Bufo Calamita Research Articles

Climate Change Could Reduce the Geographic Distribution of the Natterjack Toad in Semi-Arid Regions: A 34-Year Study in Central Spain

Amphibians are globally threatened due to pollution, infection diseases, invasive species, habitat loss and climate change. Rising temperatures and shifts in precipitation regimes can have a major impact on the persistence of amphibian populations, especially in semi-arid regions. In this study, we used a long term time series spanning 34 years in central Spain to evaluate the effect of three climatic variables on the reproductive success of the natterjack toad (Epidalea calamita) in temporary ponds. Our results showed that reproductive success was only possible for seven years (20.6%). We found that reproductive success is positively related to the accumulated spring precipitation, while negatively related to mean spring temperature and spring evaporation. This study highlights the importance of conserving and restoring amphibian breeding habitats in order to minimize the potential impacts of climate change and habitat loss.

Влияние плотности посадки на личиночное развитие камышовой жабы (Epidalea calamita) (по результатам лабораторных исследований)

The Natterjack toad (Epidalea calamita) is widespread in Europe, but it is reducing in numbers, especially in the north and east of its range. The paper presents the results of studying the effect of initial stocking density on survival, development duration and body length of E. calamita larvae in laboratory conditions. The experiments used containers with the same bottom area (1092 cm2), but with different water levels (14 and 28 cm). Larvae were grown at different stocking densities per unit volume (1–3 larvae/L) of water and per unit area of the bottom (82.4–494.5 larvae/m2). The studied larvae have very high survival rate (90.7–100.0%) and show no differences in this parameter in groups in all variants of cultivation. Animals from the lowest density groups were characterized by the shortest developmental duration. With an increase in the stocking density per unit volume of water and bottom area the larval development duration increased. There was a negative correlation of body length with the initial stocking density per unit volume of water (r=-0.52) and bottom area (r=-0.62). The stocking density per the bottom area affected greater the larval developmental duration and body length of juvenile toads, than the stocking density per unit water volume. The authors note that it is advisable to grow E. calamita larvae in laboratory conditions at maximum stocking density (3 larvae/L and 494.5 larvae/m2), as this allows for a greater number of juvenile toads.

Extensive restoration of the entire habitat complex is key to the successful recovery of threatened species: The case of the natterjack toad Epidalea calamita at the northern range margin

Increasing concerns about biodiversity loss have prompted international efforts to restore degraded habitats. Unfortunately, wildlife habitat restoration often fails due to inadequate planning, inappropriate scale, or lack of providing key elements of habitats. We investigated the effectiveness of habitat restoration of a threatened amphibian species, the natterjack toad Epidalea calamita, in Estonia, comparing two different restoration approaches – partial habitat restoration and extensive habitat restoration over 16 years. Our goal was to investigate the impact of these two different approaches on the populations, to find out which habitat components play a key role in restoration and which measures are of primary importance in the restoration of habitats for small amphibian populations. We used GIS analysis to identify changes in the area of the habitat complex and breeding sites in seven natterjack toad locations following the restoration activities. To assess population response to habitat restoration, we used the census of the natterjack toad egg-strings as a proxy to the population size. The average number of egg-strings increased by 1.2-fold after partial habitat restoration and 3.5-fold when comparing partial habitat restoration with extensive restoration. Partial habitat restoration, which consisted of improving the condition of extant habitat, had no significant effect on population size (number of egg-strings), while extensive habitat restoration, which included improving and enlarging the entire habitat complex (both terrestrial and aquatic habitats as well as migration matrix between these components) led to population growth. The habitat features that positively influenced population growth were the area of entire habitat complex and the number and size of the breeding sites. In terms of habitat restoration and conservation measures, the number of natterjack toad populations was positively affected by restoration of the entire habitat complex (terrestrial habitats and water bodies), which was supplemented by the supportive rearing of tadpoles.

Groundwater decline has negatively affected the well-preserved amphibian community of Doñana National Park (SW Spain)

Abstract The success of amphibian conservation efforts is highly dependent on the preservation of amphibian breeding habitats. Doñana National Park contains an abundance of ponds with heterogeneous characteristics, which has historically favoured its amphibian community. However, most of the park’s ponds are groundwater dependent, and aquifer overexploitation outside the park is leading to shorter hydroperiods and even desiccation in the pond network. This problem has been exacerbated by the scarcity of rain over the last decade. In 2021-2022, we surveyed the occurrence of the park’s 11 amphibian species, with a view to comparing our findings with those from the last previous survey in 2003-2004. We mapped the species occurrence within the park for the two survey periods. While all 11 amphibian species could still be found in Doñana, their occurrence had decreased across the board. In 2003-2004, 6 species were present across more than 50% of the sampling area. In contrast, 18 years later, such was the case for only two species (Pelophylax perezi and Hyla meridionalis). Declines were greatest for Epidalea calamita, followed by Pelobates cultripes, Triturus pygmaeus, and Lissotriton boscai. The mean number of species per sampling unit (i.e., grid cell) dropped from 4.3 to 3.1. To preserve Doñana’s amphibian community, it is important to restore the park’s pond network, which implies reducing regional groundwater overexploitation.

Use of agricultural fields by Natterjack toads: Implications for pesticide exposure

Land sharing conservation strategies might not work if wildlife is exposed to plant protection products. Plant protection products are used to protect crops against harmful organisms yet they can also have unwanted side effects on non-target organisms. Amphibians are one group of non-target organisms for which there is evidence that plant protection products can have negative effects on individuals and populations. Despite much research on amphibian ecotoxicology, not much is known about the exposure of amphibians to plant protection products in agricultural landscapes. Here, we study habitat use and movement behaviour of an endangered amphibian, the Natterjack toad Epidalea calamita, in two study areas in Switzerland. We placed arrays of artificial cover boards in agricultural fields and adjacent non-agricultural habitats and used a photographic mark-recapture approach to track individual toads, both adults and juveniles, during and after the reproductive season in the terrestrial habitat. We used multistate and spatial mark-recapture models to analyse the data. Toads used the agricultural fields during spring and summer and set up their home ranges within the fields but there was a great turnover of individuals. Toad densities were higher in agricultural fields than other habitat types, including a nature reserve suitable for the species. Toads preferred open soils and avoided grassy meadows typical of agricultural set-asides, suggesting that the conservation of the species in agricultural landscapes requires new types of biodiversity promotion areas. The results of the study show that toads use agricultural fields during most of the growing season and are thus likely to be exposed to plant protection products.

Can neutral genetic differentiation explain geographical variation in body size of the natterjack toad, Epidalea calamita?

Population genetic studies are crucial for evolutionary biologists because the population is the basic substrate on which evolution is forged. However little empirical evidence has been able to demonstrate the role that isolation and gene flow play in maintaining differentiation in populations at short geographic scales. Epidalea calamita exhibits a steep variation in body size and reproductive traits in southwestern Spain, associated with changes in the geological substrate. This implies a decrease of 70.9% of body mass and 28.5% in snout-vent length, on a micro-geographic scale of only 60 km. Previous results from both metamorphic and juvenile common garden experiments showed that genetic differentiation may be a causal determinant of geographic variation in adult. This study tested whether neutral genetic differentiation can explain the geographical variation in the body size observed in E. calamita. It was addressed analyzing the level of genetic structuring and gene flow among populations along the cline, comparing the genetic diversity between and within populations, as well as between ecological environments. The study showed that the geographic variation in body size observed in E. calamita has evolved in absence of geographic isolation, with moderate gene flow connecting the populations. Thus, neutral genetic differentiation cannot explain the geographical variation observed. Future studies are needed on the interaction between the genetic component with the environmental factors and will be necessary to analyze the contribution of the maternal effects in the origin and evolution of the geographical variation in the body size observed in E. calamita from southern Spain.

Landscape genetics identifies barriers to Natterjack toad metapopulation dispersal

Habitat fragmentation and loss reduce population size and connectivity, which imperils populations. Functional connectivity is key for species persistence in human-modified landscapes. To inform species conservation management, we investigated spatial genetic structure, gene flow and inferred dispersal between twelve breeding sites of the Natterjack toad (Bufo calamita); regionally Red-Listed as Endangered in Ireland. Spatial genetic structure was determined using both Bayesian and non-Bayesian clustering analysis of 13 polymorphic microsatellite loci genotyping 247 individuals. We tested the influence of geographic distance, climate, habitat, geographical features, and anthropogenic pressure on pairwise genetic distances between breeding sites using Isolation-by-distance and Isolation-by-resistance based on least-cost path and circuit theory models of functional connectivity. There was clear spatial structuring with genetic distances increasing with geographic distance. Gene flow was best explained by Isolation-by-resistance models with coniferous forestry plantations, bog, marsh, moor and heath, scrub, anthropogenic presence (Human Influence Index) and rivers (riparian density) identified as habitats with high resistance to gene flow while metapopulation connectivity was enhanced by coastal habitats (beaches, sand dunes and salt marshes) and coastal grassland. Despite substantial declines in census numbers over the past 15 years and its regional status as Endangered, the Natterjack toad population in Ireland retains high genetic diversity. If declines continue, maintaining habitat connectivity to prevent genetic erosion by management of coastal grasslands, pond construction and assisted migration through translocation will be increasingly important.

Efficacy of permanent wildlife fences as barriers to amphibian movement

To mitigate habitat fragmentation and roadkill, roads are increasingly equipped with wildlife fences and underpasses. However, the effectiveness of such fences in preventing road access for amphibians has not been tested under controlled conditions. In 2019 and 2020, we tested the efficacy of full panel fences of differing material, height, and shape (presence/absence of an overhang), to prevent road access for adult and juvenile amphibians. We selected five species according to locomotion mode: Natterjack toads (runners), European green toads (short-distance jumpers), agile frogs (proficient jumpers), American tree frogs (proficient climbers) and smooth newts (climbers). We found that Natterjack and green toads were unable to cross a concrete fence with a height of 13 and 24 cm, respectively. Addition of a 10 cm overhang reduced the height required to prevent crossing further to 10 and 17 cm, respectively. The ability of these less agile species to cross a certain fence height depended on body length. By contrast, jumping agile frogs and climbing tree frogs were not stopped by the greatest fence height tested (40 cm). However, addition of the overhang stopped the climbing tree frogs at a concrete fence height of 35 cm. An alternative metal fence (with overhang) was tested with some species and performed similar to the concrete fence (with overhang). Finally, the greatest concrete fence height passed by climbing juveniles was 20 cm (smooth newts). Hence, to stop amphibians from road crossing, we recommend the construction of durable (concrete or galvanized metal) and well-maintained fences with a minimum height of 40 cm with a 10 cm overhang.

Invasive Australian swamp stonecrop (Crassula helmsii) negatively affects spawning but accelerates larval growth of the endangered natterjack toad (Epidalea calamita)

The invasive Crassula helmsii is rapidly expanding in Europe. Several ecological effects are described, most of which focus on ecosystem functioning and native vegetation but rarely on fauna. In North-western Europe, C. helmsii often invades the breeding habitat of endangered amphibians, such as Epidalea calamita. The spawning rate, egg survival and the speed of larval growth of this toad species in relation to the presence of C. helmsii were studied. In order to unravel causal mechanisms, effects on natterjack toads were related to the effects of C. helmsii presence/absence on temperature and chemical properties of the water. Spawning and egg survival were significantly lower under C. helmsii dominance compared to bare soil conditions, and negatively affected the population size of E. calamita. However, larval growth rate was significantly higher in C. helmsii dominated treatments, which could be beneficial. Differences in water temperature and chemistry were a possible explanation for these effects. It remains unclear whether the population viability of E. calamita is negatively affected when C. helmsii is present. In many areas, however, this plant species completely overgrows and causes desiccation of waterbodies. Therefore, appropriate management measures will be required to protect this toad against this invader.

Ontogeny of the middle ear and auditory sensitivity in the Natterjack toad (Epidalea calamita).

In most anuran amphibians, acoustic communication is of prime importance for mate localization and selection. The tympanic middle ear increases auditory sensitivity and directionality and is therefore expected to be favoured by natural selection. However, especially within the family of true toads (Bufonidae) there is a tendency for species to lose parts of the middle ear apparatus and consequently have a reduced sensitivity to high-frequency sounds (above 1 kHz). Part of the explanation for this may be that development of the middle ear is especially slow in bufonids, and thus the middle ear would be more likely to be lost or non-functional in paedomorphic species. However, a timeline of development of the middle ear has not been established previously. The goal of the present study was to investigate middle ear development in a toad species that has a well-known natural history and acoustic communication behaviour. We made a detailed study of anatomy and biophysics of the middle ear with measurements of auditory sensitivity across age in post-metamorphic natterjack toads (Epidalea calamita). The tadpoles and toadlets were raised in the laboratory, so their exact age was known, their auditory sensitivity was measured using auditory brainstem responses, and middle ear development and function were assessed by anatomical studies and laser vibrometry. We found that the developmental stage of the middle ear depends on the size of the toad rather than its age. The middle ear was functional at the earliest at a snout-vent length of 40 mm, which for these toads was around 500 days post-metamorphosis, close to the time of first reproduction. The functional, adult-like middle ear was shown to have 30 dB increased sensitivity to the dominant frequency of the mating call compared with sensitivities measured in newly metamorphosed individuals.

Cytogenetic Analysis in the Toad Species Bufo spinosus, Bufotes viridis and Epidalea calamita (Anura, Bufonidae) from the Mediterranean Area.

Taxonomy in Bufonidae witnessed notable transformations. Bufotes viridis and Epidalea calamita, previously included in genus Bufo, were relocated in other genera, while the genus Bufo was restricted to members of the earlier Bufo bufo group. On the other hand, Bufo bufo sensu lato now includes four species: Bufo bufo, Bufo spinosus, Bufo verrucosissimus and Bufo eichwaldi. In this study, we examined three species of three Bufonidae genera (B. spinosus, B. viridis and E. calamita) by conventional (C-banding and Ag-NOR staining) and molecular (in situ hybridization with probes for telomeric repeats and rDNA loci, and genomic in situ hybridization (GISH)) cytogenetic methods. C-banding patterns are reported for the first time for B. spinosus and E. calamita populations from Iberian Peninsula and for B. viridis from Greece, and reveal several differences with the reported C-banded karyotypes described for other European populations of these species. Silver staining shows size heteromorphisms of the signals at the Nucleolar Organizing Region (NOR). By contrast, FISH with ribosomal probes only reveal size heteromorphism of rDNA sequences in E. calamita, suggesting that the differences observed after silver staining in B. spinosus and B. viridis should be attributed to differences in chromosomal condensation and/or gene activity rather than to differences in the copy number for ribosomal genes. Regarding telomeric repeats, E. calamita is the only species with interstitial telomeric sequences (ITS) located on centromeric regions, probably originated by accumulation of telomeric sequences in the centromeric heterochromatin. Finally, we analyzed the composition and distribution of repetitive sequences by genome in situ hybridization. These experiments reveal the accumulation of repetitive sequences in centromeric regions of the three species, although these sequences are not conserved when species from different genera are compared.

Identification and characterization of a new family of long satellite DNA, specific of true toads (Anura, Amphibia, Bufonidae)

Amphibians have some of the most variable genome sizes among vertebrates. Genome size variation has been attributed to repetitive and noncoding DNA, including satellite repeats, transposable elements, introns, and nuclear insertions of viral and organelle DNA. In vertebrates, satellite DNAs have been widely described in mammals, but few molecular studies have been carried out in amphibians. Here, we provide a detailed characterization of a new family of satellite DNA, present in all 15 examined species of the family Bufonidae. Southern-blot analysis and PCR reveal that this satellite is formed by monomers of 807 bp, is organized in tandem arrays, and has an AT-content of 57.4%. Phylogenetic analyses show that most clades exhibit species-specific variances, indicating that this satellite DNA has evolved by concerted evolution. The homogenization/fixation process is heterogeneous in Bufonidae, where the genera Bufo and Bufotes do not show species-specific differences, while populations from Rhinella marina exhibit population-specific changes. Additionally, variants of this satellite DNA have been identified in Duttaphrynus melanostictus and R. marina, supporting the ‘library hypothesis’ (a set, ‘library’, of satellite DNAs is shared by a species group). Physical mapping in Bufo bufo, Bufo spinosus, Epidalea calamita and Bufotes viridis provides evidence that this repetitive DNA is not dispersed in the karyotype, but accumulated in pericentromeric regions of some chromosomal pairs. This location, together with its presence in the transcriptomes of bufonids, could indicate a role in centromere function or heterochromatin formation and maintenance.

Histological Study of Glandular Variability in the Skin of the Natterjack Toad-Epidalea calamita (Laurenti, 1768)-Used in Spanish Historical Ethnoveterinary Medicine and Ethnomedicine.

Simple SummaryCommon toads, including the natterjack toad (Epidalea calamita), have been used since ancient times for remedies, and thus constitute excellent biological material for pharmacological and natural product research. After a previous analysis of the historical-folk therapeutic use of amphibians in Spain, a histological study was carried out to provide a complementary ethnopharmacological view through the analysis of the integumentary heterogeneity of the serous (venom) and mucous glands from two adult specimens. Plastic-embedded semi-thin sections showed that serous/venom glands are cytologically homogeneous in spite of their genetic and biochemical complexity, leading to a cocktail that remains stored until extrusion. On the contrary, mucous glands, working continuously, show a more complex cytological variation and regional heterogeneity, which suggests an adaptive variability, leading to an invisible topographic map of skin toxicity. Natterjack toad-based folk remedies are usually extracted from the whole animal as a therapeutic unit in ethnoveterinary practice. However, a new ethnopharmacological vision could emerge from the study of tegumentary regional variation.Common toads have been used since ancient times for remedies and thus constitute excellent biological material for pharmacological and natural product research. According to the results of a previous analysis of the therapeutic use of amphibians in Spain, we decided to carry out a histological study that provides a complementary view of their ethnopharmacology, through the natterjack toad (Epidalea calamita). This species possesses a characteristic integument, where the parotoid glands stand out, and it has been used in different ethnoveterinary and ethnomedical practices. This histological study of their glandular variability allow us to understand the stages through which the animal synthesises and stores a heterogeneous glandular content according to the areas of the body and the functional moment of the glands. To study tegumentary cytology, a high-resolution, plastic embedding, semi-thin (1 micron) section method was applied. Up to 20 skin patches sampled from the dorsal and ventral sides were processed from the two adult specimens collected, which were roadkill. Serous/venom glands display a genetic and biochemical complexity, leading to a cocktail that remains stored (and perhaps changes over time) until extrusion, but mucous glands, working continuously to produce a surface protection layer, also produce a set of active protein (and other) substances that dissolve into mucous material, making a biologically active covering. This study provides a better understanding of the use of traditional remedies in ethnoveterinary medicine.

EDNA Metabarcoding Benchmarked towards Conventional Survey Methods in Amphibian Monitoring.

Simple SummaryAmphibian species are declining worldwide, and precise monitoring is key to ensuring timely protection and thereby ceasing deteriorating populations. Conventional monitoring methods are invasive, time-consuming, and dependent on expert knowledge. eDNA methods have been suggested as a replacement for or supplement to conventional survey methods. The present study assessed amphibian detection of conventional survey methods and eDNA metabarcoding in Danish lakes and ponds to address how the application of eDNA surveys can supplement the currently applied methodology. The study found eDNA metabarcoding to detect five out of six species detected through conventional methods. Furthermore, it is expected that the results in the present study reflect the time of sampling for the applied methods. The findings in the present study indicate that eDNA metabarcoding detects multiple Danish amphibian species and can produce knowledge on the occurrence and distribution for amphibian species. Implementing it as a supplement for conventional survey methods in nature monitoring will enable a higher frequency of monitoring and yield knowledge of species composition.A keystone in protection work is accurate and thorough the monitoring of amphibian species, and the currently applied conventional survey methods are invasive, time-consuming, and dependent on expert knowledge. Research suggests that eDNA metabarcoding is a precise and cost-efficient method that could supplement the currently applied methods. The present study assessed the efficiency of conventional survey methods and eDNA metabarcoding in terms of species richness, the average number of detected species per site, the relative frequency of species occurrence, and the similarity of applied methods. The study found eDNA metabarcoding surveys to detect Lissotriton vulgaris (smooth newt), Triturus cristatus (great crested newt), Rana arvalis (moor frog), Rana temporaria (common frog), and Bufo bufo (common toad), as well as an average of 0.9 species per site, reflecting the species composition at the time of sampling in mid-July 2020. In addition to the species mentioned above, the conventional survey detected Epidalea calamita (natterjack toad) and an average of 1.7 species per site, reflecting the species composition at the time of sampling in early June 2020. The similarity between the methods applied in the present study was 27%, thus indicating a large number of unique observations of both eDNA metabarcoding and conventional surveys. The differences in detection can most likely be explained by the time of sampling, which was conducted a month apart. eDNA metabarcoding was efficient in detecting multiple amphibian species and produced unique observations that were not detected using conventional survey methods. Applying eDNA techniques as a supplement will most likely produce important knowledge on species distribution and presence, as well as enable more frequent monitoring due to cost efficiency and disturbance.

The relationships between toad behaviour, antipredator defences, and spatial and sexual variation in predation pressure.

BackgroundAnimal behaviour is under strong selection. Selection on behaviour, however, might not act in isolation from other fitness-related traits. Since predators represent outstanding selective forces, animal behaviour could covary with antipredator defences, such that individuals better suited against predators could afford facing the costs of riskier behaviours. Moreover, not all individuals undergo equivalent degrees of predation pressure, which can vary across sexes or habitats. Individuals under lower predation pressure might also exhibit riskier behaviours.MethodsIn this work, I tested these hypotheses on natterjack toads (Epidalea calamita). Specifically, I gauged activity time, exploratory behaviour and boldness in standard laboratory conditions, and assessed whether they correlated with body size and antipredator strategies, namely sprint speed, parotoid gland area and parotoid gland colour contrast. Additionally, I compared these traits between sexes and individuals from an agrosystem and pine grove, since there is evidence that males and agrosystem individuals are subjected to greater predation pressure.ResultsSprint speed as well as parotoid gland contrast and size appeared unrelated to the behavioural traits studied. In turn, body mass was negatively related to activity time, boldness and exploration. This trend is consistent with the fact that larger toads could be more detectable to their predators, which are mostly gape unconstrained and could easily consume them. As predicted, females exhibited riskier behaviours. Nonetheless, agrosystem toads did not differ from pine grove toads in the behavioural traits measured, despite being under stronger predation pressure.

Do Active and Passive Antipredator Defences in the Toad Epidalea calamita Differ between Males and Females from Natural Habitats and Agrosystems?

Due to their assumed costs, simultaneous antipredator strategies are expected to face trade-offs, which, however, could be milder in individuals subjected to a more intense predator pressure. In this work, I studied the relationship between locomotion and parotoid glands in the natterjack toad, Epidalea calamita. Specifically, I predicted that individuals with reduced sprint speed would rely more on their chemical defences, having larger and more aposematically coloured parotoid glands. In addition, I expected this trade-off to be more evident in females and toads from pine grove habitats, because, according to previous work, males and toads from agrosystems are under greater predator pressure. However, sprint speed showed no relationship with coloration, but toads with proportionally greater parotoid glands were also proportionally faster. Thus, the costs of these antipredator traits might not be high enough to make them interfere, or the benefits of simultaneous optimisation of sprint speed and parotoid gland size might outweigh the costs of it in some individuals. In any case, habitat and sex did not affect these relationships, so the trends detected are valid across sexes and the habitats studied.

Population genetic structure of the Natterjack toad (Epidalea calamita) in Ireland: implications for conservation management

Molecular methods can play a crucial role in species management and conservation. Despite the usefulness of genetic approaches, they are often not explicitly included as part of species recovery plans and conservation practises. The Natterjack toad (Epidalea calamita) is regionally Red-Listed as Endangered in Ireland. The species is declining and is now present at just seven sites within a highly restricted range. This study used 13 highly polymorphic microsatellite markers to analyse the population genetic diversity and structure. Genetic diversity was high with expected heterozygosity between 0.55 and 0.61 and allelic richness between 4.77 and 5.92. Effective population sizes were small (Ne < 100 individuals), but not abnormal for pond breeding amphibians. However, there was no evidence of historical or contemporary genetic bottlenecks or high levels of inbreeding. We identified a positive relationship between Ne and breeding pond surface area, suggesting that environmental factors are a key determinant of population size. Significant genetic structuring was detected throughout the species’ range, and we identified four genetic entities that should be considered in the species’ conservation strategies. Management should focus on preventing further population declines and future loss of genetic diversity overall and within genetic entities while maintaining adequate local effective population size through site-specific protection, human-mediated translocations and head-start programs. The apparent high levels of genetic variation give hope for the conservation of Ireland’s rarest amphibian if appropriately protected and managed.

Sex and habitat differences in size and coloration of an amphibian's poison glands match differential predator pressures.

Chemical defenses are frequently accompanied by salient color patterns actively avoided by predators, a phenomenon referred to as aposematism. However, the production of both chemical defenses and pigments is costly, and is thus expected to be reduced under mild predator pressure. In this work, I compared the size and coloration of parotoid glands (2 dorsal, external swollen structures that secrete toxins in toads) of male and female Epidalea calamita toads from agrosystems and from pine groves. I also quantified the predator attacks received by plasticine toad models, whose "parotoid glands" differed in size and color conspicuousness, exposed in each habitat. Predators avoided models with large and conspicuous parotoid glands, but models in agrosystems were more often attacked. Concerning actual toads, agrosystem and male individuals had larger parotoid glands, presumably implying greater production of chemical defenses than in pine grove and female conspecifics. These findings are aligned with previous research suggesting that both agrosystem toads and males in this system are subjected to a more intense predator pressure. Difference between parotoid gland and dorsum coloration was greater in agrosystem toads. A marked internal pattern could function as an aposematic signal, which could counteract increased predator pressure.

A computational model approach to assess the effect of climate change on the growth and development of tadpoles

All of the environmental conditions in nature act on an organism simultaneously. However, in experimental studies of the factors influencing metamorphosis, each factor needs to be examined individually in order to disentangle its specific effects. However, it is challenging to then build properly integrated models which include data on all of the different factors evaluated in different experiments. This study set out to develop a predictive model which could synthesize the results of several experiments on survival, development and growth of Natterjack toad (Epidalea calamita) tadpole guilds. The proposed Population Dynamic P System (PDP) model enables estimates of growth and development during the larval phase, under different environmental conditions, weather conditions, predator density, and pond characteristics and management. The architecture of the model allows the inclusion of an indefinite number of parameters and interactions, with all inputs interacting in parallel, and enables solutions to complex modeling approaches. Using the model with a range of field data, we found that the importance of predation pressure on Natterjack toad tadpole guilds exceeds the potential effects of variations in temperature and precipitation. The impact of introduced invasive predators therefore arguably poses the greatest threat to this species. This type of model holds promise as a reliable management and conservation tool for this and other species, especially where interactions between environmental factors make the impacts of individual factors difficult to predict.

Breeding behaviour of ectotherms at high latitudes: the case of the natterjack toad Epidalea calamita at its northern range limit

Abstract We explored the breeding behaviour of a threatened amphibian, the natterjack toad, at its northern range limit in Estonia, to determine the extent to which reproduction is affected by harsh and unstable climatic conditions. Using photo identification of specimens, we found that in optimal weather conditions males formed three breeding cohorts, while in adverse conditions only a single cohort occurred and under extreme conditions reproduction was skipped entirely. During the extended breeding season, larger males participated in reproduction throughout the breeding period, while smaller males appeared in later cohorts. Breeding success was related to the calling effort of a male, where larger males had greater mating success than smaller ones. We found that the natterjack toad males exhibit significant plasticity in reproductive behaviour at the northern range limit, which, given the energetic cost of reproduction and the increased risk of predation, allows them to increase their fitness at high latitudes.