Assessing the effect of oxidative enzymes and stem anatomy on adventitious rooting of Olea europaea (L.) leafy cuttings

Nikoleta-Kleio Denaxa, Peter A. Roussos, Stavros N. Vemmos, Konstantinos Fasseas


Aim of study: To assess the role of polyphenol oxidase (PPO), peroxidase (POD) and indole-3-acetic acid oxidase (IAAox) during adventitious rooting (Ar) in semi-hardwood cuttings of the easy-to-root olive cv. ‘Arbequina’ and the difficult-to-root cv. ‘Kalamata’. Simultaneously, a histological study was carried out in both cultivars to investigate the tissue related with Ar development.

Area of study: The rooting experiments were carried out in ‘Kostelenos’ nurseries (Troizinia, Greece) and in Agricultural University of Athens.

Material and methods: Plant material to set up the experiment was collected from current year shoots from 15-year-old mother plants of ‘Arbequina’ and ‘Kalamata’ at three different seasons (summer, autumn and spring). The auxin indole-3-butyric acid (IBA) at 2000 mg L-1 was used as rooting inducer.

Main results: Analysis revealed that ‘Kalamata’ had significantly higher enzymatic activities before experiment onset and during Ar compared to ‘Arbequina’. Control cuttings of both cultivars exhibited increased enzymatic activities compared to IBA treated ones. IAAox was on average three times higher in ‘Kalamata’ than in ‘Arbequina’ and exhibited significant peaks during Ar. Similar peaks of POD and PPO activities were also detected. Histological analyses in ‘Kalamata’ revealed a continuous sheath of sclerenchyma ring and increased cortex thickness. Significant cell proliferation occurred in the phloem region in ‘Arbequina’ 15 days after planting and afterwards the root initials started developing in the secondary phloem from cambial cells.

Research highlights: The differences in enzymatic activities as well as in stem anatomy could partly justify the different rooting ability of both cultivars.


callus formation; enzymatic activity; IAA oxidase; olive ‘Kalamata’; peroxidase; root initials

Full Text:



Agullό-Antόn MA, Ferrández-Ayela A, Fernández-Garcia N, Nicolás C, Albacete A, Pérez-Alfocea F, Sánchez-Bravo J, Pérez-Pérez M, Acosta M, 2014. Early steps of adventitious rooting: morphology, hormonal profiling and carbohydrate turnover in carnation stem cuttings. Physiol Plant 150 (3): 446-462.

Amissah JN, Paolillo DJ Jr, Bassuk N, 2008. Adventitious root formation in stem cuttings of Quercus bicolor and Quercus macrocarpa and its relationship to stem anatomy. J Am Soc Hort Sci 133 (4): 479-486.

Aslmoshtaghi E, Shahsavar AR, 2016. Peroxidase, polyphenol oxidase and protein changes in olives during adventitious root formation. Trakia J Sci 2: 176-182.

Avidan B, Lavee S, 1978. Physiological aspects of the rooting ability of olive cultivars. Acta Hort 79: 93-101.

Ayoub S, Qrunfleh M, 2008. A study on some physiological and anatomical aspects of rooting 'Nabali' and 'Raseei' olive semi-hardwood stem cuttings. Acta Hort 773: 221-226.

Bakr EI, Selim H, Nour GM, Gabr MF, 1977. Developmental anatomy of adventitious roots on stem cuttings of 'Wetaken' olive cultivar. Egypt J Hort 4: 91-97.

Bansal MP, Nanda KK, 1981. IAA oxidase activity in relation to adventitious root formation on stem cuttings of some forest tree species. Experientia 37: 1273-1274.

Basak U, Das A, Das P, 2000. Rooting response in stem cuttings from five species of mangrove trees: effect of auxins and enzyme activities. Mar Biol 136: 185-189.

Bradford MM, 1976. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dyebinding. Anal Biochem 72: 248-254.

Caboni E, Lauri P, Tonelli MG, Iacovacci P, Damiano C, 1997. Biochemical and molecular factors affecting in vitro rooting ability in almond. In: Biology of root formation and development; Altman A, Waisel Y (eds.). pp: 117-124. Plenum Press, NY.

Cheniany M, Ebrahimzadeh H, Masoudi-Nejad A, Vahdati K, Leslie C, 2010. Effect of endogenous phenols and some antioxidant enzyme activities on rooting of Persian walnut (Juglans regia L.). Afr J Plant Sci 4: 479-487.

De Klerk G-J, 1996. Markers of adventitious root formation. Agronomie 16: 609-616.

Denaxa N-K, Vemmos SN, Roussos PA, Kostelenos G, 2010. The effect of IBA, NAA and carbohydrates on rooting capacity of leafy cuttings in three olive cultivars (Olea europaea L.). Acta Hort 924: 101-109.

Denaxa N-K, Roussos PA, Vemmos SN, 2014. The possible role of polyamines to the recalcitrance of 'Kalamata' olive leafy cuttings to root. J Plant Growth Regul 33 (3): 579-589.

Fabbri A, 1980. The effect of various anatomical characteristics on the rooting of cuttings in olive, cv. Frangivento. Riv Ortoflorofrutt Ital 64 (4): 325-335.

Flurkey HW, Jen JJ, 1978. Peroxidase and polyphenol oxidase activities in developing peaches. J Food Sci 43: 1826-1828.

Fontanazza G, 1993. Olivicoltura intensiva meccanizzata. Edagricole eds; Bologna, Italy. 103 pp.

Gaspar T, Kevers C, Hausman JF, 1997. Indissociable chief factors in the inductive phase of adventitious rooting. In: Biology of root formation and development; Altman A, Waisel Y (eds.). pp: 55-63. Plenum Press, NY.

Gonçalves JC, Diogo G, Amâncio S, 1998. In vitro propagation of chestnut (Castanea sativa x C. crenata): Effects of rooting treatments on plant survival, peroxidase activity and anatomical changes during adventitious root formation. Sci Hort 72: 265-275.

Güneş T, 2000. Peroxidase and IAA oxidase activities during rooting in cuttings of three poplar species. Turk J Bot 24: 97-101.

Hartmann HT, Kester DE, Davies FT, Geneve RL, 2001. Plant propagation principles and practices. 5th ed; Prentice-Hall, NJ.

Husen A, 2012. Changes of soluble sugars and enzymatic activities during adventitious rooting in cuttings of Grewia optiva as affected by age of donor plants and auxin treatments. Am J Plant Physiol 7 (1): 1-16.

Husen A, Pal M, 2007. Metabolic changes during adventitious root primordium development in Tectona grandis Linn. f. (teak) cuttings as affected by age donor plants and auxin (IBA and NAA) treatment. New Forest 33: 309-323.

Jasik J, De Klerk GJ, 1997. Anatomical and ultrastructural examination of adventitious root formation in stem slices of apple. Biol Plantarum 39 (1): 79-90.

Kavrayan D, Aydemir T, 2001. Partial purification and characterization of polyphenoloxidase from peppermint (Mentha piperita). Food Chem 74: 147-154.

Konieczny R, Banas A, Surówka E, Michalec Z, Miszalski Z, Libik-Konieczny M, 2014. Pattern of antioxidant enzyme activities and hydrogen peroxide content during developmental stages of rhizogenesis from hypocotyl explants of Mesembryanthemum crystallinum L. Plant Cell Rep 33: 165-177.

Kose C, Erdal S, Kaya O, Atici O, 2011. Comparative evaluation of oxidative enzyme activities during adventitious rooting in the cuttings of grapevine rootstocks. J Sci Food Agr 91: 738-741.

Liu ZH, Hsiao IC, Pan YW, 1996. Effect of naphthaleneacetic acid on endogenous indole-3-acetic acid, peroxidase and auxin oxidase in hypocotyl cuttings of soybean during root formation. Bot Bull Acad Sinica 37 (4): 247-253.

Ludwig-Müller T, 2003. Peroxidase isoenzymes as markers for the rooting ability of easy-to-root and difficult-to-root Grevillea species and cultivars of Protea obstusifolia (Proteaceae). In Vitro Cell Dev Biol Plant 39: 377-383.

Macedo E, Vieira C, Carrizo D, Porfirio S, Hegewald H, Arnholdt-Schmitt B, Calado M, Peixe A, 2013. Adventitious root formation in olive (Olea europaea L.) microshoots: anatomical evaluation and associated biochemical changes in peroxidase and polyphenol oxidase activities. J Hortic Sci Biotechnol 88: 53-59.

Mato M, Vieitez A, 1986. Changes in auxin protectors and IAA oxidases during the rooting of chestnut shoots in vitro. Physiol Plant 66: 491-494.

Metaxas D, Syros T, Yupsanis T, Economou A, 2004. Peroxidases during adventitious rooting in cuttings of Arbutus unedo and Taxus baccata as affected by plant genotype and growth regulator treatment. Plant Growth Regul 44: 257-266.

Meudt WJ, Gaines TP, 1967. Studies on the oxidation of indole-3-acetic acid by peroxidase enzymes. I. Colorimetric determination of indole-3-acetic acid oxidation products. Plant Physiol 42: 1395-1399.

Molassiotis A, Dimassi K, Diamantidis G, Therios I, 2004. Changes in peroxidases and catalase activity during in vitro rooting. Biol Plantarum 48: 1-5.

Naija S, Nadhra E, Najoua J, Saida A, Kevers C, 2008. Anatomical and biochemical changes during adventitious rooting of apple rootstocks MM 106 cultured in vitro. C R Biol 331: 518-525.

Porfirio S, Da Silva MDR, Cabrita MJ, Azadi P, Peixe A, 2016a. Reviewing current knowledge on olive (Olea europaea L.) adventitious root formation. Sci Hort 198: 207-226.

Porfirio S, Calado ML, Noceda C, Cabrita MJ, da Silva MG, Azadi P, Peixe A, 2016b. Tracking biochemical changes during adventitious root formation in olive (Olea europaea L.). Sci Hort 204: 41-53.

Qaddoury A, Amssa M, 2004. Endogenous phenolic contents, peroxidase and polyphenol oxidase activities in date palm (Phoenix dactylifera L.) off shoots related to rooting ability. Acta Physiol Plant 25 (4): 417-421.

Ríos D, Sanchez-Olate ME, Gea MA, Revilla MA, Rodríguez R, 1997. Rooting responses in relation with PO, PPO and IAA-o activities on walnut (Juglans regia l.) explants. Acta Hort 442: 241-250.

Rout GR, Samantaray S, Das P, 2000. In vitro rooting of Psoralea corylifolia Linn: Peroxidase activity as a marker. Plant Growth Regul 305: 215-219.

Sagee O, Raviv M, Medina Sh, Becker D, Cosse A, 1992. Involvement of rooting factors and free IAA in the rootability of citrus species stem cuttings. Sci Hort 51 (3-4): 187-195.

Salama MA, Zahran MA, Hassan MM, 1987. Comparing the rooting ability of some olive cultivars propagated by leafy cuttings under mist. Annals Agri Sci 32 (1): 577-590.

Sebastiani L, Tognetti R, 2004. Growing season and hydrogen peroxide effects on root induction and development Olea europaea L. (cvs 'Frantoio' and 'Gentile di Larino') cuttings. Sci Hort 100: 75-82.

Shinshi H, Noguchi M, 1975. Relationships between peroxidase, IAA oxidase and polyphenol oxidase. Phytochemistry 14: 1255-1258.

Srivastava OmP, van Huystee RB, 1977. IAA oxidase and polyphenol oxidase activities of peanut peroxidase isozymes. Phytochemistry 16: 1527-1530.

Tchinda ND, Messi HJCM, Fotso, Nzweundji G, Oumar D, Dongmo B, Sanonne, Agbor GA, Ndoumou DO, 2013. Biochemical aspects of single-node cuttings of Ricinodendron heudelotii (Baill.) in relation with rooting. Afr J Biotechnol 12 (10): 1049-1056.

Van Hoof P, Gaspar T, 1976. Peroxidase and iso peroxidase changes in relation to root initiation of Asparagus cultured in vitro. Sci Hort 4: 27-31.

Vatulescu AD, Fortunato AS, Cláudia Sá M, Amâncio S, Ricardo CPP, Jackson PA, 2004. Cloning and characterisation of a basic IAA oxidase associated with root induction in Vitis vinifera. Plant Physiol Biochem 42: 609-615.

Wiesman Z, Lavee S, 1995. Enhancement of stimulatory effects on rooting of olive cultivar stem cuttings. Sci Hort 62: 189-198.

Wiesman Z, Riov J, Estein E, 1988. Comparison of movement and metabolism of indole-3-acetic acid in mungbean cutting. Physiol Plant 74: 556-560.

Yilmaz H, Taşkin T, Otludil B, 2003. Polyphenol oxidase activity during rooting in cuttings of grape (Vitis vinifera L.) varieties. Turk J Bot 27: 495-498.

DOI: 10.5424/sjar/2019173-14486