spatial conservation prioritization of biodiversity ... · (phenotypic, behavioural and/ or...
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ARTICLESPUBLISHED: XX XX 2017 | VOLUME: 1 | ARTICLE NUMBER: 0151
© 2017 Macmillan Publishers Limited, part of Springer Nature. All rights reserved.
NATURE ECOLOGY & EVOLUTION 1, 0151 (2017) | DOI: 10.1038/s41559-017-0151 | www.nature.com/natecolevol
Spatial conservation prioritization of biodiversity spanning the evolutionary continuumSilvia B. Carvalho1, 2*, Guillermo Velo-Antón1, Pedro Tarroso1 , Ana Paula Portela1, Mafalda Barata1, Salvador Carranza3, Craig Moritz4 and Hugh P. Possingham2, 5
Accounting for evolutionary relationships between and within species is important for biodiversity conservation planning, but is rarely considered in practice. Here we introduce a novel framework to identify priority conservation areas accounting for phylogenetic and intra-specific diversity, integrating concepts from phylogeny, phylogeography, spatial statistics and spatial conservation prioritization. The framework allows planners to incorporate and combine different levels of evolutionary diver-sity and can be applied to any taxonomic group and to any region in the world. We illustrate our approach using amphibian and reptile species occurring in a biodiversity hotspot region, the Iberian Peninsula. We found that explicitly incorporating phyloge-netic and intraspecific diversity in systematic conservation planning provides advantages in terms of maximizing overall biodi-versity representation while enhancing its persistence and evolutionary potential. Our results emphasize the need to account for the evolutionary continuum in order to efficiently implement biodiversity conservation planning decisions.
Given accelerating declines in biodiversity, a major global objective is to improve the status of biodiversity by safe-guarding species and genetic diversity in networks of pro-
tected areas (strategic goal C, Convention on Biologic Diversity (CBD) Aichi Targets; http://www.cbd.int/sp/targets). One of the targets established by the CBD is to protect at least 17% of terres-trial land by 2020. Although the extension of protected land has increased over the past century, there are still substantial shortfalls around the world1.
Many different strategies have been developed to identify pri-ority conservation areas, including biodiversity hotspots2 and key biodiversity areas3. Alternatively, systematic conservation plan-ning4 uses complementarity and efficiency principles to maximize biodiversity representation and persistence5. Typically, species have been the fundamental units of diversity used in spatial conservation prioritization — even though the importance of accounting for evolutionary relationships between and within species has long been recognized6–9.
Preserving evolutionary relationships between species is impor-tant on the premise that different species represent different amounts of evolutionary history. Additionally, phylogenetic relationships are expected to be effective surrogates of underlying feature diversity (phenotypic, behavioural and/ or ecological) and, consequently, of functional diversity10. However, there are only a few studies in which evolutionary diversity has been explicitly accounted for in spatial prioritization11–14. This is probably related to a lack of comprehen-sive phylogenies, and to major uncertainties and missing taxa in those that are available15. It is also argued that conservation areas that are selected on the basis of species distributions will adequately represent phylogenetic diversity16; as such, species distributions can be effective surrogates for evolutionary diversity, though there is empirical evidence and theoretical arguments to the contrary11,17–19.
Intraspecific genetic diversity underpins evolutionary potential (the ability of a population or species to respond to future selection
pressures) in the sense that it promotes the generation of new spe-cies, the resilience to changing environmental conditions, and the ability of a species to undergo evolutionary adaptation20–22. The inference of phylogenetic trees at the intraspecific level allows detecting independently evolving sets of populations (hereafter called lineages). When linked to spatial models, it is possible to infer the spatial distribution of each lineage23,24. This spatial information is critical for conservation planning because it allows optimizing overall representation of diversity and preserving the potential for future nascent species25. Indeed, several studies have shown that intraspecific genetic diversity is spatially structured and that areas of higher genetic diversity are often coincident among several spe-cies26, resulting in either hotspots of genetic diversity or concentra-tions of phylogeographic endemism27. However, few studies have explicitly incorporated the intraspecific phylogeography of multiple species into spatial prioritization28–30. One reason for this is the lack of such information for multiple species in the same region; another is the absence of spatially explicit genetic data31.
Here we introduce and apply a framework to identify priority areas for the conservation of biodiversity spanning the evolutionary continuum, from inter- to intraspecific diversity. The framework consists of four main stages (Fig. 1). The first stage, mapping spe-cies distributions, involves collecting data on species distributions. The second, mapping interspecific phylogenetic diversity (PD), implicates collecting tissue samples of all species and extracting, amplifying and sequencing DNA. The DNA sequences will then be used to infer interspecific phylogenetic trees that, in turn, are used to calculate and then map branch lengths, and finally calculate PD across the study area. The third stage, mapping intraspecific lin-eage diversity (LD), consists of building a georeferenced database of DNA sequences, using literature data and/or new sequences. This database will then be used to infer an intraspecific phylogenetic tree for each species, and to identify and map the main phylogenetic lineages. Finally, stage four involves using the maps of interspecific
1CIBIO/InBIO, Centro de Investigação em Biodiversidade e Recursos Genéticos da Universidade do Porto, R. Padre Armando Quintas, 4485-661 Vairão Portugal. 2The School of Biological Sciences, University of Queensland, St Lucia, Qld 4072, Australia. 3Institute of Evolutionary Biology, CSIC-Universitat Pompeu Fabra, Barcelona E-08003, Spain. 4Research School of Biology and Centre for Biodiversity Analysis, The Australian National University, Acton ACT 6201, Australia. 5Conservation Science, The Nature Conservancy, West End, Qld 4101, Australia. *e-mail [email protected]
© 2017 Macmillan Publishers Limited, part of Springer Nature. All rights reserved.
SUPPLEMENTARY INFORMATIONVOLUME: 1 | ARTICLE NUMBER: 0151
NATURE ECOLOGY & EVOLUTION | DOI: 10.1038/s41559-017-0151 | www.nature.com/natecolevol 1
Supplementary material 1 – Study Area 15
The study region is the continental Iberian Peninsula, situated in the extreme southwest of 16
Europe (bounded by 9º32’ to 3º20’E and 35º56’ to 43º55’N). With an area of 582 860 km2, it 17
includes the continental territories of Portugal, Spain and Andorra. 18
The Iberian Peninsula is located in the Mediterranean Basin, one of the largest Planet's 19
biodiversity hotspot. Three main factors have driven lineage and species diversification in this 20
region, leading to the current patterns of deep spatially structured and high intra-specific genetic 21
diversity observed for many animal groups: i) complex biogeographic histories of species 22
resulting from palaeogeographic processes, such as break-up events of the European–Iberian 23
continental margin and the reopening of the Strait of Gibraltar after the Messinian salinity crisis 24
5.3 Mya; ii) the Iberian Peninsula served as multiple climatic refugia for different taxonomic 25
groups in Europe during the Pleistocene climatic oscillations, favouring the long-term 26
persistence of lineages in different refugia and posterior population expansions; and iii) the 27
complex topographic characteristics of the Iberian Peninsula (e.g. Mountain systems and 28
rivers) acted as cradles for diversity (by promoting isolation and genetic differentiation). Thus, 29
ectotherm species, such as the Iberian amphibians and reptiles, whose evolutionary histories 30
have been studied extensively, provide a good case-study to test whether species distributions 31
are effective surrogates for evolutionary diversity. 32
33
Figure S1.1 – Study Area (Iberian Peninsula) depicting elevation, country borders, administrative regions, 34
major mountain systems and rivers. 35
Supplementary material 2 – Taxa used throughout the study 36
The taxonomy of Iberian amphibians and reptiles has changed remarkably in the last decade, 37
mostly due to increased phylogeographic and morphologic studies. Species-level taxonomy 38
used in this study concurs with the most recent expert revision of Iberian amphibians and 39
reptiles taxonomy. 40
Table S2.1 - Taxa used for the species level analysis (Sp Level), inference of inter-specific phylogenies 41
and phylogenetic diversity patterns (PD) and intra-specific diversity (LIN) are marked as “YES”. NLIN 42
indicates the number of lineages retrieved with the GMYC analysis within each analyzed taxon. 43
Taxa acron Sp Level PD LIN NLIN
Amphibians
Alytes cisternasii alycis YES YES YES 4
Alytes dickhilleni alydic YES YES NO
Alytes obstetricans alyobs YES YES YES 6
Bufo calamita1 bufcal YES YES NO
Bufo spinosus bufspi YES YES NO
Calotriton arnoldi calarn YES YES NO
Calotriton asper calasp YES YES NO
Chioglossa lusitanica chilus YES YES YES 2
Discoglossus pictus dispic YES YES NO
Discoglossus galganoi discog YES YES YES 2
Hyla meridionalis hylmer YES YES YES 2
Hyla molleri hylmol YES YES NO
Ichthyosaura alpestris mesalp YES YES YES 2
Lissotriton boscai lisbos YES YES YES 13
Lissotriton helveticus lishel YES YES YES 6
Pelobates cultripes pelcul YES YES YES 2
Pelodytes ibericus pelibe YES YES NO
Pelodytes punctatus pelpun YES YES NO
Pelodytes complex 2 pelody NO NO YES 7
Pelophylax perezi pelper YES YES NO
Pleurodeles waltl plewal YES YES YES 2
Rana dalmatina randal YES YES NO
Rana iberica ranibe YES YES NO
Rana pyrenaica ranpyr YES YES NO
Rana temporaria rantem YES YES YES 6
Salamandra salamandra salsal YES YES NO
Triturus marmoratus trimar YES YES YES 2
Triturus pygmaeus tripyg YES YES YES 2
Reptiles
Acanthodactylus erythrurus acaery YES YES YES 5
Algyroides marchi algmar YES YES NO
Anguis fragilis angfra YES YES NO
Blanus cinereus blacin YES YES YES 2
Blanus mariae blamar YES YES YES 3
Chalcides bedriagai chabed YES YES YES 3
Chalcides striatus chastr YES YES YES 2
Chamaeleo chamaeleon chacha YES YES NO
Coronella austriaca coraus YES YES YES 4
Coronella girondica corgir YES YES YES 3
Emys orbicularis emyorb YES YES YES 5
Hemidactylus turcicus hemtur YES YES YES 2
Hemorrhois hippocrepis hemhip YES YES NO
Hierophis viridiflavus hievir YES YES NO
Iberolacerta aranica ibeara YES YES NO
Iberolacerta aurelioi ibeaur YES YES NO
Iberolacerta bonnali ibebon YES YES NO
Iberolacerta cyreni ibecyr YES YES NO
Iberolacerta galani ibegal YES YES NO
Iberolacerta martinezricai ibemar YES YES NO
Iberolacerta monticola ibemon YES YES NO
Lacerta agilis lacagi YES YES NO
Lacerta bilineata lacbil YES YES NO
Lacerta schreiberi lacsch YES YES YES 2
Macroprotodon brevis macbre YES YES NO
Malpolon monspessulanus malmon YES YES NO
Mauremys leprosa maulep YES YES NO
Natrix maura natmau YES YES YES 2
Natrix natrix 3 natnat YES YES NO
Podarcis bocagei podboc YES YES NO
Podarcis carbonelli podcar YES YES YES 6
Podarcis guadarramae podgua YES YES NO
Podarcis hispanica complex4 podhis YES NO YES 8
Podarcis hispanica Albacete/ Murcia podhisalb NO YES NO
Podarcis hispanica Galera podhisgal NO YES NO
Podarcis hispanica sensu stricto podhisss NO YES NO
Podarcis liolepis podlio YES YES NO
Podarcis muralis podmur YES YES YES 2
Podarcis vaucheri podvau YES YES YES 3
Podarcis virescens podvir YES YES NO
Psammodromus algirus psaalg YES YES NO
Psammodromus edwardsianus psaedw YES YES NO
Psammodromus hispanicus complex 5 psahis NO NO YES 4
Psammodromus hispanicus psahishis YES YES NO
Psammodromus occidentalis psahisocc YES YES NO
Rhinechis scalaris rhisca YES YES NO
Tarentola mauritanica tarmau YES YES YES 2
Testudo graeca tesgra YES YES NO
Testudo hermanni tesher YES YES NO
Timon lepidus timlep YES YES YES 5
Vipera aspis vipasp YES YES NO
Vipera latastei viplat YES YES YES 5
Vipera seoanei vipseo YES YES NO
Zamenis longissimus zamlon YES YES NO
Zootoca vivipara zooviv YES YES NO 44 1 – Presently referred as Epidalea calamita 45 2 – Included Pelodytes ibericus and Pelodytes punctatus 46 3 – Presently referred as Natrix astreptophora 47 4 – Includes Podarcis guadarramae, Podarcis hispanica Albacete/ Murcia, Podarcis hispanica Galera, 48 Podarcis virescens and Podarcis hispanica sensu stricto 49 5 - Includes Psammodromus hispanicus and Psammodromus occidentalis 50
51
Supplementary material 3 – Methodological details 52
Species distributions 53
For all recently described species (Blanus mariae, Iberolacerta galani, Podarcis guadarramae, 54
P. virescens and Psammodromus occidentalis), and for the subspecies elevated to the species 55
level (Psammodromus edwarsianus) their atlas distributions were not up to date and therefore 56
the distribution of such species was accessed based on expert knowledge. One taxon 57
(Discoglossus galganoi) resulted from the merge of two previous recognized species (D. 58
galganoi and D. jeanneae) and thus the distribution used in this study was the combined 59
occurrence records of former species. Two species were excluded from this study because they 60
were recently introduced in the study area: Podarcis pityusensis and Podarcis sicula. 61
Inter–specific phylogenetic trees 62
We gathered tissue samples for all amphibian and reptile species included in the study (figure 63
1A – step 1.1) and up to six genes were PCR-amplified and sequenced in both directions (figure 64
1 – step 1.2). For amphibians, we sequenced three mitochondrial genes - 12S rRNA (12S), 16S 65
rRNA (16S), and the cytochrome b (CYTB) - and one nuclear gene, the recombination-66
activating gene 1 (RAG1). For reptiles, we sequenced the same three mitochondrial regions 67
plus three nuclear genes - the oocyte maturation factor Mos (CMOS), the melanocortin receptor 68
1 (MC1R) and the recombination-activating gene 1 (RAG1) (Table A3.1) 69
We used GENEIOUS v. R6.1.6 for assembling and editing the chromatographs. We identified 70
heterozygous positions for the nuclear coding gene fragments based on the presence of two 71
peaks of approximately equal height at a single nucleotide site in both strands and were coded 72
using IUPAC ambiguity codes. The nuclear coding fragments were translated into amino acids 73
and no stop codons were observed. DNA sequences were aligned for each gene independently 74
using the online application of MAFFT v.7 with default parameters (auto strategy, gap opening 75
penalty: 1.53, offset value: 0.0). For the 12S and 16S ribosomal fragments we applied the Q-76
INS-i strategy, in which information on the secondary structure of the RNA is considered. Very 77
poorly aligned regions in the 12S and 16S alignment of dataset 1 were eliminated with Gblocks 78
under low stringency options. 79
The data set used to infer the amphibian tree included all 27 amphibian species (plus a sample 80
of Discoglossus galganoi jeanneae which was previously considered a separate species from 81
D. galganoi) and consisted of an alignment of 3577 base pairs (bp) of concatenated 82
mitochondrial 12S (100%, 366 bp), 16S (100%, 564 bp), CYTB (100%, 1135 bp) and nuclear 83
RAG1 (75%, 1512 bp) DNA sequences. The data set used to infer the phylogenetic tree for 84
reptiles included all 50 reptile GVA3727in the Iberian Peninsula. Since the taxonomy of the 85
Iberian Podarcis complex is not well defined yet, we included in the phylogeny all the known 86
lineages within this complex: P. muralis, P. liolepis, P. carbonelli, P. hispanica Type 1A, 87
Podarcis hispanica type 1B, P. hispanica type 2, P. bocagei, P. hispanica Albacete/Murcia, P. 88
hispanica sensu stricto, P. vaucheri and P. hispanica Galera. We also included a sample from 89
the subspecies Timon lepidus nevadensis which has recently been suggested that it may 90
deserve specific status but it is not yet widely accepted, totalling 55 taxa (supplementary 91
material 2). The full dataset consisted of a concatenated alignment of 4384 base pairs (bp) for 92
12S (96%, 370 bp), 16S (98%, 450 bp), CYTB (100%, 1035 bp), CMOS (91%, 570 bp), MC1R 93
(80%, 615 bp) and RAG1 (25%, 1344 bp). Finally, we pruned the trees to remove sub-specific 94
taxa which formed monophyletic groups regarding the recognized species taxonomy. As such, 95
we removed D. g. jeanneae from the amphibian’s tree (which formed a monophyletic group with 96
the recognized species D. galganoi). From the reptiles’ tree, we removed T. l. nevadensis 97
(which formed a monophyletic group with the recognized species T. lepidus) and P. hispanica 98
type 1B (which together with P. hispanica tpe 1A forms a monophyletic group currently 99
recognized as Podarcis guadarramae). 100
We inferred best-fitting models of nucleotide evolution for both datasets using 101
PARTITIONFINDER v.1.1.1 with the following settings: branch lengths linked, only models 102
available in BEAST evaluated with BIC model selection, all partition schemes analysed. Each 103
gene was set as an independent partition. For the amphibian data set, a three-partition scheme 104
was selected: p1 12S and 16S and the GTR+I+G model of sequence evolution; p2 CYTB and 105
TrN+I+G; p3 RAG11 and GTR+I+G. For the reptile data set, a four-partition scheme was 106
selected: p1 12S and 16S and GTR+I+G; p2 CYTB and GTR+I+G; p3 CMOS and RAG1 and 107
TrN+I+G; p4 MC1R and HKY+G. 108
109
Three individual runs of 5x107 generations were carried out sampling at intervals of 10 000 110
generations. Models and prior specifications applied were as follows (otherwise by default): 111
Substitution models and clock models unlinked; the amphibian data set and for the four 112
partitions of the reptile data set as indicated above; Yule process speciation tree prior; random 113
starting tree; base substitution prior Uniform (0100); alpha prior Uniform (0,10). Posterior trace 114
plots and effective sample sizes (ESS) of the runs were monitored in TRACER v1.5 to ensure 115
convergence. The results of the individual runs were combined in LogCombiner discarding 10% 116
of the samples and the maximum clade credibility (MCC) ultrametric tree was produced with 117
TreeAnnotator (both provided with the BEAST package). Nodes were considered strongly 118
supported if they received posterior probability (pp) support values ≥ 0.95. Since branch lengths 119
are dependent on the topology of the phylogenetic tree, several constraints were applied to both 120
the reptile and amphibian BEAST analyses in order to recover the accepted topology for both 121
groups. For the reptile analysis, all the members of the clade Squamata (all the specimens 122
except the terrapins), all the members of the clade Toxicofera (the genera Anguis, Chamaeleo 123
and all the snakes), and all the members of the clade Episquamata (all the specimens except 124
the terrapins and the lizard genera Tarentola, Hemidactylus and Chalcides) were forced 125
monophyletic. For the amphibian analysis all the members of the clade Anura (all the 126
specimens except the Caudata; genera Calotriton, Triturus, Ichthyosaura, Lissotriton, 127
Pleurodeles, Salamandra and Chioglossa), all the members of the clade Neobatrachia (genera 128
Bufo, Hyla, Pelophylax and Rana), all the members of the clade formed by Neobatrachia plus 129
the genera Pelobates and Pelodytes, all the members of the clade Pleurodelinae (genera 130
Pleurodeles, Calotriton, Ichthyosaura, Lissotriton and Triturus), and all the members of the 131
clade formed by the genera Calotriton, Ichthyosaura, Lissotriton and Triturus, were forced 132
monophyletic. 133
Intra-specific phylogenetic trees 134
We used JMODELTEST v2.1.3 to select the best-fit model of evolution for each fragment, 135
following the Bayesian information criterion (BIC). When analysing concatenated sequences, 136
the dataset was partitioned by mtDNA fragment to be run under the corresponding evolutionary 137
model. We used a relaxed clock model and a coalescence constant size model as tree priors for 138
all phylogenetic inferences. Three individual runs of Markov Chain Monte Carlo (MCMC) 139
analyses were run for 1x108 generations, sampling every 10 000 generations, and discarding 140
10% trees as burn-in. Parameter convergence was verified by visual inspection of trace plots to 141
ensure good mixing of chains and by examining the effective sample sizes (ESSs) using 142
TRACER v1.6 (all parameter values of ESS were above 1000). The remaining trees were used 143
to obtain the subsequent maximum clade credibility summary tree with posterior probabilities for 144
each node using TREEANNOTATOR. All of the above mentioned trees were visualized with 145
FIGTREE 1.4.0 (http://tree.bio.ed.ac.uk/software/figtree/). Finally, we compared phylogenetic 146
trees obtained with published trees (when available) to confirm that our results did not 147
significantly differ from previous topologies. Species for which we could infer a well-supported 148
phylogenetic tree were retained for further analysis, summing up to 14 amphibian and 19 reptile 149
species (annex 2), while the remaining taxa were discarded. 150
151
Table S3.1 - Primers and Sequencing Program used for each gene 152
Gene Primers Sequencing Program Notes 12S 12Sa -AAA AAG CTT CAA
ACT GGG ATT AGA TAC CCC ACT AT- (Kocher et al. 1989)
95° - 10 min, (92º - 30’’, 50° – 40’’, 72º - 45’’) 35x, 72º - 5 min
All species - only the Fw direction was sequenced
12Sb –TGA CTG CAG AGG GTG ACG GGC GGT GTG T –
All species
ND4+tRNA-His
ND4 –CAC CTA TGA CTA CCA AAA GCT CAT GTA GAA GC- (Arevalo et al. 1994)
95° - 10 min, (92º - 30’’, 50° – 40’’, 72º - 45’’) 35x, 72º - 5 min
Leu –CAT TAC TTT TAC TTG GAT TTG CAC CA
All species - only the Rv direction was sequenced
16S 16SH –CCG GTC TGA ACT CAG ATC ACG T-
95° - 10 min, (95º - 30’’, 48° – 45’’, 72º - 1 min) 40x, 72º - 5 min
All species
16SL –CGC CTG TTT ATC AAA AAC AT-
All species - only the Fw direction was sequenced
CYTB cytb1 -CCA TCC AAC ATC TCA GCA TGA TGA AA- (modified primers from
95° - 10 min, (94º - 30’’, 50° – 30’’, 72º - 45’’) 35x, 72º - 5 min
Coronella girondica, Coronella austriaca, Psammodrommus algirus, Psammodrommus hispanicus, Lacerta schreiberi
cytb2 -CCC TCA GAA TGA TAT TTG TCC TCA - (modified primers from
Coronella girondica, Coronella austriaca, Psammodrommus algirus, Psammodrommus hispanicus, Lacerta schreiberi, Chalcides bedriagai
S1F -TTC AAC TAC AAA AAC CTA ATG ACC C-
Chalcides bedriagai
L14919 (trna GLU) -AAC CAC CGT TGT ATT TCA ACT-
Timon lepidus
H16064 (trna THR) -CTT TGG TTT ACA AGA ACA ATG CTT TA-
Natrix natrix 1, Natrix maura and Timon lepidus
CBF -AAC CTC CTC TCA GCA ATA CC-
Timon lepidus
CBR -CCT GTG GGG TTG TTT GAA-
Timon lepidus
L14724 -GAC CTG CGG TCC GAA AAA CCA-
Natrix natrix 1 and Natrix maura
MVZ15 –GAA CTA ATG GCC CAC ACW WTA CGN AA -
All amphibian species except Pelobates cultripes
MVZ16 -AAA TAG GAA RTA TCA YTC TGG TTT RAT–
All amphibian species except Pelobates cultripes
L15162 –GCA AGC TTC TAC CAT GAG GAC AAA TAT C-
Pelobates cultripes - only the Fw direction was sequenced
H15915 –GGA ATT CAT CTC TCC GGT TTA CAA GA-
Pelobates cultripes - only the Fw direction was sequenced
CR PRO –CGC CAC TGG CAC CCA AGG CCA AAA TTC T-
(D. Buckley, unpublished) 95° - 10 min, (94º - 30’’, 50° – 30’’, 72º - 45’’) 35x, 72º - 5 min
All species - only the Fw direction was sequenced
PHE –TAT CTT CAG TGC YGC GCT TTW ATT TAA- (D. Buckley, unpublished)
All species
C-MOS Lsc1 (Mos-F) –CTC TGG KGG CTT TGG KKC TGT STA CAA GG-
95° - 10 min, (94º - 1 min, 52° – 1 min, 72º - 1 min) 35x, 72º - 5 min
All species
Lsc2 (Mos-R) -GGT GAT GGC AAA NGA GTA GAT GTC TGC-
All species
MC1R MC1RF –AGG CNG CCA TYG TCA AGA ACC GGA ACC-
95° - 10 min, (94º - 30’’, 54° – 30’’, 72º - 1 min) 35x, 72º - 5 min
All species
MC1RR –ACT CCG RAA GGC RTA AAT GAT GGG GTC CAC-
All species
RAG1 L2408 –TGCACTGTGACATTGGCAA –
95° - 5 min, (94º - 30’’, 50° – 45’’, 72º - 1 min) 40x, 72º - 7 min
All species
H2920 –GCCATTCATTTTYCGAA –
All species
1 – Presently referred as Natrix astreptophora 153 154
155
General Mixed Yule-Coalescent model 156
The GMYC method optimizes the threshold node depth for an ultrametric tree denoting the 157
transition from a neutral coalescent-based model of population differentiation to species 158
diversification under a Yule pure-birth model. Finding this threshold allows identifying putative 159
independently evolving lineages. The Bayesian version of the GMYC model allows accounting 160
for uncertainty in the tree topology and branch lengths. We ran the bGYMC model using the 161
function bgmyc.multiphylo on 100 random trees from the posterior distribution generated in 162
BEAST. Simulations were set at 50 000 generations with 40 000 burn-in, sampling every 100 163
generations. The upper threshold for the number of lineages was set equal to the total number 164
of haplotypes of each tree. To identify the mean number of lineages estimated by the posterior 165
probability of the Bayesian GMYC, we used the function bgmyc.point, setting the threshold 166
equal to 0.5. 167
Phylin 168
To identify the spatial occurrence of each identified lineage within the area of occurrence of the 169
species (figure 1A, step 2.5), we used a modified method of the Kriging interpolation, 170
implemented in the R package phylin, which allows the usage of a genetic distance matrix to 171
derive a model of spatial dependence. To calculate the probability of occurrence of each intra-172
specific lineage, we first calculated a matrix of genetic distances, from the cophenetic distances 173
between distinct haplotypes in the consensus ultrametric tree for each species. Subsequently, 174
we generated a matrix of geographic distances between the locations where all tissue samples 175
were collected, by calculating euclidean distances from geographic coordinates. The genetic 176
and geographic distances matrices were then used to produce a semi-variogram with the 177
package phylin using the function gen.variogram. Next, we fitted a model to the variogram using 178
the function gv.model. Parameters of this function such as model, sill and range were manually 179
calibrated in order to optimize model fit to the data. The nugget value was set to zero, assuming 180
that samples collected in the same location are genetically identical. Finally, the model was 181
used to calculate the probability of occurrence of each lineage within the overall Iberian range of 182
the species. 183
Supplementary material 4 – GenBank Accession numbers of all genes/samples 184 included in this study 185
Table S4.1 – GenBank (https://www.ncbi.nlm.nih.gov/genbank/) accession numbers of the samples used 186
for the inter-specific phylogenetic analysis of amphibian taxa. Codes of samples in yellow represent 187
sequences obtained with the present study. Codes marked in blue represent sequences obtained from 188
previous studies. Cells in white represent missing data. 189
Taxa Family 12S 16S CYTB RAG1
Frogs
Alytes cisternasii Alytidae GU086775 GU086779 AY442019
Alytes dickhilleni Alytidae AY333672 AY333710 AY442020 DQ019494
Alytes obstetricans Alytidae
KY762015 KY762039
AY514029 AY58333
Bufo calamita 1 Bufonidae FJ882809 KY762040 L10964 EU497610
Bufo spinosus Bufonidae KY762016 KY762041 AB159262 KJ544910 Discoglossus galganoi galganoi Discoglossidae DQ283243 AY236831 NC_006690 AY583338 Discoglossus galganoi jeanneae Discoglossidae AY347472 AY333720 DQ902149 JQ626772 Discoglossus pictus Discoglossidae AY333685 AY333723 AY442085 AY364202 Hyla meridionalis Hylidae EF566953 FJ882757 FJ226925 FJ227085 Hyla molleri Hylidae KY762017 JN800891 FJ226918 FJ227101 Pelobates cultripes Pelobatidae AY364341 AY333689 GU983095 AY323758 Pelodytes ibericus Pelobatidae DQ642137 DQ642112 AY236779
Pelodytes punctatus Pelodytidae DQ283111 DQ283111 AY236783 AY364203 Pelophylax perezi Ranidae AY332763 KY762042 DQ902145
Rana dalmatina Ranidae KY762018
KY762043 AY147962 KC798654
Rana iberica Ranidae AY043043 AY147944 AY147965 KC798673
Rana pyrenaica Ranidae EU746401 AY147950 EU746403 KC798702
Rana temporaria Ranidae KY762019 KY762044 KY762161 AY323776 Salamanders
Calotriton arnoldi Salamandridae DQ092300 DQ092282 DQ092240 KC665968
Calotriton asper Salamandridae AY147258 EF107160 FJ403325 EF107283 Chioglossa lusitanica Salamandridae EU880308 EU880308 DQ821196 AY583347
Ichthyosaura alpestris Salamandridae
KY762020 KY762045 EF089335
Lissotriton boscai Salamandridae DQ092287 DQ092268 DQ821219
Lissotriton helveticus Salamandridae DQ092286 DQ092267 DQ821239
Pleurodeles waltl Salamandridae
KY762021 KY762046 KY762162 AY523736
Salamandra salamandra Salamandridae DQ283440 DQ283440 KY762163 AY583352
Triturus marmoratus Salamandridae
KY762022 DQ092231 AY583354
Triturus pygmaeus Salamandridae DQ092293 KY762047 DQ821260.1| 1 – Presently referred as Epidalea calamita 190 191
192
Table S4.2 – GenBank (https://www.ncbi.nlm.nih.gov/genbank/) accession numbers of the samples used for the inter-specific phylogenetic analysis of reptile taxa. Codes of 193
samples in yellow represent sequences obtained with the present study. Codes marked in blue represent sequences obtained from previous studies. Cells in white represent 194
missing data. 195
Taxa Family 12S 16S CMOS CYTB MC1R RAG1
Tortoises
Emys orbicularis Emydidae KY762023 KY762048 KY762067 KY762164 KY762180
Mauremys leprosa Geoemydidae KY762024 KY762049 KY762068 KY762165
Testudo graeca Testudinidae KY762025 KY762050 KY762069 KY762166
Testudo hermanni Testudinidae KY762026 KY762051 KY762070 KY762167
Geckos
Tarentola mauritanica Phyllodactylidae EU443255 HM014589 AF363566 AF364327 KY762213 KY762181
Hemidactylus turcicus Gekkonidae HQ675926 KY762052 AF363540 AF364319 KY762214 KY762182
Skinks
Chalcides bedriagai Scincidae EU277909 EU278041 KY762071 EU278148
Chalcides striatus Scincidae EU277987 EU278068 AY234232 EU278232 KY762215 KY762183
Lizards
Acanthodactylus erythrurus Lacertidae HQ616540 AY633436 HQ616540 HQ616540 HQ616540 HQ616540
Algyroides marchi Lacertidae GQ142080 GQ142103 GQ142146 GQ142133 KY762216 GQ142156
Iberolacerta aranica Lacertidae AY151955 AF440612 AY152029 AY267239 KY762217 KY762184
Iberolacerta aurelioi Lacertidae KY762027 AF440610 AY152025 AY267238 KY762218 KY762185
Iberolacerta bonnali Lacertidae AY151970 AF080292 AY152035 AY267240 KY762219 KY762186
Iberolacerta cyreni Lacertidae AY151928 KY762053 AY152009 AY267232 KY762220 HQ616539
Iberolacerta galani Lacertidae DQ497135 KY762054 DQ497120 DQ497074 KY762221 KY762187
Iberolacerta martinezricai Lacertidae PHSAS1_ab1 AF440609 KY762072 KY762168 KY762222 KY762188
Iberolacerta monticola Lacertidae AY151940 AF440604 DQ097146 KY762169 KY762223 EF632220
Lacerta agilis Lacertidae DQ097096 DQ658846 EU365405 GQ142118 KY762224 EF632222
Lacerta bilineata Lacertidae KY762055 KY762073 KY762170 KY762225 KY762189
Lacerta schreiberi Lacertidae AF206591 DQ097097 EU365406 AF372119 KY762226 KY762190
Podarcis bocagei Lacertidae AF469425 DQ081077 AF315399 AF469426 KY762227 KY762191
Podarcis carbonelli Lacertidae AY214449 DQ081080 KY762074 DQ081140 KY762228 KY762192
Podarcis guadarramae guadarramae Lacertidae KY762029 KY762056 KY762075 KY762171 KY762229
Podarcis guadarramae lusitanica Lacertidae KY762030 KY762057 AF372084 KY762230
Podarcis hispanica Albacete/ Murcia Lacertidae HQ898190 HQ898056 HQ898037
Podarcis hispanica Galera Lacertidae DQ081070 DQ081095 DQ081146
Podarcis hispanica sensu stricto Lacertidae AY134712 AY134677
Podarcis liolepis Lacertidae KY762031 KY762058 KY762076 KY762172
Podarcis muralis Lacertidae AJ001470 AY896190 EF632282 DQ646343 KY762231 KY762193
Podarcis vaucheri Lacertidae HQ898232 HQ898060 HQ898041
Podarcis virescens Lacertidae KY762032 KY762059 KY762077 KY762173 KY762232 KY762194
Psammodromus algirus Lacertidae DQ298635 AY217970 AY151998 EU116517 KY762233 KY762195
Psammodromus edwardsianus Lacertidae DQ298607 DQ298677 EF632285 FJ587571 EF632242
Psammodromus hispanicus Lacertidae KY762033 KY762060 KY762078 KY762174 KY762234 KY762196
Psammodromus occidentalis Lacertidae FJ587746_1
Timon lepidus Lacertidae AF206595 GQ142094 AY151994 GQ142119 KY762235 EF632247
Timon nevadensis Lacertidae AF378944 AF378949 EU365408 AF379006 JF732931
Zootoca vivipara Lacertidae KY762035 KY762064 KY762080 KY762177 KY762244 KY762205
Worm Lizards
Blanus cinereus Blanidae NC_012433 EF036388 DQ324864 NC_012433 KJ624894
Blanus mariae Blanidae KJ624872 AY444019 KY762175 KY762236 KY762204
Snakes
Coronella austriaca Colubridae AY122836 EU022640 AY486954 AY122752 KY762237 KY762197
Coronella girondica Colubridae AY122835 EU022641 AF471113 AF471088 KY762238 KY762198
Hemorrhois hippocrepis Colubridae DQ451992 102 AY486940 DQ451987 KY762239 KY762199
Hierophis viridiflavus Colubridae AY541505 AY376774 AY486949 AY486925 KY762240 KY762200
Macroprotodon brevis Colubridae KY762034 KY762061 KY762079 KY762176 KY762241 KY762201
Rhinechis scalaris Colubridae AY122802 KY762062 AY486956 AY122718 KY762242 KY762202
Zamenis longissimus Colubridae AY122783 KY762063 DQ902072 DQ902114 KY762243 KY762203
Natrix maura Natricidae
AF402623 KY762065 KY762081 AY866530 030145M_F_
KY762245 KY762206
Natrix natrix 1 Natricidae AF158461 AF158530 AF471121 AY487756 KF258660 KY762207
Malpolon monspessulanus Psammophiidae KY762036 AY643354 KY762082 KY762178 KY762246 KY762208
Vipera aspis Viperidae JN870190 KY762083 AY321099 KY762247 KY762209
Vipera latastei Viperidae KY762037 KY762066 AY321094
Vipera seoanei Viperidae KY762038 AJ275782 KY762084 DQ186030 KY762248 KY762212
Limbless lizards
Anguis fragilis Anguidae EU443256 EU443256 AY099972 AY099996 KY762249 KY762210
Chameleons
Chamaeleo chamaeleon Chamaeleonidae AF372133 KY762179 KY762250 KY762211
1 – Presently referred as Natrix astreptophora196
Supplementary material 5 – Data Compilation Details 197
Table S5.1 – List of amphibian species, number of locations from where DNA was sampled (LOC), 198 number of total sequences compiled from literature PubSeq), number of mtDNA sequences obtained in 199 this project (LabSeq) and respective Genbank accession numbers (GB Acession n), and references from 200 published sequences. 201
Species Gene LOC Lit. Seq. Lab. Seq.
GB Acession n References
Frogs
Alytes cisternasii ND4 38 70 - 1
Alytes dickhilleni Not enough data
Alytes obstetricans ND4 19 27 - 2
Bufo spinosus No intra-specific variation found 3,4
Bufo calamita 1 No intra-specific variation found 5
Discoglossus galganoi
Cytb 71 86 - 6-10
ND4 71 69 -
Hyla molleri No intra-specific variation found 11,12
Hyla meridionalis COI 23 28 - 13,14
Pelobates cultripes Cytb 20 49 1 KY762129 15-17
Pelodytes sp. Cytb 138 367 -
17-19 ND4 137 340 -
Pelophylax perezi Not enough data
Rana dalmatina Not enough data
Rana iberica Not enough data
Rana pyrenaica No intra-specific variation found 20
Rana temporaria 16S 2 2
21 Cytb 51 304 2 KY762154 - KY762155
Salamanders
Chioglossa lusitanica
Cytb 18 44 1 KY762088 22
Calotriton asper No intra-specific variation found 23
Calotriton arnoldi No intra-specific variation found 23
Lissotriton boscai ND4 65 103 - 24
CR 65 103 -
Lissotriton helveticus
COI 32 96 - 25
CR 32 96 -
Ichthyosaura alpestris
16S 7 25 - 26
ND4 7 25 -
Pleurodeles waltl Cytb 35 24 24 KY762130 - KY762153 27,28
Salamandra salamandra
Cytb 54 104 - 29,30
Phylogenetic tree not well supported
Triturus marmoratus
ND4 41 235 7 KY762255 - KY762261 31-33
Triturus pygmaeus ND4 51 438 - 31-33 1 – Presently referred as Epidalea calamita 202
203
204
Table S5. 2 – List of reptile species, number of locations from where DNA was sampled (LOC), number of 205 total sequences compiled from literature PubSeq), number of mtDNA sequences obtained in this project 206 (LabSeq) and respective Genbank accession numbers (GB Acession n), and references from published 207 sequences. 208
Species Gene LOC Pub Seq Lab Seq GB Acession n References
Tortoises
Emys orbicularis No intra-specific variation found
Mauremys leprosa Cytb 51 131 - 38,39
Testudo graeca No intra-specific variation found
Testudo hermanni No intra-specific variation found
Geckos
Tarentola mauritanica 12S 87 107 3
KY761999 - KY762001 42-46
16S 87 107 3 KY762012 - KY762014
Hemidactylus turcicus
Cytb 15 16 - 47-49
12S 15 16 -
Skinks
16S 3 3 -
50 Chalcides bedriagai Cytb 37 23 11 KY761983 - KY761993
12S 29 23 4 KY761995 - KY761998
16S 2 2 -
50 Chalcides striatus Cytb 14 14 -
12S 14 14 -
Lizards
Acanthodactylus erythrurus
12S 11 9 1 KY761994
51-53
16S 11 8 3
KY762002 - KY762004
Algyroides marchi Not enough data
Iberolacerta aranica No intra-specific variation found
Iberolacerta aurelioi No intra-specific variation found 54
Iberolacerta bonnali No intra-specific variation found 54
Iberolacerta cyreni No intra-specific variation found 54
Iberolacerta galani No intra-specific variation found 54
Iberolacerta martinezricai
No intra-specific variation found 54
Iberolacerta monticola
No intra-specific variation found 54
Lacerta agilis Not enough data
Lacerta bilineata Not enough data
Lacerta schreiberi Cytb 20 2 KY762089 - KY762090 55-57
Podarcis bocagei
ND4 20 0 58-62
Podarcis carbonelli ND4 15 - 58-62
Podarcis hispanica 12S 133 -
58-61,63 CR 45 -
Podarcis muralis Cytb 34 8 KY762121 - KY762128 54,60
ND4 28 4 KY762251 - KY762254
Podarcis vaucheri ND4 16 - 60,61,64-66
Psammodromus algirus Phylogenetic tree not well supported
67,68
Psammodromus hispanicus
Cytb 11 278 4 69
ND4 10 48 0
Timon lepidus Cytb 107 302 5 KY762156 - KY762160 70
Zootoca vivipara No intra-specific variation found
Worm Lizards
Blanus cinereus ND4 56 80 - 74,75
Blanus mariae ND4 14 19 - 76
Snakes
Coronella austriaca Cytb 23 26 - 77
Coronella girondica Cytb 62 64 4 KY762085 - KY762087
78 16S 63 66 8 KY762004 - KY762011
Hemorrhois hippocrepis
No intra-specific variation found
Hierophis viridiflavus No intra-specific variation found
Macroprotodon brevis No intra-specific variation found
Rhinechis scalaris No intra-specific variation found
Zamenis longissimus Not enough data
Natrix maura Cytb 55 93 30 KY762091 - KY762120
85
Natrix natrix 1 No intra-specific variation found
Malpolon monspessulanus
No intra-specific variation found
Vipera aspis No intra-specific variation found
Vipera latastei cytb 58 58 -
88 ND4 58 58 -
Vipera seoanei No intra-specific variation found
Limbless Lizards
Anguis fragilis Not enough data
Chameleons
Chamaeleo chamaeleon
Not enough data 91
1 – Presently referred as Natrix astreptophora 209 210
211
212
Supplementary material 6 – Phylogenetic Trees 213
As a result of both reptile and amphibian phylogenetic trees being restricted to only Iberian taxa, 214
with the consequent lack of a backbone including representatives of many relevant reptile and 215
amphibian lineages, several nodes of both trees had to be constrained as part of the inference 216
process in BEAST in order to recover the accepted topologies inferred using very complete 217
datasets 92,93 218
The amphibian tree (Figure A6.1) was fairly asymmetric, with one long root-descending branch 219
ramifying into 10 Caudata (Salamanders) taxa, and another shorter root-descending branch 220
ramifying in 17 Anura (Frogs) taxa. The tree contained many short terminal branches 221
representing recent speciation events. The reptile tree (Figure A6.2) was also asymmetric, 222
containing a long root-descending branch ramifying into the four non-Squamate Testudine 223
(Tortoises) taxa and another shorter descending branch ramifying into two groups: one 224
containing two Gekkota (Geckos) taxa, and the other ramifying into the remaining Squamate 225
taxa (skinks, lizards, worm lizards, snakes, chameleons and limbless lizards). There were some 226
clades containing several short terminal branches, which also reflect recent radiations. For 227
instance, the genera Psammodromus, Iberolacerta and Podarcis are known for having relatively 228
recent and complex speciation events. 229
230
231
232
233
234
235
236
Figure S6.1 – Bayesian consensus phylogenetic tree inferred for Iberian Amphibian species based on an alignment of 3577 base pairs of concatenated mitochondrial (366 bp 237
of 12S; 564 of 16S; 1135 bp of CYTB) and nuclear (1512 of RAG1) DNA sequences. Scale bar represents branch length. Black circles on the nodes represent highly supported 238
clades (Bayesian posterior probabilities > 0.95). 239
240
241 242 Figure S6.2 – Bayesian consensus phylogenetic tree inferred for Iberian Reptile species based on an alignment of 4384 base pairs of concatenated mitochondrial (370 bp of 243
12S; 450 of 16S; 1035 bp of CYTB) and nuclear (570 bp of CMOS; 1344 of rag1; 615 of RAG2) DNA sequences. Scale bar represents branch length. Black circles on the 244
nodes represent highly supported clades (Bayesian posterior probabilities > 0.95). 245
Supplementary material 7 – Phylogenetic trees obtained for each species, number of lineagesidentified within each tree, location of tissue samples collected, and predicted spatialoccurrence of each lineage.
This supplementary material presents the results obtained for the intra‐specific phylogenetic analysis including 14amphibian and 19 reptile species. In each page it is presented the topology of the phylogenetic tree obtained. Thetips of the tree represent unique haplotypes, which are classified (and colored) according to one of the lineagesidentified using the bGMYC approach. The maps represent the overall species distribution. In each map, part of thespecies distribution is colored with one of the colors of the tree tips, representing the predicted occurrence of therespective lineage. The remaining species distribution is colored in dark grey. Light grey areas represent areaswhere the species does not occur. Black dots represent the locations where tissue samples assigned to therespective lineage were collected.
Lin1
Lin2 Lin3 Lin4
Alytes cisternasii
Figure S7.1 – Bayesian phylogenetic tree obtained for Alytes cisternasii and maps of the predicted occurrence of each identified lineage (L1‐L4).
Lin1 Lin2 Lin3
Lin4
Alytes obstetricans
Lin5 Lin6
Figure S7.2 – Bayesian phylogenetic tree obtained for Alytes obstetricans, and maps of the predicted occurrence of each identified lineage (L1‐L6).
Lin1 Lin2
Chioglossa lusitanica
Figure S7.3 – Bayesian phylogenetic tree obtained for Chioglossa lusitanica, and maps of the predicted occurrence of each identified lineage (L1‐L2).
Lin1 Lin2
Discoglossus galganoi
Figure S7.4 – Bayesian phylogenetic tree obtained for Discoglossus galganoi , and maps of the predicted occurrence of each identified lineage (L1‐L2).
Lin1 Lin2
Hyla meridionalis
Figure S7.5 – Bayesian phylogenetic tree obtained for Hyla meridionalis, and maps of the predicted occurrence of each identified lineage (L1‐L2).
Lin1 Lin2
Ichthyosaura alpestris
Figure S7.6 – Bayesian phylogenetic tree obtained for Ichthyosaura alpestris, and maps of the predicted occurrence of each identified lineage (L1‐L2).
Lin1 Lin2
Lin3
Lin5
Lin8 Lin9 Lin10
Lin11 Lin12 Lin13
Lin4
Lissotriton boscai
Lin6
Lin7
Figure S7.7 – Bayesian phylogenetic tree obtained for Lissotriton boscai, and maps of the predicted occurrence of each identified lineage (L1‐L13).
Lin1
Lin2
Lissotriton helveticus
Lin3
Lin4
Lin5
Lin6 Figure S7.8– Bayesian phylogenetic tree obtained for Lissotriton helveticus, and maps of the predicted occurrence of each identified lineage (L1‐L6).
Lin1 Lin2
Pelobates cultripes
Figure S7.9– Bayesian phylogenetic tree obtained for Pelobates cultripes, and maps of the predicted occurrence of each identified lineage (L1‐L2).
Lin1
Lin2
Pelodytes complex
Lin3
Lin4
Lin5
Lin6
Lin7
Figure S7.10– Bayesian phylogenetic tree obtained forPelodytescomplex , and maps of the predicted occurrence of each identified lineage (L1‐L7).
Lin1 Lin2
Pleurodeles waltl
Lin3
Lin4 Lin5
Figure S7.11– Bayesian phylogenetic tree obtained forPleurodeles waltl, and maps of the predicted occurrence of each identified lineage (L1‐L5).
Lin1 Lin2
Rana temporaria
Lin3
Lin4 Lin5 Lin6
Figure S7.12– Bayesian phylogenetic tree obtained for Rana temporaria, and maps of the predicted occurrence of each identified lineage (L1‐L6).
Lin1 Lin2
Triturus marmoratus
Figure S7.13– Bayesian phylogenetic tree obtained for Triturus marmoratus, and maps of the predicted occurrence of each identified lineage (L1‐L2).
Lin1 Lin2
Triturus pygmaeus
Figure S7.14– Bayesian phylogenetic tree obtained for Triturus pygmaeus, and maps of the predicted occurrence of each identified lineage (L1‐L2).
Lin1 Lin2
Acanthodactylus erythrurus
Lin3
Lin4 Lin5
Figure S7.15– Bayesian phylogenetic tree obtained for Acanthodactylus erythrurus, and maps of the predicted occurrence of each identified lineage (L1‐L5).
Lin1 Lin2
Blanus cinereus
Figure S7.16– Bayesian phylogenetic tree obtained for Blanus cinereus and maps of the predicted occurrence of each identified lineage (L1‐L2).
Lin1 Lin2
Blanus mariae
Lin3
Figure S7.17– Bayesian phylogenetic tree obtained for Blanus mariae and maps of the predicted occurrence of each identified lineage (L1‐L3).
Lin1 Lin2
Chalcides bedriagai
Lin3
Figure S7.18– Bayesian phylogenetic tree obtained for Chalcides bedriagai and maps of the predicted occurrence of each identified lineage (L1‐L3).
Lin1 Lin2
Chalcides striatus
Figure S7.19– Bayesian phylogenetic tree obtained for Chalcides striatus and maps of the predicted occurrence of each identified lineage (L1‐L2).
Lin1 Lin2
Coronella austriaca
Lin3
Lin4
Figure S7.20– Bayesian phylogenetic tree obtained for Coronella austriaca and maps of the predicted occurrence of each identified lineage (L1‐L4).
Lin1 Lin2
Coronella girondica
Lin3
Figure S7.21– Bayesian phylogenetic tree obtained for Coronella girondica and maps of the predicted occurrence of each identified lineage (L1‐L3).
Lin1 Lin2
Emys orbicularis
Lin3
Lin4 Lin5
Figure S7.22– Bayesian phylogenetic tree obtained for Emys orbicularis and maps of the predicted occurrence of each identified lineage (L1‐L5).
Lin1 Lin2
Hemidactylus turcicus
Figure S7.23– Bayesian phylogenetic tree obtained for Hemidactylus turcicus and maps of the predicted occurrence of each identified lineage (L1‐L2).
Lin1 Lin2
Lacerta schreiberi
Figure S7.24– Bayesian phylogenetic tree obtained for Lacerta schreiberi and maps of the predicted occurrence of each identified lineage (L1‐L2).
Lin1 Lin2
Natrix maura
Figure S7.25– Bayesian phylogenetic tree obtained for Natrix maura and maps of the predicted occurrence of each identified lineage (L1‐L2).
Lin1 Lin2
Podarcis carbonelli
Lin3
Lin4 Lin5 Lin6
Figure S7.26– Bayesian phylogenetic tree obtained for Podarcis carbonelli and maps of the predicted occurrence of each identified lineage (L1‐L6).
Lin1
Lin2
Podarcis hispanica complex
Lin3
Lin4
Lin5 Lin6
Lin7 Lin8
Figure S7.27– Bayesian phylogenetic tree obtained for Podarcis hispanica complex and maps of the predicted occurrence of each identified lineage (L1‐L8).
Lin1 Lin2
Podarcis muralis
Figure S7.28– Bayesian phylogenetic tree obtained for Podarcis muralis and maps of the predicted occurrence of each identified lineage (L1‐L2).
Lin1 Lin2
Podarcis vaucheri
Lin3
Figure S7.29– Bayesian phylogenetic tree obtained for Podarcis vaucheri and maps of the predicted occurrence of each identified lineage (L1‐L3).
Lin1
Lin2
Psammodromus hispanicus complex
Lin3
Lin4
Figure S7.30– Bayesian phylogenetic tree obtained for Psammodromus hispanicus complex and maps of the predicted occurrence of each identified lineage (L1‐L4).
Lin1 Lin2
Tarentola mauritanica
Figure S7.31– Bayesian phylogenetic tree obtained for Tarentola mauritanica and maps of the predicted occurrence of each identified lineage (L1‐L2).
Lin1
Lin2
Timon lepidus
Lin3
Lin4
Lin5
Figure S7.32– Bayesian phylogenetic tree obtained for Timon lepidus and maps of the predicted occurrence of each identified lineage (L1‐L5).
Lin1 Lin2
Vipera latastei
Lin3
Lin4 Lin5
Figure S7.33– Bayesian phylogenetic tree obtained for Vipera latestei and maps of the predicted occurrence of each identified lineage (L1‐L5).
5�Refer Supplementary material 8 - Spatial Patterns of Diversity – Null models 246�
247�
Figure S8.1 - Spatial patterns of diversity for the amphibian/ reptile species: standardized effect size (A/D) 248�
and respective p-values (B/E). The maps in C/F illustrate the standardized effect size in grid cells where 249�
the null hypothesis was accepted (i.e. with significant p-values (>0.95). 250�
251�
252�
Supplementary material 9 - Least Square regression between species richness 253�
(SR) and lineage richness (LR) for amphibians and reptiles 254�
Since the distributions of intra- specific lineages of a given species are generally allopatric, the 255�
expectation is that lineage richness co-varies linearly with species richness. Exceptions may 256�
occur in contact zones (where two or more lineages co-occur) or in areas where the occurring 257�
lineage could not be assigned using Phylin (possible due to some areas being under sampled, 258�
or to the estimated probability of lineage occurrence being lower than 0.4). In order to analyze 259�
deviations to the expectation, we used ordinary least square regression (OLS). Since not all 260�
species have data for phylogeograpical lineages, we restricted the analysis of species richness 261�
to only the species accounted in the estimation of lineage richness (see supplementary material 262�
2). To identify the areas with deviations from the expected, we mapped OLS’s residuals. 263�
264�
Figure S9.1 – Spatial patterns of OLS residuals for Amphibians (A) and Reptiles (B) 265�
266�
267�
268�
269�270�
References 271� 272�
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