We studied the genetic structure and diversity of a vulnerable population of the Halys pit viper (Gloydius halys) living at the northern distribution limit of the species (Novosibirsk Region, West Siberia, Russia) and fairly isolated from the nearest known locations (180-200 km). 157 snakes from three locations (separated by 5-11 km) were genotyped using 8 microsatellite loci. In addition, nucleotide sequences of a mitochondrial marker gene (ND4) were obtained from 30 individuals. Two mitochondrial haplotypes differing by one synonymous substitution could be detected. Haplotype H1 had a prevalence of 96.7%. High levels of genetic diversity (mean ) were detected and no evidence of a recent bottleneck in any sampling site could be discovered. We found that G. halys exhibits a low, although significant, level of genetic differentiation on a fine geographic scale (overall FST = 0.013). The degree of differentiation is comparably low with regard to values reported for other vipers and more similar to those in Colubridae. In addition, using local based FST estimates, we detected a significant difference in the extent of genetic drift between the centrally located and marginal local populations as predicted by the stepping-stone model of migration. Thus, from a short-term perspective, primary attention should be given to the peripheral local populations, due to limited immigration and stronger influence of genetic drift. Since all habitat patches are situated along the river and have a linear order it is important to preserve each habitat site to maintain gene flow across the metapopulation.
Evgeniy Simonov and Michael Wink
Peter Lenk, Michael Wink and Ulrich Joger
In order to elucidate the phylogenetic relationships in European ratsnakes of the genus Elaphe, we analyzed a 597 bp part of the mitochondrial cytochrome b gene of eight West Eurasian and one East Asian species. Lampropeltis served as outgroup. Maximum parsimony and maximum likelihood suggest the existence of four lineages: 1) E. scalaris; 2) the E. longissima species group comprising E. longissima, E. lineata, E. situla, E. hohenackeri, and E. persica; 3) E. quatuorlineata and 4) E. dione as a sister group to 3). Elaphe scalaris is basal and shows no closer affiliation with any other analyzed species. The Middle Eastern E. persica and E. hohenackeri appear basal within the E. longissima group. Elaphe lineata differs by 8% nucleotide substitutions from E. longissima, supporting the hypothesis that both taxa represent distinct species. Elaphe situla is associated with Elaphe longissima and E. lineata. Three analyzed subspecies of E. quatuorlineata are represented by distinct haplotypes. The extent of divergence gives reason to assign species status to the taxon sauromates. Besides, we found two very distinct haplotypes within the range of E. (q.) sauromates, indicating the existence of a third, so far unidentified, species within the E. quatuorlineata complex. The East Asian E. porphyracea clusters with the E. longissima group. This, as well as comparisons with supplementary sequences of Asian Elaphe species, document the multiple origins of European Elaphe. Um die phylogenetischen Beziehungen europäischer Kletternattern der Gattung Elaphe zu erhellen, sequenzierten und analysierten wir ein 597 Nukleotide messendes Stück des mitochondrialen Cytochrom b Gens von acht west-eurasischen und einer ostasiatischen Art. Lampropeltis diente als Außengruppe. Maximum parsimony und maximum likelihood Berechnungen zeigen die Existenz von vier genetischen Linien auf: a) E. scalaris; b) die E. longissima Artengruppe mit E. longissima , E. lineata, E. situla, E. hohenackeri, und E. persica; c) E. quatuorlineata und d) E. dione als Schwestergruppe zu c). Elaphe scalaris nimmt eine basale Position ein und scheint mit keiner untersuchten Art näher verwandt zu sein. Die westasiatischen Arten E. persica und E. hohenackeri sind die Schwestergruppe zu E. longissima, E. lineata und E. situla. Elaphe lineata unterscheidet sich durch 8% Nukleotidaustausche von E. longissima, wodurch der Artstatus von E. lineata unterstützt wird. Elaphe longissima, E. lineata und E. situla bilden eine Abstammungsgemeinschaft. Auch die Haplotypen der drei untersuchten E. quatuorlineata-Unterarten unterscheiden sich so stark, daß Artstatus für das Taxon sauromates zu fordern ist. Darüberhinaus fanden wir im Gebiet von E. (q.) sauromates zwei stark unterschiedliche Haplotypen, die die Existenz einer dritten bislang unidentifizierten Art nahelegen. Die ostasiatische E. porphyracea scheint der E. longissima-Gruppe nahe zu stehen. Dies, sowie der Vergleich mit ergänzenden Sequenzdaten asiatischer Elaphe-Arten, belegt den mehrfachen Ursprung europäischer Elaphe.
Ana Paunović, Zoltán Nagy, Michael Wink, Zoltán Korsós and Mátyás Bellaagh
The Caspian whipsnake (Squamata: Serpentes: Dolichophis caspius) reaches its northwestern distribution limit in Hungary where it is the single representative of its genus. Recently, several new localities of the species were discovered in Hungary. Focusing on these edge populations and on other ones in the western parts of the distribution, a phylogeographic study based on cytochrome b sequences was carried out. Thereby, a clear mitochondrial divergence was observed in Europe, with eastern and western haplotype groups found by analysing a number of specimens and populations. Our haplotype network analysis suggests a rather rapid postglacial (re)colonization of Europe by this snake species. In Hungary, three different haplotypes were detected.
Pavel Široký, Uwe Fritz, Oğuz Türkozan, Michael Wink, Jan Lehmann, Lyudmila Mazanaeva, Markus Auer, Hajigholi Kami and Anna Hundsdörfer
Tortoises of the Testudo graeca complex inhabit a patchy range that covers part of three continents (Africa, Europe, Asia). It extends approximately 6500 km in an east-west direction from eastern Iran to the Moroccan Atlantic coast and about 1600 km in a north-south direction from the Danube Delta to the Libyan Cyrenaica Peninsula. Recent years have seen a rapid increase of recognized taxa. Based on morphological investigations, it was suggested that this group consists of as many as 20 distinct species and is paraphyletic with respect to T. kleinmanni sensu lato and T. marginata. Based on samples from representative localities of the entire range, we sequenced the mitochondrial cytochrome b gene and conducted nuclear genomic fingerprinting with ISSR PCR. The T. graeca complex is monophyletic and sister to a taxon consisting of T. kleinmanni sensu lato and T. marginata. The T. graeca complex comprises six well-supported mtDNA clades (A-F). Highest diversity is found in the Caucasian Region, where four clades occur in close neighbourhood. This suggests, in agreement with the fossil record, the Caucasian Region as a radiation centre. Clade A corresponds to haplotypes from the East Caucasus. It is the sister group of another clade (B) from North Africa and western Mediterranean islands. Clade C includes haplotypes from western Asia Minor, the southeastern Balkans and the western and central Caucasus Region. Its sister group is a fourth, widely distributed clade (D) from southern and eastern Asia Minor and the Levantine Region (Near East). Two further clades are distributed in Iran (E, northwestern and central Iran; F, eastern Iran). Distinctness of these six clades and sister group relationships of (A + B) and (C + D) are well-supported; however, the phylogeny of the resulting four clades (A + B), (C + D), E and F is poorly resolved. While in a previous study (Fritz et al., 2005a) all traditionally recognized Testudo species were highly distinct using mtDNA sequences and ISSR fingerprints, we detected within the T. graeca complex no nuclear genomic differentiation paralleling mtDNA clades. We conclude that all studied populations of the T. graeca complex are conspecific under the Biological Species Concept. There is major incongruence between mtDNA clades and morphologically defined taxa. Morphologically well-defined taxa, like T. g. armeniaca or T. g. floweri, nest within clades comprising also geographically neighbouring, but morphologically distinctive populations of other taxa (clade A: T. g. armeniaca, T. g. ibera, T. g. pallasi ; clade D: T. g. anamurensis, T. g. antakyensis, T. g. floweri, T. g. ibera, T. g. terrestris), while sequences of morphologically similar tortoises of the same subspecies (T. g. ibera sensu stricto or T. g. ibera sensu lato) scatter over two or three genetically distinct clades (A, C or A, C, D, respectively). This implies that pronounced morphological plasticity, resulting in phenotypes shaped by environmental pressure, masks genetic differentiation. To achieve a more realistic taxonomic arrangement reflecting mtDNA clades, we propose reducing the number of T. graeca subspecies considerably and regard in the eastern part of the range five subspecies as valid (T. g. armeniaca, T. g. buxtoni, T. g. ibera, T. g. terrestris, T. g. zarudnyi). As not all North African taxa were included in the present study, we refrain from synonymizing North African taxa with T. g. graeca (mtDNA clade B) that represents a further valid subspecies.
Mathieu Détaint, Anthony Olivier, Marc Cheylan, Christophe Coïc, Uwe Fritz, Daniela Guicking, Antoine Cadi, Peter Lenk, Ulrich Joger, Michael Wink and Elisabeth Rosecchi
The European pond turtle, Emys orbicularis, is a wide ranging species, distributed from Northwest Africa over a large part of Europe and Asia Minor to the Caspian and Aral Seas. For 106 pond turtles from France mtDNA sequence variation has been assessed, using a 1031 bp portion of the mitochondrial cytochrome b gene. Three of nine haploclades currently known from the entire species' range were found in France. One clade (II) is represented with four very similar haplotypes, differing by one mutation, and the two other clades (V, VI) are represented with one haplotype each. A syntopic occurrence of clades II and V is reported for the first time for the Camargue. Besides, clade II occurs in the French regions Aquitaine, Centre-Val de Loire, and Rhône-Alpes. Outside of France, it is found mainly in the catchment areas of the Danube and Oder rivers and in the Balkans. Haploclade V, which is also known from the Apennine peninsula, Sardinia, and the northern Mediterranean coast of Spain, is restricted in France to Corsica and the Provence-Alpes-Côte d'Azur region. A single individual bearing a haplotype of an Iberian and North African clade (VI) was found in Aquitaine near Pau. This could indicate gene flow between the Iberian peninsula and West France, if the specimen is native. The distribution of the distinct haploclades in France probably reflects Holocene range expansions, especially of haploclade II turtles. In the postglacial, haploclade II terrapins arrived from the east and spread over the Rhône corridor to the Mediterranean coast. In the southern Rhône area they met and hybridized with haploclade V turtles. Further research is needed to clarify whether this hybridization is a locally restricted phenomenon.
Antonia Celani, Peter Havaš, Sandro Tripepi, Viner Khabibullin, Andrey Bakiev, Soumia Fahd, Uwe Fritz, Guillermo Velo Antón, Pavel Široký, Hajigholi Kami, Daniela Guicking, Aitor Valdeón Vélez, Markus Auer, Michael Wink, Ulrich Joger, Marine Arakelyan, Georg Džukić, César Ayres Fernández, Lyudmila Mazanaeva and Dinçer Ayaz
Based on more than 1100 samples of Emys orbicularis and E. trinacris, data on mtDNA diversity and distribution of haplotypes are provided, including for the first time data for Armenia, Georgia, Iran, and the Volga, Ural and Turgay River Basins of Russia and Kazakhstan. Eight mitochondrial lineages comprising 51 individual haplotypes occur in E. orbicularis, a ninth lineage with five haplotypes corresponds to E. trinacris. A high diversity of distinct mtDNA lineages and haplotypes occurs in the south, in the regions where putative glacial refuges were located. More northerly parts of Europe and adjacent Asia, which were recolonized by E. orbicularis in the Holocene, display distinctly less variation; most refuges did not contribute to northern recolonizations. Also in certain southern European lineages a decrease of haplotype diversity is observed with increasing latitude, suggestive of Holocene range expansions on a smaller scale.