Conservation Genetics

, Volume 20, Issue 3, pp 571–584 | Cite as

Paternity analysis reveals constraints on hybridization potential between native and introduced bluebells (Hyacinthoides)

  • D. D. KohnEmail author
  • M. Ruhsam
  • P. E. Hulme
  • S. C. H. Barrett
  • P. M. Hollingsworth
Research Article


The native UK bluebell Hyacinthoides non-scripta is considered to be at risk from hybridization with naturalised non-native bluebells. The non-natives, likely to be hybrid themselves (H. x massartiana) between H. non-scripta and H. hispanica, occur in relatively small numbers throughout the UK range of natives. Full interfertility between taxa has been repeatedly asserted in reporting spread of non-natives and predicting genetic erosion or assimilation of the native. However, there have been no published data to support suppositions that non-natives arose from in-situ hybridization, or that natural hybridization represents an ongoing threat to the native bluebell. Here we first investigated hybridization potential via reciprocal hand-crosses and observed overlap in flowering periods of native and non-native bluebells, finding that flowering was largely synchronous and that seed set and early seedling survival were equivalent for between-taxon crosses. We then established an experimental array allowing natural pollination to occur among flowering plants, and determined the paternity of offspring using microsatellite markers. We found that natives were more likely to produce seeds than non-natives, and that paternities approximated three native to one non-native, regardless of the identity of the maternal parent. Our results demonstrate that hybridization in natural populations and introgression between natives and non-natives are possible. However, lower reproductive success of non-natives coupled with the massive numerical advantage of natives represents a substantial constraint against ‘extinction-by-hybridization’ of H. non-scripta in the UK.


Invasive species Congenerics Flowering phenology Genetic erosion Introgression Paternity analysis 



This work was supported by a Daphne Jackson Fellowship to DDK funded by the Natural Environment Research Council, the European Union FP 6 Integrated Project ALARM: Assessing LArgescale environmental Risks for biodiversity with tested Methods (GOCE-CT-2003-506675), the Botanical Society of the British Isles, the National Botanic Gardens of Scotland Membership Programme Small Projects Fund, and a Discovery Grant from the Natural Sciences and Engineering Research Council of Canada to SCHB. Our thanks to Jane Squirrell for work on pollen and chromosomes at RBGE, to Alex Clark for translating Geerinck (1996), and to landowners for permissions. The Royal Botanic Garden Edinburgh is supported by the Scottish Government’s Rural and Environmental Science and Analytical Services Division.

Supplementary material

10592_2019_1158_MOESM1_ESM.docx (53 kb)
Supplementary Fig. 1 Design of the experimental array established at the Royal Botanic Garden Edinburgh to investigate natural crossing between native and non-native bluebells. (A) A group of 16 flowering plants, showing pattern of individually-potted natives (8) and non-natives (8). The triploid plant was a non-native. (B) Schematic field layout of experimental array (not to scale). The seven groups (shaded boxes), of 16 bluebells each, occupied a total area of 75 m x 40 m, with each group 23.5-35 m from the nearest other group (DOCX 53 KB)
10592_2019_1158_MOESM2_ESM.docx (823 kb)
Supplementary Fig. 2. Example of stained and unstained bluebell pollen, showing 100% viability (native, left) and 14% viability (non-native, right). Size bars on the images are 100 µm at lower magnification (left) and 50 µm at the higher magnification (right) (DOCX 822 KB)
10592_2019_1158_MOESM3_ESM.docx (18 kb)
Supplementary Table 1 Statistics and results (in bold) for interfertility and early seedling survival in experimental hand-crosses between native and non-native Hyacinthoides taxa. See Figure 4 for plots of these data. Crosses are maternal taxon x paternal taxon; crosses in 2009 and 2010 comprised native maternal plants only. (DOCX 17 KB)


  1. Beatty GE, Barker L, Chen P-P, Kelleher CT, Provan J (2015) Cryptic introgression into the kidney saxifrage (Saxifraga hirsuta) from its more abundant sympatric congener Saxifraga spathularis, and the potential risk of genetic assimilation. Ann Bot 115:179–186CrossRefPubMedGoogle Scholar
  2. Blackman GE, Rutter AJ (1954) Endymion [Hyacinthoides] non-scriptus (L.) Garcke [non-scripta (L.) Chouard ex Rothm.]. J Ecol 42:629–638CrossRefGoogle Scholar
  3. Bleeker W, Schmitz U, Ristow M (2007) Interspecific hybridisation between alien and native plant species in Germany and its consequences for native biodiversity. Biol Conserv 137:248–253CrossRefGoogle Scholar
  4. Bonin A, Bellemain E, Eidesen PB, Pompanon F, Brochmann C, Taberlet P (2004) How to track and assess genotyping errors in population genetics studies. Mol Ecol 13:3261–3273CrossRefPubMedGoogle Scholar
  5. Branquard E, Vanderhoeven S, Van Landuyt W, Van Rossum F, Verloove F (2010) Harmonia database: Hyacinthoides hispanica. Harmonia version 1.2, Belgian Forum on Invasive Species, Accessed 1 Oct 2013 from:
  6. Cheung KW, Razeq FM, Sauder CA, James T, Martin SL (2015) Bidirectional but asymmetrical sexual hybridization between Brassica carinata and Sinapis arvensis (Brassicaceae). J Plant Res 128:469–480CrossRefPubMedGoogle Scholar
  7. Corbet SA (1998) Fruit and seed production in relation to pollination and resources in bluebell Hyacinthoides non-scripta. Oecologia 114:349–360CrossRefPubMedGoogle Scholar
  8. Corbet SA (1999) Spatiotemporal patterns in the flowering of bluebell, Hyacinthoides non-scripta (Hyacinthaceae). Flora 194:345–356CrossRefGoogle Scholar
  9. Corbet SA, Tiley CF (1999) Insect visitors to flowers of bluebell (Hyacinthoides non-scripta). Entomol Monthly Mag 135(1620–1623):133–141Google Scholar
  10. Crawley MJ (2005) The Flora of Berkshire. Harpenden. Brambleby Books, HertsGoogle Scholar
  11. Crawley MJ (2007) The R Book. Wiley, ChichesterCrossRefGoogle Scholar
  12. Cruzan MB, Barrett SCH (1996) Postpollination mechanisms influencing mating patterns and fecundity: an example from Eichhornia paniculata. Am Nat 147:576–598CrossRefGoogle Scholar
  13. Currat M, Ruedi M, Petit RJ, Excoffier L (2008) The hidden side of invasions: massive introgression by local genes. Evolution 62:1908–1920PubMedGoogle Scholar
  14. Doyle JJ, Doyle JL (1990) Isolation of plant DNA from fresh tissue. Focus 12:13–15Google Scholar
  15. Ellstrand NC, Schierenbeck KA (2000) Hybridization as a stimulus for the evolution of invasiveness in plants? Proc Natl Acad Sci 97:7043–7050CrossRefPubMedGoogle Scholar
  16. Geerinck D (1996) An epithet for the hybrid Hyacinthoides hispanica (Mill.) Rothm x Hyacinthoides non-scripta. (L.) Chouard ex Rothm: Hyacinthoides x massartiana Geerinck (Liliaceae). Belg J Bot 129:83–85Google Scholar
  17. Genovart M (2009) Natural hybridization and conservation. Biodivers Conserv 18:1435–1439CrossRefGoogle Scholar
  18. Grosser C, Potts BM, Vaillancourt RE (2010) Microsatellite based paternity analysis in a clonal Eucalyptus nitens seed orchard. Silvae Genet 59:57–62CrossRefGoogle Scholar
  19. Grundmann M, Rumsey FJ, Ansell SW, Russell SJ, Darwin SC, Vogel JC, Spencer M, Squirrell J, Hollingsworth PM, Ortiz S, Schneider H (2010) Phylogeny and taxonomy of Hyacinthoides. Taxon 59:68–82CrossRefGoogle Scholar
  20. Hallmann CA, Sorg M, Jongejans E, Siepel H, Hofland N, Schwan H, Stenmans W, Müller A, Sumser H, Hörren T, Goulson D, de Kroon H (2017) More than 75 percent decline over 27 years in total flying insect biomass in protected areas. PLoS ONE 12(10):e0185809. CrossRefPubMedPubMedCentralGoogle Scholar
  21. Hersch EI, Roy BA (2007) Context-dependent pollinator behavior: an explanation for patterns of hybridization among three species of Indian paintbrush. Evolution 61:111–124CrossRefPubMedGoogle Scholar
  22. Hodkinson DJ, Thompson K (1997) Plant dispersal: the role of man. J Appl Ecol 34:1484–1496CrossRefGoogle Scholar
  23. Huxel GR (1999) Rapid displacement of native species by invasive species: effects of hybridization. Biol Conserv 89:143–152CrossRefGoogle Scholar
  24. Ietswaart JH, de Smet SJM, Lubbers JPM (1983) Hybridisation between Scilla non-scripta. and S. hispanica (Liliaceae) in the Netherlands. Acta Bot Neerl 32:467–480CrossRefGoogle Scholar
  25. Ingrouille M (1995) Historical ecology of the British flora. Chapman and Hall, LondonCrossRefGoogle Scholar
  26. Johnson MG, Lang K, Manos P, Golet GH, Schierenbeck KA (2016) Evidence for genetic erosion of a California native tree, Platanus racemosa, via recent, ongoing introgressive hybridization with an introduced ornamental species. Conserv Genet 17:593–602CrossRefGoogle Scholar
  27. Johnston JA, Arnold ML, Donovan LA (2003) High hybrid fitness at seed and seedling life history stages in Louisiana irises. J Ecol 91:438–446CrossRefGoogle Scholar
  28. Kalinowski ST, Taper ML, Marshall TC (2007) Revising how the computer program CERVUS accommodates genotyping error increases success in paternity assignment. Mol Ecol 16:1099–1106CrossRefGoogle Scholar
  29. Klein EK, Lagache-Navarro L, Petit RJ (2017) Demographic and spatial determinants of hybridization rate. J Ecol 105:29–38. CrossRefGoogle Scholar
  30. Knight GH (1964) Some factors affecting the distribution of Endymion nonscriptus (L.) Garcke in Warwickshire woods. J Ecol 52:405–421CrossRefGoogle Scholar
  31. Kohn DD, Hulme PE, Hollingsworth PM, Butler A (2009) Are native bluebells (Hyacinthoides non-scripta) at risk from alien congenerics? Evidence from distributions and co-occurrence in Scotland. Biol Conserv 142:61–74CrossRefGoogle Scholar
  32. Kowarik I (2003) Human agency in biological invasions: secondary releases foster naturalisation and expansion of alien plant species. Biol Invasions 5:293–312Google Scholar
  33. Larcombe MJ, Holland B, Steane DA, Jones RC, Nicolle D, Vaillancourt RE, Potts BM (2015) Patterns of reproductive isolation in Eucalyptus—a phylogenetic perspective. Mol Biol Evol 32:1833–1846CrossRefPubMedGoogle Scholar
  34. Leslie AC (1987) Flora of surrey supplement and checklist. AC & P Leslie, Guildford, p 43Google Scholar
  35. Lewontin RC, Birch LC (1966) Hybridization as a source of variation for adaptation to new environments. Evolution 20:315–336CrossRefPubMedGoogle Scholar
  36. Li TJ, Xu LL, Liao L, Deng HS, Han XJ (2014) Patterns of hybridization in a multispecies hybrid zone in the Ranunculus cantoniensis complex (Ranunculaceae). Bot J Linn Soc 174:227–239CrossRefGoogle Scholar
  37. Littlemore J, Barker S (2001) The ecological response of forest ground flora and soils to experimental trampling in British urban woodlands. Urban Ecosyst 5:257–276CrossRefGoogle Scholar
  38. Mallet J (2005) Hybridization as an invasion of the genome. Trends Ecol Evol 20:229–237CrossRefGoogle Scholar
  39. Mallet J (2008) Hybridization, ecological races and the nature of species: empirical evidence for the ease of speciation. Philos Trans R Soc London B 363:2971–2986CrossRefGoogle Scholar
  40. Marshall TC, Slate J, Kruuk LEB, Pemberton JM (1998) Statistical confidence for likelihood-based paternity inference in natural populations. Mol Ecol 7:639–655CrossRefPubMedGoogle Scholar
  41. Marutani M, Sheffer RD, Kamemoto H (1993) Cytological analysis of Anthurium andraeanum (Araceae), its related taxa and their hybrids. Am J Bot 80:93–103CrossRefGoogle Scholar
  42. McKellar MA, Quesenberry KH (1992) Chromosome pairing and pollen viability in Desmodium ovalifolium Wall X Desmodium heterocarpon (L) Dc hybrids. Aust J Bot 40:243–247CrossRefGoogle Scholar
  43. Merryweather J, Fitter A (1995) Arbuscular mycorrhiza and phosphorus as controlling factors in the life history of Hyacinthoides non-scripta (L.) Chouard ex Rothm. New Phytol 129:629–636CrossRefGoogle Scholar
  44. Natalis LC, Wesselingh RA (2012) Post-pollination barriers and their role in asymmetric hybridization in Rhinanthus (Orobanchaceae). Am J Bot 99:1847–1856CrossRefPubMedGoogle Scholar
  45. Page KW (1987) Hybrid bluebells. Bot Soc Br Isles News 46:9Google Scholar
  46. Pilgrim E, Hutchinson N (2004) Bluebells for Britain—the report of the 2003 Bluebells for Britain survey. In: Vines G (ed). Salisbury: PlantlifeGoogle Scholar
  47. Plume O, Raimondo FM, Troia A (2015) Hybridization and competition between the endangered sea marigold (Calendula maritima, Asteraceae) and a more common congener. Plant Biosyst 149:68–77CrossRefGoogle Scholar
  48. Prentis PJ, White EM, Radford IJ, Lowe AJ, Clarke AR (2007) Can hybridization cause local extinction: A case for demographic swamping of the Australian native Senecio pinnatifolius by the invasive Senecio madagascariensis? New Phytol 176:902–912. CrossRefPubMedGoogle Scholar
  49. Preston CD, Pearman DA, Dines TD (eds) (2002) New atlas of the British Flora. Oxford University Press, Oxford, p 817Google Scholar
  50. Quené-Boterenbrood AJ (1984) Over het voorkomen van Scilla non-scripta (L.) Hoffmanns, Link S. hispanica Miller en hun hybride in Nederland. Gorteria 12:91–104Google Scholar
  51. R Core Team (2013) R: A language and environment for statistical computing. R Foundation for Statistical Computing, Vienna, Austria. URL
  52. Rands SA, Whitney HM (2011) Field margins, foraging distances and their impacts on nesting pollinator success. PLoS ONE 6(10):e25971. CrossRefPubMedPubMedCentralGoogle Scholar
  53. Rhymer JM, Simberloff D (1996) Extinction by hybridization and introgression. Annu Rev Ecol Syst 27:83–109CrossRefGoogle Scholar
  54. Rieseberg LH, Archer MA, Wayne RK (1999) Transgressive segregation, adaptation and speciation. Heredity 83:363–372CrossRefPubMedGoogle Scholar
  55. Rix M (2004) Plate 481 Hyacinthoides non-scripta, Hyacinthaceae. Curtis’s Bot Mag 21:20–25CrossRefGoogle Scholar
  56. Simon M, Durand S, Pluta N, Gobron N, Botran L, Ricou A, Camilleri C, Budar F (2016) Genomic conflicts that cause pollen mortality and raise reproductive barriers in Arabidopsis thaliana. Genetics 203:1353–1367CrossRefPubMedPubMedCentralGoogle Scholar
  57. Srikanth S, Rao MV, Mallikarjuna N (2014) Interspecific hybridization between Cajanus cajan (L.) Millsp and C. lanceolatus (WV Fitgz) van der Maesen. Plant Genet Resour 12:255–258CrossRefGoogle Scholar
  58. Stace CA, Preston CD, Pearman DA (2015) Hybrid Flora of the British Isles. Botanical Society of Britain & Ireland, LondonGoogle Scholar
  59. Stebbins GL (1950) Variation and evolution in plants. Columbia University press, New York.USACrossRefGoogle Scholar
  60. Thompson PA, Cox SA (1978) Germination of the bluebell Hyacinthoides non-scripta in relation to its distribution and habitat. Ann Bot 42:51–62CrossRefGoogle Scholar
  61. Thompson K, Grime JP (1979) Seasonal variation in the seed banks of herbaceous species in ten contrasting habitats. J Ecol 67:893–921CrossRefGoogle Scholar
  62. Turrill WB (1951) Some problems of plant range and distribution. J Ecol 39:205–227CrossRefGoogle Scholar
  63. Urbanek S, Bibiko H-J (2012) GUI 1.62 (6558 Snow Leopard build), © R Foundation for Statistical Computing, 2012. R: Copyright © 2004–2013, The R Foundation for Statistical Computing;
  64. Vandelook F, Van Assche JA (2008) Temperature requirements for seed germination and seedling development determine timing of seedling emergence of three monocotyledonous temperate forest spring geophytes. Ann Bot 102:865–875CrossRefPubMedPubMedCentralGoogle Scholar
  65. Warburg EF (1962) Endymion. p 974 in: Clapham AR, Tutin TG, Warburg EF (eds) Flora of the British Isles. Cambridge: Cambridge University PressGoogle Scholar
  66. Wildlife and Countryside Act (1981)
  67. Wilson JY (1956) Polyploidy in bluebells (Endymion nonscriptus and E. hispanicus). Nature 178:195CrossRefGoogle Scholar
  68. Wilson JY (1959) Vegetative reproduction in the bluebell Endymion nonscriptus (L.) Garcke. New Phytol 58:155–163CrossRefGoogle Scholar
  69. Wolf DE, Takebayashi N, Rieseberg LH (2001) Predicting the risk of extinction through hybridization. Conserv Biol 15:1039–1053CrossRefGoogle Scholar
  70. Zlonis KJ, Gross BL (2018) Genetic structure, diversity, and hybridization in populations of the rare arctic relict Euphrasia hudsoniana (Orobanchaceae) and its invasive congener. Euphrasia stricta. Conserv Genet 19:43–55. CrossRefGoogle Scholar

Copyright information

© Springer Nature B.V. 2019

Authors and Affiliations

  1. 1.Royal Botanic Garden EdinburghEdinburghUK
  2. 2.Bio-Protection Research CentreLincoln UniversityChristchurchNew Zealand
  3. 3.Department of Ecology and Evolutionary BiologyUniversity of TorontoTorontoCanada

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