European Journal of Forest Research

, Volume 132, Issue 2, pp 195–209 | Cite as

Assignment testing reveals multiple introduced source populations including potential ash hybrids (Fraxinus excelsior × F. angustifolia) in Ireland

  • Muriel Thomasset
  • Juan F. Fernández-Manjarrés
  • Gerry C. Douglas
  • Paola Bertolino
  • Nathalie Frascaria-Lacoste
  • Trevor R. Hodkinson
Original Paper


Large-scale reforestation programmes are a major source of unwarranted gene flow and can have profound consequences on local genetic diversity. Recently, ash has been introduced to Ireland from continental Europe to stock plantations but has often exhibited poor stem form. It was not known whether these trees were Fraxinus excelsior, Fraxinus angustifolia or interspecific hybrids that are known to occur in continental Europe. F. excelsior is the only native ash in Ireland, so the introduced populations represent a potential threat for the genetic diversity of native populations. We characterized the introduced trees within two plantations (Clonee and Kildalkey) using morphological characters and six microsatellite markers. Samples from continental Europe were included for comparison. Plantations exhibited higher genetic diversity than control populations because our data suggest they contain a mixture of several provenances. There was a small but significant differentiation between plantations and control populations (ФCT = 0.0211). Bayesian analysis to infer population structure and to assign introduced individuals to reference populations clearly demonstrated the presence of hybrid individuals within the plantations. The percentage of trees detected with potential hybrid origin ranged from 28 to 58 % depending on the plantation and the threshold data analysis level chosen. Most hybrids could be considered cryptic because there was a lack of intermediate morphology for hybrid individuals that mainly clustered with F. excelsior. The results indicated that the source of material at the two plantation sites differed. Management options to minimize the impact of these introduced populations are discussed.


Assignment test Cryptic hybrids Fraxinus Hybridization Leaf morphology Microsatellites 



This work forms part of Muriel Thomasset’s PhD research and was supported by COFORD and a Teagasc Walsh Fellowship. We thank the owners of the two plantation sites Mr. Gerald Potterton and Mr. Brian White for the access to their land. Special thanks to Bridget Brankin for her help with the morphological measurements and to Martina Temunovic for access to the genotypes of her samples from Croatia and Bulgaria.

Supplementary material

10342_2012_667_MOESM1_ESM.doc (142 kb)
Supplementary material 1 (DOC 142 kb)


  1. Allendorf FW, Leary RF, Spruell P, Wenburg JK (2001) The problems with hybrids: setting conservation guidelines. Trends Ecol Evol 16(11):613–622CrossRefGoogle Scholar
  2. Anderson E, Stebbins J (1954) Hybridization as an evolutionary stimulus. Evolution 8:378–388CrossRefGoogle Scholar
  3. Anderson EC, Thompson EA (2002) A model-based method for identifying species hybrids using multilocus genetic data. Genetics 160:1217–1229PubMedGoogle Scholar
  4. Anttila CK, Daehler CC, Rank NE, Strong DR (1998) Greater male fitness of a rare invader (Spartina alterniflora, Poaceae) threatens a common native (Spartina foliosa) with hybridization. Am J Bot 85(11):1597–1601PubMedCrossRefGoogle Scholar
  5. Arnold ML (1992) Natural hybridization as an evolutionary process. Annu Rev Ecol Syst 23(1):237–261CrossRefGoogle Scholar
  6. Arnold ML (1997) Natural hybridization and evolution. Oxford University Press, OxfordGoogle Scholar
  7. Arnold ML (2004) Natural hybridization and the evolution of domesticated, pest and disease organisms. Mol Ecol 13(5):997–1007PubMedCrossRefGoogle Scholar
  8. Barbour RC, Potts BM, Vaillancourt RE, Tibbits WN, Wiltshire RJE (2002) Gene flow between introduced and native Eucalyptus species. New For 23:177–191CrossRefGoogle Scholar
  9. Bartlett MS (1937) Properties of sufficiency and statistical tests. Royal Statistical Society. pp 268–282Google Scholar
  10. Brachet S, Jubier MF, Richard M, Jung-Muller B, Frascaria-Lacoste N (1999) Rapid identification of microsatellite loci using 5′ anchored PCR in the common ash Fraxinus excelsior. Mol Biol 8:157–169Google Scholar
  11. Craft KJ, Ashley MV, Koenig WD (2002) Limited hybridization between Quercus lobata and Quercus douglasii (Fagaceae) in a mixed stand in central coastal California. Am J Bot 89(11):1792–1798PubMedCrossRefGoogle Scholar
  12. Daniels R, Sheail RE (1999) Genetic pollution: concepts, concerns and transgenic crops. In: Lutman PW (ed) Gene flow and agriculture: relevance to transgenic crops. British Crop Protection Council, Farnham, ROYAUME-UNI, pp 65–72Google Scholar
  13. Douglas GC, Fernandez-Manjarres JF, Hodkinson TR, Frascaria-Lacoste N, Thomasset M (2009) Keeping Irish trees pure. Sci Spin 31:10–11Google Scholar
  14. Earl DA (2009) Structure Harvester v0.3, from website:
  15. Ellstrand NC (1992) Gene flow by pollen: implications for plant conservation genetics. Oikos 63(1):77–86CrossRefGoogle Scholar
  16. Ellstrand NC, Elam DR (1993) Population genetic consequences of small population size: implications for plant conservation. Annu Rev Ecol Syst 24:217–242CrossRefGoogle Scholar
  17. Ellstrand NC, Schierenbeck KA (2000) Hybridization as a stimulus for the evolution of invasiveness in plants? Euphytica 148(1–2):35–46Google Scholar
  18. Ellstrand NC, Prentice HC, Hancock JF (1999) Gene flow and introgression from domesticated plants into their wild relatives. Annu Rev Ecol Syst 30:539–563CrossRefGoogle Scholar
  19. Evanno G, Regnaut S, Goudet J (2005) Detecting the number of clusters of individuals using the software structure: a simulation study. Mol Ecol 14(8):2611–2620PubMedCrossRefGoogle Scholar
  20. Excoffier L, Smouse PE, Quattro JM (1992) Analysis of molecular variance inferred from metric distances among DNA haplotypes: application to human mitochondrial DNA restriction data. Genetics 131:479–491PubMedGoogle Scholar
  21. Excoffier L, Laval G, Schneider S (2005) Arlequin ver. 3.0: an integrated software package for population genetics data analysis. Evol Bioinformatics Online 1:47–50Google Scholar
  22. Fernández-Manjarres JF, Gerard PR, Dufour J, Raquin C, Frascaria-Lacoste N (2006) Differential patterns of morphological and molecular hybridization between Fraxinus excelsior L. and Fraxinus angustifolia Vahl (Oleaceae) in eastern and western France. Mol Ecol 15:3245–3257PubMedCrossRefGoogle Scholar
  23. Genton BJ, Shykoff JA, Giraud T (2005) High genetic diversity in French invasive populations of common ragweed, Ambrosia artemisiifolia, as a result of multiple sources of introduction. Mol Ecol 14(14):4275–4285PubMedCrossRefGoogle Scholar
  24. Gérard PR, Fernández-Manjarrés JF, Frascaria-Lacoste N (2006a) Temporal cline in a hybrid zone population between Fraxinus excelsior L. and Fraxinus angustifolia Vahl. Mol Ecol 15:3655–3667PubMedCrossRefGoogle Scholar
  25. Gérard PR, Klein EK, Austerlitz F, Fernández-Manjarrés JF, Frascaria-Lacoste N (2006b) Assortative mating and differential male mating success in an ash hybrid zone population. BioMed Central Evol Biol 6:96–110CrossRefGoogle Scholar
  26. Goudet J (2001) FSTAT, a program to estimate and test gene diversities and fixation indices (version 2.9.3). Available from Updated from Goudet (1995)
  27. Guo SW, Thompson EA (1992) Performing the exact test of Hardy-Weinberg proportion for multiple alleles. Biometrics 48(2):361–372PubMedCrossRefGoogle Scholar
  28. Harbourne ME (2005) The characterisation of genetic diversity of common ash (Fraxinus excelsior L.) in Ireland and around Europe. Trinity College, DublinGoogle Scholar
  29. Heuertz M, Hausman J-F, Tsvetkov I, Frascaria-Lacoste N, Vekemans X (2001) Assessment of genetic structure within and among Bulgarian populations of the common ash (Fraxinus excelsior L.). Mol Ecol 10:1615–1623PubMedCrossRefGoogle Scholar
  30. Heuertz M, Fineschi S, Anzidei M, Pastorelli R, Salvini D, Paule L, Frascaria-Lacoste N, Hardy OJ, Vekemans X, Vendramin GG (2004a) Chloroplast DNA variation and postglacial recolonization of common ash (Fraxinus excelsior L.) in Europe. Mol Ecol 13:3437–3452PubMedCrossRefGoogle Scholar
  31. Heuertz M, Hausman J-F, Hardy OJ, Vendramin GG, Frascaria-Lacoste N, Vekemans X (2004b) Nuclear microsatellites reveal contrasting patterns of genetic structure between western and southeastern european populations of the common ash (Fraxinus excelsior L.). Evolution 58(5):976–988PubMedGoogle Scholar
  32. Heuertz M, Carnevale S, Fineschi S, Sebastiani F, Hausman JF, Paule L, Vendramin GG (2006) Chloroplast DNA phylogeography of European ashes, Fraxinus sp. (Oleaceae): roles of hybridization and life history traits. Mol Ecol 15(8):2131–2140PubMedCrossRefGoogle Scholar
  33. Hodkinson TR, Ní Chonghaile G, Sungkaew S, Chase MW, Salamin N, Stapleton CMA (2010) Phylogenetic analyses of plastid and nuclear DNA sequences indicate a rapid late Miocene radiation of the temperate bamboo tribe Arundinarieae (Poaceae, Bambusoideae). Plant Ecol Divers 3:109–120CrossRefGoogle Scholar
  34. Jato V, Rodriguez-Rajo FJ, Dacosta N, Aira MJ (2004) Heat and chill requirements of Fraxinus flowering in Galicia (NW Spain). Grana 43:217–223CrossRefGoogle Scholar
  35. Jeandroz S, Frascaria-Lacoste N, Bousquet J (1996) Molecular recognition of the closely related Fraxinus excelsior and F. oxyphylla (Oleaceae) by RAPD markers. For Genet 3(4):237–242Google Scholar
  36. Johnston JA, Donovan LA, Arnold ML (2004) Novel phenotypes among early generation hybrids of two Louisiana iris species: flooding experiments. J Ecol 92:967–976CrossRefGoogle Scholar
  37. Kalinowski ST, Taper ML, Marshall TC (2007) Revising how the computer program CERVUS accommodates genotyping error increases success in paternity assignment. Mol Ecol 16:1099–1106PubMedCrossRefGoogle Scholar
  38. Kowarik I (1995) Time-lags in biological invasions. In: Pyšek P, Prach K, Rejmánek M, Wade W (eds) Plant invasions: general aspects and special problems. SPB Academic Publ, Amsterdam, pp 15–38Google Scholar
  39. Kowarik I, von der Lippe M (2007) Pathways in plant invasions. In: Nentwig W (ed) Biological invasions, vol 193. Ecological studies, vol 193. Springer, Berlin, pp 29–47CrossRefGoogle Scholar
  40. Lefort F, Brachet S, Frascaria-Lacoste N, Edwards KJ, Douglas GC (1999) Identification and characterization of microsatellite loci in ash (Fraxinus excelsior L.) and their conservation in the olive family (Oleaceae). Mol Ecol 8:1088–1090CrossRefGoogle Scholar
  41. Legendre P, Legendre L (1998) Numerical ecology. Elsevier sciences, AmsterdamGoogle Scholar
  42. Lopez GA, Potts BM, Tilyard PA (2000) F1 hybrid inviability in Eucalyptus: the case of E-ovata x E-globulus. Heredity 85(3):242–250PubMedCrossRefGoogle Scholar
  43. Mack RN, Lonsdale WM (2001) Humans as global plant dispersers: getting more than we bargained for. Bioscience 51(2):95–102. doi:10.1641/0006-3568(2001)051[0095:HAGPDG]2.0.CO;2CrossRefGoogle Scholar
  44. Marigo G, Peltier JP, Girel J, Pautou G (2000) Success in the demographic expansion of Fraxinus excelsior L. Trees 15:1–13CrossRefGoogle Scholar
  45. Marshall TC, Slate J, Kruuk LEB, Pemberton JM (1998) Statistical confidence for likelihood-based paternity inference in natural populations. Mol Ecol 7(5):639–655PubMedCrossRefGoogle Scholar
  46. McGrath S, Hodkinson TR, Barth S (2007) Extremely high cytoplasmic diversity in natural and breeding populations of Lolium (Poaceae). Heredity 99:531–544PubMedCrossRefGoogle Scholar
  47. Meirmans PG, Hedrick PW (2011) Assessing population structure: FST and related measures. Mol Ecol Resour 11(1):5–18PubMedCrossRefGoogle Scholar
  48. Meirmans PG, Van Tienderen PH (2004) GENOTYPE and GENODIVE: two programs for the analysis of genetic diversity of asexual organisms. Mol Ecol Notes 4:792–794CrossRefGoogle Scholar
  49. Morand M-E, Brachet S, Rossignol P, Dufour J, Frascaria-Lacoste N (2002) A generalized heterozygote deficiency assessed with microsatellites in French common ash populations. Mol Ecol 11:377–385PubMedCrossRefGoogle Scholar
  50. Muller E, Guilloy-Froget H, Barsoum N, Brocheton L (2002) Populus nigra L. in the Garonne Valley: legacy of the past and present constraints. C R biol 325:1129–1141PubMedCrossRefGoogle Scholar
  51. Nei M (1987) Molecular evolutionary genetics. Columbia University Press, New YorkGoogle Scholar
  52. Nielsen EE, Bach LA, Kotlicki P (2006) hybridlab (version 1.0): a program for generating simulated hybrids from population samples. Mol Ecol Notes 6(4):971–973CrossRefGoogle Scholar
  53. Peakall R, Smouse PE (2006) GENALEX 6: genetic analysis in Excel. Population genetic software for teaching and research. Mol Ecol Notes 6:288–295Google Scholar
  54. Peterken GF (2001) Ecological effects of introduced tree species in Britain. For Ecol Manag 141(1–2):31–42CrossRefGoogle Scholar
  55. Potts BM, Barbour RC, Hingston AB, Vaillancourt RE (2003) Genetic pollution of native eucalypt gene pools—identifying the risks. Aust J Bot 51(1):1–25CrossRefGoogle Scholar
  56. Pritchard JK, Stephens M, Donnelly P (2000) Inference of population structure using multilocus genotype data. Genetics 155(2):945–959PubMedGoogle Scholar
  57. Raquin C, Brachet S, Jeandroz S, Vedel F, Frascaria-Lacoste N (2002) Combined analyses of microsatellite and RAPD markers demonstrate possible hybridisation between Fraxinus excelsior L. and Fraxinus angustifolia Vahl. For Genet 9(2):111–114Google Scholar
  58. Raymond M, Rousset F (1995) (version 1.2): population genetics software for exact tests and ecumenicism. J Hered 86:248–249Google Scholar
  59. Rhymer JM, Simberloff D (1996) Extinction by hybridization and introgression. Annu Rev Ecol Syst 27:83–109CrossRefGoogle Scholar
  60. Richardson DM, Pyšek P, Rejmánek M, Barbour MG, Panetta FD, West CJ (2000) Naturalization and invasion of alien plants: concepts and definitions. Divers Distrib 6(2):93–107CrossRefGoogle Scholar
  61. Rieseberg LH, Ellstrand NC (1993) What can molecular and morphological markers tell us about plant hybridization? Crit Rev Plant Sci 12(3):213–241Google Scholar
  62. Rieseberg LH, Willis JH (2007) Plant speciation. Science 317(5840):910–914PubMedCrossRefGoogle Scholar
  63. Rieseberg LH, Sinervo B, Linder CR, Ungerer MC, Arias DM (1996) Role of gene interactions in hybrid speciation: evidence from ancient and experimental hybrids. Science 272(5262):741–745. doi: 10.1126/science.272.5262.741 PubMedCrossRefGoogle Scholar
  64. Rieseberg LH, Raymond O, Rosenthal DM, Lai Z, Livingstone K, Nakazato T, Durphy JL, Schwarzbach AE, Donovan LA, Lexer C (2003) Major ecological transitions in wild sunflowers facilitated by hybridization. Science 301(5637):1211–1216PubMedCrossRefGoogle Scholar
  65. Sakai AK, Allendorf FW, Holt JS, Lodge DM, Molofsky J, With KA, Baughman S, Cabin RJ, Cohen JE, Ellstrand NC, McCauley DE, O’Neil P, Parker IM, Thompson JN, Weller SG (2001) The population biology of invasive species. Annu Rev Ecol Syst 32(1):305–332. doi: 10.1146/annurev.ecolsys.32.081501.114037 CrossRefGoogle Scholar
  66. Sala OE, Chapin FS III, Armesto JJ, Berlow E, Bloomfield J, Dirzo R, Huber-Sanwald E, Huenneke LF, Jackson RB, Kinzig A, Leemans R, Lodge DM, Mooney HA, Oesterheld M, Poff NL, Sykes MT, Walker BH, Walker M, Wall DH (2000) Global biodiversity scenarios for the year 2100. Science 287(5459):1770–1774PubMedCrossRefGoogle Scholar
  67. Schierenbeck KA, Symonds VV, Gallagher KG, Bell J (2005) Genetic variation and phylogeographic analyses of two species of Carpobrotus and their hybrids in California. Mol Ecol 14:539–547PubMedCrossRefGoogle Scholar
  68. Shapiro SS, Wilk MB (1965) An analysis of variance test for normality (complete samples). Biometrika 52(3–4):591–611Google Scholar
  69. Squirrell J, Hollingsworth PM, Bateman RM, Dickson JH, Light MHS, MacConaill M, Tebbitt MC (2001) Partitioning and diversity of nuclear and organelle markers in native and introduced populations of Epipactis helleborine (Orchidaceae). Am J Bot 88(8):1409–1418PubMedCrossRefGoogle Scholar
  70. Stace CA (1975) Hybridization and the Flora of the British Isles. Academic press, LondonGoogle Scholar
  71. StatSoft, Inc (2004) Electronic statistics textbook. Tulsa, OK, StatSoft.
  72. Sutherland BG, Belaj A, Nier S, Cottrell JE, Vaughan SP, Hubert J, Russell K (2010) Molecular biodiversity and population structure in common ash (Fraxinus excelsior L.) in Britain: implications for conservation. Mol Ecol 19:2196–2211PubMedCrossRefGoogle Scholar
  73. Thomasset M (2011) Introduced hybrid ash: Fraxinus excelsior × F. angustifolia in Ireland and its potential for interbreeding with native ash. Trinity College, DublinGoogle Scholar
  74. Thomasset M, Fernández-Manjarres JF, Douglas GC, Frascaria- Lacoste N, Raquin C, Hodkinson TR (2011a) Molecular and morphological characterization of reciprocal F1 hybrid ash (Fraxinus excelsior × F. angustifolia, Oleaceae) and parental species reveals asymmetric character inheritance. Int J Plant Sci 172:423–433Google Scholar
  75. Thomasset M, Fernández-Manjarrés JF, Douglas GC, Frascaria-Lacoste N, Hodkinson TR (2011b) Hybridisation, introgression and climate change: a case study for the tree genus Fraxinus (Oleaceae). In: Hodkinson TR, Jones MB, Waldren S, Parnell JAN (eds) Climate change, ecology and systematics. Cambridge University Press, CambridgeGoogle Scholar
  76. Tovar-Sanchez E, Oyama K (2004) Natural hybridization and hybrid zones between Quercus crassifolia and Quercus crassipes (Fagaceae) in Mexico: morphological and molecular evidence. Am J Bot 91(9):1352–1363. doi: 10.3732/ajb.91.9.1352 PubMedCrossRefGoogle Scholar
  77. Usher AV, Whelan RJ, Ayre DJ (2010) Window of opportunity: an episode of recruitment in a Banksia hybrid zone demonstrates continuing hybridization and phenotypic plasticity. Ann Bot 105(3):419–429PubMedCrossRefGoogle Scholar
  78. Vähä J-P, Primmer CR (2006) Efficiency of model-based Bayesian methods for detecting hybrid individuals different hybridization scenarios and with different numbers of loci. Mol Ecol 15:63–72PubMedCrossRefGoogle Scholar
  79. Valbuena-Carabanã M, Gonzàlez-Martinez SC, Sork VL, Collada C, Soto A, Goicoechea PG, Gil L (2005) Gene flow and hybridisation in a mixed oak forest (Quercus pyrenaica Willd. and Quercus petraea (Matts.) Liebl.) in central Spain. Heredity 95:457–465PubMedCrossRefGoogle Scholar
  80. Vanden Broeck A, Villar M, Van Bockstaelle E, Van Slycken J (2005) Natural hybridization between cultivated poplars and their wild relatives: evidence and consequences for native poplar populations. Ann For Sci 62:601–613CrossRefGoogle Scholar
  81. Wallander E (2008) Systematics of Fraxinus (Oleaceae) and evolution of dioecy. Plant Syst Evol 273:25–49CrossRefGoogle Scholar
  82. Weir BS, Cockerham CC (1984) Estimating F-statistics for the analysis of population structure. Evolution 38:1358–1370CrossRefGoogle Scholar
  83. Williamson MH, Fitter AH (1996) The characters of successful invaders. Biol Conserv 78:163–170CrossRefGoogle Scholar
  84. Ziegenhagen B, Gneuss S, Rathmacher G, Leyer I, Bialozyt R, Heinze B, Liepelt S (2008) A fast and simple genetic survey reveals the spread of poplar hybrids at a natural Elbe river site. Conserv Genet 9(2):373–379CrossRefGoogle Scholar
  85. Zobel BJ, van Wyk G, Stahl P (1987) Growing exotic forests. USA, New YorkGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2012

Authors and Affiliations

  • Muriel Thomasset
    • 1
    • 2
  • Juan F. Fernández-Manjarrés
    • 3
    • 4
    • 5
  • Gerry C. Douglas
    • 2
  • Paola Bertolino
    • 3
    • 4
    • 5
  • Nathalie Frascaria-Lacoste
    • 4
    • 3
    • 5
  • Trevor R. Hodkinson
    • 1
    • 6
  1. 1.School of Natural Sciences, Trinity College DublinUniversity of DublinDublin 2Ireland
  2. 2.Kinsealy Research CentreTeagascDublin 17Ireland
  3. 3.CNRS, UMR 8079OrsayFrance
  4. 4.AgroParisTechParisFrance
  5. 5.UMR 8079Université Paris-SudOrsayFrance
  6. 6.Trinity Centre for Biodiversity Research, Trinity College DublinUniversity of DublinDublin 2Ireland

Personalised recommendations