In Europe, southern peninsulas served as refugia during cold periods in the Pleistocene, acting both as centres of origin of endemisms and as sources from which formerly glaciated areas were recolonized during interglacial periods. Previous studies have revealed that within the main refugial areas, intraspecific lineages often survived in allopatric refugia. We analysed two mitochondrial markers (nad4, control region, approximately 1.4 kb) in 103 individuals representing the entire distribution of Lissotriton boscai, a newt endemic to the western Iberian Peninsula. We inferred the evolutionary history of the species through phylogenetic, phylogeographic and historical demographic analyses. The results revealed unexpected, deep levels of geographically structured genetic variability. We identified two main evolutionary lineages, each containing three well-supported clades. The first historical split involved populations from central-southwestern coastal Portugal and the ancestor of all the remaining populations around 5.8 million years ago. Both lineages were subsequently fragmented into different population groups between 2.5 and 1.2 million years ago. According to nested clade analysis, at lower hierarchical levels the patterns suggest restricted gene flow with isolation by distance, whereas at higher levels the clades exhibit signatures of contiguous range expansion. Bayesian Skyline Plots show recent bottlenecks, followed by demographic expansions in all lineages. The significant genetic structure found is consistent with long-term survival of populations in allopatric refugia, supporting the 'refugia-within-refugia' scenario for southern European peninsulas. The comparison of our results with other co-distributed species highlights the generality of this hypothesis for the Iberian herpetofauna and suggests that Mediterranean refuges had more relevance for the composition and distribution of present biodiversity patterns than currently acknowledged. We briefly discuss the taxonomic and conservation implications of our results.

Quaternary climatic oscillations and geographic barriers have strongly influenced the distribution and diversification of thermophilic species occurring in the Mediterranean Basin. The Western Mediterranean pond turtle, Mauremys leprosa, is widely distributed throughout the Iberian Peninsula, southern France and most of the Maghreb region, with two subspecies currently recognized. In this work, we used 566 samples, including 259 new individuals, across the species range, and sequenced two mitochondrial markers (cytochrome b gene and control region; 163 samples in a concatenated mtDNA dataset) and one nuclear intron (R35; 23 samples representing all identified sublineages) to study the evolutionary history of M. leprosa. We combined phylogenetic methods and phylogeographic continuous diffusion models with spatial analysis. Our results (1) show a high level of genetic structure in Morocco originated during the Pleistocene; (2) reveal two independent population expansion waves from Morocco to Tunisia and to southern Europe, which later expanded throughout the Iberian Peninsula, and (3) identify several secondary contact zones in Morocco. Our study also sheds new light on the role of geographical features (Moroccan mountains ranges and the Strait of Gibraltar) and Pleistocene climatic oscillations in shaping genetic diversity and structure of M. leprosa, and underlines the importance of the Maghreb as a differentiation centre harbouring distinct glacial refugia.

Ponds provide vital ecological services. They are biodiversity hotspots and important breading sites for rare and endangered species, including amphibians and dragonflies. Nevertheless, their number is decreasing due to habitat degradation caused by human activities. The “Ponds with Life” environmental education project was developed to raise public awareness and engagement in the study of ponds by promoting the direct contact between the public and nature, researchers and pedagogical hands-on exploration activities. A pre-post- project survey was set-up to assess the effects of the project on the environmental consciousness, knowledge and attitude changes towards ponds and the associated biodiversity of school students aged 15 to 18. The survey questions were based on Likert scales and their pre-post project comparisons used an innovative multivariate hypothesis testing approach. The results showed that the project improved the students’ knowledge and attitudes towards ponds and associated biodiversity, especially the amphibians. Ponds can be found or constructed in urban areas and despite small sized, they proved to be interesting model habitats and living laboratories to foster environmental education, by encompassing a high number of species and a fast ecological succession.

The power of phylogeographic inference resides in its ability to integrate information from multiple sources in an iterative hypothesis-testing framework. In this paper, we build upon previous mtDNA-based hypotheses about the evolutionary history of the Iberian newt Lissotriton boscai using sequences of the highly variable nuclear β-fibrinogen intron 7. In addition to the nuclear sequences, we produced new mtDNA data across the species range to delineate contact zones and test the congruence between nuclear and mitochondrial datasets at the same level of spatial organization. Through a combination of phylogenetic, phylogeographic continuous diffusion, and genetic landscape modelling analyses, we infer the evolutionary history of the species. We found notable congruence between nuclear and mtDNA datasets, which confirms deep and consistent differentiation between two major lineages that originated in the Miocene. Additionally, we found a new nuclear haplogroup with no mtDNA counterpart, roughly circumscribed to the Iberian Sistema Central mountains, and extensive areas of nuclear admixture across mtDNA lineages. We describe potential historical dispersal routes from an ancestral hypothetical refugium in the western end of the Sistema Central in central Portugal and highlight how deep phylogeographic breaks do not necessarily indicate cryptic speciation events.

Chioglossa lusitanica is A streamside salamander endemic to the northwestern corner of the Iberian Peninsula. In an extensive Held survey we found the species to be widely distributed across northwestern Portugal. The distribution data were used to identify macroenvironmental parameters related to the presence of the species in Portugal, and to produce a descriptive distribution model for C. lusitanica. We modelled the species distribution based on five environmental parameters (precipitation, relief, July temperature, number of frost months and hardness of water) selected by logistic regression analysis. The model yielded a high score of correct classification (95%). Excluding parameters unavailable for Spain the model included four parameters (precipitation, relief, July temperature and altitude) and had a 93% correct classification score. Extrapolation of the model to Spain yielded a correct classification of 92% and served as a validation of the modelling technique in an independent area. The models indicate the existence of several potentially suitable areas outside the known distribution of the species, providing clues about the species historical biogeography. The models, through the identification of highly suitable and stable protection regions, may contribute to informed conservation planning.