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Abstract Predator–prey interaction is a major force driving natural selection. Yet, the identification of species preying on, or consuming, aposematic species is largely unknown. Here, I conduct a study evaluating the role of the exotic Rattus rattus as a consumer and possible predator of the aposematic and toxic Salamandra salamandra. I used camera traps to investigate the response of R. rattus towards S. salamandra carcasses in two insular populations, Ons and San Martiño (NW Spain), which show remarkable contrasting behaviour (nocturnal vs. diurnal activity) and demographic and phenotypic differences. This study unveils R. rattus consumes S. salamandra despite its aposematic colour pattern and toxicity. The high number of salamander carcasses consumed or taken by rats throughout each island (90%–100%) and the lack of other possible predator–prey interactions points to R. rattus as an efficient consumer of S. salamandra in these insular environments, which might exert a high predation pressure on both islands. Yet, the drivers underlying the behavioural and phenotypic differences in these insular populations should be further investigated.

Abstract Aquatic environmental DNA is increasingly used for biodiversity monitoring, such as surveying threatened and invasive species. Mainstreaming these methods in practical applications, however, still requires significant standardization and optimisation, namely regarding DNA capture methods. Here we evaluated how filter type (standard disc filters vs high‐capacity capsules), number of sampling sites, volume of water filtered and environmental factors affected amphibian detection in Mediterranean temporary ponds. The study involved water filtering until clogging at one (capsules) and five (discs) sites from 16 small and shallow ponds, where three urodele and seven anuran species were recorded through sweep‐netting and adult observations. Detection probabilities were estimated from site occupancy models based on replicate sampling and from an adaptation of time‐to‐detection models relating detection probability to volume of water filtered. Discs filtered relatively small volumes (15–1250 mL), with detection probabilities of the two abundant species (Pelobates cultripes, Hyla meridionalis) increasing rapidly with sample size and water volume, reaching almost perfect detection (0.95) at four and seven discs, and 420 mL and 1860 mL, respectively. However, reaching high detection probabilities for rare species (Pelodytes atlanticus, Pleurodeles waltl, Triturus pygmaeus) would require larger sampling effort than that used in our study. Despite filtering much larger volumes (600–5300 mL), filtering with capsules at a single site per pond provided lower detection probabilities for abundant species than filtering with discs at five sites. Rarer species showed no difference between methods, which may be due to small sample sizes and reduced statistical power for species with few detections. The effect of conductivity on species detectability was largely negative, while the influence of water clarity varied across species, and pH had no effects. Overall, our results suggest that eDNA amphibian surveys in Mediterranean temporary ponds need to consider filter clogging, heterogeneous DNA distribution, and highly conductive waters.

Measurement(s) reptile diversity Technology Type(s) DNA barcoding Factor Type(s) DNA barcode sequencing Sample Characteristic - Organism reptiles Sample Characteristic - Environment terrestrial Sample Characteristic - Location Western Sahara-Sahel

Abstract Explicitly accounting for phenotypic differentiation together with environmental heterogeneity is crucial to understand the evolutionary dynamics in hybrid zones. Species showing intra-specific variation in phenotypic traits that meet across environmentally heterogeneous regions constitute excellent natural settings to study the role of phenotypic differentiation and environmental factors in shaping the spatial extent and patterns of admixture in hybrid zones. We studied three environmentally distinct contact zones where morphologically and reproductively divergent subspecies of Salamandra salamandra co-occur: the pueriparous S. s. bernardezi that is mostly parapatric to its three larviparous subspecies neighbours. We used a landscape genetics framework to: (i) characterise the spatial location and extent of each contact zone; (ii) assess patterns of introgression and hybridization between subspecies pairs; and (iii) examine the role of environmental heterogeneity in the evolutionary dynamics of hybrid zones. We found high levels of introgression between parity modes, and between distinct phenotypes, thus demonstrating the evolution to pueriparity alone or morphological differentiation do not lead to reproductive isolation between these highly divergent S. salamandra morphotypes. However, we detected substantial variation in patterns of hybridization across contact zones, being lower in the contact zone located on a topographically complex area. We highlight the importance of accounting for spatial environmental heterogeneity when studying evolutionary dynamics of hybrid zones.

Understanding how hybridization influences the morphology and fitness of hybrids is essential for studying adaptive evolution and ecological speciation. Secondary contact zones, where separately evolving populations meet and hybridize, offer valuable insights into the evolutionary processes driving speciation and provide an excellent system to address these questions. In this study, we investigate patterns of morphological and genetic variation of two congeneric viper species, Vipera aspis and V. latastei, across a contact zone in northern Spain (Oja‐Tirón), where vipers with mixed morphology are often detected, but genetic studies addressing hybridization and relating patterns of genetic and morphological admixture are lacking. Using nine morphological traits (scalation and colouration) and 18 microsatellite markers, we (1) estimated the extent of hybridization, (2) morphologically characterized parental species and hybrids, and (3) evaluated the correlation between patterns of genetic and morphological admixture. Analyses revealed a bimodal hybrid zone with high rate of hybridization (22%) and prevalence of late‐generation hybrids (F2 and backcrosses). Morphological analyses differentiated the two parental species, and a positive correlation (r = 0.95) was found between morphological and genetic patterns. The hybrid group displayed on average an intermediate morphology between the parentals, yet morphologically intermediate hybrids were rare in our dataset. Instead, most hybrids resembled the parental species with whom they share most of the genetic background. Notably, the hybrid group exhibited greater morphological variation than the parental groups. Traits with adaptative value, such as ventral scales and dorsal marks, showed significant differences between hybrids and the two parental species. Introgression of these traits may confer ecological advantages to hybrids, enhancing local adaptation. Overall, this study reveals a positive correlation between patterns of morphological and genetic variation across a hybrid zone and provides insights into the phenotypic consequences of hybridization on these viper species. [ABSTRACT FROM AUTHOR]

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The Atlantic Meridional Overturning Circulation entails vigorous thermohaline transformations in the subpolar North Atlantic Ocean (SPNA). There, warm and saline waters originating in the (sub)tropics are converted into cooler and fresher waters by a combination of surface fluxes and sub‐surface mixing. Using microstructure measurements and a small‐scale variance conservation framework, we quantify the diapycnal and isopycnal contributions –by microscale turbulence and mesoscale eddies, respectively– to thermohaline mixing within the eastern SPNA. Isopycnal stirring is found to account for the majority of thermal (65%) and haline (84%) variance dissipation in the upper 400 m of the eastern SPNA. A simple dimensional analysis suggests that isopycnal stirring could account for O $\mathcal{O}$(5–10) Sv of diahaline volume flux, suggesting an important role of such stirring in regional water‐mass transformations. Our mixing measurements are thus consistent with recent indirect estimates in highlighting the importance of isopycnal stirring for North Atlantic overturning. Plain Language Summary: The North Atlantic hosts an ocean circulation system called the Atlantic Meridional Overturning Circulation (AMOC). It is often likened to a giant conveyor belt in the ocean, moving warm, salty waters from south to north and transforming them into cold, fresh waters that flow back southward at greater depth. The AMOC is a crucial element of the Earth's climate, and if it were to slow down, it could lead to major climatic changes. For a long time, scientists thought that the AMOC was mainly driven by cooling in the North Atlantic. But recently, we have discovered that the mixing of different water masses is also important. In our study, we used small‐scale measurements of ocean properties to examine the processes behind this mixing. Our findings show that large swirling flows known as mesoscale eddies, which are tens to hundreds of kilometers wide and hundreds of meters deep, play a dominant role in mixing heat and salt in the North Atlantic. This discovery helps us to better understand the AMOC and its future behavior. [ABSTRACT FROM AUTHOR]

Abstract Environmental DNA (eDNA) is increasingly used for biodiversity monitoring, particularly in aquatic systems. However, each step, from sample collection to bioinformatic analysis, can introduce biases and influence the reliability of results. While much effort has been put into the optimization of laboratory methods, less attention has been devoted to estimate the impacts of eDNA capture methods. To address this issue, water samples were collected at nine small ponds and puddles where up to 10 amphibian species occur, using precipitation, disc filters, and capsules. We focused on targeted detection of an amphibian species, Salamandra salamandra, and on the composition of the whole amphibian community. Species detection was performed using a novel qPCR assay for S. salamandra and high‐throughput sequencing, combined with stringent versus relaxed PCR replication thresholds. Filtration techniques (disc filters and capsules) outperformed precipitation, generating a higher number of detections of S. salamandra and higher amounts of captured eDNA, while species detection was identical between disc filters and capsules. There were no significant differences between capture methods regarding amphibian community composition. The variation in detection success associated with capture methods was far higher than that associated with PCR replication, regardless of the detection method used. Our results highlight the importance of choosing a suitable capture method for eDNA studies and suggest that the choice of capture method outweighs the choice of detection method used. To the best of our knowledge, this is the first study to compare high‐capacity capsules with common eDNA methods for water samples, such as precipitation and standard disc filters.

Abstract Reduced representation genome sequencing has popularized the application of single nucleotide polymorphisms (SNPs) to address evolutionary and conservation questions in nonmodel organisms. Patterns of genetic structure and diversity based on SNPs often diverge from those obtained with microsatellites to different degrees, but few studies have explicitly compared their performance under similar sampling regimes in a shared analytical framework. We compared range‐wide patterns of genetic structure and diversity in two amphibians endemic to the Iberian Peninsula: Hyla molleri and Pelobates cultripes, based on microsatellite (18 and 14 loci) and SNP (15,412 and 33,140 loci) datasets of comparable sample size and spatial extent. Model‐based clustering analyses with STRUCTURE revealed minor differences in genetic structure between marker types, but inconsistent values of the optimal number of populations (K) inferred. SNPs yielded more repeatable and less admixed ancestries with increasing K compared to microsatellites. Genetic diversity was weakly correlated between marker types, with SNPs providing a better representation of southern refugia and of gradients of genetic diversity congruent with the demographic history of both species. Our results suggest that the larger number of loci in a SNP dataset can provide more reliable inferences of patterns of genetic structure and diversity than a typical microsatellite dataset, at least at the spatial and temporal scales investigated.

Traditional methods of phylogenetic reconstruction and species delimitation may be impeded by frequent hybridization among lineages. In this study, we conducted phylogenetic and clustering analyses of ddRAD genomic data on the entire genus Salamandra, which includes six species and over 25 subspecies of terrestrial salamanders. We expanded previous datasets to include missing subspecies and incorporated new samples, with an emphasis on secondary contact zones. Results obtained from a full dataset of 392 individuals (356,874 bp; 24,192 SNPs) were compared with those obtained after excluding substantially admixed individuals (n = 95; 835,467 bp; 51,557 SNPs) to explore the consequences of introgression on phylogenetic inference and taxonomic arrangement of subspecies. We found conflicting phylogenetic placements for taxa represented by many admixed individuals (identified by clustering ancestries). In contrast, a time-calibrated tree constructed without hybrids largely agrees with previous phylogenetic hypotheses. Within S. atra, we found paraphyly of S. atra atra, suggesting an additional candidate subspecies. Within S. infraimmaculata, two lineages are assignable to known subspecies and we additionally identified a third, deeply diverged lineage sampled near the Turkish/Syrian border. In S. algira, we found limited admixture between the subspecies S. a. tingitana and S. a. splendens despite their geographic proximity. Finally, within S. salamandra, we detected significant levels of hybridization between subspecies, which blurred their phylogenetic relationships, although the removal of admixed samples in subset analyses clarified the situation in most cases. Monophyly was recovered for subspecies that were previously found paraphyletic, including S. s. salamandra, S. s. gallaica, and S. s. fastuosa. Salamandra s. "alfredschmidti" was confirmed to be a junior synonym of S. s. bernardezi. Previously disputed subspecies, like S. s. "molleri" and S. s. "hispanica", correspond to separated lineages but are affected by admixture with other lineages. Further newly identified candidate subspecies in S. salamandra included a southern lineage within S. s. wer-neri and a western lineage within S. s. bernardezi. Finally, we re-evaluate the status of recognized subspecies in Salamandra, based on evidence from multiple delimitation criteria. Given that the evolutionary history could not be resolved for all subspecies, we highlight taxa within Salamandra that warrant further molecular examination and taxonomic revision, notably within the S. s. gallaica/"molleri"/bejarae complex. This study illustrates the impact of hybridization in phylogenetic analyses and its downstream effects in the identification of conservation units and their naming in the Linnean classification. [ABSTRACT FROM AUTHOR]

DNA metabarcoding has proven to be an accessible, cost-effective, and non-invasive tool for dietary analysis of predators in situ. Although DNA metabarcoding provides numerous benefits in characterizing diet—such as detecting prey animals that are difficult to visually identify—this method has seen limited application in amphibian species. Here, we used DNA metabarcoding to characterize the diet of fire salamanders (Salamandra salamandra) (Linnaeus, 1758) in three distinct regions across the northwestern Iberian Peninsula. To test the efficiency of COI-based metabarcoding in determining salamanders’ diet diversity, we compared our COI-based results with results from traditional diet studies from neighboring and distant populations, as well as with recent findings obtained in a DNA metabarcoding study using 18S. Two COI primers were used in combination to investigate the potential impact of primer bias in prey detection. Our COI metabarcoding approach increased taxonomic resolution and supported a generalist diet in S. salamandra. Between primers, there were no significant differences in the diversity and richness of prey detected. We observed differences in the prevalence of prey identified between sampling regions both in our study and in other studies of S. salamandra diet. This COI metabarcoding study provides recommendations and resources for subsequent research using DNA metabarcoding to study amphibian diets.

The recent introduction of the chytrid fungus Batrachochytrium salamandrivorans into northeastern Spain threatens salamander diversity on the Iberian Peninsula. We assessed the current epidemiological situation with extensive field sampling of urodele populations. We then sought to delineate priority regions and identify conservation units for the Iberian Peninsula by estimating the susceptibility of Iberian urodeles using laboratory experiments, evidence from mortality events in nature and captivity and inference from phylogeny. None of the 1395 field samples, collected between 2015 and 2021 were positive for Bsal and no Bsal-associated mortality events were recorded, in contrast to the confirmed occurrence of Bsal outbreak previously described in 2018. We classified five of eleven Iberian urodele species as highly susceptible, predicting elevated mortality and population declines following potential Bsal emergence in the wild, five species as intermediately susceptible with variable disease outcomes and one species as resistant to disease and mortality. We identified the six conservation units (i.e., species or lineages within species) at highest risk and propose priority areas for active disease surveillance and field biosecurity measures. The magnitude of the disease threat identified here emphasizes the need for region-tailored disease abatement plans that couple active disease surveillance to rapid and drastic actions.