AimThe ecological dimension of evolutionary processes has been scarcely addressed in phylogeographic studies. We reconstruct the historical biogeography of Western Mediterranean vipers to discover the role of climate in fostering diversification.LocationWestern Mediterranean Basin.TaxonVipera aspis and Vipera latastei‐monticola complex.MethodsWe used nearly range‐wide phylogeographic analyses of three mitochondrial genes followed by geographic assignment of 4,056 records to genetic units to test phylogenetic niche conservatism, under a 3D hypervolume approach, and reconstruct paleoclimatic scenarios for the diversification of main lineages during Pleistocene.ResultsBayesian inferences from mtDNA recovered three Miocene clades and nine Pliocene lineages that diversified during the Pleistocene. Diversification was mostly restricted to southern regions of Iberian and Italian peninsulas and to the north‐western African mountains until the late Pliocene. Some lineages expanded northwards during the Pleistocene. Accordingly, genetic diversity was higher in southern regions. Ecological niche tests mostly supported allopatric diversification with niche conservatism, although niche shifts occurred with two divergence events. Palaeoclimatic models identified particular requirements for the current distribution of main lineages and distinct responses to the cooling and warming events of the Pleistocene. Areas of climatic stability during the Pleistocene were identified for main lineages; however, climatic stability was weakly correlated with haplotype diversity.Main conclusionsIn the Western Mediterranean biodiversity hotspot, palaeo‐tectonic and palaeo‐climatic factors drove diversification since the Miocene. Comparisons among patterns of diversification, haplotype diversity, and climatic stability suggest that southern Iberian and Italian peninsulas, and north‐western African mountains acted as refugia since the Pliocene, while some northern areas favoured population persistence during the Pleistocene. Climate adaptation likely played a secondary role in the diversification of some lineages.

Aim Climate variability is a major force affecting diversification processes and restricting species to specific areas, and thus, it has important impacts on species biogeographic patterns. This study aims to infer the role of climate in the evolutionary history of the endemic Iberian adder Vipera seoanei.Location Northern Iberian Peninsula and south-western France.Methods We combined genetic analyses with ecological niche-based modelling. Genetic analyses, based on sequencing of mitochondrial markers (cyt b, ND4), include phylogenetic and phylogeographic analyses, spatial interpolations of genetic variability and diversity, and identification of putative geographical origin of the most recent common ancestor of the species. Ecological modelling involved the combination of six modelling algorithms and projections to past conditions (Last Interglacial -LIG, Last Glacial Maximum -LGM) and the identification of climatic stable areas. ResultsThe species shows a shallow phylogeographic structure, dated at middle-upper Pleistocene, and low haplotype diversity, with the highest genetic diversity located in north-western Iberia. This region is identified as the putative origin of the ancestral populations. Projections to past periods spatially fit genetic results, indicating range contractions to north-western Iberia during the LIG and expansions during the LGM.Main conclusions This study exemplifies how the combination of phylogeographic and ecological niche-based models is a powerful tool for inferring evolutionary scenarios and responses of species to Pleistocene climatic oscillations. Vipera seoanei responded accordingly to a cold temperate model and fits a simplified example of 'R' type species where interglacial warming periods during the Pleistocene probably caused major range reductions with persistence in a single refuge in north-western Iberia. The single mtDNA lineage observed in this study does not support the differentiation at subspecific level in V. seoanei. Our work highlights the importance of climate in explaining evolutionary processes and current biogeographical patterns of species with restrictive ranges.

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Aim Climatic fluctuations in northern Africa substantially changed the extent of the Sahara desert and Saharan species’ ranges. Yet, the region contained areas of climatic stability. We test the hypothesis that the Atlantic Sahara was a stable corridor, connecting ecoregions, for the spiny‐footed lizard Acanthodactylus aureus. Location Africa, Saharan Atlantic coastal desert. Methods We combined ecological modelling and phylogeographic and population genetic analyses. Ecological models for past and current conditions were used to predict climatically stable areas for the species over time. Genetic analysis, including for three mitochondrial fragments (12S, Cytb and COI), one nuclear gene (C‐mos) and 18 microsatellite markers, were used to unveil patterns of genetic structure and diversity, and gene flow dynamics within A. aureus. Results Three mtDNA allopatric lineages diversified during the Pliocene‐Pleistocene along the Atlantic Sahara. Two main areas of high climatic stability largely fit the regions with highest mtDNA diversity. Mito‐nuclear discordances along some coastal regions indicate evidence of gene flow between lineages, which are likely mediated by population expansions and male‐biased dispersal. Several geographical barriers to gene flow were also identified. Main conclusion This study highlights the role of the Atlantic Sahara ecoregion both as a centre of lineage diversification and as a occasional suitable corridor within the Sahara desert. Population retractions and expansions resulting from climatic oscillations during the Pleistocene, facilitated allopatric diversification and genetic introgression processes along this region, whereas stable geographical barriers limited gene flow dynamics.

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.