Annals of Forest Science

, 75:90 | Cite as

Geometric morphometric analyses of leaf shapes in two sympatric Chinese oaks: Quercus dentata Thunberg and Quercus aliena Blume (Fagaceae)

  • Yuan Liu
  • Yuejuan Li
  • Jialin Song
  • Ruipu Zhang
  • Yu Yan
  • Yuyao Wang
  • Fang K. DuEmail author
Research Paper


Key message

Geometric morphometric analyses (GMMs) of the leaf shape can distinguish two congeneric oak species Quercus dentata Thunberg and Quercus aliena Blume in sympatric areas.


High genetic and morphological variation in different Quercus species hinder efforts to distinguish them. In China, Q. dentata and Q. aliena are generally sympatrically distributed in warm temperate forests, and share some leaf morphological characteristics.


The aim of this study was to use the morphometric methods to discriminate these sympatric Chinese oaks preliminarily identified from molecular markers.


Three hundred sixty-seven trees of seven sympatric Q. dentata and Q. aliena populations were genetically assigned to one of the two species or hybrids using Bayesian clustering analysis based on nSSR. This grouping served as a priori classification of the trees. Shapes of 1835 leaves from the 367 trees were analyzed in terms of 13 characters (landmarks) by GMMs. Correlations between environmental and leaf morphology parameters were studied using linear regression analyses.


The two species were efficiently discriminated by the leaf morphology analyses (96.9 and 95.9% of sampled Q. aliena trees and Q. dentata trees were correctly identified), while putative hybrids between the two species were found to be morphologically intermediate. Moreover, we demonstrated that the leaf morphological variations of Q. aliena, Q. dentata, and their putative hybrids are correlated with environmental factors, possibly because the variation of leaf morphology is part of the response to different habitats and environmental disturbances.


GMMs were able to correctly classify individuals from the two species preliminary identified as Q. dentata or Q. aliena by nSSR. The high degree of classification accuracy provided by this approach may be exploited to discriminate other problematic species and highlight its utility in plant ecology and evolution studies.


Geometric morphometrics Genetic assignment Leaf morphology Quercus Sympatric distribution 



We thank anonymous reviewers for helpful comments on a previous version of this manuscript. We thank Dr. Antoine Kremer of INRA, France; Saneyoshi Ueno of Forestry and Forest Products Research Institute, Japan; and Dr. Nian Wang of Shandong Agricultural University, China, for the improvement of the manuscript. We would also like to thank Dr. Juqing Kang of Shanxi Normal University and Shangfang Mountain National Forest Park, Beijing, P.R. China, for assisting us during the field sampling.


This research was supported by the Fundamental Research Funds for the Central Universities (grant no. 2015ZCQ-LX-03), the National Science Foundation of China (grant no. 41671039), and the Beijing Nova Program for FKD (grant no. Z151100000315056).

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Supplementary material

13595_2018_770_MOESM1_ESM.docx (1.2 mb)
ESM 1 (DOCX 1227 kb)


  1. Abràmoff MD, Magalhães PJ, Ram SJ (2005) Image processing with Image J Part II. Biophoton Int 11:36–43. CrossRefGoogle Scholar
  2. Adams DC, Slice DE, Rohlf FJ (2004) Geometric morphometrics: ten years of progress following the ‘revolution’. Ital J Zool 71:5–16.
  3. Arnold ML, Ballerini ES, Brothers AN (2012) Hybrid fitness, adaptation and evolutionary diversification: lessons learned from Louisiana irises. Heredity 108:159–166. CrossRefPubMedGoogle Scholar
  4. Bacilieri R, Ducousso A, Kremer A (1996) Comparison of morphological characters and molecular markers for the analysis of hybridization in sessile and pedunculate oak. Ann Sci For 53:79–91.
  5. Benzécri JP (1992) Correspondence analysis handbook. Biometrics 49:672. CrossRefGoogle Scholar
  6. Bookstein FL (1996) Combining the tools of geometric morphometrics. In: Advances in Morphometrics. Springer, Boston, pp 131–151CrossRefGoogle Scholar
  7. Box EO, Fujiwara K (2015). Warm-temperate deciduous forests: concept and global overview. In: Warm-temperate deciduous forests around the Northern hemisphere. Geobotany studies. Springer Cham, pp 7–26Google Scholar
  8. Bresson CC, Vitasse Y, Kremer A, Delzon S (2011) To what extent is altitudinal variation of functional traits driven by genetic adaptation in European oak and beech? Tree Physiol 31:1164–1174. CrossRefPubMedGoogle Scholar
  9. Chatterjee S, Hadi AS, Price B (2006) Simple linear regression. Regression analysis by example. John Wiley and Sons, New York, pp 21–51Google Scholar
  10. Costa C, Paglia G, Salvador FR, Lolletti D, Rimatori V, Menesatti P (2009) Hazelnut cultivar identification with leaf morphometric analysis: preliminary results. Acta Hortic (845):245–248.
  11. Dormann CF, Elith J, Bacher S, Buchmann C, Carl G, Carré G (2013) Collinearity: a review of methods to deal with it and a simulation study evaluating their performance. Ecography 36:27–46. CrossRefGoogle Scholar
  12. Dow BD, Ashley MV, Howe HF (1995) Characterization of highly variable (GA/CT) n microsatellites in the bur oak, Quercus macrocarpa. Theor Appl Genet 91:137–141. CrossRefPubMedGoogle Scholar
  13. Du FK, Peng XL, Liu JQ, Lascoux M, Hu FS, Petit RJ (2011) Direction and extent of organelle DNA introgression between two spruce species in the Qinghai-Tibetan Plateau. New Phytol 192:1024–1033. CrossRefPubMedGoogle Scholar
  14. Du FK, Xu F, Qu H, Feng S, Tang J, Wu R (2013) Exploiting the transcriptome of euphrates poplar, Populus euphratica (Salicaceae) to develop and characterize new EST-SSR markers and construct an EST-SSR database. PLoS One 8:e61337. CrossRefPubMedPubMedCentralGoogle Scholar
  15. Durand J, Bodénès C, Chancerel E, Frigerio JM, Vendramin G, Sebastiani F (2010) A fast and cost-effective approach to develop and map EST-SSR markers: oak as a case study. BMC Genomics 11:570. CrossRefPubMedPubMedCentralGoogle Scholar
  16. Earl DA, Vonholdt BM (2012) STRUCTURE HARVESTER: a website and program for visualizing structure output and implementing the Evanno method. Conserv Genet Resour 4:359–361 CrossRefGoogle Scholar
  17. Eaton DA, Hipp AL, González-Rodríguez A, Cavender-Bares J (2015) Historical introgression among the American live oaks and the comparative nature of tests for introgression. Evolution 69:2587–2601. CrossRefPubMedGoogle Scholar
  18. Elias TS (1980) The complete trees of North America. Van Nostrand Reinhold Co., New YorkGoogle 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 14:2611–2620. CrossRefPubMedGoogle Scholar
  20. Fujiwara K, Harada A (2015) Character of warm-temperate Quercus forests in Asia. Warm-temperate deciduous forests around the northern hemisphere. Springer, Cham, pp 27–80Google Scholar
  21. Gerber S, Chadoeuf J, Gugerli F, Lascoux M, Buiteveld J, Cottrell J, Goicoechea PG (2014) High rates of gene flow by pollen and seed in oak populations across Europe. PLoS One 9:e85130. CrossRefPubMedPubMedCentralGoogle Scholar
  22. Gower JC (1975) Generalised Procrustes analysis. Psychometrika 40:33–51CrossRefGoogle Scholar
  23. Graham JH, Raz S, HelOr H, Nevo E (2010) Fluctuating asymmetry: methods, theory, and applications. Symmetry 2:466–540. CrossRefGoogle Scholar
  24. Gugerli F, Walser JC, Dounavi K, Holderegger R, Finkeldey R (2007) Coincidence of small-scale spatial discontinuities in leaf morphology and nuclear microsatellite variation of Quercus petraea and Q. robur in a mixed forest. Ann Bot 99:713–722. CrossRefPubMedPubMedCentralGoogle Scholar
  25. Guichoux E, Lagache L, Wagner S, Petit R (2011) Current trends in microsatellites genotyping. Mol Ecol Resour 11:591–611. CrossRefPubMedGoogle Scholar
  26. Huang CJ, Zhang YT, Bruce B (1999a) Fagaceae. Flora of China, vol 4. Science Press, Beijing, pp 314–400Google Scholar
  27. Huang CJ, Zhang YT, Bartholomew B (1999b) Fagaceae. In: Wu ZY, Raven PH (eds) Flora of China Vol. 4 (in English). Science Press, Beijing, pp 370–380Google Scholar
  28. Hubert F, Grimm GW, Jousselin E, Berry V, Franc V, Kremer A (2014) Multiple nuclear genes stabilize the phylogenetic backbone of the genus Quercus. Syst Biodivers 12:405–423. CrossRefGoogle Scholar
  29. Janes JK, Miller JM, Dupuis JR, Malenfant RM, Gorrell JC, Cullingham CI, Andrew RL (2017) The K=2 conundrum. Mol Ecol 26:3594–3602. CrossRefPubMedGoogle Scholar
  30. Jaramillo-Correa JP, Aguirre-Planter E, Khasa DP, Eguiarte LE, Piñero D, Furnier GR (2008) Ancestry and divergence of subtropical montane forest isolates: molecular biogeography the genus Abies (Pinaceae) in southern Mexico and Guatemala. Mol Ecol 17:2476–2490. CrossRefPubMedGoogle Scholar
  31. Jensen RJ, Ciofani KM, Miramontes LC (2002) Lines, outlines, and landmarks: morphometric analyses of leaves of Acer rubrum, Acer saccharinum (Aceraceae) and their hybrid. Taxon 51:475–492.
  32. Klingenberg CP (1998) Heterochrony and allometry: the analysis of evolutionary change in ontogeny. Biol Rev 73:79–123. CrossRefPubMedGoogle Scholar
  33. Klingenberg CP (2003) A developmental perspective on developmental instability: theory, models and mechanisms. In: Polak M, ed. Developmental instability: causes and consequences. Camb Law J, New York pp 14–34Google Scholar
  34. Klingenberg CP (2011) MorphoJ: an integrated software package for geometric morphometrics. Mol Ecol Resour 11:353–357. CrossRefPubMedGoogle Scholar
  35. Klingenberg CP, Monteirob LR (2005) Distances and directions in multidimensional shape spaces: implications for morphometric applications. Syst Biol 54:678–688. CrossRefPubMedGoogle Scholar
  36. Klingenberg CP, Barluenga M, Meyer A (2002) Shape analysis of symmetric structures: quantifying variation among individuals and asymmetry. Evolution 56:1909–1920. CrossRefPubMedGoogle Scholar
  37. Kremer A, Dupouey LJ, Deans JD, Cottrell J, Csaikl U, Finkeldey R, Ducousso A (2002) Leaf morphological variation in mixed oak stands (Quercus robur and Quercus petraea) in stable western European population. Ann For Sci 59:777–787. CrossRefGoogle Scholar
  38. Lepais O, Petit RJ, Guichoux E, Lavabre JE, Alberto F, Kremer A (2009) Species relative abundance and direction of introgression in oaks. Mol Ecol 18:2228–2242. CrossRefPubMedGoogle Scholar
  39. Lyu J, Song J, Liu Y, Wang Y, Li J, Du FK (2018) Species boundaries between three sympatric oak species: Quercus aliena, Q. dentata, and Q. variabilis at the northern edge of their distribution in China. Front Plant Sci 9:414. CrossRefPubMedPubMedCentralGoogle Scholar
  40. MacLeod N, Forey PL (2002). Introduction: morphology, shape, and phylogenetics. In: Morphology, shape, and phylogeny. Taylor & Francis, London, pp 1–7Google Scholar
  41. Manos PS, Doyle JJ, Nixon KC (1999) Phylogeny, biogeography, and processes of molecular differentiation in Quercus subgenus Quercus (Fagaceae). Mol Phylogenet Evol 12:333–349. CrossRefPubMedGoogle Scholar
  42. Mitteroecker P, Gunz P (2009) Advances in geometric morphometrics. Evol Biol 36:235–247. CrossRefGoogle Scholar
  43. Nixon KC (1993) Infrageneric classification of Quercus (Fagaceae) and typification of sectional names. Ann For Sci 50:25s–34s.
  44. Nixon KC (1997) Quercus. In: Flora of North America editorial committee (eds) vol. 3. Oxford University Press, New York, pp 445–447Google Scholar
  45. Nugroho A, Song BM, Su HS, Choi JS, Choi J, Choi JY (2016) HPLC analysis of phenolic substances and anti-Alzheimer’s activity of Korean Quercus species. Nat Prod Sci 22:299. CrossRefGoogle Scholar
  46. Peñaloza-Ramírez JM, González-Rodríguez A, Mendoza-Cuenca L, Caron H, Kremer A, Oyama K (2010) Interspecific gene flow in a multispecies oak hybrid zone in the Sierra Tarahumara of Mexico. Ann Bot 105:389–399. CrossRefPubMedPubMedCentralGoogle Scholar
  47. Peng YS, Chen L, Li JQ (2007) Study on numerical taxonomy of Quercus L. (Fagaceae) in China. J Wuhan Bot Res 25:149–157. CrossRefGoogle Scholar
  48. Petit RJ, Duminil J, Fineschi S, Hampe A, Salvini D, Vendramin GG (2005) Comparative organization of chloroplast, mitochondrial and nuclear diversity in plant populations. Mol Ecol 14:689–701. CrossRefPubMedGoogle Scholar
  49. Pritchard JK, Stephens M, Donnelly P (2000) Inference of population structure using multilocus genotype data. Genetics 155: 945-959.Google Scholar
  50. Rellstab C, Bühler A, Graf R, Folly C, Gugerli F (2016) Using joint multivariate analyses of leaf morphology and molecular-genetic markers for taxon identification in three hybridizing European white oak species ( Quercus, spp.). Ann For Sci 73:1–11. CrossRefGoogle Scholar
  51. Ren XW, Wang LM (1985) Geographic distribution of deciduous oaks in China. Journal of Beijing Forestry University 2:57–69Google Scholar
  52. Rohlf FJ (2010) tpsDig, digitize landmarks and outlines, ver. 2.16. Department of Ecology and Evolution, State University of New York at Stony Brook, NJ. life.
  53. Rohlf FJ, Slice DE (1990) Extensions of the Procrustes method for the optimal superimposition of landmarks. Syst Zool 39:40–59.
  54. Rushton BS (1993) Natural hybridization within the genus Quercus L. Ann For Sci 50:73–90. CrossRefGoogle Scholar
  55. Savriama Y, Klingenberg CP (2011) Beyond bilateral symmetry: geometric morphometric methods for any type of symmetry. BMC Evol Biol 11:280. CrossRefPubMedPubMedCentralGoogle Scholar
  56. Slice DE (1996) A glossary for geometric morphometrics. Advances in morphometrics, pp 531–551Google Scholar
  57. Souza SMF, Moreira DAI, Joseph MS (2012) Geometric morphometrics of leaf blade shape in Montrichardia linifera (Araceae) populations from the Rio Parnaíba delta, north-east Brazil. Bot J Linn Soc 170:554–572. CrossRefGoogle Scholar
  58. Stace CA (1982) Plant taxonomy and biosystematics. Brittonia 34:80–80CrossRefGoogle Scholar
  59. Stephan JM, Teeny PW, Vessella F, Schirone B (2018) Oak morphological traits: between taxa and environmental variability. Flora 243:32–44. CrossRefGoogle Scholar
  60. Suarez - Gonzalez A, Hefer CA, Lexer C, Douglas CJ, Cronk Q (2018) Introgression from Populus balsamifera underlies adaptively significant variation and range boundaries in P. trichocarpa. New Phytol 217:416–427. CrossRefPubMedGoogle Scholar
  61. Tsukaia H (2005) Leaf shape: genetic controls and environmental factors. Int J Dev Biol 49:547–555. CrossRefGoogle Scholar
  62. Tucić B, Budečević S, Manitašević SJ, Vuleta A, Klingenberg CP (2018) Phenotypic plasticity in response to environmental heterogeneity contributes to fluctuating asymmetry in plants: first empirical evidence. J Evol Biol 31:197–210. CrossRefPubMedGoogle Scholar
  63. Ueno S, Taguchi Y, Tsumura Y (2008) Microsatellite markers derived from Quercus mongolica var. crispula (Fagaceae) inner bark expressed sequence tags. Genes Genet Syst 83:179–187. CrossRefPubMedGoogle Scholar
  64. Ueno S, Aoki K, Tsumura Y (2009) Generation of expressed sequence tags and development of microsatellite markers for Castanopsis sieboldii var. sieboldii (Fagaceae). Ann For Sci 66:509. CrossRefGoogle Scholar
  65. Vähä JP, Primmer CR (2006) Efficiency of model-based Bayesian methods for detecting hybrid individuals under different hybridization scenarios and with different numbers of loci. Mol Ecol 15:63–72. CrossRefPubMedGoogle Scholar
  66. Viscosi V (2015) Geometric morphometrics and leaf phenotypic plasticity: assessing fluctuating asymmetry and allometry in European white oaks (Quercus). Bot J Linn Soc 179:335–348. CrossRefGoogle Scholar
  67. Viscosi V, Cardini A (2011) Leaf morphology, taxonomy and geometric morphometrics: a simplified protocol for beginners. PLoS ONE 6:e25630.
  68. Viscosi V, Fortini P (2011) Leaf shape variation and differentiation in three sympatric white oak species revealed by elliptic Fourier analysis. Nord J Bot 29: 632-640. doi: 10.1111/j.1756-1051.2011.01098.xGoogle Scholar
  69. Viscosi V, Fortini P, Slice DE, Loy A, Blasi C (2009) Geometric morphometric analyses of leaf variation in four oak species of the subgenus Quercus (Fagaceae). Plant Biosyst 143:575–587. CrossRefGoogle Scholar
  70. Viscosi V, Antonecchia G, Lepais O, Fortini P, Gerber S, Loy A (2012) Leaf shape and size differentiation in white oaks: assessment of allometric relationships among three sympatric species and their hybrids. Int J Plant Sci 173:875–884. CrossRefGoogle Scholar
  71. Vitasse Y, Bresson CC, Kremer A (2010) Quantifying phenological plasticity to temperature in two temperate tree species. Funct Ecol 24:1211–1218. CrossRefGoogle Scholar
  72. Yoshioka Y, Iwata H, Ohsawa R, Ninomiya S (2004) Analysis of petal shape variation of Primula sieboldii by elliptic Fourier descriptors and principal component analysis. Ann Bot 94:657–664. CrossRefPubMedPubMedCentralGoogle Scholar
  73. Zelditch ML, Swiderski DL, Sheets HD (2012) Glossary-geometric morphometrics for biologists, (2nd edn). Geometric morphometrics for biologists, pp 455–469Google Scholar
  74. Zeng YF, Liao WJ, Petit RJ (2010) Exploring species limits in two closely related Chinese oaks. PLoS One 5:e15529. CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© INRA and Springer-Verlag France SAS, part of Springer Nature 2018

Authors and Affiliations

  1. 1.College of ForestryBeijing Forestry UniversityBeijingPeople’s Republic of China
  2. 2.School of Biological Science and TechnologyUniversity of JinanJinanPeople’s Republic of China
  3. 3.Kunyushan National Forest ParkYantaiPeople’s Republic of China

Personalised recommendations