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Comparative studies of mortality in the wild are necessary to understand the evolution of aging; yet, ectothermic tetrapods are underrepresented in this comparative landscape, despite their suitability for testing evolutionary hypotheses. We present a study of aging rates and longevity across wild tetrapod ectotherms, using data from 107 populations (77 species) of nonavian reptiles and amphibians. We test hypotheses of how thermoregulatory mode, environmental temperature, protective phenotypes, and pace of life history contribute to demographic aging. Controlling for phylogeny and body size, ectotherms display a higher diversity of aging rates compared with endotherms and include phylogenetically widespread evidence of negligible aging. Protective phenotypes and life-history strategies further explain macroevolutionary patterns of aging. Analyzing ectothermic tetrapods in a comparative context enhances our understanding of the evolution of aging.

The following protocol is intended to respond to the requirements set by the European Union's Marine Strategy Framework Directives (MSFD) for the D10C3 Criteria reported in the Commission Decision (EU), related to the amount of litter ingested by marine animals. Standardized methodologies for extracting litter items ingested from dead sea turtles along with guidelines on data analysis are provided. The protocol starts with the collection of dead sea turtles and classification of samples according to the decomposition status. Turtle necropsy must be performed in authorized centers and the protocol described here explains the best procedure for gastrointestinal (GI) tract isolation. The three parts of the GI (esophagus, stomach, intestine) should be separated, opened lengthways and contents filtered using a 1 mm mesh sieve. The article describes the classification and quantification of ingested litter, classifying GI contents into seven different categories of marine litter and two categories of natural remains. The quantity of ingested litter should be reported as total dry mass (weight in grams, with two decimal places) and abundance (number of items). The protocol proposes two possible scenarios to achieve the Good Environmental Status (GES). First: "There should be less than X% of sea turtles having Y g or more plastic in the GI in samples of 50-100 dead turtles from each sub-region", where Y is the average weight of plastic ingested and X% is the percentage of sea turtles with more weight (in grams) of plastic than Y. The second one, which considers the food remain versus plastic as a proxy of individual health, is: "There should be less than X% of sea turtles having more weight of plastic (in grams) than food remains in the GI in samples of 50-100 dead turtles from each sub-region". C1 [Matiddi, Marco; Silvestri, Cecilia; Piermarini, Raffaella; Daffina, Roberto; Pisapia, Marco; Genta, Daniela] Italian Natl Inst Environm Protect & Res ISPRA, Rome, Italy. [deLucia, Giuseppe A.; Camedda, Andrea] Inst Coastal Marine Environm Natl Res Council IAM, Rome, Italy. [Darmon, Gaelle; Gambaiani, Delphine; Miaud, Claude] Univ P Valery, PSL Res Univ, UMR 5175 CE3FE,EPHE, CNRS,UM,IRD,INRA,SupAgro,Biogeog & Ecol Vertebras, Montpellier, France. [Tomas, Jesus; Revuelta, Ohiana] Univ Valencia, Cavanilles Inst Biodivers & Evolutionary Biol, Valencia, Spain. [Pham, Christopher K.; Vandeperre, Frederic; Rodriguez, Yasmina] Univ Acores, Dept Oceanog & Pescas, Inst Mar Okeanos, Ponta Delgada, Portugal. [Vandeperre, Frederic] Univ Acores, MARE Marine & Environm Sci Ctr, Ponta Delgada, Portugal. [Claro, Francoise; Moussier, Judicaelle] Museum Natl Hist Nat, Paris, France. [Kaska, Yakup; Sozbilen, Dogan] Pamukkale Univ, Sea Turtle Res & Applicat Ctr DEKAMER, Pamukkale, Turkey. [Kaberi, Helen; Tsangaris, Catherine] Hellenic Ctr Marine Res, Inst Oceanografy, Anavyssos, Greece. [Bradai, Mohamed N.; Chaieb, Olfa] INSTM, Salammbo, Tunisia. [Loza, Ana L.] Univ Las Palmas Gran Canaria, Las Palmas Gran Canaria, Spain.

Parasitic chytrid fungi have emerged as a significant threat to amphibian species worldwide, necessitating the development of techniques to isolate these pathogens into sterile culture for research purposes. However, early methods of isolating chytrids from their hosts relied on killing amphibians. We modified a pre-existing protocol for isolating chytrids from infected animals to use toe clips and biopsies from toe webbing rather than euthanizing hosts, and distributed the protocol to interested researchers worldwide as part of the BiodivERsA project RACE; here called the RML protocol. In tandem, we developed a lethal procedure for isolating chytrids from tadpole mouthparts. Reviewing a database of use a decade after their inception, we find that these methods have been widely applied across at least 5 continents, 23 countries and in 62 amphibian species, and have been successfully used to isolate chytrids in remote field locations. Isolation of chytrids by the non-lethal RML protocol occured in 18% of attempts with 207 fungal isolates and three species of chytrid being recovered. Isolation of chytrids from tadpoles occured in 43% of attempts with 334 fungal isolates of one species (Batrachochytrium dendrobatidis) being recovered. Together, these methods have resulted in a significant reduction and refinement of our use of threatened amphibian species and have improved our ability to work with this important group of emerging fungal pathogens.

Summary. — Life-history variation within and among populations of widespread amphibian species has attracted much attention so far. In relation to a variety of problems (e. g. climate change, large-scale clines, conservation), this information is needed not only on broad geographic scale, but also at local level and from year to year. In this study, we investigated variation in body size and life history traits, including age, fecundity and egg size, in two neighbouring populations of a widespread amphibian, the Common Toad Bufo bufo, during a 3-year period. Body size varied significantly between populations in males only, but not between years. Age was assessed by skeletochronology, and osteometrical analyses revealed a complete endosteal resorption of the first line of arrested growth in 1.1 % of the individuals and a partial resorption in 77 % of the individuals. Age structures did not differ significantly between sexes, populations, nor between years. Age at first reproduction was one year later in females than in males in both populations (4 vs 3 years), but adult survival rates did not differ between sexes. Fecundity did not differ between populations or years, but egg size varied between years. Fecundity increased with size, but not with age of females. This lack of significant interpopulation variation in age structure and reproductive traits is important for monitoring local populations which can be exposed to various potential disturbance factors. On the other hand, these results stress the importance of analysing temporal variation in life history traits because one-year data may not represent properly the reproductive features of a population, and this issue is becoming more so important with the growing interest in climate changes and its possible effects on amphibian populations., Résumé. — Variation interannuelle des traits d’histoire de vie entre des populations voisines de l’amphibien à grande aire de répartition, Bufo bufo.— Les variations intra-et interpopulationnelles des traits d’histoire de vie chez les amphibiens à grande aire de répartition font l’objet d’une attention soutenue. Pour de nombreux problèmes (e. g. les changements climatiques, variations clinales, conservation), cette information est nécessaire, non seulement à une large échelle géographique, mais aussi au niveau local et d’une année à une autre. Dans cette étude nous avons étudié durant trois années successives les variations de la taille corporelle, de l’âge et des traits d’histoire de vie, y compris la fécondité et la taille des oeufs, dans deux populations voisines du Crapaud commun Bufo bufo, une espèce à grande aire de répartition. La taille corporelle des mâles varie significativement entre les populations mais pas d’une année à une autre. L’âge a été déterminé par squelettochronologie et les analyses ostéologiques révèlent une résorption endostéale complète de la première ligne d’arrêt de croissance chez 1,1 % des individus et une résorption partielle chez 77 % des individus. Les structures d’âge ne diffèrent pas entre les sexes, les populations, ni entre les années. Les femelles se reproduisent pour la première fois un an après les mâles dans les deux populations (4 vs 3 ans), mais le taux de survie des adultes ne diffère pas entre les sexes. La fécondité ne varie pas entre les populations ou les années, mais la taille des oeufs change d’une année à une autre. La fécondité augmente avec la taille, mais pas avec l’âge des femelles. Cette absence de variation de la structure d’âge et des paramètres de la reproduction est importante pour la surveillance des populations locales qui peuvent être exposées à divers facteurs de perturbation potentiels. Ces résultats soulignent également l’importance d’analyser les variations temporelles des traits d’histoire de vie car les données d’une année peuvent ne pas représenter correctement les caractéristiques de la reproduction d’une population, cette question étant exacerbée par l’intérêt croissant sur l’impact possible des changements climatiques sur les populations d’amphibiens., Toma Šević Nataša,Cvetkoviović Dragana,Miaud Claude,Aleksic Ivan,Crnobrnja-Isailović Jelka. Interannual variation in life history traits between neighbouring populations of the widespread amphibian Bufo bufo. In: Revue d'Écologie (La Terre et La Vie), tome 63, n°4, 2008. pp. 371-381.

[Departement_IRSTEA]Eaux [TR1_IRSTEA]QUASARE; International audience; Global biodiversity in freshwater and the oceans is declining at high rates. Reliable tools for assessing and monitoring aquatic biodiversity, especially for rare and secretive species, are important for efficient and timely management. Recent advances in DNA sequencing have provided a new tool for species detection from DNA present in the environment. In this study, we tested whether an environmental DNA (eDNA) metabarcoding approach, using water samples, can be used for addressing significant questions in ecology and conservation. Two key aquatic vertebrate groups were targeted: amphibians and bony fish. The reliability of this method was cautiously validated in silico, invitro and insitu. When compared with traditional surveys or historical data, eDNA metabarcoding showed a much better detection probability overall. For amphibians, the detection probability with eDNA metabarcoding was 0.97 (CI=0.90-0.99) vs. 0.58 (CI=0.50-0.63) for traditional surveys. For fish, in 89% of the studied sites, the number of taxa detected using the eDNA metabarcoding approach was higher or identical to the number detected using traditional methods. We argue that the proposed DNA-based approach has the potential to become the next-generation tool for ecological studies and standardized biodiversity monitoring in a wide range of aquatic ecosystems. see also the Perspective by Hoffmann, Schubert and Calvignac-Spencer.

High-altitude lakes are vulnerable ecosystems that require protection and sustainability management, although their overallfunctioning is still poorly understood. In France protected area managers and scientists are cooperating to address this problem.Their results show the huge diversity of these altitude lakes and imply specificities in their functioning and in the way they respondto stressors. Multidisciplinary studies on these ecosystems, on their individual history and the stressors they face, need to be developedin a number of lakes, alongside long-term monitoring surveys. This is the objective of the Sentinel lakes network.

International audience; Landscape connectivity is a key issue for biodiversity conservation. Many species have to refrain to move between scattered resources patches. This is particularly the case for the common frog, a widespread amphibian migrating between forest and aquatic habitats for breeding. Face to the growing need for maintaining connectivity between amphibians' habitat patches, the aim of this study is to provide a method based on habitat suitability modelling and graph theory to explore and analyze ecological networks. We first used the maximum entropy modelling with environmental variables based on forest patches distribution to predict habitat patches distribution. Then, with considerations about landscape permeability, we applied graph theory in order to highlight the main habitat patches influencing habitat availability and connectivity by the use of the software's Conefor Sensinode 2.2 and Guidos. The use of the JRC Forest/Non Forest European map for the characterisation of common frog terrestrial habitat distribution combined with the maximum entropy modelling gives promising results for the identification of habitat discontinuities within a regional perspective. This approach should provide an operational tool for the identification of the effects of landscape barriers and corridors on populations structure. Then, the method appears as a promising tool for landscape planning.

International audience; Landscape connectivity is considered a key issue for biodiversity conservation and for the maintenance of natural ecosystems stability and integrity. Landscape connectivity defines the degree to which the landscape facilitates or impedes movement among resource patches. A wide range of methodological approaches can be involved in such studies. Spatial distribution analyses are common tools but can hardly integrate connectivity. We do here suggestions to apply graph theory and least cost path approaches in a specific application related to common frog habitats connectivity. Amphibian's life cycle involve seasonal migrations between terrestrial and aquatic habitats which constrain them to regularly cross an inhospitable fragmented landscape matrix. Thus, there is a growing need for maintaining and restoring landscape connectivity between their habitat patches. This is especially the case for the common frog Rana temporaria, a widespread amphibian in Europe occurring in various habitat types and migrating between forest and aquatic habitats for breeding. The aim of preliminary study is to explore a method based on habitat suitability modeling and graph theory in order to analyze an ecological network. In order to assess in which manner habitat patches distribution can affect landscape connectivity between ponds, we use both configuration and distributions of suitable forest patches as model inputs. The link between common frog occurrence and forest patches configuration and distribution is defined with a probabilistic model from sampled data and relevant indices. Especially, elevation, land use distribution, distances to forest patches, distance to rivers, and landscape indices computed from forest patches distribution were shown being the main significant environmental variables influencing habitat patches distribution. In our application, we obtained then a suitable habitat patches distribution map by the use of ponds occupancy location data and maximum entropy modelling. Then, we applied least cost path modelling and graph theory approach in order to highlight the connected ponds and their importance for regional connectivity. These results emphasize the potential of maximum entropy modelling, and graph theory approach for integrating connectivity in landscape planning. The quantification of landscape matrix permeability in relation with the common frog dispersion patterns appears as limited in order to quantify edge between nodes for the design of a graph integrating ponds as nodes for a regional perspective. Nevertheless, this method combined with the use of genetic markers may be useful to assess main barriers and corridors for the common frog from a regional to a local perspective for planning. In this context, the use of genetic distances could be considered as a good surrogate to the use of least cost path as edges in a graph theoretical approach for studying connectivity.

International audience; Propagule pressure is considered the main determinant of success of biological invasions: when a large number of individuals are introduced into an area, the species is more likely to establish and become invasive. Nevertheless, precise data on propagule pressure exist only for a small sample of invasive species, usually voluntarily introduced. We studied the invasion of the American bullfrog, Rana catesbeiana, into Europe, a species that is considered a major cause of decline for native amphibians. For this major invader with scarce historical data, we used population genetics data (a partial sequence of the mitochondrial cytochrome b gene) to infer the invasion history and to estimate the number of founders of non-native populations. Based on differences between populations, at least six independent introductions from the native range occurred in Europe, followed by secondary translocations. Genetic diversity was strongly reduced in non-native populations, indicating a very strong bottleneck during colonization. We used simulations to estimate the precise number of founders and found that most non-native populations derive from less than six females. This capability of invasion from a very small number of propagules challenges usual management strategies; species with such ability should be identified at an early stage of introduction.

International audience; We estimated the age, size and growth patterns of a mountain ranid, Rana holtzi, from Central Anatolia, using skeletochronology. We used lines of arrested growth (LAGs) recorded in phalanges to estimate the age of juveniles and adults. Results showed that age at maturity in this population was 4-5 years (median age of juveniles was 3.5 yrs and minimum age for adults was 4.0 yrs for both sexes). Median ages of males and females were similar (6 yrs) and longevity reached 8.0 yrs for males and 10.5 yrs for females. Mean body and tibia length of males was significantly larger than that of females. Growth of the Anatolian mountain frog was well described by the von Bertalanffy model, and males and females did not differ in growth parameters. These demographic parameters are compared with those obtained with other amphibians, especially ranids. The sexual size dimorphism observed-larger males than females-has rarely been observed previously and should prompt observers to collect other biological information on this rare endemic species.

International audience; We used skeletochronology to determine the ages of 149 (74 females, 44 males, and 31 juveniles) Mountain Yellow-legged Frogs (Rana muscosa) from 13 locations (elevation 1509-3501 m) throughout their current range in the Sierra Nevada mountains of California. Lines of arrested growth (LAGs) from excised toe bones were distinct in these high elevation frogs, and each LAG was assumed to represent one year of age. Females ranged in age from 0-10 years (mean = 4.1 years) and males from 0-8 years (mean = 4.0 years). The skeletochronological age was that of the post-metamorphic frog and did not include the tadpole stage. Mountain Yellow-legged Frogs spend 3-4 years as tadpoles, but no age markers are found in their cartilaginous skeletons; thus, their total age, if both tadpole and post-metamorphic stages were included, would range up to 14 years. Females were significantly longer (snout-vent length: SVL) than males and had greater mean mass, but there was no difference in the mean ages. juvenile frogs of unknown sex ranged in age from 0-3. The von Bertalanffy growth curve demonstrated that female SVLs were larger than males for all ages. Using a semi-parametric growth model, we also found that elevation within the Sierra Nevada range was an important variable in the relationship between SVL and age; frogs from lower elevation sites were consistently larger at a given age when compared to higher elevation sites. For each increase of 1000 or in elevation, the estimated length (on average) decreases by 8.7 mm. This is the first age determination study of a Sierra Nevada amphibian, and compared to other anuran species, Mountain Yellow-legged Frogs were found to be relatively long-lived, which will have implications for restoration and recovery plans.

ISI Document Delivery No.: 821GR Times Cited: 6 Cited Reference Count: 14 English Article 0268-0130; International audience; This study describes a non-destructive DNA sampling method for genetic studies on amphibians using buccal swabs. We assessed the quantity and quality of DNA collected in each species by amplifying a part of the cytochrome b gene (381-1060 bp) and microsatellite markers. Buccal swab sampling is a useful alternative method for DNA sampling for both mtDNA and nDNA markers in amphibians. However, only frozen storage allowed microsatellite genotyping. We conclude that this method could greatly increase the accessibility of genetic studies in small vertebrates and could be preferred in the field of conservation genetics.

Times Cited: 5; International audience; The black salamander, Salamandra lanzai, exists in isolated and fragmented populations in the Cottian Alps. To examine the degree of intra- and interpopulation divergence, mitochondrial cytochrome b sequences were obtained from samples representing all populations of S. lanzai. No genetic variation was observed either within or among populations. The presence of a single lineage suggests historical explanations closely related to Pleistocene events.

Times Cited: 11; International audience; Fifty individuals of the endemic Alpine salamander, Salamandra atra, representing 13 populations throughout the range of the two currently recognized subspecies, atra and aurorae, were examined for sequence variation in a large portion (1050 bp) of the mitochondrial cytochrome b gene. We revealed a large number of mitochondrial DNA (mtDNA) haplotypes (10). Interpopulation sequence divergence was very low, ranging from 0 to 3.1%. The relationships among haplotypes were poorly resolved. The divergence time estimate between several mtDNA haplotypes suggested a pre-Pleistocene differentiation approximately 3 million years ago. Moreover, the impact of the Pleistocene glaciations on the phylogeographical patterns appears to have been secondary, although a somewhat reduced genetic variability was found in populations living in areas that were directly affected by the glaciation.

International audience

International audience; We studied a population of the alpine newt, Triturus alpestris, living at the upper limit of its altitudinal range (2200 m in the northwestern Alps) during the aquatic activity period. Age was estimated by skeletochronology applied to phalanges and body size was measured. Age structures were similar between sexes, with age at maturity at 9-11 years and longevity up to 20 years. Annual survival was 0.81. Females were significantly larger than males. Growth, modelled according to van Bertalanffy's growth model, was higher in females during the juvenile period. A comparative analysis of age and size at maturity, mean size, and longevity in several European populations distributed from sea level to high altitudes shows a significant increase in these phenotypic variables with the decrease of the duration of the activity period. The growth coefficient IC indicating the shape of the curve, is smaller in highland than in lowland populations, and the asymptotic size is smaller in the plain than at higher altitude. If adult size increases with altitude, age at maturity is extensively delayed in short activity period environments. The discussion is focussed on the patterns of variation of these traits. However, age and size of newts can also depend on traits of the previous stages of the life cycle, and larval stages in response to environmentalgradients.

La technique squelettochronologique est appliquée pour déterminer l'âge d'adultes #Triturus helveticus$, #Triturus alpestris$ et #Triturus cristatus$ capturés dans leurs sites de reproduction dans le sud-est de la France. L'os utilisé pour cette méthode histologique est une phalange pour les deux plus grande espèces (#T. c$ et #T. a$) et un humérus pour la plus petite (#T. h$). Les coupes transversales réalisées ont 15 à 20 microns d'épaisseur et sont colorées à l'hématoxyline de Ehrlich. Les lignes d'arrêt de croissance (LAC) sont bien visibles et l'âge peut être estimé pour 100ù des #T. helveticus$, 84% des #T. cristatus$ et 75% des #T. alpestris$... La faisabilité de la méthode (utilisation de différents os), la précision des estimations des âges individuels et des problèmes de lecture des LAC (lignes effacées ou surnuméraires, résorption endostéale) sont discutés. Les structures d'âge de différentes populations et de différentes espèces de #Triturus$ en Europe, obtenues à l'aide de squelettochronologie, sont comparées

Batrachochytrium dendrobatidis is a pathogen infecting amphibians at the global scale and causing their decline, but knowledge of the distribution of this pathogen is far from complete. We sampled amphibians from three species (Hyla intermedia, Rana dalmatina and Pelophylax synklepton esculentus) to evaluate whether B. dendrobatidis infects amphibians in the Po River Delta Natural Park, Northern Italy. We detected the pathogen in one population of P. sk. esculentus (prevalence: 0.33). These findings expand the known distribution of B. dendrobatidis in Italy and add further concern to the conservation of amphibians in this area., Acta Herpetologica, Vol 6 No 2 (2011)

International audience; Several experiments have shown that ambient ultraviolet-B radiation (UV-B) has negative effects on the development of amphibians'embtyos. We studied the effects of UV-B radiation on development, survival and frequency of deformity during egg development in the Natterjack toad (Bufo calamita) from a semiarid region of Lleida (Catalonia, Spain). Eggs exposed to ambient levels of UV-B and those protected from UV-B with a filter exhibited similar developmental rate, mortality rate and frequency of developmental anomalies. These experiments show that eggs of Bufo calamita of the studied population are able to develop normally during embryonic period when exposed to current high levels of UV-B observed in Catalonia. These results will be used as reference for future studies on geographic variation in UV-B tolerance in this species.

Biological invasions and land-use changes are two major causes of the global modifications of biodiversity. Habitat suitability models are the tools of choice to predict potential distributions of invasive species. Although land-use is a key driver of alien species invasions, it is often assumed that land-use is constant in time. Here we combine historical and present day information, to evaluate whether land-use changes could explain the dynamic of invasion of the American bullfrog Rana catesbeiana ( 5Lithobathes catesbeianus) in Northern Italy, from the 1950s to present-day. We used MAXENT to build habitat suitability models, on the basis of past (1960s, 1980s) and present-day data on land-uses and species distribution. For example, we used models built using the 1960s data to predict distribution in the 1980s, and so on. Furthermore, we used land-use scenarios to project suitability in the future. Habitat suitability models predicted well the spread of bullfrogs in the subsequent temporal step. Models considering land-use changes predicted invasion dynamics better than models assuming constant land-use over the last 50 years. Scenarios of future land-use suggest that suitability will remain similar in the next years. Habitat suitability models can help to understand and predict the dynamics of invasions; however, land-use is not constant in time: land-use modifications can strongly affect invasions; furthermore, both land management and the suitability of a given land-use class may vary in time. An integration of land-use changes in studies of biological invasions can help to improve management strategies.