Mitochondrial DNA diversity of the alpine newt (Ichthyosaura alpestris) in the Carpathian Basin: evidence for multiple cryptic lineages associated with Pleistocene refugia

  • Judit Vörös Department of Zoology, Hungarian Natural History Museum, H-1088 Budapest, Baross u. 13, Hungary https://orcid.org/0000-0001-9790-1443
  • Zoltán Varga Department of Evolutionary Zoology and Human Biology, University of Debrecen, H-4032 Debrecen, Egyetem tér 1., Hungary
  • Iñigo Martínez-Solano Museo Nacional de Ciencias Naturales, CSIC, c/ José Gutiérrez Abascal 2, 28006 Madrid, Spain https://orcid.org/0000-0002-2260-226X
  • Krisztián Szabó Department of Ecology, University of Veterinary Medicine, H-1077 Budapest, Rottenbiller u. 50, Hungary https://orcid.org/0000-0002-5110-8279
Keywords: Caudata, phylogeography, genetic diversity, Salamandridae, molecular taxonomy, Pleistocene refugia, alpine newt, Ichthyosaura alpestris

Abstract

The phylogeography and molecular taxonomy of the Alpine newt, Ichthyosaura alpestris, has been intensively studied in the past. However, previous studies did not include a comprehensive sampling from the Carpathian Basin, possibly a key region in the evolution of the species. We used a 1251 bp long fragment of the mitochondrial genome to infer the species’ evolutionary history in central-eastern Europe by assigning isolated Carpathian Basin populations from 6 regions to previously defined mtDNA lineages. We also revised the morphology-based intraspecific taxonomy of the species in the light of new genetic data. Alpine newt populations from the Carpathian Basin represented two different mitochondrial lineages. The Mátra, Bükk and Zemplén Mts populations can be assigned to the Western lineage of the nominotypical subspecies. Bakony and Őrség populations showed high haplotype diversity and formed a separate clade within the Western lineage, suggesting that the Carpathian Basin might have provided cryptic refugia for Alpine newt populations in their cold-continental forest-steppe landscapes during the younger Pleistocene. Newts from Apuseni Mts were related to the Eastern lineage but formed a distinct clade within this lineage. Considering the morphological and genetic differentiation of the Bakony and Őrség populations, consistent with a long independent evolutionary history, we propose these populations be referred to as Ichthyosaura alpestris bakonyiensis (Dely, 1964). We provide a redescription of this poorly known subspecies.

References

Andreone, F. & Tripepi, S. (2006): Triturus alpestris. Pp. 236–239. In: Sindaco, R., Doria, G., Razzetti, E. & Bernini, F. (eds): Atlante degli Anfibi e dei Rettili d’Italia/Atlas of Italian Amphibians and Reptiles. – Societas Herpetologica Italica, Ed. Polistampa, Firenze.

Arano, B. & Arntzen, J. W. (1987): Genetic differentiation in the alpine newt, Triturus alpestris. Pp. 21–24. In: van Gelder, J. J., Strijbosch, H. & Bergers P. J. M. (eds): Proceedings of the Fourth Ordinary General Meeting of the Societas Europea Herpetologica. – Faculty of Sciences Nijmegen Press, Nijmegen.

Arévalo, E., Davis, S. K. & Sites, J. W. (1994): Mitochondrial DNA sequence divergence and phylogenetic relationships among eight chromosome races of the Sceloporus grammicus complex (Phrynosomatidae) in central Mexico. – Systematic Biology 43: 387–418. https://doi.org/10.1093/sysbio/43.3.387

Babik, W., Branicki, W., Sandera, M., Litvinchuk, S., Borkin, L. J., Irwin, J. T. & Rafinski, J. (2004): Mitochondrial phylogeography of the moor frog, Rana arvalis. – Molecular Ecology 13: 1469–1480. https://doi.org.10.1111/j.1365-294X.2004.02157.x

Bandelt, H. J., Forster, P. & Röhl, A. (1999): Median-joining networks for inferring intraspecific phylogenies. – Molecular Biology and Evolution 16: 37–48. https://doi.org/10.1093/oxfordjournals.molbev.a026036

Bálint, M., Ujvárosi, L., Theissinger, K., Lehrian, S., Mészáros, N. & Pauls, S. U. (2011): The Carpathians as a major diversity hotspot in Europe. Pp 189–205. In: Zachos, F. E. & Habel, J. C. (eds): Biodiversity hotspots. – Springer, London. https://doi.org/10.1007/978-3-642-20992-5_11

Chenna, R., Sugawara, H., Koike, T., Lopez, R., Gibson, T. J., Higgins, D. G. & Thompson, J. D. (2003): Multiple sequence alignment with the Clustal series of programs. – Nucleic Acids Research 31: 497–500. https://doi.org/10.1093/nar/gkg500

Dely, O. Gy. (1959): Examen du Triton alpestre (Triturus alpestris), spécialement en vue des populations de la Hongrie et des Carpathes. – Acta Zoologica Academiae Scientiarum Hungaricae 5: 255–315.

Dely, O. Gy. (1964): Környezettani és rendszertani vizsgálatok az alpesi gőtén. [Ecological and taxonomic investigations on the alpine newt.]. Dissertation for Candidate of Science degree, Budapest, 369 pp.

Dely, O. Gy. (1967): Kétéltűek–Amphibia, Magyarország Állatvilága. – Fauna Hungariae 20(3), Akadémia Kiadó, Budapest, 78 pp. [in Hungarian]

Denoël, M. (1994): Le Triton alpestre, Triturus alpestris (Laurenti, 1768). – Les naturalistes Belges 75: 47–64.

Dubois, A. & Raffaëlli, J. (2009): A new ergotaxonomy of the family Salamandridae Goldfuss, 1820 (Amphibia, Urodela). – Alytes 26: 1–85.

Herrero, P. & Arano, B. (1986): Cytogenetic and morphological studies on Triturus alpestris cyreni. Pp. 151–154. In: Roček, Z. (ed.): Studies in Herpetology. – Charles University Press, Praha.

Hollós, A., Pecsenye, K., Bereczki, J., Bátori, E., Rákosy, L. & Varga, Z. (2012): Pattern of genetic and morphometric differentiation in Maculinea nausithous (Lepidoptera: Lycaenidae) in the Carpathian Basin. – Acta Zoologica Academiae Scientiarum Hungaricae 58: 87–103.

Lanfear, R., Calcott, B., Ho, S. Y. W. & Guindon, S. (2012): PartitionFinder: combined selection of partitioning schemes and substitution models for phylogenetic analyses. – Molecular Biology and Evolution 29: 1695–1701. https://doi.org/10.1093/molbev/mss020

Librado, P. & Rozas, J. (2009): DnaSP v5: A software for comprehensive analysis of DNA polymorphism data. – Bioinformatics 25: 1451–1452. https://doi.org/10.1093/bioinformatics/btp187

Lužnik, M., Kryštufek, B. & Bužan, E. (2011): Mitochondrial sequences do not support the independent taxonomic position of the extinct Alpine newt subspecies Mesotriton alpestris lacusnigri. – Amphibia-Reptilia 32: 435–440. https://doi.org/10.1163/017353711X588191

Molnár, P. (2001): Alpesi gőte (Triturus alpestris Laurenti, 1768) populációk morfometriai és élőhelypreferencia vizsgálata hazai előfordulási területein. – MSc thesis, University of Debrecen, 50 pp. [in Hungarian]

Mráz, P. & Ronikier, M. (2016): Biogeography of the Carpathians: evolutionary and spatial facets of biodiversity. – Biological Journal of the Linnean Society 119: 528–559. https://doi.org/10.1111/bij.12918

Palumbi, S. R., Martin, A. P., Romano, S. L., McMillan, W. O., Stice, L. & Grabowski, G. (1991): The Simple Fool’s Guide to PCR. – Department of Zoology, University of Hawaii, Honolulu, 24 pp.

Pecsenye, K., Tóth, J., Bereczki, J., Szolnoki, N. & Varga, Z. (2015): Genetic structure of Parnassius mnemosyne (Lepidoptera: Papilionidae) populations in the Carpathian Basin. – Organisms, Diversity, Evolution 16: 809–819. https://doi.org/10.1007/s13127-016-0281-7

Pecsenye, K., Tóth, A., Bereczki, J., Tóth, J. P., Katona, G. & Varga, Z. (2018): Surprising diversity in the Pannonian populations of Marsh Fritillary (Euphydryas aurinia, Lepi­doptera: Nymphalidae): Morphometric and molecular aspects. – Journal of Zoological Systematics and Evolutionary Research. https://doi.org/10.1111/jzs.12227

Pop, A. A., Pop, V. V. & Csúzdi, Cs. (2010): Significance of the Apuseni Mountains (the Carpathians) in the origin and distribution of Central European earthworm fauna (Oligochaeta: Lumbricidae). – Zoology Middle East 51:89–110. https://doi.org/10.1080/09397140.2010.10638462

Popart 1.7 [Online] (available at http://popart.otago.ac.nz.).

Posada, D. (2011): Collapse: Describing haplotypes from sequence alignments. [Online]. (Available at http://darwin.uvigo.es/software/collapse.html).

Raffaëlli, J. (2018): Proposal for a new taxonomic arrangement of Ichthyosaura alpestris (Laurenti, 1768) (Urodela, Salamandridae), an iconic species with a complex phylogenetic structure. – Alytes 36: 178.

Recuero, E. & Martinez-Solano, I. (2002): Triturus alpestris (Laurenti, 1768). Triton alpino. Pp. 58–60. In: Pleguezuelos, J. M., Marquez, R. & Lizana, M. (eds): Atlas y libro rojo de los anfibios y reptiles de España. – Direccion General de Conservacion de la Naturaleza-Asociacion Herpetologica Española, Madrid.

Recuero, E., Buckley, D., García-París, M., Arntzen, J. W., Cogâlniceanu, D. & Martínez-Solano, I. (2014): Evolutionary history of Ichthyosaura alpestris (Caudata, Salamandridae) inferred from the combined analysis of nuclear and mitochondrial markers. – Molecular Phylogenetics and Evolution 81: 207–220. https://doi.org/10.1016/j.ympev.2014.09.014

Roček, Z. (1974): Biometrical investigations of Central European populations of the Alpine newt, Triturus alpestris alpestris (Laurenti, 1768) (Amphibia: Urodela). – Acta Universitatis Carolinae, Biologica 5–6: 295–373.

Roček, Z., Joly, P. & Grossenbacher, K. (2003): Triturus alpestris (Laurenti, 1768) – Bergmolch. Pp. 607–656. In: Grossenbacher, K. & Tiesmeier, B. (eds): Handbuch der Reptilien und Amphibien Europas; Band 4/IIA Schwanzlurche (Urodela) IIA Salamandridae II: Triturus I. – Aula-Verlag, Wiebelsheim.

Ronquist, F. & Huelsenbeck, J. P. (2003): MrBayes 3: Bayesian phylogenetic inference under mixed models. – Bioinformatics 19: 1572–1574. https://doi.org/10.1093/bioinformatics/btg180

Schmitt, T. & Seitz, A. (2001): Intraspecific allozymatic differentiation reveals the glacial refugia and the postglacial expansions of European Erebia medusa (Lepidoptera: Nymphalidae). – Biological Journal of the Linnean Society 74: 429–458. https://doi.org/10.1006/bijl.2001.0584

Schmitt, T. & Varga, Z. (2012): Extra-Mediterranean refugia: The rule and not the exception? – Frontiers in Zoology 9: 22. https://frontiersinzoology.biomedcentral.com/articles/10.1186/1742-9994-9-22

Sotiropoulos, K., Eleftherakos, K., Džukić, G., Kalezić, M. L., Legakis, A. & Polymeni, R. M. (2007): Phylogeny and biogeography of the Alpine newt Mesotriton alpestris (Salamandridae, Caudata), inferred from mtDNA sequences. – Molecular Phylogenetics and Evolution 45: 211–226. https://doi.org/10.1016/j.ympev.2007.03.012

Sotiropoulos, K., Eleftherakos, K., Kalezić, M. L., Legakis, A. & Polymeni, R. M. (2008): Genetic structure of the alpine newt, Mesotriton alpestris (Salamandridae, Caudata), in the southern limit of its distribution: Implications for conservation. – Biochemical Systematics and Ecology 36: 297–311. https://doi.org/10.1016/j.bse.2007.10.002

Speybroeck, J., Beukema, W. & Crochet, P.-A. (2010): A tentative species list of the European herpetofauna (Amphibia and Reptilia) – an update. – Zootaxa 2492: 1–27. https://doi.org/10.5281/zenodo.195659

Speybroeck, J., Beukema, W., Dufresnes, C., Fritz, U., Jablonski, D., Lymberakis, P., Martínez-Solano, I., Razzetti, E., Vamberger, M., Vences, M., Vörös, J. & Crochet, P-A. (2020): Species list of the European herpetofauna – 2020 update by the Taxonomic Committee of the Societas Europaea Herpetologica. – Amphibia-Reptilia 41: 139–189. https://doi.org/10.1163/15685381-bja10010

Szabó, K. & Vörös, J. (2014): Distribution and hybridisation of Anguis fragilis and A. colchica in Hungary. – Amphibia-Reptilia 35: 135–140. https://doi.org/10.1163/15685381-00002927

Tamura, K., Stecher, G., Peterson, D., Filipski, A., & Kumar, S. (2013): MEGA6: Molecular Evolutionary Genetics Analysis version 6.0. – Molecular Biology and Evolution 30: 2725–2729. https://doi.org/10.1093/molbev/mst197

Ursenbacher, S., Carlsson, M., Helfer, V., Tegelstrom, H. & Fumagalli, L. (2006): Phylogeography and Pleistocene refugia of the adder (Vipera berus) as inferred from mitochondrial DNA sequence data. – Molecular Ecology 15: 3425–3437. https://doi.org/10.1111/j.1365-294X.2006.03031.x

Varga, Z. (1995): Geographical patterns of biodiversity in the Palearctic and in the Carpathian Basin. – Acta Zoologica Academiae Scientiarum Hungaricae 41: 71–92.

Varga, Z. (2010): Extra-Mediterranean Refugia, Post-Glacial Vegetation History and Area Dynamics in Eastern Central Europe. Pp. 57–87. In: Habel J.C. & Assmann T. (eds): Relict species: Phylogeography and conservation biology. – Springer-Verlag Berlin, Heidelberg. https://doi.org/10.1007/978-3-540-92160-8_3

Varga, Z. (2019): Biogeográfia – az élet földrajza. [Biogeography – the geography of life.] – Pars Kft., Nagykovácsi, 610 pp.

Vörös, J., Alcobendas, M., Martínez-Solano, I. & García-París, M. (2006): Evolution of Bombina bombina and Bombina variegata (Anura: Discoglossidae) in the Carpathian Basin: a history of repeated mt-DNA introgression across species. – Molecular Phylogenetics and Evolution 38: 705–715. https://doi.org/10.1016/j.ympev.2005.08.010

Vörös, J., Herczeg, D., Fülöp, A., Gál, J. T., Dán, Á., Harmos, K. & Bosch, J. (2018): Batrachochytrium dendrobatidis in Hungary: a review of recent and historical occurrence. – Acta Herpetologica 13: 125–40. https://doi.org/10.13128/Acta_Herpetol-22611

Vörös, J., Herczeg, D., Papp, T., Monsalve-Carcano, C. & Bosch, J. (2020): First detection of Ranavirus infection in Hungary: A survey of amphibians. – Herpetology Notes 13: 213–217.

Vörös, J., Kiss, I. & Puky, M. (2014): Amphibian declines and conservation in Hungary. Pp. 99–130. In: Heatwole, H. & Wilkinson, J. W. (eds): Status of decline of Amphibians: Eastern Hemisphere. Amphibian Biology Vol. 11. – Pelagic Publishing, Exeter.

Vörös, J., Mikulíček, P., Major, Á., Recuero, E. & Arntzen, J. W. (2016a): Phylogeographic analysis reveals northerly refugia for the riverine amphibian Triturus dobrogicus (Caudata: Salamandridae). – Biological Journal of the Linnean Society 119: 974–991. ­https://doi.org/10.1111/bij.12866

Vörös, J., Ursenbacher, S., Kiss, I., Jelic, D., Schweiger, S. & Szabó, K. (2016b): Increased genetic structuring of isolated Salamandra salamandra populations (Caudata: Salamandridae) at the margins of the Carpathian Mountains. – Journal of Zoological Systematics and Evolutionary Research 55: 138–149. https://doi.org/10.1111/jzs.12157

Williams, P., Humphries, C., & Araújo, M. (1999): Mapping Europe’s biodiversity. Pp. 12–20. In: Delbaere, B. (ed.): Facts and figures on Europe’s biodiversity, state and trends 1998–1999. – ECNC, Tilburg.

Willis, K. J., Braun, M., Sümegi, P. & Tóth, A. (1997): Does soil change cause vegetation change or vice versa? A temporal perspective from Hungary. – Ecology 78: 740–750. https://doi.org/10.1890/0012-9658(1997)078[0740:DSCCVC]2.0.CO;2

Willis, K. J. & van Andel, T. H. (2004) Trees or no trees? The environments of Central and Eastern Europe during the last glaciation. – Quaternary Science Reviews 23: 2369–2387. https://doi.org/10.1016/j.quascirev.2004.06.002

Zuiderwijk, A. (1997): Triturus alpestris (Laurenti, 1768). Pp. 72–73. In: Gasc, J. P., Cabela, A., Crnobrnja-Isailović, J., Dolmen, D., Grossenbacher, K., Haffner, P., Lescure, J., Martens, H., Martínez Rica, J. P., Maurin, H., Oliveira, M. E., Sofianidou, T. S., Veith, M. & Zuiderwijk, A. (eds): Atlas of amphibians and reptiles in Europe. – Societas Europaea Herpetologica, Muséum National d’Histoire Naturelle, Paris.

Published
2021-05-19
How to Cite
VörösJ., VargaZ., Martínez-SolanoI., & SzabóK. (2021). Mitochondrial DNA diversity of the alpine newt (Ichthyosaura alpestris) in the Carpathian Basin: evidence for multiple cryptic lineages associated with Pleistocene refugia. Acta Zoologica Academiae Scientiarum Hungaricae, 67(2), 177–197. https://doi.org/10.17109/AZH.67.2.177.2021