New genetic lineages within Moroccan day geckos Quedenfeldtia (Sphaerodactylidae) revealed by mitochondrial and nuclear DNA sequence data

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DOI: 10.1163/15685381-00003088
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Abstract
The genus Quedenfeldtia is composed of two species, Q. moerens and Q. trachyblepharus, both endemic to the Atlas Mountains region of Morocco. Previous studies recovered two main genetic lineages within each Quedenfeldtia species, although sampling did not cover a substantial portion of their known distribution. In this study we collected individuals from previously unsampled localities of Quedenfeldtia and carried out genetic analyses in order to assess the range of previously identified lineages and the occurrence of additional lineages. Phylogenetic reconstruction based on both mitochondrial (12S and ND4 + tRNA) and nuclear (MC1R) markers revealed that while the new individuals of Q. moerens belong to previously described lineages, two new lineages of Q. trachyblepharus were uncovered from the northern and southern parts of the range. Genetic divergence of these new lineages (8-9% ND4 + tRNA p-distance) was higher than values observed between other lizard sister species. In the future a thorough morphological assessment is needed to complement this study and allow a taxonomic revision of these taxa. The results of this study highlight the importance of biodiversity assessments in mountainous regions characterized by high endemicity but which are difficult to access.
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(A) Bayesian tree based on mitochondrial DNA (mtDNA) sequences (12S and ND4 + tRNA) of Quedenfeldtia. Bayesian posterior probabilities (>0.9) are shown above branches; bootstrap values for Maximum Likelihood (>50) are shown below branches. Main lineages within each species are represented by distinct grey tones (colours online). New samples used in this study are in bold. (B) Parsimony Network based on nuclear DNA sequences (MC1R). Haplotypes are represented by circle with size proportional to their frequency and coloured according to the mitochondrial lineages as defined in the tree. (C) Map with known localities of Quedenfeldtia moerens (small black circles) and Quedenfeldtia trachyblepharus (small white circles) based on Bons and Geniez (1996). Samples analysed in genetic analyses by Barata et al. (2012a) and this study are represented by dark or light grey symbols (coloured online) for Q. moerens and Q. trachyblepharus respectively. Samples analysed in Barata et al. (2012a) are represented by small symbols and new samples analysed in this study by large symbols. Symbols correspond to lineages uncovered by the phylogenetic tree and network for each species: dark grey triangles (Q. moerens, North; light green online), dark grey squares (Q. moerens, South; dark green online), light grey circles (Q. trachyblepharus, Jebel Sirwa; light blue online), light grey triangles (Q. trachyblepharus, Oukaimeden; light purple online), light grey squares (Q. trachyblepharus, North; dark blue online), light grey diamond (Q. trachyblepharus, South; dark purple online). This figure is published in colour in the online version.
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Amphibia-Reptilia 38 (2017): 97-101
Short Notes
New genetic lineages within Moroccan day geckos Quedenfeldtia
(Sphaerodactylidae) revealed by mitochondrial and nuclear
DNA sequence data
David James Harris1, Daniela Rosado1, Raquel Xavier1, Daniele Salvi1,2,
Abstract. The genus Quedenfeldtia is composed of two species, Q. moerens and Q.trachyblepharus, both endemic to the
Atlas Mountains region of Morocco. Previous studies recovered two main genetic lineages within each Quedenfeldtia species,
although sampling did not cover a substantial portion of their known distribution. In this study we collected individuals from
previously unsampled localities of Quedenfeldtia and carried out genetic analyses in order to assess the range of previously
identified lineages and the occurrence of additional lineages. Phylogenetic reconstruction based on both mitochondrial (12S
and ND4 +tRNA) and nuclear (MC1R) markers revealed that while the new individuals of Q. moerens belong to previously
described lineages, two new lineages of Q. trachyblepharus were uncovered from the northern and southern parts of the
range. Genetic divergence of these new lineages (8-9% ND4 +tRNA p-distance) was higher than values observed between
other lizard sister species. In the future a thorough morphological assessment is needed to complement this study and allow a
taxonomic revision of these taxa. The results of this study highlight the importance of biodiversity assessments in mountainous
regions characterized by high endemicity but which are difficult to access.
Keywords: cryptic diversity, genetic diversity, Morocco, mountain endemic, Quedenfeldtia moerens,Quedenfeldtia trachy-
blepharus.
Quedenfeldtia Boettger, 1883 is a genus of di-
urnal geckos endemic to the Atlas Mountain re-
gion in Morocco. Two species are currently rec-
ognized, Quedenfeldtia moerens (Chabanaud,
1916) and Quedenfeldtia trachyblepharus
(Boettger, 1874), which are easy to distin-
guish based on colour patterns and morpholo-
gical traits (Arnold, 1990). While Q. moerens
is more widespread and can be found down
to sea level, Q. trachyblepharus is only known
from high mountain regions (Bons and Geniez,
1996; Harris et al., 2010; Rosado et al., 2016).
Both species present high genetic variation with
1 - CIBIO-InBIO, Centro de Investigação em Biodiversi-
dade e Recursos Genéticos, Campus Agrário de Vairão,
4485-661 Vairão, Portugal
2 - Department of Health, Life and Environmental Sci-
ences, University of L’Aquila, 67100 Coppito, L’Aquila,
Italy
Corresponding author;
e-mail: danielesalvi.bio@gmail.com
two highly divergent lineages each (ND4 +
tRNA genetic p-distance >8%; Barata et al.,
2012a). Such levels of divergence are higher
than values observed between lizard species
(e.g. Ahmadzadeh et al., 2013). Quedenfeldtia
moerens was divided into a northern (Agoudal)
and a southern lineage, with the latter split-
ting into two sublineages [Tafraoute (south) and
Ida-ou-Bouzia (centre)] (Barata et al., 2012a).
These lineages are also partially differentiated
through morphological characters such as head
dimension and trunk length, pholidotic traits
and colour patterns although these differences
are difficult to appreciate in the field (Barata
et al., 2012a). The two main lineages of Q.
moerens are geographically separated by more
than 250 km, and although individuals from the
genus Quedenfeldtia have been reported in the
intermediate area by Bons and Geniez (1996), it
is not known whether these populations belong
to Q. moerens or to Q. trachyblepharus. Results
©Koninklijke Brill NV, Leiden, 2017. DOI:10.1163/15685381-00003088
98 Short Notes
from ecological niche modelling by Barata et
al. (2012a) suggest that the habitat in this re-
gion is suitable for both species, but this has not
yet been confirmed through field observations.
In the case of Q. trachyblepharus, the two di-
vergent genetic lineages were both found in the
southern-central part of the species range (Jebel
Sirwa and Oukaimeden), however neither north-
ern populations nor extreme southern popula-
tions were sampled (Barata et al., 2012a). Mor-
phologically, the lineages within Q. trachyble-
pharus are more difficult to distinguish, with
some biometric traits being partially diagnos-
tic for male specimens (Barata et al., 2012a).
Despite the relatively wide sampling of Barata
et al. (2012a), the genetic affinities of popula-
tions from large portions of the ranges of both
Quedenfeldtia species are still unknown, and
given the high genetic divergence found so far
it would not be unexpected to find similar levels
of divergence throughout the remaining distri-
butional range.
The aims of this study were to investigate
the phylogenetic relationships of Quedenfeld-
tia from previously unsampled regions and to
collect additional distribution data for these
species.
We collected tissues of Quedenfeldtia spp. during spring
2015 and 2016 from previously unsampled localities. We
captured individuals by hand or with a noose, remov-
ing small tail tips which were stored in 96% ethanol in
the tissue collection of CIBIO-InBIO (Centro de Investi-
gação em Biodiversidade e Recursos Genéticos, Univer-
sidade do Porto). After we took digital photographs fol-
lowing procedures of Barata et al. (2012b) the animals
were released at the collection site. For genetic analysis
we used three genetic markers previously employed by
Barata et al. (2012a): partial 12S rRNA and NADH dehy-
drogenase 4 with flanking tRNA-histidine (ND4 +tRNA)
mitochondrial genes, and partial melanocortin receptor 1
(MC1R) nuclear gene. We performed DNA extraction using
a high salt protocol and amplification following Barata et
al. (2012a). Sequences generated in this studied for the12S,
ND4 +tRNA and MC1R fragments were combined with
those downloaded from Genbank from Barata et al. (2012a)
andalignedinGENEIOUS6.0(www.geneious.com)us-
ing the Geneious Alignment algorithm. New sequences
were submitted to GenBank (table 1). Haplotype diversity
for each marker was calculated in DnaSP v.5.5.5 (Librado
and Rozas, 2009). Phylogenetic relationships among mito-
chondrial sequences were inferred using Maximum Like-
lihood (ML) and Bayesian inferences (BI) methods us-
ing, respectively, the software RaxML v.7.4.2 (Stamatakis,
2006), by means of the graphical front-end RAxML GUI
v.1.1.3 (Silvestro and Michalak, 2012), and the BEAST
v.1.7.5 package (Drummond et al., 2012). Trees were rooted
with sequences from three Saurodactylus species (Gen-
bank accession numbers S. mauritanicus EU014316.1; S.
mauritanicus EU014345.1; S. fasciatus EU014299.1; S.
fasciatus EU014343.1) including two previously unpub-
lished sequences of S. brosseti (Genbank accession numbers
KY467067 and KY467053). We implemented the GTR +
G model in RaxML and the following partition scheme se-
lected by PartitionFinder 1.1.1 (Lanfear et al., 2012) under
the Bayesian Information Criterion (BIC): (12S), (ND4 1st
+2nd codon positions), (ND4 3rd codon position +tRNA
histidine). The same partition scheme and the following sub-
stitution models were selected for Bayesian analyses: the
Tab le 1 . Quedenfeldtia samples used in the study. Sample codes (according to CIBIO-InBio’s tissue collection, see methods
section), species, lineage, geographic coordinates and GenBank accession numbers are reported.
Sample Species Lineage Latitude Longitude Accession Numbers
(12S/ND4 +
tRNA/MC1R)
DB981 Q. moerens North 32.127 5.304 KY467054/KY467040/KY467027
DB25222 Q. moerens North 31.878 5.476 KY467061/KY467047/KY467034
DB25183 Q. moerens North 31.858 5.468 KY467060/KY467046/KY467033
DB24164 Q. moerens North 31.621 5.560 KY467059/KY467045/KY467032
DB14751 Q. moerens North 31.133 5.491 KY467057/KY467043/KY467030
DB25247 Q. moerens South 30.457 7.673 KY467062/KY467048/KY467035
DB7996 Q. moerens South 31.236 9.183 KY467065/KY467051/KY467038
DB2024 Q. moerens South 31.156 9.279 KY467058/KY467044/KY467031
DB11939 Q. trachyblepharus North 32.202 6.001 KY467056/KY467042/KY467029
DB9396 Q. trachyblepharus North 31.758 6.288 KY467066/KY467052/KY467039
DB11721 Q. trachyblepharus North 31.796 6.379 KY467055/KY467041/KY467028
DB5054 Q. trachyblepharus Jebel Sirwa 31.301 7.410 KY467063/KY467049/KY467036
DB6830 Q. trachyblepharus South 30.788 8.846 KY467064/KY467050/KY467037
Short Notes 99
HKY +G model for the (12S) and (ND4 1st +2nd codon
positions) partitions and TrN +I+G for the (ND4 3rd
codon position +tRNA histidine) partition. For ML analy-
sis, support was estimated using 1000 bootstrap replicates.
BI analysis was run for 30 million generations under a re-
laxed (lognormal) clock model. We used the software Tracer
1.6 (Rambaut et al., 2014) to check for proper mixing and
convergence of the run. Trees from a stationarity distribu-
tion (burnin =25%) were used to calculate a Maximum
Clade Credibility tree in TreeAnnotator 1.7.5 (Drummond
et al., 2012). We checked electropherograms of MC1R for
polymorphic positions in GENEIOUS, and we inferred hap-
lotypes using PHASE 2.1 (Stephens et al., 2001). Phyloge-
netic relationships between MC1R haplotypes were inferred
by a parsimony network under the 95% probability crite-
rion for a parsimonious connection using the software TCS
1.21 (Clement et al., 2000), and the output was edited using
TCSbu 1.0 (Santos et al., 2016).
Thirteen individuals were newly sequenced
for all three gene regions, and combined with
sequences of the individuals previously ana-
lyzed by Barata et al. (2012a). Lenghts of
the alignments were 400 base pairs (bp) for
12S, 587 bp for ND4 +tRNA (ND4: 519 bp;
tRNAHis: 68 bp) and 602 bp for MC1R. Haplo-
type diversity for the 12S, ND4 +tRNA and
MC1R fragments was 0.978 (±0.007), 0.993
(±0.005), 0.951 (±0.012) respectively. Within
Q. moerens, Bayesian phylogenetic reconstruc-
tion recovered the two previously described
southern and northern lineages. Within Q. tra-
chyblepharus, instead of two lineages, four dis-
tinct lineages were now identified (fig. 1). One
new lineage represents a northern population
that had not been included in earlier assess-
ments (fig. 1C: light grey squares; dark blue
Figure 1. (A) Bayesian tree based on mitochondrial DNA (mtDNA) sequences (12S and ND4 +tRNA) of Quedenfeldtia.
Bayesian posterior probabilities (>0.9) are shown above branches; bootstrap values for Maximum Likelihood (>50) are
shown below branches. Main lineages within each species are represented by distinct grey tones (colours online). New
samples used in this study are in bold. (B) Parsimony Network based on nuclear DNA sequences (MC1R). Haplotypes are
represented by circle with size proportional to their frequency and coloured according to the mitochondrial lineages as defined
in the tree. (C) Map with known localities of Quedenfeldtia moerens (small black circles) and Quedenfeldtia trachyblepharus
(small white circles) based on Bons and Geniez (1996). Samples analysed in genetic analyses by Barata et al. (2012a) and this
study are represented by dark or light grey symbols (coloured online) for Q. moerens and Q. trachyblepharus respectively.
Samples analysed in Barata et al. (2012a) are represented by small symbols and new samples analysed in this study by
large symbols. Symbols correspond to lineages uncovered by the phylogenetic tree and network for each species: dark grey
triangles (Q. moerens, North; light green online), dark grey squares (Q. moerens, South; dark green online), light grey circles
(Q. trachyblepharus, Jebel Sirwa; light blue online), light grey triangles (Q. trachyblepharus, Oukaimeden; light purple
online), light grey squares (Q. trachyblepharus, North; dark blue online), light grey diamond (Q. trachyblepharus, South;
dark purple online). This figure is published in colour in the online version.
100 Short Notes
online), and the other is represented by a sin-
gle individual from the far south of the range
(fig. 1C: light grey diamond; dark purple on-
line). Genetic divergence among these new lin-
eages were high: the new Q. trachyblepharus
northern lineage was 8% divergent (ND4 +
tRNA sequences, p-distance) from its sister
taxa, the Jebel Sirwa lineage, while the ex-
treme southern lineage diverged by 9% from the
Oukaimeden lineage (ND4 +tRNA sequences,
p-distance). The phylogenetic network based
on MC1R haplotypes showed that also at the nu-
clear level the two lineages of Q. moerens and
the four lineages of Q. trachyblepharus are dis-
tinct and do not share haplotypes, thus confirm-
ing phylogenetic results based on mitochondrial
sequences (fig. 1A, B). Divergence between lin-
eages at MC1R (average MC1R p-distances be-
tween groups: 1%) is as high, or even higher,
than divergences found between other proposed
cryptic species of geckos (e.g. Rocha et al.,
2011; fig. 1B). On the other hand, limited ge-
netic variation within lineages is observed both
at mitochondrial and nuclear loci (fig. 1A, B;
Barata et al., 2012a). According to Barata et al.
(2012a) while lineage divergence started during
the Middle Miocene the low intra-lineage vari-
ation could be explained by demographic and
range contraction of lineages in glacial refugia
during the Pliocene-Pleistocene.
All samples retrieved from the 250 km region
separating the two allopatric Q. moerens lin-
eages were identified both morphologically (in
the field) and genetically as Q. trachyblepharus.
Thus, although ecological niche modelling re-
sults of Barata et al. (2012a) suggest a possible
sympatry between Q. moerens and Q. trachy-
blepharus, so far only Q. trachyblepharus has
been found in this region. Moreover, the same
models also predicted the existence of a region
with unfavorable habitat for the occurrence of
Q. moerens that would potentially isolate popu-
lations from the Jbel Saghro region, located in
the southeastern part of the species range. How-
ever, the single sample from the Jbel Saghro re-
gion analysed in this study (DB14751), while
representing a new haplotype, exhibited shal-
low divergence to samples from the northern
lineage, suggesting recent isolation.
The distribution of the genetic lineages of Q.
trachyblepharus remains less clear. So far Q.
trachyblepharus has only been reported from
high altitudes in the Middle and High Atlas
Mountains. Interestingly, some of the highly
distinct lineages have been found in relatively
close proximity, and with apparently suitable
habitat between them (personal field observa-
tion). A similar pattern has been observed in the
high altitude lizard Atlantolacerta andreanskyi
which shows seven highly divergent lineages
separated by as little as 10 km (Barata et al.,
2012b). Because of this, extensive assessments
across such regions would be valuable to accu-
rately delimit the ranges of these lineages and
to determine the possibility of gene flow or the
occurrence of hybrids. Additionally, a thorough
morphological assessment is needed for the two
new lineages of Q. trachyblepharus identified in
this study as although Arnold (1990) and Barata
et al. (2012a) collected extensive data on the
species, all examined specimens came from the
central-southern regions.
To conclude, complementary sampling of
Quedenfeldtia has uncovered two additional ge-
netic lineages within Q. trachyblepharus, which
may correspond to distinct species. These data
have helped to delimit the ranges of Queden-
feldtia and indirectly supported some of the re-
sults from ecological niche models for the two
species (Barata et al., 2012a). However, a tax-
onomic revision of the Quedenfeldtia genus
would still require additional data collection to
determine the levels of morphological and eco-
logical variation between lineages. Finally, this
work highlights the need for further biodiversity
assessments in the High Atlas Mountains, sup-
porting the hypothesis that biodiversity is under-
estimated is such remote regions (Ficetola et al.,
2013).
Short Notes 101
Acknowledgements. We thank all the colleagues from
CIBIO who helped with the fieldwork in Morocco, in parti-
cular Mafalda Barata, Ana Perera, and Fernando Martínez-
Freiría. Fieldwork was supported by the British Herpetolog-
ical Society (to DR in 2014) and by the Explorers Fund (to
DR in 2015). Lizards were captured and handled under per-
mit of the Haut Commisariat aux Eaux and Forets of Mo-
rocco (HCEFLCD/DLCDPN/DPRN/DFF N°14/2010). This
work was partially supported by FEDER through the COM-
PETE program, Portuguese national funds through the FCT
(Fundação para a CiênciaeaTecnologia,Portugal).DJH,
RX, and DS are supported by the FCT under the Programa
Operacional Potencial Humano – Quadro de Referência Es-
tratégico Nacional funds from the European Social Fund
and Portuguese Ministério da Educação e Ciência: DJH,
IF-contract IF/01627/2014; RX, IF-contract IF/00359/2015;
DS, post-doctoral grant SFRH/BPD/105274/2014. DS is
currently supported by the program ‘Rita Levi Montalcini’
for the recruitment of young researchers at the University of
L’Aquila.
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  • ... Salvi et al., 2013Salvi et al., , 2014Senczuk et al., 2018). On the other hand, marked differentiation at these nuclear fragments is frequent in potential cryptic reptile species (Sampaio et al., 2015;Harris et al., 2017;Rosado et al., 2017;Mendes et al., 2018). For example, MC1R showed high diversity and almost no haplotype sharing between divergent lineages in Saurodactylus brosseti , and in A. andreanskyi (Barata, Carranza and Harris, 2012). ...
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    Quedenfeldtia (Boettger, 1883) is a genus of diurnal geckos, endemic to the Atlas Mountains in Morocco, with two species being recognized: Quedenfeldtia moerens and Quedenfeldtia trachyblepharus. Quedenfeldtia moerens is found across a wide variety of habitats, from sea level to 3000 m a.s.l., whereas Q. trachyblepharus occupies exclusively high mountain regions reaching up to 4000 m a.s.l. This differentiation, offers an interesting model for study biogeographical patterns and evolutionary scenarios in a North African endemic. Analysis of two mitochondrial (12S rRNA and ND4) and four nuclear (ACM4, MC1R, PDC, and Rag1) DNA markers revealed high genetic variation, consistent with other recent phylogeographical studies, and with the two currently described species. However, within each species, a subdivision into two groups with geographical consistence was found. Multivariate morphological analyses confirmed the existence of two main phenotypes, whereas ecological niche modelling identified various environmental variables associated with the distribution of each species, and helped to predict occurrences outside the confirmed ranges. The results obtained in the present study indicate the possible existence of additional ‘cryptic’ species within this genus, a condition found in many North African reptiles, and particularly common in geckos. In general, North African montane fauna appears to reflect the occurrence of diverse palaeoendemics, as seen in Central Africa Mountain systems, rather than the pattern of recent postglacial recolonization observed in Europe. © 2012 The Linnean Society of London, Biological Journal of the Linnean Society, 2012, ••, ••–••.
  • Article
    While traditionally species recognition has been based solely on morphological differences either typological or quantitative, several newly developed methods can be used for a more objective and integrative approach on species delimitation. This may be especially relevant when dealing with cryptic species or species complexes, where high overall resemblance between species is coupled with comparatively high morphological variation within populations. Rock lizards, genus Darevskia, are such an example, as many of its members offer few diagnostic morphological features. Herein, we use a combination of genetic, morphological and ecological criteria to delimit cryptic species within two species complexes, D. chlorogaster and D. defilippii, both distributed in northern Iran. Our analyses are based on molecular information from two nuclear and two mitochondrial genes, morphological data (15 morphometric, 16 meristic and four categorical characters) and eleven newly calculated spatial environmental predictors. The phylogeny inferred for Darevskia confirmed monophyly of each species complex, with each of them comprising several highly divergent clades, especially when compared to other congeners. We identified seven candidate species within each complex, of which three and four species were supported by Bayesian species delimitation within D. chlorogaster and D. defilippii, respectively. Trained with genetically determined clades, Ecological Niche Modeling provided additional support for these cryptic species. Especially those within the D. defilippii-complex exhibit well-differentiated niches. Due to overall morphological resemblance, in a first approach PCA with mixed variables only showed the separation between the two complexes. However, MANCOVA and subsequent Discriminant Analysis performed separately for both complexes allowed for distinction of the species when sample size was large enough, namely within the D. chlorogaster-complex. In conclusion, the results support four new species, which are described herein.
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    Estimating genealogical relationships among genes at the population level presents a number of difficulties to traditional methods of phylogeny reconstruction. These traditional methods such as parsimony, neighbour-joining, and maximum-likelihood make assumptions that are invalid at the population level. In this note, we announce the availability of a new software package, TCS, to estimate genealogical relationships among sequences using the method of Templeton et al. (1992) .
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    Aim - The incompleteness of information on biodiversity distribution is a major issue for ecology and conservation. Researchers have made many attempts to quantify the amount of biodiversity that still remains unknown. We evaluated whether models that integrate ecogeographical variables with measures of the effectiveness of sampling can be used to estimate biodiversity patterns (species richness) of reptiles in remote areas that have received limited surveys. Location - The Western Palaearctic (Europe, Northern Africa, the Middle East and Central Asia). Methods We gathered data on the distribution of turtles, amphisbaenians and lizards. We used regression models integrating spatial autocorrelation (spatial eigenvector mapping and Bayesian autoregressive models) to analyse species richness, and identified relationships between species richness, ecogeographical features and large-scale measures of accessibility. Results - The two regression techniques were in agreement. Known species richness was dependent on ecogeographical factors, peaking in areas with high temperature and annual actual evapotraspiration, and intermediate cover of natural vegetation. However, richness declined sharply in the least accessible areas. Our models revealed regions where reptile richness is likely to be higher than currently known, particularly in the biodiversity hotspots in the south of the Arabian Peninsula, the Irano-Anatolian region, and the Central Asian mountains. An independent validation data set, with distribution data collected recently throughout the study region, confirmed that combining accessibility measures with ecogeographical variables allows a good estimate of reptile richness, even in remote areas that have received limited monitoring so far. Some remote regions that support very rich communities are covered very little by protected areas. Main conclusions - Integrating accessibility measures into species distribution models allows biologists to identify areas where current knowledge underestimates the actual richness of reptiles. Our study identifies regions requiring future biodiversity research, proposes a novel approach to biodiversity prediction in poorly studied areas, and identifies potential regions for conservation.
  • Article
    The genus Quedenfeldtia of Morocco is generally considered monotypic but actually contains two well-defined species, Q. trachyblepharus and Q. moerens, which can be distinguished by both external and osteological characters. The lowland Q. moerens has a number of apomorphies not found in Q. trachyblepharus which has a restricted, mainly highland, range and is more generally primitive in its anatomy. Because of this, Q. trachyblepharus cannot have arisen from an isolated population of the widespread Q. moerens, and the two forms are likely to have had substantially separate histories.
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    Background Atlantolacerta andreanskyi is an enigmatic lacertid lizard that, according to the most recent molecular analyses, belongs to the tribe Eremiadini, family Lacertidae. It is a mountain specialist, restricted to areas above 2400 m of the High Atlas Mountains of Morocco with apparently no connection between the different populations. In order to investigate its phylogeography, 92 specimens of A. andreanskyi were analyzed from eight different populations across the distribution range of the species for up to 1108 base pairs of mitochondrial DNA (12S, ND4 and flanking tRNA-His) and 2585 base pairs of nuclear DNA including five loci (PDC, ACM4, C-MOS, RAG1, MC1R). Results The results obtained with both concatenated and coalescent approaches and clustering methods, clearly show that all the populations analyzed present a very high level of genetic differentiation for the mitochondrial markers used and are also generally differentiated at the nuclear level. Conclusions These results indicate that A. andreanskyi is an additional example of a montane species complex.
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    Additional data on the distribution of terrestrial herpetofauna from Morocco are pre-sented, based on fieldwork carried out in March and May 2008. Thirty-eight species were recorded from 78 localities. Some of these represent considerable range extensions for the species, indicating that more prospection is needed to complement the existing knowledge of herpetofauna from this country.