Genetic Variability Within the Oudri's Fan-Footed Gecko Ptyodactylus
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YMPEV 3409 No. of Pages 6, Model 5G ARTICLE IN PRESS 30 October 2009 Molecular Phylogenetics and Evolution xxx (2009) xxx–xxx 1 Contents lists available at ScienceDirect Molecular Phylogenetics and Evolution journal homepage: www.elsevier.com/locate/ympev 2 Short Communication 3 Genetic variability within the Oudri’s fan-footed gecko Ptyodactylus oudrii 4 in North Africa assessed using mitochondrial and nuclear DNA sequences a, a,b 5 Ana Perera *, D. James Harris 6 a CIBIO-UP, Centro de Investigação em Biodiversidade e Recursos Genéticos, Campus Agrário de Vairão, 4485-661 Vairão, Portugal 7 b Departamento de Zoologia e Antropologia, Faculdade de Ciências da Universidade do Porto, 4099-002 Porto, Portugal 8 article info abstract 2410 11 Article history: We analyse for the first time the genetic diversity within Ptyodactylus oudrii across the Maghreb. Two 25 12 Received 12 March 2009 mitochondrial (12s rRNA and 16s rRNA) and two nuclear (C-mos and ACM4) markers are used. The 26 13 Revised 24 August 2009 results confirm the specific status of P. oudrii and show high levels of intraspecific variability, indicative 27 14 Accepted 9 October 2009 of species complex. Lineages found are geographically concordant, and show similar patterns to that 28 15 Available online xxxx found in other species from the region. The study highlights once more the importance of the region 29 as a source of genetic diversity. 30 16 Keywords: Ó 2009 Published by Elsevier Inc. 31 17 Ptyodactylus 18 Maghreb 19 Gecko 20 Genetic variability 21 Nuclear markers 22 Mitochondrial markers 23 32 33 34 1. Introduction Ptyodactylus is one of the most characteristic genera of geckos, 56 easily recognisable by their widely dilated toes formed by two diver- 57 35 The traditional method of delimiting species, based on morpho- gent series of lamellae, similar to a fan (Schleich et al., 1996). Up to 58 36 logical characters changed drastically in the end of the seventies now, six species have been described on the basis of their morphol- 59 37 with the development of molecular techniques (Avise et al., ogy and distribution (Sindaco and Jeremcenko, 2008). Three of them 60 38 1979). Since then many new taxa have been redefined and cryptic are found in the Western Asia region (P. puiseuxi, P. guttatus, P. has- 61 39 species identified based on genetic variation (see for example, Ba- selquistii), one (P. homolepis) is restricted to Pakistan, and the 62 40 ker and Bradley, 2006). Such taxonomic reassessments are more remaining ones (P. ragazzii and P. oudrii) are found in North and Cen- 63 41 evident in groups that have suffered phenomena of morphological tral Africa (Sindaco and Jeremcenko, 2008). This genus has several 64 42 convergence or parallelism or that are morphologically more con- exceptional characteristics, including a high intraspecific morpho- 65 43 servative. A good example among reptiles are geckos. Most of the logical variability, and a wide range of ecological habits (Heimes, 66 44 species are very similar in general morphology and habits, and tax- 1987; Werner and Sivan, 1993, 1994). Such variability has compli- 67 45 onomy is based on some characters, such as digit morphology or cated attempts to settle the taxonomy of this group (Werner and Si- 68 46 pupil characteristics, that have been proved to be homoplastic van, 1994; Sindaco and Jeremcenko, 2008). Morphological studies 69 47 (Russell and Bauer, 2002; Gamble et al., 2008, and references on the three species living in Israel showed relatively high variation 70 48 therein). Studies on gecko phylogeny have revealed up to now at both intra and inter specific levels (Werner and Sivan, 1993). 71 49 the presence of cryptic species that were not detected by tradi- However, with the exception of a general analysis on geckos includ- 72 50 tional morphology based methods (Gamble et al., 2008) or even ing a couple of samples of Ptyodactylus (Gamble et al., 2008), no de- 73 51 species wrongly assigned to a genus (Carranza et al., 2002). Addi- tailed phylogenetic study has been performed to decipher the 74 52 tionally, most of the studiesUNCORRECTED show that geckos have exceptional genetic variation PROOF within this group. 75 53 levels of intraspecific variation in mtDNA (Harris et al., 2004a; Je- The Oudri’s fan-footed gecko P. oudrii is the most recent taxo- 76 54 Q1 sus et al., 2008) that might be a consequence of comparatively fas- nomically redefined species within this genus (Heimes, 1987). 77 55 ter rates of mtDNA evolution (Harris et al., 2004a). Considered a subspecies of P. hasselquistii, it was recently raised 78 to specific level based on electrophoretic and morphological evi- 79 dences as well as its completely different distribution pattern (Hei- 80 * Corresponding author. Fax: + 351 252 661 780. mes, 1987). P. oudrii is a saharien species distributed across the 81 E-mail addresses: [email protected] (A. Perera), [email protected] (D. Maghreb, including South Morocco, Central-Northern Algeria and 82 James Harris). 1055-7903/$ - see front matter Ó 2009 Published by Elsevier Inc. doi:10.1016/j.ympev.2009.10.020 Please cite this article in press as: Perera, A., James Harris, D. Genetic variability within the Oudri’s fan-footed gecko Ptyodactylus oudrii in North Africa assessed using mitochondrial and nuclear DNA sequences. Mol. Phylogenet. Evol. (2009), doi:10.1016/j.ympev.2009.10.020 YMPEV 3409 No. of Pages 6, Model 5G ARTICLE IN PRESS 30 October 2009 2 A. Perera, D. James Harris / Molecular Phylogenetics and Evolution xxx (2009) xxx–xxx 83 Western Tunisia (Bons and Geniez, 1996). This region has several species, we included two individuals from P. hasselquistii and three 111 84 geological, topographic, and climatic characteristics well differen- from P. ragazzii, plus the only four Ptyodactylus sequences available 112 85 tiated from the rest of the continent (Bons and Geniez, 1996), giv- from GenBank (two C-mos and two ACM4 sequences from P. has- 113 86 ing rise to complex patterns of diversity (Harris et al., 2004a; selquistii and P. guttatus; GenBank Accession Numbers EU293658, 114 87 Perera et al., 2007; Rato and Harris, 2008). In effect, most of the EU293659, EU293681 and EU293682; Gamble et al., 2008). Mito- 115 88 reptiles studied phylogeographically in this area have revealed chondrial sequences of Phyllodactylus xanti (Accession Numbers 116 89 unexpected levels of mitochondrial DNA variation and the exis- AY763273 and AY763284, Weiss and Hedges, 2007) and Tarentola 117 90 tence, in many cases, of cryptic taxa (Pinho et al., 2006; Mendonça mauritanica (Accession Numbers AY828481 and AY828456, Harris 118 91 and Harris, 2007; Perera et al., 2007; Barata et al., 2008). In partic- et al., 2004b) were used as outgroups. 119 92 ular, the two geckos from this region analysed up to now, Tarentola Total genomic DNA was extracted from small pieces of tail using 120 93 (Harris et al., 2004a) and Saurodactylus (Rato and Harris, 2008), standard saline methods (Sambrook et al., 1989). Fragments of two 121 94 show high levels of genetic variability. mitochondrial regions (12s rRNA and 16s rRNA) and two nuclear 122 95 In this study, we analyse for the first time the genetic variation regions (C-mos and ACM4) were analysed. Primers used in both 123 96 within the Oudri’s fan-footed gecko P. oudrii across the Maghreb. amplification and sequencing were 12sa and 12sb for the 12s rRNA 124 97 We use both mitochondrial (12s rRNA and 16s rRNA) and nuclear (Kocher et al., 1989), 16sa and 16sb for the 16s rRNA (Hedges et al., 125 98 markers (C-mos and ACM4) with different rates of evolution to 1991), TgF and TgR for the ACM4 (Gamble et al., 2008), and G73 126 99 (i) assess the genetic diversity within Ptyodactylus and to confirm and G74 for the C-mos (Gamble et al., 2008). PCR mix was carried 127 100 the recent taxonomic differentiation between P. oudrii and P. has- out in a 25 ll total volume, following conditions described in Har- 128 101 selquistii, ii) analyse the genetic variation within P. oudrii and the ris et al. (1998) for the mitochondrial markers and Gamble et al. 129 102 geographic distribution of the lineages, and (iii) corroborate the (2008) for the nuclear ones. Amplified fragments were sequenced 130 103 possible existence of faster rates of mtDNA evolution observed in on a 310 Applied Biosystem DNA sequencing apparatus. 131 104 gecko species (Harris et al., 2004a; Jesus et al., 2006; Rato and Har- 105 ris, 2008). 2.2. Phylogenetic analyses 132 106 2. Materials and methods Corrected sequencesPROOF were opened in MEGA4 software (Tamura 133 et al., 2007) Sequences were first aligned using ClustalW with de- 134 107 2.1. DNA extraction, amplification and sequencing fault parameters and then adjusted by hand, taking into account 135 known secondary structures (Kjer, 1995). 136 108 A total of 29 samples of P. oudrii from 12 localities were ana- Aligned mtDNA sequences were concatenated. Combined 137 109 Q3 lysed (Table 1 and Fig. 1). In order to evaluate the relative position mtDNA sequences were 885 bp (371 for 12s rRNA and 514 for 138 110 of P. oudrii and its taxonomic value relative to other Ptyodactylus 16s rRNA). Prior to phylogenetic analysis, mtDNA sequences were 139 Table 1 Codes, species, sampling localities and GenBank Accession Numbers for the samples included in this study. ‘‘–” indicates samples that were not analysed genetically. Code Species Locality Country GenBank Accession Numbers 12s 16s c-mos ACM4 P1 P. oudrii Batna Algeria * * * * P2 P. oudrii Erfoud Morocco * * * * P3 P. oudrii Tayahiat Morocco * * * * P4 P. oudrii Tiguert Morocco * * * * P5 P. oudrii Talsinnt Morocco * * * * P6 P.