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Cytomolecular diversity of the subtribe Laeliinae (Epidendroidae, Orchidaceae) suggests no relationship between genome size and heterochromatin abundance

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The subtribe Laeliinae (Orchidaceae) comprises approximately 2000 species and 38 genera, with numerically stable karyotypes of 2n = 40 (x = 20) in most species. Cytomolecular and chromosome banding analyses have not yet been combined with phylogenetic studies, making the understanding of chromosomal evolution in the group more difficult. The present work sought to perform comparative cytogenetic analyses of representatives of the subtribe Laeliinae, followed by the interpretation of that data within an evolutive context. We analyzed 23 species from seven genera for their chromosome numbers and morphology, the distribution of CMA and DAPI bands, fluorescent in situ hybridization (FISH) with 5S and 35S rDNA and DNA quantification by flow cytometry. All of the species showed 2n = 40, suggesting an apparent macrostructural karyotypic stability, except the polyploids Encyclia alboxanthina, E. jenischiana, E. seidelii, Laelia gouldiana and Prosthechea faresiana (2n = 80). At least two CMA+/DAPI terminal bands were observed in all analyzed species. Additionally, CMA+/DAPI blocks were observed in the pericentromeric region of species of the genera Brassavola, Cattleya (except C. trianae) and Laelia. Terminal CMA/DAPI+ bands were observed in representatives of Brassavola, Laelia and Guarianthe. The clade Brassavola + Cattleya + Laelia, that presented high number of heterochromatic bands, also showed expansions of the numbers of 5S rDNA sites. DNA contents varied from 2C = 2.85 pg (Prosthechea fragrans) to 2C = 5.76 pg (Laelia gouldiana) without relation with number of CMA/DAPI bands or 5S rDNA sites. The relationship between the amplification of repetitive DNAs (viewed here as rDNA sites or heterochromatic bands) and genome size variation is discussed and compared with other plant groups.

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  1. Acosta MC, Moscone EA, Cocucci AA (2016) Using chromosomal data in the phylogenetic and molecular dating framework: karyotype evolution and diversification in Nierembergia (Solanaceae) influenced by historical changes in sea level. Pl Biol 18:514–526. https://doi.org/10.1111/plb.12430

  2. Altinkut A, Raskina O, Nevo E, Belyayev A (2006) En/Spm-like transposons in Poaceae species: transposase sequence variability and chromosomal distribution. Cell Molec Biol Lett 11:214–230. https://doi.org/10.2478/s11658-006-0017-3

  3. Ambrožová K, Mandáková T, Bureš P, Neumann P, Leitch IJ et al (2011) Diverse retrotransposon families and an AT-rich satellite DNA revealed in giant genomes of Fritillaria lilies. Ann Bot (Oxford) 107:255–268. https://doi.org/10.1093/aob/mcq235

  4. Assis FNM, Souza BCQ, Medeiros-Neto E, Pinheiro F, Silva AEB, Felix LP (2013) Karyology of the genus Epidendrum (Orchidaceae: Laeliinae) with emphasis on subgenus Amphiglottium and chromosome number variability in Epidendrum secundum. Bot J Linn Soc 172:329–344

  5. Baltisberger M, Hörandl E (2016) Karyotype evolution supports the molecular phylogeny in the genus Ranunculus (Ranunculaceae). Perspect Pl Ecol Evol Syst 18:1–14. https://doi.org/10.1016/j.ppees.2015.11.001

  6. Barros F, Vinhos F, Rodrigues VT, Barberena FFVA, Fraga CN, Pessoa EM, Forster W, Menini Neto L, Furtado SG, Nardy C, Azevedo CO, Guimarães LRS (2015) Orchidaceae in Lista de Espécies da Flora do Brasil. Jardim Botânico do Rio de Janeiro, Rio de Janeiro. Available at: http://floradobrasil.jbrj.gov.br/jabot/floradobrasil/FB11329

  7. Barros-e-Silva AE, Guerra M (2010) The meaning of DAPI bands observed after C-banding and FISH procedures. Biotech Histochem 85:115–125. https://doi.org/10.3109/10520290903149596

  8. Barros-e-Silva AE, Marques A, Santos KGB, Guerra M (2010) The evolution of CMA bands in Citrus and related genera. Chromosome Res 18:503–514. https://doi.org/10.1007/s10577-010-9130-2

  9. Bastos CA (2014) Filogenia do Gênero Encyclia Hook. (Orchidaceae—Laeliinae) e Revisão Taxonômica das Espécies Brasileiras. PhD Thesis, Universidade Estadual de Feira de Santana, Feira de Santana

  10. Begum R, Alam SS, Menzel G, Schmidt T (2009) Comparative molecular cytogenetics of major repetitive sequence families of three Dendrobium species (Orchidaceae) from Bangladesh. Ann Bot (Oxford) 104:863–872. https://doi.org/10.1093/aob/mcp178

  11. Bennett MD, Leitch IJ (2005) Nuclear DNA amounts in Angiosperms: progress, problems and prospects. Ann Bot (Oxford) 95:45–90. https://doi.org/10.1093/aob/mci003

  12. Bennetzen JL, Ma J, Devos KM (2005) Mechanisms of recent genome size variation in flowering plants. Ann Bot (Oxford) 95:127–132. https://doi.org/10.1093/aob/mci008

  13. Cabral JS, Felix LP, Guerra M (2006) Heterochromatin diversity and its co-localization with 5S and 45S rDNA sites in chromosomes of four Maxillaria species (Orchidaceae). Genet Molec Biol 29:659–664. https://doi.org/10.1590/S1415-47572006000400015

  14. Chase MW, Cameron KM, Barrett RL, Freudenstein JV, Pridgeon AM, Salazar G, van den Berg C, Schuiteman A (2015) An updated classification of Orchidaceae. Bot J Linn Soc 177:151–174. https://doi.org/10.1111/boj.12234

  15. Chiron GR, Castro VP (2002) Révision dês espèces brésiliènnes du genre Laelia Lindley. Richardiana 2:4–28. https://doi.org/10.13140/2.1.4273.6320

  16. D’Emerico S, Pignone D, Bartolo G, Pulvirenti S, Terrasi C, Stuto S, Scrugli A (2005) Karyomorphology, heterochromatin patterns and evolution in the genus Ophrys (Orchidaceae). Bot J Linn Soc 148:87–99. https://doi.org/10.1111/j.1095-8339.2005.00393.x

  17. Datson PM, Murray BG (2006) Ribosomal DNA locus evolution in Nemesia: transposition rather than structural rearrangement as the key mechanism? Chromosome Res 14:845–857. https://doi.org/10.1007/s10577-006-1092-z

  18. Deanna R, Smith SD, Särkinen T, Chiarini F (2018) Patterns of chromosomal evolution in the florally diverse Andean clade Iochrominae (Solanaceae). Perspec Plant Ecol Evol Syst 35:31–43. https://doi.org/10.1016/j.ppees.2018.09.004

  19. Demerico SD, Galasso I, Pignone D, Scrugli A (2001) Localization of rDNA loci by fluorescent in situ hybridization in some wild orchids from Italy (Orchidaceae). Caryologia 54:31–36. https://doi.org/10.1080/00087114.2001.10589211

  20. Doležel J, Sgorbati S, Lucretti S (1992) Comparison of three DNA fluorochromes for flow cytometric estimation of nuclear DNA content in plants. Physiol Pl 85:625–631. https://doi.org/10.1111/j.1399-3054.1992.tb04764.x

  21. Dressler RL (1981) The orchids: natural history and classification. Harvard University Press, Harvard. https://doi.org/10.2307/1219717

  22. Dressler RL (1993) Phylogeny and classification of the orchid family. Dioscorides Press, Portland

  23. Felix LP, Guerra M (2010) Variation in chromosome number and the basic number of subfamily Epidendroideae (Orchidaceae). Bot J Linn Soc 163:234–278. https://doi.org/10.1111/j.1095-8339.2010.01059.x

  24. Govaerts R, Bernet P, Kratochvil K, Gerlach G, Carr G, Alrich P, Pridgeon AM, Pfahl J, Campacci MA, Baptista H, Tigges H, Shaw J, Cribb J, George A, Kreuz K, Wood J (2015) World checklist of Orchidaceae. Board of Trustees of the Royal Botanic Gardens, Kew. Available at: http://www.kew.org/wcsp/monocots/. Accessed 6 Nov 2015

  25. Guerra MS (1986) Reviewing the chromosome nomenclature of Levan et. Brazil J Genet 9:741–743

  26. Hall KJ, Parker JS (1995) Stable chromosome fission associated with rDNA mobility. Chromosome Res 3:417–422. https://doi.org/10.1007/BF00713891

  27. Hanson RE, Islam-Faridi MN, Percival EA, Crane CF, Ji Y, McKnight TD, Stelly DM, Price HJ (1996) Distribution of 5S and 18S-28S rDNA loci in a tetraploid cotton (Gossypium hirsutum L.) and its putative diploid ancestors. Chromosoma 105:55–61. https://doi.org/10.1007/BF02510039

  28. Hoffmann V, Verboom GA, Cotterill FP (2015) Dated plant phylogenies resolve neogene climate and landscape evolution in the Cape Floristic Region. PLoS ONE 10:e0137847. https://doi.org/10.1371/journal.pone.0137847

  29. Jones HG (1975) Nomenclatural revision of the genus Brassavola R. BR. of the Orchidaceae. Ann Nathist Mus Wien 79:9–22

  30. Kao YY, Chang SB, Lin TY, Hsieh CH, Chen YH, Chen WH, Chen CC (2001) Differential accumulation of heterochromatin as a cause for karyotype variation in Phalaenopsis orchids. Ann Bot (Oxford) 873:387–395. https://doi.org/10.1006/anbo.2000.1348

  31. Kirov I, Khrustaleva L, Van Laere K, Soloviev A, Meeus S, Romanov D, Fesenko I (2017) DRAWID: user-friendly java software for chromosome measurements and idiogram drawing. Comp Cytogenet 11:747–757. https://doi.org/10.3897/CompCytogen.v11i4.20830

  32. Koehler S, Cabral JS, Whitten WM, Williams NH, Singer RB, Neubig KM, Guerra M, Souza AP, Amaral MC (2008) Molecular phylogeny of the neotropical genus Christensonella (Orchidaceae, Maxillariinae): species delimitation and insights into chromosome evolution. Ann Bot (Oxford) 102:491–507. https://doi.org/10.1093/aob/mcn128

  33. Kondo K, Hoshi Y, Tanaka R (1994) Somatic chromosome differentiation in Cypripedium segawai Masamune and C. japonicum Thunberg. Cytologia 59:115–120. https://doi.org/10.1508/cytologia.59.115

  34. Kovarik A, Dadejova M, Lim YK, Chase MW, Clarkson JJ, Knapp S, Leitch AR (2008) Evolution of rDNA in Nicotiana allopolyploids: a potential link between rDNA homogenization and epigenetics. Ann Bot (Oxford) 101:815–823. https://doi.org/10.1093/aob/mcn019

  35. Lan T, Albert VA (2011) Dynamic distribution patterns of ribosomal DNA and chromosomal evolution in Paphiopedilum, a lady’s slipper orchid. BMC Pl Biol 11:126. https://doi.org/10.1186/1471-2229-11-126

  36. Las Peñas ML, Kiesling R, Bernardello G (2019) Phylogenetic reconstruction of the genus Tephrocactus (Cactaceae) based on molecular, morphological, and cytogenetical data. Taxon 68:714–730. https://doi.org/10.1002/tax.12092

  37. Lee YI, Yap JW, Izan S, Leitch IJ, Fay MF, Lee YC, Hidalgo O, Dodsworth S, Smulders M, Gravendeel B, Leitch Leitch AR (2018) Satellite DNA in Paphiopedilum subgenus Parvisepalum as revealed by high-throughput sequencing and fluorescent in situ hybridization. BMC Genom 19:578. https://doi.org/10.1186/s12864-018-4956-7

  38. Leitch IJ, Bennett MD (2004) Genome downsizing in polyploid plants. Biol J Linn Soc 82:651–663. https://doi.org/10.1111/j.1095-8312.2004.00349.x

  39. Leitch IJ, Kahandawala I, Suda J, Hanson L, Ingrouille MJ, Chase MW, Fayet MF (2009) Genome size diversity in orchids: consequences and evolution. Ann Bot (Oxford) 104:469–481. https://doi.org/10.1093/aob/mcp003

  40. Liu B, Davis T (2011) Conservation and loss of ribosomal RNA gene sites in diploid and polyploid Fragaria (Rosaceae). BMC Pl Biol 11:157. https://doi.org/10.1186/1471-2229-11-157

  41. Loureiro J, Rodriguez E, Doležel J, Conceição S (2006) Comparasion of four nuclear isolation buffers for plant DNA flow cytometry. Ann Bot (Oxford) 98:679–689. https://doi.org/10.1093/aob/mcl141

  42. Macas J, Novak P, Pellicer J, Cizkova J, Koblizkova A, Neumann P, Fukova I, Dolezel J, Kelly LJ, Leitch IJ (2015) In depth characterization of repetitive DNA in 23 plant genomes reveals sources of genome size variation in the legume tribe Fabeae. PLoS ONE 10:e0143424. https://doi.org/10.1371/journal.pone.0143424

  43. Moraes AP, Leitch I, Lia J, Leitch AR (2012) Chromosome studies in Orchidaceae: karyotype divergence in Neotropical genera in subtribe Maxillariinae. Bot J Linn Soc 170:29–39. https://doi.org/10.1111/j.1095-8339.2012.01266.x

  44. Moraes AP, Chinaglia M, Palma-Silva C, Pinheiro F (2013) Interploidy hybridization in sympatric zones: the formation of Epidendrum fulgens x E. puniceoluteum hybrids (Epidendroideae, Orchidaceae). Ecol Evol 3:3824–3837. https://doi.org/10.1002/ece3.752

  45. Moreno NC, Amarilla LD, Las peñas ML, Bernardello G (2015) Molecular cytogenetic insights into the evolution of the epiphytic genus Lepismium (Cactaceae) and related genera. Bot J Linn Soc 177:263–277. https://doi.org/10.1111/boj.12242

  46. Moscone EA, Samuel R, Schwarzacher T, Schweizer D, Pedrosa-Harand A (2007) Complex rearrangements are involved in Cephalanthera (Orchidaceae) chromosome evolution. Chromosome Res 15:931–943. https://doi.org/10.1007/s10577-007-1174-6

  47. Oliveira IG, Moraes A, Almeida EM, Assis F, Cabral JS, Barros F, Felix LP (2015) Chromosomal evolution in Pleurothallidinae (Orchidaceae: Epidendroideae) with an emphasis on the genus Acianthera: chromosome numbers and heterochromatin: chromosomal evolution in Pleurothallidinae. Bot J Linn Soc 178:102–120. https://doi.org/10.1111/boj.12273

  48. Pabst GFJ, Dungs F (1975) Orchidaceae Brasiliensis, vol. 1. Brücke-Verlag Kurt Schmersow, Hildesheim

  49. Pabst GFJ, Dungs F (1977) Orchidaceae Brasiliensis, vol. 2. Brücke-Verlag Kurt Schmersow, Hildesheim

  50. Pedrosa A, Sandal N, Stougaard J, Schweiser D, Bachmair A (2002) Chromosomal map of the model Legume Lotus japonicus. Genetics 161:1661–1672

  51. Pridgeon AM, Cribb PJ, Chase MW, Rasmussen FN (2005) Genera Orchidacearum, vol. 4. Epidendroideae (Part 1). Oxford University Press, New York

  52. Raskina O, Belyayev A, Nevo E (2004) Activity of the En/Spm-like transposons in meiosis as a base for chromosome repatterning in a small, isolated, peripheral population of Aegilops speltoides Tausch. Chromosome Res 12:153. https://doi.org/10.1023/B:CHRO.0000013168.61359.43

  53. Raskina O, Barber JC, Nevo E, Belyayev A (2008) Repetitive DNA and chromosomal rearrangements: speciation-related events in plant genomes. Cytogenet Genome Res 120:351–357. https://doi.org/10.1159/000121084

  54. Schubert I, Wobus U (1985) In situ hybridization confirms jumping nucleolus organizing regions in Allium. Chromosoma 92:143–148. https://doi.org/10.1007/BF00328466

  55. Siljak-Yakovlev S, Peccenini S, Muratovic E, Zoldos V, Robin O, Vallès J (2003) Chromosomal differentiation and genome size in three European mountain Lilium species. Pl Syst Evol 236:165–173. https://doi.org/10.1007/s00606-002-0240-y

  56. Souza G, Costa L, Guignard MS, Van-Lume B, Pellicer J, Gagnon E, Leitch IJ, Lewis GP (2019) Do tropical plants have smaller genomes? Correlation between genome size and climatic variables in the Caesalpinia Group (Caesalpinioideae, Leguminosae). Perspect Pl Ecol Evol Syst 38:13–23. https://doi.org/10.1016/j.ppees.2019.03.002

  57. Thomas HM, Harper JA, Meredith MR, Morgan WG, Thomas ID, Timms E, King IP (1996) Comparison of ribosomal DNA sites in Lolium species by fluorescence in situ hybridization. Chromosome Res 4:486–490. https://doi.org/10.1007/BF02261775

  58. van den Berg C (2008) New combinations in the genus Cattleya (Orchidaceae). Neodiversity 3:3–12

  59. van den Berg C (2009) Phylogeny and systematics of Cattleya and Sophronitis. In: Sauleda RP, SandowLA (eds) The 19th world orchid conference. American Printing Arts, Miami, pp 319–323

  60. van den Berg C (2014) Reaching a compromise between conflicting nuclear and plastid phylogenetic trees: a new classification for the genus Cattleya (Epidendreae; Epidendroideae; Orchidaceae). Phytotaxa 186:75–86

  61. van den Berg C, Chase MW (2000) Nomenclatural notes on Laeliinae—I. Lindleyana 15:115–119

  62. van den Berg C, Chase MW (2001) Nomenclatural notes on Laeliinae—II additional combinations and notes. Lindleyana 16:109–112

  63. van den Berg C, Higgins WE, Dressler RL, Whitten WM, Soto Arenas A, Culham A, Chase MW (2000) A phylogenetic analysis of Laeliinae (Orchidaceae) based on sequence data from internal transcribed spacers (ITS) of Nuclear Ribosomal DNA. Lindleyana 15:96–114

  64. van den Berg C, Higgins WE, Dressler RL, Whitten WM, Sotoarenas MA, Chase MW (2009) A phylogenetic study of Laeliinae (Orchidaceae) based on combined nuclear and plastid DNA sequences. Ann Bot (Oxford) 104:417–430. https://doi.org/10.1093/aob/mcp101

  65. Van-Lume B, Esposito T, Diniz-Filho JAF, Gagnon E, Lewis GP, Souza G (2017) Heterochromatic and cytomolecular diversification in the Caesalpinia group (Leguminosae): relationships between phylogenetic and cytogeographical data. Perspect Pl Ecol Evol Syst 29:51–63. https://doi.org/10.1016/j.ppees.2017.11.004

  66. Zoldos V, Papes D, Cerbah M, Panaud O, Besenborfer V, Siljak Y (1999) Molecular-cytogenetic studies of ribosomal genes and heterochromatin reveal conserved genome organization among 11 Quercus species. Theor Appl Genet 99:969–977. https://doi.org/10.1007/s001220051404

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The authors wish to thank the Brazilian agencies Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) (Process Number 307879/2016-0) for financial assistance and INSA (Instituto Nacional do Semiárido) for providing transport during various new collection outings. G.S. and L.P.F received a productivity fellowship from CNPq (Processes Number PQ-310693/2018-7 and PQ-400209/2014-4, respectively).

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Correspondence to Gustavo Souza.

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Online Resource 1. List of Brazilian analyzed taxa of the subtribe Laeliinae, voucher, provenances and analyzed individuals per species.

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Querino, B.C., Ferraz, M.E., Mata-Sucre, Y. et al. Cytomolecular diversity of the subtribe Laeliinae (Epidendroidae, Orchidaceae) suggests no relationship between genome size and heterochromatin abundance. Plant Syst Evol 306, 19 (2020). https://doi.org/10.1007/s00606-020-01650-2

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  • Cytotaxonomy
  • Genome size
  • Laeliinae
  • rDNA sites