Given the hybrid genomic constitutions and increased ploidy of many asexual animals, the identification of processes governing the origin and maintenance of clonal diversity provides useful information about the evolutionary consequences of interspecific hybridization, asexuality and polyploidy. In order to understand the processes driving observed diversity of biotypes and clones in the Cobitis taenia hybrid complex, we performed fine-scale genetic analysis of Central European hybrid zone between two sexual species using microsatellite genotyping and mtDNA sequencing. We found that the hybrid zone is populated by an assemblage of clonally (gynogenetically) reproducing di-, tri- and tetraploid hybrid lineages and that successful clones, which are able of spatial expansion, recruit from two ploidy levels, i.e. diploid and triploid. We further compared the distribution of observed estimates of clonal ages to theoretical distributions simulated under various assumptions and showed that new clones are most likely continuously recruited from ancestral populations. This suggests that the clonal diversity is maintained by dynamic equilibrium between origination and extinction of clonal lineages. On the other hand, an interclonal selection is implied by nonrandom spatial distribution of individual clones with respect to the coexisting sexual species. Importantly, there was no evidence for sexually reproducing hybrids or clonally reproducing non-hybrid forms. Together with previous successful laboratory synthesis of clonal Cobitis hybrids, our data thus provide the most compelling evidence that 1) the origin of asexuality is causally linked to interspecific hybridization; 2) successful establishment of clones is not restricted to one specific ploidy level and 3) the initiation of clonality and polyploidy may be dynamic and continuous in asexual complexes.

Department of Vertebrate Evolutionary Biology and Genetics Institute of Animal Physiology and Genetics AS CR vvi Liběchov Czech Republic

Zobrazit více v PubMed

Janko K, Drozd P, Flegr J, Pannell JR (2008) Clonal turnover versus clonal decay: a null model for observed patterns of asexual longevity, diversity and distribution. Evolution 62: 1264–1270. PubMed

Law JH, Crespi BJ (2002) Recent and Ancient Asexuality in PubMed DOI

Moritz C, Brown WM, Densmore LD, Wright JW, Vyas D, et al. (1989) Genetic diversity and the dynamics of hybrid parthenogenesis in

D’souza TG, Storhas M, Michiels NK (2005) The effect of ploidy level on fitness in parthenogenetic flatworms. Biological Journal of the Linnean Society 85: 191–198 doi:10.1111/j.1095–8312.2005.00482.x. DOI

Lampert KP, Lamatsch DK, Epplen JT, Schartl M (2005) Evidence for a monophyletic origin of triploid clones of the Amazon molly, PubMed

Grimanelli D, Leblanc O, Perotti E, Grossniklaus U (2001) Developmental genetics of gametophytic apomixis. Trends Genet 17: 597–604. PubMed

Whitton J, Sears CJ, Baack EJ, Otto SP (2008) The Dynamic Nature of Apomixis in the Angiosperms. International Journal of Plant Sciences 169: 169–182 doi:10.1086/523369. DOI

Stöck M, Lampert KP, Möller D, Schlupp I, Schartl M (2010) Monophyletic origin of multiple clonal lineages in an asexual fish ( PubMed DOI

Choleva L, Janko K, De Gelas K, Bohlen J, Šlechtová V, et al. (2012) Synthesis of clonality and polyploidy in vertebrate animals by hybridization between two sexual species. Evolution 66–7: 2191–2203 doi:10.1111/j.1558–5646.2012.01589.x. PubMed DOI

Corley LS, Moore AJ (1999) Fitness of alternative modes of reproduction: developmental constraints and the evolutionary maintenance of sex. Proc Biol Sci 266: 471 doi:10.1098/rspb.1999.0661. DOI

Schwander T, Crespi BJ (2009) Twigs on the tree of life? Neutral and selective models for integrating macroevolutionary patterns with microevolutionary processes in the analysis of asexuality. Molecular Ecology 18: 28–42 doi:10.1111/j.1365-294X.2008.03992.x. PubMed DOI

Angers B, Schlosser IJ (2007) The origin of PubMed DOI

Janko K, Culling MA, Rab P, Kotlik P (2005) Ice age cloning-comparison of the Quaternary evolutionary histories of sexual and clonal forms of spiny loaches ( PubMed

Kearney M, Fujita MK, Ridenour J (2009) Lost sex in the reptiles: constraints and correlations. Lost Sex. Dordrecht Netherlands: Springer. 447–474.

Polihronakis M, Caterino MS, Chatzimanolis S (2010) Elucidating the phylogeographic structure among a mosaic of unisexual and bisexual populations of the weevil DOI

Johnson S, Leefe W (1999) Clonal diversity and polyphyletic origins of hybrid and spontaneous parthenogenetic PubMed DOI

Sinclair EA, Pramuk JB, Bezy RL, Crandall KA, Sites JW Jr (2010) DNA evidence for nonhybrid origins of parthenogenesis in natural populations of vertebrates. Evolution 64: 1346–1357 doi:10.1111/j.1558–5646.2009.00893.x. PubMed DOI

Simon J -c, Delmotte F, Rispe C, Crease T (2003) Phylogenetic relationships between parthenogens and their sexual relatives: the possible routes to parthenogenesis in animals. Biological Journal of the Linnean Society 79: 151–163 doi:10.1046/j.1095–8312.2003.00175.x. DOI

Kraus F, Miyamoto MM (1990) Mitochondrial genotype of a unisexual salamander of hybrid origin is unrelated to either of its nuclear haplotypes. Proc Natl Acad Sci U S A 87: 2235–2238. PubMed PMC

Sousa-Santos C, Collares-Pereira MJ, Almada VC (2006) Evidence of extensive mitochondrial introgression with nearly complete substitution of the typical DOI

Janko K, Flajšhans M, Choleva L, Bohlen J, Šlechtová V, et al. (2007) Diversity of European spined loaches (genus

Saat (1991) Reproduction of the diploid and polyploid Spinous Loach (

Vasil’ev VP, Akimova NV, Emel’yanova NG, Pavlov DA, Vasil’eva ED (2003) Reproductive capacities in the polyploid males of spined loaches from the unisexual-bisexual complex, occurred in the Moscow River. Folia Biol (Krakow) 51 Suppl: 67–73 PubMed

Janko K, Bohlen J, Lamatsch D, Flajšhans M, Epplen JT, et al. (2007) The gynogenetic reproduction of diploid and triploid hybrid spined loaches ( PubMed

Maciak S, Janko K, Kotusz J, Choleva L, Boroń A, et al. (2011) Standard Metabolic Rate (SMR) is inversely related to erythrocyte and genome size in allopolyploid fish of the DOI

Boroń A (2003) Karyotypes and cytogenetic diversity of the genus PubMed

De Gelas K, Janko K, Volckaert FAM, De Charleroy D, Van Houdt JKJ (2008) Development of nine polymorphic microsatellite loci in the spined loach, PubMed

Kotusz J (2008) Morphological Relationships Between Polyploid Hybrid Spined Loaches of the Genus DOI

Janko K, Kotlik P, Ráb P (2003) Evolutionary history of asexual hybrid loaches ( PubMed

Choleva L, Apostolou A, Ráb P, Janko K (2008) Making it on their own: sperm-dependent hybrid fishes ( PubMed PMC

Templeton AR, Crandall KA, Sing CF (1992) A Cladistic Analysis of Phenotypic Associations with Haplotypes Inferred from Restriction Endonuclease Mapping and DNA Sequence Data. III. Cladogram Estimation. Genetics 132: 619–633. PubMed PMC

Clement M, Posada D, Crandall KA (2000) TCS: a computer program to estimate gene genealogies. Molecular Ecology 9: 1657–1659 doi:10.1046/j.1365-294x.2000.01020.x. PubMed DOI

Anderson EC, Thompson EA (2002) A Model-Based Method for Identifying Species Hybrids Using Multilocus Genetic Data. Genetics 160: 1217–1229. PubMed PMC

Balloux F, Lehmann L, de Meeûs T (2003) The Population Genetics of Clonal and Partially Clonal Diploids. Genetics 164: 1635–1644. PubMed PMC

Peakall ROD, Smouse PE (2006) GENALEX 6: genetic analysis in Excel. Population genetic software for teaching and research. Molecular Ecology Notes 6: 288–295. PubMed PMC

Arnaud-Haond S, Belkhir K (2007) GENCLONE: a computer program to analyse genotypic data, test for clonality and describe spatial clonal organization. Molecular Ecology Notes 7: 15–17.

Arnaud-Haond S, Duarte CM, Alberto F, Serrao EA (2007) Standardizing methods to address clonality in population studies. Molecular Ecology 16: 5115–5139. PubMed

Rogstad SH, Keane B, Beresh J (2002) Genetic variation across VNTR loci in central North American Taraxacum surveyed at different spatial scales. Plant Ecology 161: 111–121.

Douhovnikoff V, Dodd RS (2003) Intra-clonal variation and a similarity threshold for identification of clones: application PubMed

Meirmans PG, Van Tienderen PH (2004) GENOTYPE and GENODIVE: two programs for the analysis of genetic diversity of asexual organisms. Molecular Ecology Notes 4: 792–794.

Guldbrandtsen B, Tomiuk J, Loeschcke V (2000) POPDIST, version 1.1.1: a program to calculate population genetic distance and identity measures. J Hered 91: 178–179. PubMed

Tomiuk J, Loeschcke V (1991) A New Measure of Genetic Identity between Populations of Sexual and Asexual Species. Evolution 45: 1685–1694 doi:10.2307/2409789. PubMed DOI

Otto SP, Whitton J (2000) Polyploid Incidence and Evolution. Annual Review of Genetics 34: 401–437 doi:10.1146/annurev.genet.34.1.401. PubMed DOI

Yue G, David L, Orban L (2007) Mutation rate and pattern of microsatellites in common carp ( PubMed DOI

Hartigan JA, Hartigan P M (1985) The Dip Test of Unimodality. Ann Statist 13: 70–84 doi:10.1214/aos/1176346577. DOI

Culling MA, Janko K, Boron A, Vasil’ev VP, Cote IM, et al. (2006) European colonization by the spined loach ( PubMed

Hotz H, Mancino G, Bucciinnocenti S, Ragghianti M, Berger L, et al. (1985) DOI

Hernández Chávez C, Turgeon J (2007) Asexual and sexual hybrids between PubMed DOI

Vasil’ev VP, Lebedeva EB, Vasil’eva ED, Ryskov P (2007) Monoclonal and de novo arising tetraploid forms of the genus PubMed

Hakoyama H, Nishimura T, Matsubara N, Iguchi K (2001) Difference in parasite load and nonspecific immune reaction between sexual and gynogenetic forms of DOI

Juchno D, Boroń A, Gołaszewski J (2007) Comparative morphology and histology of the ovaries of the spined loach DOI

King KC, Jokela J, Lively CM (2011) Parasites, Sex, and Clonal Diversity in Natural Snail Populations. Evolution 65: 1474–1481 doi:10.1111/j.1558–5646.2010.01215.x. PubMed DOI

Vrijenhoek RC (1979) Factors Affecting Clonal Diversity and Coexistence. American Zoologist 19: 787–797 doi:10.1093/icb/19.3.787. DOI

Butlin RK, Schön I, Martens K (1999) Origin, age and diversity of clones. Journal of Evolutionary Biology 12: 1020–1022 doi:10.1046/j.1420–9101.1999.00126.x. DOI

Pantel JH, Juenger TE, Leibold MA (2011) Environmental gradients structure PubMed DOI

Beukeboom LW, Vrijenhoek RC (1998) Evolutionary genetics and ecology of sperm-dependent parthenogenesis. Journal of Evolutionary Biology 11: 755–782 doi:10.1046/j.1420–9101.1998.11060755.x. DOI

Lowe WH, Allendorf FW (2010) What can genetics tell us about population connectivity? Molecular Ecology 19: 3038–3051 doi:10.1111/j.1365-294X.2010.04688.x. PubMed DOI

Neigel JE, Ball RM, Avise JC (1991) Estimation of Single Generation Migration Distances from Geographic Variation in Animal Mitochondrial DNA. Evolution 45: 423–432 doi:10.2307/2409675. PubMed DOI

Moritz C, Densmore L, Wright J, Brown W (1992) Molecular evidence for constraints on the origin and evolution of parthenogenetic whiptail lizards ( PubMed

Janko K, Drozd P, Eisner J (2011) Do clones degenerate over time? Explaining the genetic variability of asexuals through population genetic models. Biology Direct 6: 17. PubMed PMC

Neiman M, Meirmans S, Meirmans PG (2009) What Can Asexual Lineage Age Tell Us about the Maintenance of Sex? Annals of the New York Academy of Sciences 1168: 185–200 doi:10.1111/j.1749–6632.2009.04572.x. PubMed DOI

Martens K, Loxdale HD, Schön I (2009) The Elusive Clone – In Search of Its True Nature and Identity. Lost Sex: The Evolutionary Biology of Parthenogenesis. Springer-Verlag. 187–200.

Little TJ, Hebert PD (1996) Ancient asexuals: scandal or artifact? Trends Ecol Evol (Amst) 11: 296. PubMed

Janko K, Vasil’ev VP, Rab P, Rabova M, Slechtova V, et al. (2005) Genetic and morphological analyses of 50-chromosome spined loaches (

The Influence of Genome Composition and Temperature on the Hatching Success and Development of the Offspring of Allotriploid Cobitis (Pisces: Cobitidae) Females

Genetic and karyotype divergence between parents affect clonality and sterility in hybrids

Sperm-dependent asexual species and their role in ecology and evolution

Genome Fractionation and Loss of Heterozygosity in Hybrids and Polyploids: Mechanisms, Consequences for Selection, and Link to Gene Function

Challenges and Costs of Asexuality: Variation in Premeiotic Genome Duplication in Gynogenetic Hybrids from Cobitis taenia Complex

No evidence for accumulation of deleterious mutations and fitness degradation in clonal fish hybrids: Abandoning sex without regrets

Cytogenetic Characterization of Seven Novel satDNA Markers in Two Species of Spined Loaches (Cobitis) and Their Clonal Hybrids

The Legacy of Sexual Ancestors in Phenotypic Variability, Gene Expression, and Homoeolog Regulation of Asexual Hybrids and Polyploids

Fine-scale genetic structure of the European bitterling at the intersection of three major European watersheds

All-male hybrids of a tetrapod Pelophylax esculentus share its origin and genetics of maintenance

Hybrid asexuality as a primary postzygotic barrier between nascent species: On the interconnection between asexuality, hybridization and speciation

Asexual Reproduction Does Not Apparently Increase the Rate of Chromosomal Evolution: Karyotype Stability in Diploid and Triploid Clonal Hybrid Fish (Cobitis, Cypriniformes, Teleostei)

Distinguishing between incomplete lineage sorting and genomic introgressions: complete fixation of allospecific mitochondrial DNA in a sexually reproducing fish (Cobitis; Teleostei), despite clonal reproduction of hybrids