Jana Bakan, Lavadinović Vukan, Zoran Popović, Ladislav Paule


 Genetic differentiation of grey wolf (Canis lupus) populations from the Balkans (Serbia and Bulgaria) and the Western Carpathians was studied by means of using tissue and bone samples from legally hunted wolves. In total, 339 samples of tissues and bones and small number of feaces, blood and urine samples were used. Genotyping was based on 15 microsatellites of nuclear DNA. Genetic differentiation was studied using Bayesian approach (structure software). Obtained results showed the differenetiation between the Balkan and the Carpathian populations as well as between the Bulgarian population and both populations from Serbia and Slovakia. There are indications of migration towards west. Although these results are based on the nuclear markers, it would be advised to reanalyze the samples using mtDNA markers.


Carpathians, Balkan, genetic differentiation, Canis lupus

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Belkhir, K., Borsa, P., Chikhi, L., Raufauste, N., Bnhomme, F. (2001): GENETIX 4.02,logiciel sous WindowsTM pour la génétique des populations. Laboratoire Génome, Populations, Interactions. CNRS UMR 5000. Université de Montpellier II, Montpellier (France). Available from URL:

Cournuet, J.M., Luikart, G. (1996): Description and power analysis of two tests for detecting recent population bottlenecks from allele frequency data. Genetics, 144:2001–2014.

Czarnomska, S.D., Jędrzejewska, B., Borowik, T., Niedziałkowska, M., Stronen, A.V., Nowak, S., Mysłajek, R.W., Okarma, H., Konopiński, M., Pilot, M., Śmietana, W., Caniglia, R., Fabbri, E., Randi, E. (2013): Concordant mitochondrial and microsatellite DNA structuring between Polish lowland and Carpathian Mountain wolves. Conservation Genetics, 14: 573–588.

Dalén, L., Fuglei, E., Hersteinsson, P., Kapel, C.M.O., Roth, J.D., Samelius, G., Tannerfeldt, M., Angerbjörn, A. (2005): Population history and genetic structure of a circumpolar species: the arctic fox. Biological Journal of Linnean Society, 84: 79–89.

Di Rienzo, A., Peterson, A.C., Garza, J.C., Valdes, A.M., Slatkin, M., Freimer, N.B. (1994): Mutational processes of simple-sequence repeat loci in human populations. Proceeding of the National Academy of Sciences USA, 91: 3166–3170.

Djan, M., Maletić, V., Trbojević, I., Popović, D., Veličković, N., Burazerović, J., Ćirović, D. (2014): Genetic diversity and structuring of the grey wolf population from the Central Balkans based on mitochondrial DNA variation. Mammalian Biology, 79: 277–282.

Dolf, G., Schläpfer, J., Gaillard, C., Randi, E., Lucchini, V., Breintenmoser, U., Stahlberger–Saitbekova, N. (2000): Differentiation of the Italian wolf and the domestic dog based on microsatellite analysis. Genetics, Selection, Evolution, 32(5): 533–541.

Doyle, J.J., Doyle, J.L. (1987): A rapid DNA isolation procedure for small quantities of fresh leaf tissue. Phytochemical Bulletin, 19: 11–15.

Ehrich, D., Gaudeul, M., Assefa, A. (2007): Genetic consequences of Pleistocene range shifts: Contrast between the Arctic, the Alps and the East African mountains. Molecular Ecology,16: 2542–2559.

Evanno, G., Regnaut, S., Goudet, J. (2005): Detecting the number of clusters of individuals using the software STRUCTURE: a simulation study. Molecular Ecology, 14: 2611–2620.

Excoffier, L., Lava, G., Schneider, S. (2006): Arlequin ver 3.1. An integral software package for population genetics data analysis. University of Bern, Switzerland.

Falush, D., Stephens, M., Pritchard, J.K. (2003): Inference of population structure using multilocus genotype data: linked loci and correlated allele frequencies. Genetics, 164: 1567–1587.

Francisco, L.V., Langsten, A.A., Mellersh, C.S., Neal, C.L., Ostrander, E.A., 1996: A class of highly polymorphic tetranucleotide repeats for canine genetic mapping. Mammalian Genome, 7: 359–362.

Fredholm, M., Wintero, A.K., 1995: Variation of short tandem repeats within and between species belonging to the Canidae family. Mammalian Genome, 6: 11–18.

Gomerčić, T., Sindičić, M., Galov, A., Arbanasić, H., Kusak, J. (2010): High genetic variability of grey wolf (Canis lupus L.) population from Croatia as revealed by mitochondrial DNA control region sequences. Zoologiccal Studies, 49: 816–823.

Goudet, J. (2001): FSTAT, a program to estimate and test gene diversities and fixation indices (version 2.9.3). Available from from Goudet (1995).

Gruber, B., Adamack, A. (2013): Introduction to PopGenReport. Available from

Linnell, J., Salvatori, V., Boitani, L. (2007): Guidelines for population level management plans for large carnivores. Large Carnivore Initiative for Europe report prepared for the European Commission. Rome, Italy.

Lucchini, V., Galov, A., Randi, E. (2004): Evidence of genetic distinction and long-term population decline in wolves (Canis lupus) in the Italian Apennines. Molecular Ecology, 13: 523–536.

Muñoz-Fuentes, V., Darimont, C.T., Wayne, R.K., Paquet, P.C., Leonard, J.A. (2009): Ecological factors drive differentiation in wolves from British Columbia. Journal of Biogeography, 36: 1516–1531.

Oliveira, C.G., Martinez, R.A., Gaiotto, F.A. (2007): DNA extraction from bristles and quills of Chaetomys subspinosus (Rodentia: Erethizontidae) using a novel protocol. Genetic and Molecular Research,6: 657–666.

Pilot, M., Branicki, W., Jędrzejewski, W., Goszczyński, J., Jędrzejewska, B., Dykyy, I., Shkvyrya, M., Tsingarska, E. (2010): Phylogeographic history of grey wolves in Europe. BMC Evolutionary Biology, 10: 104.

Pilot, M., Jędrzejewski, W.J., Branicki, W., Sidorovich, V.E., Jędrzejewska, B., Stachura, K., Funk, S.M. (2006): Ecological factors influence population genetic structure of European grey wolves. Molecular Ecology, 15: 4533–4553.

Pritchard, J.K., Wen, W., and Falush, D. (2007): Documentation for STRUCTURE software: Version 2.2. University of Chicago, Chicago: 36 pp.

Randi, E. (2011): Genetics and conservation of wolves Canis lupus in Europe. Mammal Review, 41: 99–111.

Rosenberg, N.A., Pritchard, J.K., Weber, J.L., Cann, H.M., Kidd, K.K., Zhivotovsky, L.A., Feldman, M.W. (2002): Genetic structure of human populations. Science, 298: 2381–2385.

Rueness, E.K., Stenseth, C., O’Donoghue, M., Boutin, S., Ellegren, H., Jacobsen, S. (2003): Ecological and genetic spatial structuring in the Canadian lynx. Nature, 425: 69–72.

Scandura, M., Iacolina, L., Capitani, C., Gazzola, A., Mattioli, L., Apollonio, M. (2011): Fine-scale genetic structure suggests low levels of short-range gene flow in a wolf population of the Italian Apennines. European Journal of Wildlife Research 57: 949–958.

Sneath, P.H.A., Sokal, R.R. (1973): Numerical Taxonomy. The Principles and Practice of Numerical Classification. ISBN 0-7167-0697-0, 573 pp.

Souto, L., Ferreira, E., Fonseca, C. (2004). Microsatellite analysis of wild boar populations in Portugal by multiplex PCR. QIAGEN News, 3: 63–64.

Stronen, A.V., Jedrzejewska, B., Pertoldi, C., Demontis, D., Randi, E. (2013): North-south differentiation and a region of high diversity in European wolves (Canis lupus). PLoS ONE, 8(10): e76454.

Sundqvist, A.K., Ellegren, H., Vilà, C. (2007): Wolf or dog? Genetic identification of the predator from saliva collected around bite wounds on prey. Conservation Genetics, 9: 1275–1279.

van Oosterhout, C., Hitchinson, W.F., Wills, D.P.M., Shipley, P. (2004): MICRO-CHECKER: software for identifying and correcting genotyping errors in microsatellite data. Molecular Ecology, 4: 535–538.

Vilá, C., Amorim, I.R., Leonard, J.A., Posada, D., Castroviejo, J. (1999): Mitochondrial DNA phylogeography and population history of the grey wolf Canis lupus. Molecular Ecology, 8: 2089–2103.

Vilà, C., Sundqvist, A.K., Flagstad, Ø., Seddon, J., Bjornerfeldt, S., Koyola, I., Casulli, A., Sand, H., Wabakken, P., Ellegren, H. (2003): Rescue of a severely bottlenecked wolf (Canis lupus) population by a single immigrant. Proceedings of the Royal Society of London, Series B, 270: 91–97.

Walsh S.P., Metzger D.A., Higuchi R. (1991): Chelex 100 as a medium for simple extraction of DNA for PCR based typing from forensic material. Biotechniques 10(4): 506–513.

Weir, B.S., Cockerham, C.C. (1984): Estimating F-statistics for the analysis of population structure. Evolution, 38: 1358–1370.

Wright, S. (1969): Evolution and the genetics of populations. The theory of gene frequencies. University of Chicago Press.



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ISSN: 2335-0113

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