Advertisement

Plant Cell, Tissue and Organ Culture (PCTOC)

, Volume 136, Issue 1, pp 15–27 | Cite as

Structural and ultrastructural differences between field, micropropagated and acclimated leaves and stems of two Leucospermum cultivars (Proteaceae)

  • Emma SuárezEmail author
  • Carmen Alfayate
  • Juan Felipe Pérez-Francés
  • Juan Alberto Rodríguez-Pérez
Original Article
  • 141 Downloads

Abstract

The anatomy of field, in vitro and acclimatized shoots (leaves and stems) of two cultivars of Leucospermum (L. cordifolium ‘Flame Spike’ and L. ‘Tango’) was compared using light, scanning and transmission electron microscopy. Field plants showed several scleromorphic anatomical structures related to excess solar radiation such as: cuticle thickness, subepidermal collenchyma and sclerenchyma. Furthermore, a large quantity of phenolic deposits present in the cell lumen of various tissues is also a scleromorphic feature. The special conditions during in vitro culture result in plantlets with abnormal morphology and anatomy. These disorders are associated with the gaseous environment in the culture vessels, low irradiance in the incubation chamber and the addition of sucrose, nutrients and growth regulators to the culture medium. After transfer from in vitro to ex vitro conditions, substantial changes in leaf and stem anatomy were observed, above all in cuticle thickness, epidermal characteristics (stomatal and trichome index, and stomatal and pore size), differentiation of leaf mesophyll, chloroplast structure, and amount and localization of phenolic deposits. These changes allowed the plants to adapt to the new environmental conditions. The study of anatomical features of in vitro shoots facilitated adapting the acclimation protocol to predict which plantlet would survive the critical acclimation stage.

Keywords

Adaptation In vitro tissue culture Leucospermum Plant anatomy Proteaceae Tissue features 

Notes

Author contributions

ES and CA planned and designed the research, JAR-P contributed plant material, ES performed the experiments and collected data. ES, CA, JFP-F and JAR-P analysed the data, ES wrote the manuscript and CA supervised the writing. All the co-authors reviewed the manuscript before submission.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

References

  1. Apóstolo N, Brutti C, Llorente B (2005) Leaf anatomy of Cynara scolymus L. in successive micropropagation stages. In vitro Cell Dev Biol 41:307–313CrossRefGoogle Scholar
  2. Ben-Jaacov J, Jacobs G (1986) Establishing Protea, Leucospermum and Serruria in vitro. Acta Hortic 185:39–52CrossRefGoogle Scholar
  3. Bergmann DC (2004) Integrating signals in stomatal development. Curr Opin Plant Biol 7:26–32CrossRefGoogle Scholar
  4. Bisbis B, Kevers C, Crevecoeur M, Dommes J, Gaspar T (2003) Restart of lignification in micropropagated walnut shoots coincides with rooting induction. Biol Plant 47(1):1–5CrossRefGoogle Scholar
  5. Carpenter BJ, Hill RS, Jordan RS (2005) Leaf cuticular morphology links Platanaceae and Proteaceae. Int J Plant Sci 166(5):843–855CrossRefGoogle Scholar
  6. Conner LN, Conner AJ (1984) Comparative water loss from leaves of Solanum laciniatum plants cultured in vitro and in vivo. Plant Sci Lett 36:241–246CrossRefGoogle Scholar
  7. Dias Ferreira C, Dias JD, Canhot JM (2003) In vitro propagation of Leucadendron laureolum x L. salignum cv. Safari Sunset: ultrastructural and anatomical studies of regenerated plantlets. Acta Hortic 602:29–38CrossRefGoogle Scholar
  8. Dickinson WC (2000) Integrative plant anatomy. Academic Press, New YorkGoogle Scholar
  9. Ďurkovič J, Mišalová A (2009) Wood formation during ex vitro acclimatisation in micropropagated true service tree (Sorbus domestica L.). Plant Cell Tissue Organ Cult 96:343–348CrossRefGoogle Scholar
  10. Evert RF (2006) Esau’s plant anatomy: meristems, cells and tissues of the plant body: their structure, function, and development, 3rd edn. Wiley, HobokenCrossRefGoogle Scholar
  11. Gan Y, Zhou L, Shen ZJ, Shen ZX, Zhang YQ, Wang GX (2010) Stomatal clustering, a new marker for environmental perception and adaptation in terrestrial plants. Bot Stud 51:325–336Google Scholar
  12. George EF, Hall MA, De Klerk G (2008) Plant propagation by tissue culture, 3rd edn. Exegetics, BasingstokeGoogle Scholar
  13. Hatzilazarou SP, Syros TD, Yupsanis TA, Bosabalidis A, Economou AS (2006) Peroxidases, lignin and anatomy during in vitro and ex vitro rooting of gardenia (Gardenia jasminoides Ellis) microshoots. J Plant Physiol 163:827–836CrossRefGoogle Scholar
  14. Hazarika BN (2006) Morpho-physiological disorders in in vitro culture of plants. Sci Hortic 108:105–120CrossRefGoogle Scholar
  15. Jacobs G, Steenkamp JC (1976) Rooting stem cuttings of Leucospermum cordifolium and some of its hybrids under mist. Farming in South Africa, series: flowers, ornamental shrubs and trees, B.7. Department Agricultural Technical Services, PretoriaGoogle Scholar
  16. Jausoro V, Llorente BE, Apóstolo NM (2010) Structural differences between hyperhydric and normal in vitro shoots of Handroanthus impetiginosus (Mart. ex DC) Mattos (Bignoniaceae). Plant Cell Tissue Organ Cult 101:183–191CrossRefGoogle Scholar
  17. Jeffree CE (2006) The fine structure of the plant cuticle. In: Riederer M, Muller C (eds) Biology of the plant cuticle. Blackwell Publishing, Oxford, pp 11–125CrossRefGoogle Scholar
  18. Johansen DA (1940) Plant microtechniques. Mc Graw-Hill Book Co. Inc., New YorkGoogle Scholar
  19. Jordaan A, Theunissen JD (1992) Phenolic deposits and tannin in the leaves of five xerophytic species from southern Africa. Bot Bull Acad Sin 33:55–61Google Scholar
  20. Jordan GJ, Dillon RA, Weston PH (2005) Solar radiation as a factor in the evolution of scleromorphic leaf anatomy in Proteaceae. Am J Bot 92(5):789–796CrossRefGoogle Scholar
  21. Kunisaki JT (1989) In vitro propagation of Leucospermum hybrid, ‘Hawaii Gold’. HortScience 24(4):686–687Google Scholar
  22. Kunisaki JT (1990) Micropropagation of Leucospermum. Acta Hortic 264:45–48CrossRefGoogle Scholar
  23. Ladygin VG, Semenova GA (1993) The influence of iron deficiency on the composition of chlorophyll-protein complexes and the ultrastructure of pea chloroplasts. Russ J Plant Physiol 40:723–731Google Scholar
  24. Ladygin VG, Bondarev NI, Semenova GA, Smolov AA, Reshetnyak OV, Nosov AM (2008) Chloroplast ultrastructure, photosynthetic apparatus activities and production of steviol glycosides in Stevia rebaudiana in vivo and in vitro. Biol Plant 52(1):9–16CrossRefGoogle Scholar
  25. León F, Alfayate C, Vera Batista C, López A (2014) Phenolic compounds, antioxidant activity and ultrastructural study from Protea hybrid ‘Susara’. Ind Crop Prod 55:230–237CrossRefGoogle Scholar
  26. Louro RP, Santiago LJM, dos Santos AV, Machado RD (2003) Ultrastructure of Eucalyptus grandis x E. urophylla plants cultivated ex vitro in greenhouse and field conditions. Trees 17:11–22CrossRefGoogle Scholar
  27. Lucchesini M, Monteforti G, Mensuali-Sodi A, Serra G (2006) Leaf ultrastructure, photosynthetis rate and growth of myrtle plantlets under different in vitro culture conditions. Biol Plant 50(2):161–168CrossRefGoogle Scholar
  28. Majada JP, Sierra MI, Sánchez-Tamés R (2001) Air exchange rate affects the in vitro developed leaf cuticle of carnation. Sci Hortic 87:121–130CrossRefGoogle Scholar
  29. Majada JP, Fal MA, Tadeo F, Sánchez-Tamés R (2002) Effects of natural ventilation of leaf ultrastructure of Dianthus caryophyllus L. cultured in vitro. In Vitro Cell Dev Biol-Plant 38:272–278CrossRefGoogle Scholar
  30. Matthews LJ (2002) The protea book. A guide to cultivated Proteaceae. Canterbury University Press, New ZealandGoogle Scholar
  31. Murashige T, Skoog F (1962) A revised medium for rapid for rapid growth and bioassays with tobacco tissue culture. Physiol Plant 15:473–497CrossRefGoogle Scholar
  32. Oliveira M, Leandro MJ, Figueiredo E (2012) Factors affecting the success of the rooting process in some Proteaceae. Acta Hortic 937:817–824CrossRefGoogle Scholar
  33. Pérez-Francés JF, Raya-Tamayo V, Rodríguez-Pérez JA (2001) Micropropagation of Leucospermum ‘Sunrise’ (Proteaceae). Acta Hortic 545:161–165CrossRefGoogle Scholar
  34. Pospíšilová J, Tichá I, Kadleček P, Haisel D, Plzáková Š (1999) Acclimatization of micropropagated plants to ex vitro conditions. Biol Plant 42(4):481–497CrossRefGoogle Scholar
  35. Rodríguez-Pérez JA, Vera Batista MC, de León Hernández AM, Rodríguez Hernández I (2009) Vegetative cutting propagation of Protea Hybrid ‘Susara’. Eur J Hortic Sci 74(4):175–179Google Scholar
  36. Rodríguez-Pérez JA, Vera Batista MC, de León Hernández AM, Rodríguez Hernández I (2011) Use of proleptic shoots in the cutting propagation of Protea ‘Susara’ (Proteaceae). Span J Agric Res 9(2):565–569CrossRefGoogle Scholar
  37. Rodríguez-Pérez JA, Vera Batista MC, de León Hernández AM, Rodríguez Hernández I, Rodríguez Hernández H (2014) The effect of cutting position, wounding and IBA on the rooting of Leucospermum ‘Spider’. Acta Hortic 1031:77–81CrossRefGoogle Scholar
  38. Rourke JP (1972) Taxonomic studies of Leucospermum R. Br S Afr J Bot 8:194Google Scholar
  39. Rugge BA, Jacobbs G, Theron KI (1989) Factors affecting bud sprouting in multinodal stem segments of Leucospermum cv. Red Sunset in vitro. J Hortic Sci 65(1):55–58CrossRefGoogle Scholar
  40. Salatino A, Monteiro WR, Bomtempi N (1988) Histochemical localization of phenolic deposits in shoot apices of common species of Asteraceae. Ann Bot 61(5):557–559CrossRefGoogle Scholar
  41. Salisbury EJ (1929) On the causes and ecological significance of stomatal frequency, with special reference to the woodland flora. Philos Trans R Soc Lond B 216:1–65CrossRefGoogle Scholar
  42. Serna L, Fenoll C (1997) Tracing the ontogeny of stomatal clusters in Arabidopsis with molecular markers. Plant J 12(4):747–755CrossRefGoogle Scholar
  43. Serna L, Fenoll C (2000) Stomatal development in Arabidopsis: how to make a functional pattern. Trends Plant Sci 5:458–460CrossRefGoogle Scholar
  44. Serna L, Fenoll C (2002) Reinforcing the idea of signalling in the stomatal pathway. Trends Genet 18:597–600CrossRefGoogle Scholar
  45. Serret MD, Trillas MI (2000) Effects of light and sucrose levels on the anatomy, ultrastructure, and photosynthesis of Gardenia jasminoides Ellis leaflets cultured in vitro. Int J Plant Sci 161(2):281–289CrossRefGoogle Scholar
  46. Skelton PR, Midgley JJ, Nyaga JM, Johnson SD, Cramer MD (2012) Is leaf pubescence of Cape Proteaceae a xeromorphic or radiation-protective trait? Aust J Bot 60:104–113CrossRefGoogle Scholar
  47. Spurr AR (1969) A low-viscosity epoxy resin embedding medium for electron microscopy. J Ultrastruct Res 26(1):31–43CrossRefGoogle Scholar
  48. Stefanova M, Koleva D, Ganeva T (2013) Effect of plant growth regulators on chloroplast ultrastructure in Lamium album plantlets. Bul J Agri Sci 19(6):1208–1212Google Scholar
  49. Stoyanova-Koleva D, Stefanova M, Zhiponova M, Kapchina-Toteva V (2012) Effect of N6-benzyladenine and índole-3-butyric acid on photosynthetic apparatus of Orthosiphon stamineus plants grown in vitro. Biol Plant 56(4):607–612CrossRefGoogle Scholar
  50. Suárez E, Pérez-Francés JF, Rodríguez-Pérez JA (2010) Use of multinodal explants for micropropagation of Leucadendron ‘Safari Sunset’. Span J Agric Res 8(3):790–796CrossRefGoogle Scholar
  51. Suárez E, Alfayate C, Pérez-Francés JF, Rodríguez-Pérez JA (2018) Structural and ultrastructural variations in in vitro and ex vitro rooting of microcuttings from two micropropagated Leucospermum (Proteaceae). Sci Hortic 239:300–307CrossRefGoogle Scholar
  52. Sutter E (1984) Chemical composition of epicuticular wax in cabbage plants grown in vitro. Can J Bot 62(1):74–77CrossRefGoogle Scholar
  53. Tal E, Ben-Jaacov J, Watad AA (1992) Hardering and in vivo establishment of micropropagated Grevillea and Leucospermum. Acta Hortic 316:63–67CrossRefGoogle Scholar
  54. Tang M, Hu YX, Lin JX (2002) Developmental mechanism and distribution pattern of stomatal clusters in Begonia eltatifolia. Acta Bot Sin 44:24–33Google Scholar
  55. Theunissen JD, Jordaan A (1990) Histochemical localization of phenolic deposits in leaf blades of Eragrostis racemose. Ann Bot 65(2):633–636CrossRefGoogle Scholar
  56. Thillerot M, Choix F, Poupet A, Montarone M (2006) Micropropagation of Leucospermum ‘High Gold’ and three cultivars of Protea. Acta Hortic 716:17–24CrossRefGoogle Scholar
  57. van Staden J, Bornman CH (1976) Initiation and growth of Leucospermum cordifolium callus. J S Afr Bot 42(1):17–23Google Scholar
  58. Vera Batista MC (2016) Contribución al conocimiento de la propagación por estacas de algunas especies y cultivares de proteas. Thesis. Universidad de La Laguna, San Cristóbal de La Laguna, TenerifeGoogle Scholar
  59. Zhao X, Yang Y, Shen Z, Zhang H, Wang G, Gan Y (2006a) Stomatal clustering in Cinnamomum camphora. S Afr J Bot 72:565–569CrossRefGoogle Scholar
  60. Zhao Y, Zhou Y, Grout BWW (2006b) Variation in leaf structure of micropropagated rhubarb (Rheum rhaponticum L.) PC49. Plant Cell Tissue Organ Cult 85:115–121CrossRefGoogle Scholar
  61. Zobayed SMA, Armstrong J, Armstrong W (2001) Leaf anatomy of in vitro tobacco and cauliflower plantlets as affected by different types of ventilation. Plant Sci 161:537–548CrossRefGoogle Scholar
  62. Zucker WV (1983) Tannins: does structure determine function? An ecological perspective. Am Nat 12:335–365CrossRefGoogle Scholar

Copyright information

© Springer Nature B.V. 2018

Authors and Affiliations

  • Emma Suárez
    • 1
    Email author
  • Carmen Alfayate
    • 2
  • Juan Felipe Pérez-Francés
    • 1
  • Juan Alberto Rodríguez-Pérez
    • 3
  1. 1.Pharmacy Section, Department of Botany, Ecology and Plant Physiology, Faculty of Health SciencesUniversity of La LagunaTenerifeSpain
  2. 2.Biology Section, Department of Biochemistry, Microbiology, Cell Biology and Genetics, Faculty of SciencesUniversity of La LagunaTenerifeSpain
  3. 3.Agricultural Engineering Section, Department of Agricultural, Nautical, Civil and Marine Engineering, Higher Polytechnic School of EngineeringUniversity of La LagunaTenerifeSpain

Personalised recommendations