduction and longevity among 11 common frog (Rana temporaria) populations sampled along a 1,600-km-long latitudinal gradient across Scandinavia. Mean age, age at first reproduction and longevity increased linearly with decreasing growth season length. Lifetime activity (i.e. the estimated number of active days during life-time) was highest at mid-latitudes and females had on average more active days throughout their lives than males. Variation in body size was due to differences in lifetime activity among populations--individuals (especially females) were largest where they had the longest cumulative activity period--as well as to differences between populations in skeletal growth rate as determined by skeletochronological analyses. Especially, males grew faster at intermediate latitudes. While life-history trait variation was strongly associated with latitude, the direction and shape of these relationships were sex- and trait-specific. These context-dependent relationships may be the result of life-history trade-offs enforced by differences in future reproductive opportunities and time constraints among the populations. Thus, seasonality appears to be an important environmental factor shaping life-history trait variation in common frogs.

Testing the role of phenotypic plasticity for local adaptation: growth and development in time-constrained Rana temporaria populations.

Phenotypic plasticity can be important for local adaptation, because it enables individuals to survive in a novel environment until genetic changes have been accumulated by genetic accommodation. By analysing the relationship between development rate and growth rate, it can be determined whether plasticity in life-history traits is caused by changed physiology or behaviour. We extended this to examine whether plasticity had been aiding local adaptation, by investigating whether the plastic response had been fixed in locally adapted populations. Tadpoles from island populations of Rana temporaria, locally adapted to different pool-drying regimes, were monitored in a common garden. Individual differences in development rate were caused by different foraging efficiency. However, developmental plasticity was physiologically mediated by trading off growth against development rate. Surprisingly, plasticity has not aided local adaptation to time-stressed environments, because local adaptation was not caused by genetic assimilation but on selection on the standing genetic variation in development time.

Brain plasticity over the metamorphic boundary: carry-over effect of larval environment on froglet brain development.

Brain development shows high plasticity in response to environmental heterogeneity. However, it is unknown how environmental variation during development may affect brain architecture across life history switch points in species with complex life cycles. Previously, we showed that predation and competition affect brain development in common frog (Rana temporaria) tadpoles. Here, we studied whether larval environment had carry-over effects in brains of metamorphs. Tadpoles grown at high density had large optic tecta at metamorphosis, whereas tadpoles grown under predation risk had small diencephala. We found that larval density had a carry-over effect on froglet optic tectum size, whereas the effect of larval predation risk had vanished by metamorphosis. We discuss the possibility that the observed changes may be adaptive, reflecting the needs of an organism in given environmental and developmental contexts.

Predation- and competition-mediated brain plasticity in Rana temporaria tadpoles.

An increasing number of studies have demonstrated phenotypic plasticity in brain size and architecture in response to environmental variation. However, our knowledge on how brain architecture is affected by commonplace ecological interactions is rudimentary. For example, while intraspecific competition and risk of predation are known to induce adaptive plastic modifications in morphology and behaviour in a wide variety of organisms, their effects on brain development have not been studied. We studied experimentally the influence of density and predation risk on brain development in common frog (Rana temporaria) tadpoles. Tadpoles grown at low density and under predation risk developed smaller brains than tadpoles at the other treatment combinations. Further, at high densities, tadpoles developed larger optic tecta and smaller medulla oblongata than those grown at low densities. These results demonstrate that ecological interactions - like intraspecific competition and predation risk - can have strong effects on brain development in lower vertebrates.

Mobile phone mast effects on common frog (Rana temporaria) tadpoles: the city turned into a laboratory.

An experiment has been made exposing eggs and tadpoles of the common frog (Rana temporaria) to electromagnetic radiation from several mobile (cell) phone antennae located at a distance of 140 meters. The experiment lasted two months, from the egg phase until an advanced phase of tadpole prior to metamorphosis. Measurements of electric field intensity (radiofrequencies and microwaves) in V/m obtained with three different devices were 1.8 to 3.5 V/m. In the exposed group (n = 70), low coordination of movements, an asynchronous growth, resulting in both big and small tadpoles, and a high mortality (90%) was observed. Regarding the control group (n = 70) under the same conditions but inside a Faraday cage, the coordination of movements was normal, the development was synchronous, and a mortality of 4.2% was obtained. These results indicate that radiation emitted by phone masts in a real situation may affect the development and may cause an increase in mortality of exposed tadpoles. This research may have huge implications for the natural world, which is now exposed to high microwave radiation levels from a multitude of phone masts.

[Mast cells of lymph hearts during ontogenesis of frogs Rana temporaria].

Electron microscopic observations of the lymph hearts of tadpoles and yearling frogs of Rana temporaria showed that mast cells (MCs) were present not only between muscle fibers (population of resident MCs), but in the cavities of lymph heart (population of circulating MCs), too. There were some differences in the ultrastructure of the resident MCs at each studied stage of larval development. The first recognizable MCs were revealed in the lymph hearts at premetamorphosis (stages 39-41). MCs presented as mononuclear relatively small and slightly elongated cells with a few immature secretory granules and numerous free ribosomes, polysomes and short cisternae of rough endoplasmic reticulum (RER) in the cytoplasm. Chromatin of their nuclei was poorly condensed; the Golgi apparatus was moderately developed. At pro-metamorphosis (stages 44-45), we revealed MCs at different levels of their differentiation. Some MCs demonstrated an active process of granulogenesis in their cytoplasm. Among densely packed cytoplasmic organelles, immature secretory granules were closely associated with cisternae of RER and free ribosomes. Other MCs appeared as more differentiated cells. They were characterized by a predominantly heterochromatic nuclei and cytoplasm filled with polymorphic and heterogeneous granules. MCs also showed a reduction in the number of free ribosomes and cisternae of RER in the cytoplasm. On the contrary, the Golgi apparatus was well developed. Stacks of Golgi cisternae, detaching vacuoles, and progranules occupied the perinuclear region. The majority of the outlines above ultrastructural features of differentiated MCs were typical for MCs of yearling frogs. At metamorphic climax (stages 52-53), MCs often tightly contacted with macrophages. We did not reveal apoptotic MCs. However, some MCs exhibited morphological features typical for programmed necrosis-like death, which was characterized by mitochondria swelling, dilatation of cisternae of RER and nuclear envelope, plasma membrane rupture and subsequent loss of intracellular contents. Electron microscopical immunocytochemistry revealed the localization of atrial natriuretic peptide (ANP), substance S (SP) and heat shock protein (Hsp70) in the secretory granules of the resident and circulating MCs at different stages of tadpole development and in yearling frogs.

Post-metamorphic carry-over effects of altered thyroid hormone level and developmental temperature: physiological plasticity and body condition at two life stages in Rana temporaria.

Environmental stress induced by natural and anthropogenic processes including climate change may threaten the productivity of species and persistence of populations. Ectotherms can potentially cope with stressful conditions such as extremes in temperature by exhibiting physiological plasticity. Amphibian larvae experiencing stressful environments display altered thyroid hormone (TH) status with potential implications for physiological traits and acclimation capacity. We investigated how developmental temperature (Tdev) and altered TH levels (simulating proximate effects of environmental stress) influence the standard metabolic rate (SMR), body condition (BC), and thermal tolerance in metamorphic and post-metamorphic anuran larvae of the common frog (Rana temporaria) reared at five constant temperatures (14-28 °C). At metamorphosis, larvae that developed at higher temperatures had higher maximum thermal limits but narrower ranges in thermal tolerance. Mean CTmax was 37.63 °C ± 0.14 (low TH), 36.49 °C ± 0.31 (control), and 36.43 °C ± 0.68 (high TH) in larvae acclimated to different temperatures. Larvae were able to acclimate to higher Tdev by adjusting their thermal tolerance, but not their SMR, and this effect was not impaired by altered TH levels. BC was reduced by 80% (metamorphic) and by 85% (post-metamorphic) at highest Tdev. The effect of stressful larval conditions (i.e., different developmental temperatures and, to some extent, altered TH levels) on SMR and particularly on BC at the onset of metamorphosis was carried over to froglets at the end of metamorphic climax. This has far reaching consequences, since body condition at metamorphosis is known to determine metamorphic success and, thus, is indirectly linked to individual fitness in later life stages.