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FIGURE 9. a–c in Integration of molecular, bioacoustical and morphological data reveals two new cryptic species of Pelodytes (Anura, Pelodytidae) from the Iberian Peninsula

FIGURE 9. a–c, Mouthparts of preserved tadpoles of three species of Pelodytes. Note the absence of a central gap in the third anterior labial tooth row in P. ibericus. d, mouthpart, and e, lateral view of a living tadpole of P. ibericus from near Jerez de la Frontera.

FIGURE 6 in Integration of molecular, bioacoustical and morphological data reveals two new cryptic species of Pelodytes (Anura, Pelodytidae) from the Iberian Peninsula

FIGURE 6. Morphometric differentiation among the four western Pelodytes lineages. The upper graph is a scatterplot of first and second principal components from a PCA of raw morphometric values in 117 male specimens of Pelodytes. Note that PC1 corresponds mostly to body size (Table 5). The lower graphs are boxplots of SVL, relative tympanum diameter and relative hindlimb length in male Pelodytes, with median (dot), 25% percentiles (box) and ranges, illustrating the smaller body size of P. atlanticus sp. nov. and P. ibericus, and larger tympanum and shorter limbs of P. ibericus. All specimens of P. ibericus, P. atlanticus, and P. hespericus were measured by JDR, and specimens of P. punctatus were measured by MV.

FIGURE 4 in Integration of molecular, bioacoustical and morphological data reveals two new cryptic species of Pelodytes (Anura, Pelodytidae) from the Iberian Peninsula

FIGURE 4. Species tree calculated from the analysis of DNA sequences of two mitochondrial and six nuclear gene segments (left), after exclusion of admixed/hybrid samples identified from the nDNA networks in Fig. 2 and Bayesian assignment analyses in Fig. 3. The tree on the right shows the same topology as recovered in a species tree analysis based only on the six nuclear gene segments.

FIGURE 1 in Integration of molecular, bioacoustical and morphological data reveals two new cryptic species of Pelodytes (Anura, Pelodytidae) from the Iberian Peninsula

FIGURE 1. Location of 45 populations of Pelodytes sampled for molecular analysis (indicative map unscaled). Colored dots represent the four species described: green, lineage A (P. atlanticus sp. nov.); orange, lineage B (P. ibericus); blue, lineage C (P. hespericus sp. nov.); and purple, lineage D (P. punctatus). In some populations (7, 8, 9, 10, 11, 22, 23, 36 and 37) we found evidence for admixture (double-colored dots). Background colors represent the ranges of P. punctatus (blue) and P. ibericus (orange) according to current knowledge (www.iucn.org).

FIGURE 12 in Integration of molecular, bioacoustical and morphological data reveals two new cryptic species of Pelodytes (Anura, Pelodytidae) from the Iberian Peninsula

FIGURE 12. Dorsal, ventral and dorsolateral view, and close up of ventral side of forelimb, of living Pelodytes hespericus sp. nov.: (a-b) male holotype JDR 2012-1, (c-f); male paratype JDR 2012-2. Not to scale.

FIGURE 5 in Integration of molecular, bioacoustical and morphological data reveals two new cryptic species of Pelodytes (Anura, Pelodytidae) from the Iberian Peninsula

FIGURE 5. Ventral views of specimen hands (palmar surface) of the four western Pelodytes lineages. The pictures show variation in the size and arrangement of the subarticular and metacarpal tubercles within species and populations, suggesting that no fully diagnostic interspecific differences in this character exist. All photos from specimens in life. M, male, F, female. Asterisks denote mirrored images. Not to scale.

FIGURE 3 in Integration of molecular, bioacoustical and morphological data reveals two new cryptic species of Pelodytes (Anura, Pelodytidae) from the Iberian Peninsula

FIGURE 3. Clustering results of Bayesian assignment analyses using Structure, based on nuclear DNA sequences with the number of clusters K ranging from 1 to 7. Each individual is represented as a vertical bar partitioned into K colored segments, whose length is proportional to the individual's estimated group membership coefficient. The enlarged figure shows the results for K=4, with population numbers as in Fig. 1 and Supplementary Table 1. Population numbers on the top are shown to highlight individuals with mixed ancestry, and the mitochondrial haplogroup (A-D) for each included individual is shown in the bottom. Individuals in the graph are ordered as in Supplementary Table S1. Note that the comparatively large number of putative hybrids is due to the overrepresentation of populations in the contact zones in our analysis, as compared with Díaz- Rodríguez et al. (2015).

FIGURE 11 in Integration of molecular, bioacoustical and morphological data reveals two new cryptic species of Pelodytes (Anura, Pelodytidae) from the Iberian Peninsula

FIGURE 11. (a,b) Dorsal and ventral views of preserved Pelodytes atlanticus sp. nov. holotype (male; specimen GLA-03). (c,d) Dorsolateral view of paratype (GLA-02) and holotype (GLA-03) in life. (e-i) dorsolateral, dorsal, and ventral view, as well as ventral view of hand and foot of male paratype ZSM 194/2016. Not to scale.

FIGURE 10 in Integration of molecular, bioacoustical and morphological data reveals two new cryptic species of Pelodytes (Anura, Pelodytidae) from the Iberian Peninsula

FIGURE 10. Comparative photos of Pelodytes ibericus in life, male above, female below, from near Jerez de la Frontera, Spain. Specimens not collected. Images not to scale.

FIGURE 8 in Integration of molecular, bioacoustical and morphological data reveals two new cryptic species of Pelodytes (Anura, Pelodytidae) from the Iberian Peninsula

FIGURE 8. Comparative photos of Pelodytes punctatus in life. The upper two photos show a male and a female from near Montpellier, France; the lower two pictures show a male and a female from near Vic, Girona, Spain. Specimens not collected. Images not to scale. Identity of P. punctatus individuals assessed by geographical location; specimens not DNA barcoded.

FIGURE 2 in Integration of molecular, bioacoustical and morphological data reveals two new cryptic species of Pelodytes (Anura, Pelodytidae) from the Iberian Peninsula

FIGURE 2. Median-joining networks of Western Palearctic Pelodytes based on six nuclear markers and mtDNA (mt-ND4+mt- CYB). Samples in the nDNA networks are colored according to their assignment to the four major clusters in the mtDNA network, corresponding to the species as color coded in Fig. 1. Each circle represents a different haplotype and its size is proportional to its relative frequency. Black dots represent inferred unsampled or extinct haplotypes. Markers RPL3 and Cherpint7 are represented at the same scale.

FIGURE 7 in Integration of molecular, bioacoustical and morphological data reveals two new cryptic species of Pelodytes (Anura, Pelodytidae) from the Iberian Peninsula

FIGURE 7. Bioacoustic differences between lineages of Pelodytes occurring in the Iberian Peninsula. For raw data, see Supplementary Table S3. The boxplot shows the average number of B notes per call series, i.e, every call consisting of one A note and one or more B notes. Average values were first computed per individual, and then summarized across species. Differences are significant between P. hespericus sp. nov. and all other species, and between P. atlanticus sp. nov. and P. ibericus (ANOVA with Tukey's post-hoc test; see Results).

FIG. 4 in Age structure and growth in an isolated population of Pelodytes punctatus in northern Spain

FIG. 4. Growth in length body (SVL) of P. punctatus in Burgos, Spain. Mean and range are given at each age. Curves fitted to the Von Bertalanffy equation.

FIG. 1 in Age structure and growth in an isolated population of Pelodytes punctatus in northern Spain

FIG. 1. Cross-sections at the diaphysis of phalanges for P. punctatus in Burgos, Spain. (A) Male, 40.3 mm SVL, six LAGs; (B) female, 45.5 mm SVL, five LAGs. Arrows, LAGs; ml, metamorphosis line; eb, endosteal bone.

FIG. 3 in Age structure and growth in an isolated population of Pelodytes punctatus in northern Spain

FIG. 3. Snout–vent length distribution of P. punctatus in Burgos, Spain. Black bars, males; grey bars, females.

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Data from: Phenotypic plasticity allows the Mediterranean parsley frog Pelodytes punctatus to exploit two contrasted temporal niches under continuous gene flow

Environmental changes, such as climate change, lead to the opening of new niches. In such situations, species that adapt to new niches can survive and/or expand their ranges. However, gene flow can hamper genetic adaptation to new environments. Alternatively, recent models have highlighted the importance of phenotypic plasticity in tracking environmental change. In this study, we illustrate how plasticity allows an amphibian species to exploit two very different climatic niches under continuous gene flow. In the Mediterranean region, the parsley frog Pelodytes punctatus breeds both in spring, as do most other species, and in autumn, a temporal niche not exploited by most other species, but which may become increasingly important with global warming. Conditions of development are dramatically different between the two seasons and deeply impact tadpole life-history traits. To determine whether these temporal niches are exploited by two genetically differentiated sub-populations, or whether the bimodal phenology arises in a panmictic population displaying plastic life-history traits, we use two complementary approaches. We measure both molecular genetic differentiation and quantitative-trait differentiation between spring and autumn cohorts, using microsatellites an common garden experiments respectively. Seasonal cohorts were not genetically differentiated and differences in tadpole life-history between cohorts were not maintained in laboratory conditions. We conclude that phenotypic plasticity, rather than genetic adaptation, allows Parsley frog to exploit two contrasted temporal niches.

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Data from: Phenotypic plasticity allows the Mediterranean parsley frog Pelodytes punctatus to exploit two contrasted temporal niches under continuous gene flow

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FIG. 2 in Age structure and growth in an isolated population of Pelodytes punctatus in northern Spain

FIG. 2. Age distribution of P. punctatus in Burgos, Spain. Black bars, males; grey bars, females.