Search

1. Wetland ecosystems worldwide are threatened by habitat alteration, climate change and the introduction of invasive species, even within protected areas. Unravelling the reliance of sensitive wetland-dwelling species, such as amphibians, on habitat characteristics is thus essential to identify conservation targets. 2. Here we assess the distribution of genetic diversity of two strongly aquatic amphibians (Pelophylax perezi and Pleurodeles waltl) in association with habitat features across the most extensive, protected wetland of the Iberian Peninsula: Doñana National Park. 3. Despite inhabiting a protected area free from anthropogenic barriers, the genetic diversity of P. perezi and P. waltl is not homogeneously distributed across the wetland, but instead concentrates in core areas, mainly in the northern zone. Both genetic diversity and connectivity (as opposed to genetic differentiation) showed significant positive associations with the area of the breeding sites and the flooded area surrounding the breeding sites within the dispersal potential of either species, that is nearby pond availability. 4. Large water bodies connected to abundant temporary ponds are key for the maintenance of amphibian genetic diversity. Nevertheless, the core populations of our target species, which show markedly aquatic habits, are concentrated in areas colonised by invasive species, which could compromise their long-term viability. 5. Our results highlight that maintaining widely connected arrays of ponds of different hydroperiods, including large breeding sites free from invasive predators and competitors, is paramount for amphibian conservation in Mediterranean wetlands.

Phenotypic plasticity allows organisms to improve the match between their phenotype and heterogeneous environments. Theoretical models have argued that costs of maintaining the sensory and response machinery necessary for adaptive phenotypic plasticity are important determinants to the evolution of plasticity. Despite recurrent arguments invoking putative metabolic costs associated with maintenance of cellular machinery, no studies have yet attempted to quantify it from a molecular standpoint. Here we experimentally examine physiological differences across genotypes (sibships) of spadefoot toad larvae with different degrees of plasticity in response to predator cues. We observed marked differences across sibships in developmental, growth and morphological responses to predators, and tested whether increased plasticity was associated with oxidative stress or immune suppression. We observed that more plastic sibships experienced higher antioxidant enzymatic activity when reared in the absence of predator cues, i.e. not expressing their plastic responses. The degree of plasticity was also associated with higher lipid peroxidation and slightly greater granulocyte-to-lymphocyte ratio. Higher antioxidant activity in highly plastic sibships suggests that maintenance of phenotypic plasticity may be linked to energy demanding metabolic processes. Our findings suggest that having the potential to produce plastic responses may incur oxidative and immunological costs. In the long term, such maintenance costs may erode individual fitness and even constrain the evolution of plasticity. To our knowledge, this is the first empirical evidence indicating the existence of a physiological cost to the maintenance of phenotypic plasticity.

Among invasive species, floating plants have a great impact on aquatic ecosystems, forming dense mats that cause a great alteration of the physical and chemical water features. We experimentally assessed the effect of Azolla filiculoides mats on Mediterranean temporary waters using mesocosms with sediments from an invaded marsh and amphibian larvae from surrounding wetlands. Azolla filiculoides formed a dense mat over the water surface in the experimental tanks, which decreased pH and oxygen concentration, and increased nutrients, nitrogen and phosphorus compounds in the water compared to mesocosms lacking the exotic fern. Macrophyte abundance and richness were reduced under the Azolla mat. Invaded mesocosms also had higher phytoplankton abundance and different zooplankton composition, which was mainly characterized by higher abundance of juvenile copepods. Regarding amphibian development, tadpoles requiring a long developmental period, the western spadefoot toad, Pelobates cultripes, had significantly lower survival in the presence of A. filiculoides (4.8% compared to 60% survival in fern-free mesocosms). For tadpoles of the Mediterranean tree frog, Hyla meridionalis, which develop faster, we did not detect effects on survival, but on body size and physical condition of metamorphs. The formation of A. filiculoides mats produced a deterioration of the trophic web, as it caused most sources to be depleted in 13C and 15N. Primary consumers, tadpoles and zooplankton, were also 13C depleted, although the diet of tadpoles from Azolla mesocosms was mainly composed of the invasive fern. We therefore conclude that A. filiculoides has a negative impact on physical and chemical water features, reduces the richness and biomass of the submerged macrophyte guild and reduces survival and body condition of tadpoles. Overall, A. filiculoides causes substantial deterioration of the food web of the aquatic ecosystem.

In temporary ponds, seed germination largely determines how well aquatic plant assemblages recover after dry periods. Some aquatic plants have terrestrial morphotypes that can produce seeds even in dry years. Here, we performed an experiment to compare germination patterns for seeds produced by aquatic and terrestrial morphotypes of Ranunculus peltatus subsp. saniculifolius over the course of five inundation events. During the first inundation event, percent germination was higher for terrestrial morphotype seeds (36.1%) than for aquatic morphotype seeds (6.1%). Seed germination peaked for both groups during the second inundation event (terrestrial morphotype: 47%; aquatic morphotype: 34%). Even after all five events, some viable seeds had not yet germinated (terrestrial morphotype: 0.6%; aquatic morphotype: 5%). We also compared germination patterns for the two morphotypes in Callitriche brutia: the percent germination was higher for terrestrial morphotype seeds (79.5%) than for aquatic morphotype seeds (41.9%). Both aquatic plant species use two complementary strategies to ensure population persistence despite the unpredictable conditions of temporary ponds. First, plants can produce seeds with different dormancy periods that germinate during different inundation periods. Second, plants can produce terrestrial morphotypes, which generate more seeds during dry periods, allowing for re-establishment when conditions are once again favorable.

In temporary ponds, seed germination largely determines how well aquatic plant assemblages recover after dry periods. Some aquatic plants have terrestrial morphotypes that can produce seeds even in dry years. Here, we performed an experiment to compare germination patterns for seeds produced by aquatic and terrestrial morphotypes of Ranunculus peltatus subsp. saniculifolius over the course of five inundation events. During the first inundation event, percent germination was higher for terrestrial morphotype seeds (36.1%) than for aquatic morphotype seeds (6.1%). Seed germination peaked for both groups during the second inundation event (terrestrial morphotype: 47%; aquatic morphotype: 34%). Even after all five events, some viable seeds had not yet germinated (terrestrial morphotype: 0.6%; aquatic morphotype: 5%). We also compared germination patterns for the two morphotypes in Callitriche brutia: the percent germination was higher for terrestrial morphotype seeds (79.5%) than for aquatic morphotype seeds (41.9%). Both aquatic plant species use two complementary strategies to ensure population persistence despite the unpredictable conditions of temporary ponds. First, plants can produce seeds with different dormancy periods that germinate during different inundation periods. Second, plants can produce terrestrial morphotypes, which generate more seeds during dry periods, allowing for re-establishment when conditions are once again favorable.

In temporary ponds, submerged plants have adapted to recurrent periods of drought. These ponds often lack the impact of large herbivores but can harbor large numbers of herbivorous tadpoles. Tadpoles of the Iberian spadefoot toad (Pelobates cultripes) may reach large body size and consume large quantities of macrophytes during their long larval period, which is shortened with risk of pond drying. We conducted a mesocosm experiment to analyze the interplay between hydroperiod and impact of herbivorous tadpoles on the abundance and life history of 3 species of submerged plants common to temporary ponds. We observed differences in growth among plant species resulting in lags in timing of maximum cover, probably related to interspecific competitive interactions. Some plants responded to pond drying with a remarkable increase in the production of flowers. Tadpoles greatly reduced the plant cover, although their impact varied across plant species. Differential plant consumption by these large tadpoles can substantially modify the internal structure and complexity of temporary ponds, and tadpoles can also interfere with plant responses to shortened hydroperiod.

This dataset consists of a single excel file split into distinct sheets for different types of data: climate, population genetics, stable isotopes, standard metabolic rate, skeletochronology, telomere length, male advertisement calls, and female preference tests. The climatic data is split into sheets for NDVI, Worldclim Bio19, and edaphology. The population genetics data is split into sheets for basic statistics, genetic differentiation indicators, isolation by distance, and effective population size.

Telomere shortening with age has been documented in many organisms, butfew studies have reported telomere length measurements in amphibians, andno information is available for growth after metamorphosis, nor in wild popu-lations. We provide both cross-sectional and longitudinal evidence of nettelomere attrition with age in a wild amphibian population of natterjacktoads (Epidalea calamita). Based on age-estimation by skeletochronology andqPCR telomere length measurements in the framework of an individual-based monitoring programme, we confirmed telomere attrition in recapturedmales. Our results support that toads experience telomere attrition throughouttheir ontogeny, and that most attrition occurs during the first 1–2 years. We didnot find associations between telomere length and inbreeding or body con-dition. Our results on telomere length dynamics under natural conditionsconfirm telomere shortening with age in amphibians and provide quantifi-cation of wide telomere length variation within and among age-classes in awild breeding population.

Ample variation in body size is common in vertebrates over extensive geographical distances, or in isolated populations, where effective geographical barriers may cause dwarfism or gigantism. Here we study potential causes of extreme size reduction in continental populations of amphibians within a short geographical distance and in the absence of geographical barriers. Natterjack toads Epidalea calamita in Doñana National Park (Spain) experience up to 2.1-fold difference in body mass in as little as 37 km. Studying six populations divergent in body size, we tested for genetic isolation of the dwarf populations using multilocus genotypes (16 microsatellites), and explored whether populations differ in trophic status (through stable isotope analysis), standard metabolic rate and growth pattern, senescence and age structure (conducting telomere length assays and skeletochronology). We also recorded advertisement calls across populations and experimentally tested for behavioural reinforcement of the body size variation through female preferences. Local dwarfism in these populations occurs in the absence of genetic isolation while maintaining relatively high effective population sizes. Dwarf populations, however, are exposed to drier and warmer climatic conditions, have different trophic status, show lower mass-specific metabolic rate, and male advertisement calls with a higher dominant frequency. Juvenile growth differed among populations, reaching the adult stage at different body sizes. Altogether, our results suggest a significant influence of environmental conditions on the physiology and ecology of the Doñana E. calamita populations, mainly affecting toads between metamorphosis and sexual maturity. Further experimental and genomic studies focusing on these early life stages are necessary to dissect the relative roles of the environment and adaptive genetic differentiation on this phenomenon.

Understanding the movement of individuals across natural habitats is important to determine fundamental aspects of their ecology, from dispersal patterns and habitat selection to interactions with predators, pathogens or conspecifics. Amphibian larvae are key to aquatic systems, playing fundamental roles as both consumers and prey, affecting nutrient cycling and even physico-chemical properties of the water bodies they occupy. Nevertheless, despite an extensive body of literature on most other aspects of their ecology, we know very little about spatial movements of amphibian larvae in their natural habitats, and the little information we have is restricted to lotic systems. Here we use a mark-and-recapture procedure using elastomers to determine movement patterns and microhabitat selection of western spadefoot toad tadpoles (Pelobates cultripes) in two large ponds in southwestern Spain. We observed maximum distances moved by these tadpoles of >67 m and >245 m, depending on the pond. These distances coincided with the maximum cross distance in each pond, suggesting that they could potentially move even farther in larger wetlands. Mean distances moved were 29.7 and 105 m in each pond. We sampled at two, four and nine days after release of tagged individuals, and we observed a stable, not expanding, movement pattern across sampling dates, suggesting that tadpoles resumed normal activity immediately after release. Tadpoles showed preference for deeper and more densely vegetated areas of the ponds, especially associated with plant species such as Ranunculus peltatus and Potamogeton pectinatus

Non-native turtles are susceptible to pathogenic infections that may be transmitted to native species. We performed hematological, biochemical, histopathological, and microbiological analyses in two invasive populations of red-eared sliders (Trachemys scripta elegans), which were living alongside native turtle species in southwestern Spain. The red-eared sliders that were captured had a healthy external appearance. However, the histopathological analyses revealed that up to 88% of these turtles had internal pathologies. The most common were hepatic lipidosis and chronic nephritis, which frequently co-occurred with each other or with pulmonary or pancreatic lesions. A high proportion of turtles were susceptible to infections caused by common bacteria in these habitats. We detected Herpesvirus, Mycoplasma spp. and more than 18 Gram-negative bacteria. The high prevalence of disease recorded in the two populations suggests that red-eared sliders are poorly suited to the conditions in their non-native range.

Invasive species have major impacts on biodiversity and are one of the primary causes of amphibian decline and extinction. Unlike other top ant invaders that negatively affect larger fauna via chemical defensive compounds, the Argentine ant (Linepithema humile) does not have a functional sting. Nonetheless, it deploys defensive compounds against competitors and adversaries. We estimated levels of ant aggression toward 3 native terrestrial amphibians by challenging juveniles in field ant trails and in lab ant foraging arenas. We measured the composition and quantities of toxin in L. humile by analyzing pygidial glands and whole-body contents. We examined the mechanisms of toxicity in juvenile amphibians by quantifying the toxin in amphibian tissues, searching for histological damages, and calculating toxic doses for each amphibian species. To determine the potential scope of the threat to amphibians, we used global databases to estimate the number, ranges, and conservation status of terrestrial amphibian species with ranges that overlap those of L. humile. Juvenile amphibians co-occurring spatially and temporally with L. humile die when they encounter L. humile on an ant trail. In the lab, when a juvenile amphibian came in contact with L. humile the ants reacted quickly to spray pygidial-gland venom onto the juveniles. Iridomyrmecin was the toxic compound in the spray. Following absorption, it accumulated in brain, kidney, and liver tissue. Toxic dose for amphibian was species dependent. Worldwide, an estimated 817 terrestrial amphibian species overlap in range with L. humile, and 6.2% of them are classified as threatened. Our findings highlight the high potential of L. humile venom to negatively affect amphibian juveniles and provide a basis for exploring the largely overlooked impacts this ant has in its wide invasive range.

In Mediterranean temporary ponds, the timing of annual flooding is highly variable and depends on heavy seasonal rains. Ponds can flood in the autumn, winter, or spring, and thus the environmental conditions faced by emerging zooplankton can be very different. We performed an experiment in a climatic chamber where we simulated annual variation in natural temperature and light conditions to study how differences in pond-filling season affected zooplankton assemblage composition. We sampled sediments from a temporary pond and placed them in aquariums that were filled with water during three different seasons: autumn (October), winter (January), and spring (March). Zooplankton abundance, species richness, diversity, and assemblage composition differed significantly among treatments, and post-inundation temperature and pH appeared to be the main drivers of these differences. Diversity was highest in the winter treatment. It was lower in the autumn treatment and the spring treatment, and no indicator species were present in the latter. Our results suggest that interannual variability in initial inundation conditions favours the emergence of different species and thus contributes to high species richness in the egg bank. However, climate change and/or groundwater drawdown could delay pond flooding, impoverishing the zooplankton assemblage in the long term.