Neogene climatic oscillations shape the biogeography and evolutionary history of the Eurasian blindsnake P. Kornilios a,⇑ , Ç. Ilgaz b , Y. Kumlutasß b , P. Lymberakis c , J. Moravec d , R. Sindaco e , N. Rastegar-Pouyani f , M. Afroosheh f , S. Giokas a , S. Fraguedakis-Tsolis a , B. Chondropoulos a a Section of Animal Biology, Department of Biology, School of Natural Sciences, University of Patras, GR-26500 Patras, Greece b Department of Biology, Faculty of Science, Dokuz Eylül University, 35160 Buca- _ Izmir, Turkey c Natural History Museum of Crete, University of Crete, Knossou Ave., 71409 Irakleio, Crete, Greece d Department of Zoology, National Museum, 115 79 Prague1, Czech Republic e Istituto per le Piante da Legno e l’ Ambiente, Corso Casale 476, 10132 Torino, Italy f Department of Biology, Faculty of Science, Razi University, 67149 Kermanshah, Iran article info Article history: Received 12 May 2011 Revised 23 November 2011 Accepted 30 November 2011 Available online 13 December 2011 Keywords: Aridification Aridity Greek blindsnake Molecular clock Phylogeography Typhlops vermicularis abstract Typhlops vermicularis is the only extant scolecophidian representative occurring in Europe. Its main dis- tribution area, the eastern Mediterranean, has a complicated geological and climatic history that has left an imprint on the phylogenies and biogeography of many taxa, especially amphibians and reptiles. Since reptiles are sensitive indicators of palaeogeographical and palaeoclimatic events, we investigated the intraspecific genealogy of T. vermicularis in a phylogeographical framework. A total of 130 specimens were analyzed, while the use of formalin and ethanol as preservatives called for a special treatment of the samples. Partial sequences of two mitochondrial (12S and ND2) and one nuclear (PRLR) marker were targeted and the results of the phylogenetic analyses (NJ, ML and BI) and the parsimony-network revealed the existence of 10 evolutionary significant units within this species. In combination with the results of the dispersal-vicariance analysis, we may conclude that the Eurasian blindsnake has encoun- tered a sequence of extinction events, followed by secondary expansion from refugia. Estimation of diver- gence times showed that severe climatic changes between significantly wetter and drier conditions in the Late Neogene have played a key role on the evolutionary and biogeographical history of T. vermicularis. Additionally, both markers (mtDNA and nDNA) distinguished a largely-differentiated evolutionary lineage (Jordan and south Syria), which could even be reckoned as a full species. Our study reveals the existence of cryptic evolutionary lineages within T. vermicularis, which calls for further attention both on the protection of intraspecific varieties and the respective geographic areas that hold them. Ó 2011 Elsevier Inc. All rights reserved. 1. Introduction Of the two major divisions of snakes (Alethinophidia, i.e. ‘‘true snakes’’ and Scolecophidia, i.e. ‘‘wormsnakes’’), scolecophidians are the most poorly known in terms of species diversity, phylog- eny, biogeography and ecology (Greene, 1997; Vidal et al., 2010). They include approximately 400 known species, which, due to their fossorial living, are characterized by extreme morphological uniformity and include cryptic genetic varieties revealed recently by DNA studies (Adalsteinsson et al., 2009; Thomas and Hedges, 2007). Scolecophidians are distributed on all continents except Antarctica, with most species inhabiting the southern hemisphere and tropical islands (Zug et al., 2001). The only extant scolecophidian representative occurring in Europe, and specifically in the southern Balkans, is Typhlops vermicularis Merrem, 1820, also known as Greek or Eurasian blind- snake. The fossil record confirms its occurrence, or the occurrence of other scolecophidians, in most parts of Europe in the past. Spe- cifically, they have been found throughout the Eocene (Rage, 1984) but they are absent through most of the Oligocene, probably due to a dramatic global climate change resulting in a shift from tropical forest habitats to open environments (Badiola et al., 2009 and references therein). Wormsnakes reappear towards the end of the Oligocene, while during the Miocene the group extended from western to eastern Europe (Szyndlar, 1991). During the Pliocene they were restricted to the southern parts of Europe, from Spain to Greece (Szyndlar, 1991). Known scolecophidian fossil remains are restricted to precaudal vertebrae, whose simple and homoge- neous morphology within the group renders identification impossible, even at family level. Szyndlar (1991), however, states that the latest scolecophidian fossils from Ukraine (Late Miocene), continental Greece (Late Pliocene and Middle Pleistocene) and the island of Rhodos (Early Pliocene) do not differ from each other or 1055-7903/$ - see front matter Ó 2011 Elsevier Inc. All rights reserved. doi:10.1016/j.ympev.2011.11.035 ⇑ Corresponding author. E-mail address: korniliospan@yahoo.gr (P. Kornilios). Molecular Phylogenetics and Evolution 62 (2012) 856–873 Contents lists available at SciVerse ScienceDirect Molecular Phylogenetics and Evolution journal homepage: www.elsevier.com/locate/ympev