Pollen Ultrastructural Image Analysis among Ancient Native Olive Genotypes in the Central Eastern Tunisia

Authors
1 High Agronomic Institute, IRESA-University of Sousse, 4042 Chott-Mariem, Tunisia.
2 Olive Tree Institute, B.P. 40, 4061 Sousse, Tunisia.
3 Center of Electron Microscopy “Laura Bonzi”, ICCOM, National Research Council (CNR), Firenze, Italy.
4 Regional Research Center for Horticulture and Organic Agriculture (CRRHAB), B. P. 57, 4042 Chott Mariem, Sousse, Tunisia.
Abstract
The olive tree (Olea europaea L.) is considered as one of the oldest and the most important fruit crops of the Mediterranean basin, which is characterized by the existence of a considerable number of different olive cultivars. Therefore, the olive cultivar identification is crucial to safeguard the genetic patrimony of this species. Different morphological and molecular markers were used to discriminate among cultivars. The aim of the present work was to describe different pollen morphological and ultrastructural parameters (shape, size and exine pattern) as an additional tool for the identification of olive cultivars. Observations were carried on seventy centennial olive accessions grown in the Central Eastern part of Tunisia using Scanning Electron Microscopy (SEM) and Image analysis (ImageJ). Pollen were three-zonocolpate and elliptical-prolate or subprolate. Pollen morphological qualitative traits revealed specific differences among the studied genotypes including variation in whole grain shape and also exine pattern ornamentation as meshes profile and regularity and muri thickness. The quantitatively measured traits were significantly different among pollen from diverse genotypes. Polar and equatorial diameters varied from 21.80 to 29.88 µm and from 14.47 to 21.14 µm, respectively, while the pollen area ranged between 274.58 and 466.35 µm2. Frequency distributions of most measured pollen parameters depicted a normal distribution. The three principal components of the Principal Component Analysis (PCA) accounted for more than 97% of the total variation. The first Principal Component (PC1) was correlated to pollen size. The second (PC2) and the third (PC3) were correlated to exine texture and to pollen shape, respectively. Both morphometric features and exine pattern observations were potentially relevant tools to discriminate among the studied genotypes. Further combination between pollen ultrastructural analysis, morphological and molecular markers is fully desirable, in subsequent work, to improve both reliability and discriminative ability for cultivars classification.

Keywords


1. Al-Watban, A. A., Doaigey, A. R. and El-Zaidy, M. 2015. Pollen Morphology of Six Species of Subfamily Stachyoideae (Lamiaceae) in Saudi Arabia. Afr. J. Plant Sci., 9(5): 239-243.
2. Arzani, K., Nejatian, M. A. and Karimzadeh, G. 2005. Apricot (Prunus armeniaca) Pollen Morphological Characterization through Scanning Electron Microscopy, Using Multivariate Analysis. New Zeland J. Crop Hort. Sci., 33: 381–388.
3. Bartolini, G., Prevost, G., Messeri, C. and Carignani, G. 2005. Olive Germplasm: Cultivars and World-wide Collections. FAO/Plant Production and Protection Division, Rome. Available at: http://www.apps3.fao.org/wiews/olive/oliv.jsp [Accessed Dec. 28, 2008].
4. Bartolini, S., Minnocci, A. and Vitagliano, C. 1992. Morphological Studies on Pollen in Some Clones of Olive cv. ‘Leccino’. Agric. Med., 122:282-286.
5. Beris, F.del B., Sanchez, C. P., Gilabert, C. E. and Castillo, M. E. C. 1993. The Pollen Morphology of Citrus Lemon cv. ‘Verna’ from the Murcia Region, S.E. Spain. Anales-de-Biologia, 19:63-69.
6. Currie, A. J., Noiton, D. A., Lawes, G. S. and Bailey, D. 1997. Preliminary Results of Differentiating Apple Sports by Pollen Ultrastructure. Euphytica, 98(3): 155–161.
7. D’hallewin, G., Mulas, M., Nieddu, G. and Gitmta, F. 1990. Analysis of Pollen Morphology to Distinguish Olive Cultivars in a Germplasm Collection. Agric. Med., 120: 339-346.
8. Dezhong, T., Boaming, W., Gaixiu, D. and Xiaofung, F. 1995. Studies on the Pollen Morphology and Ultrastructure of Cultivated Varieties of Apricot (Armenica vulgaris Lam). Acta Hort., 403:144-144.
9. Erdtman, G. 1966. Pollen Morphology and Plant Taxonomy. I. In: “Angiosperms: An Introduction to Palinology”. Hefner Publ. Co., New York and London.
10. Erdtman, G. 1971. Pollen Morphology and Plant Taxonomy. 1. “Angyosperms”. Hefner, NewYork, USA, 553 PP.
11. Gomes, S., Martins-Lopes, P. and Guedes-Pinto, H. 2012. Olive Tree Genetic Resources Characterization through Molecular Markers. In: “Genetic Diversity in Plants”, (Ed.): Caliskan, M. ISBN: 978-953-51-0185-7, InTech.
12. Hannachi, H., Msallem, M., Ben Elhadj, S. and El Gazzah, M. 2007. Influence du Site Géographique sur les Potentialités Agronomiques et Technologiques de L'olivier (Olea europaea L.) en Tunisie. Comptes Rendus Biologies, 330: 135-142.
13. Heslop-Harrison, J. 1971. Wall Pattern Development in Angiosperm Microsporogenesis. Symp. Soc. Expt. Biol., 25: 277-300.
14. ImageJ/FIJI 1.46. 2012. Image J 1.46 Guide. Revised Version by: Ferreira, T. and Rasband, W. http://imagrj.nih.gov/ij/docs/guide.
15. Inceoglu, O., Pinar, M. and Oybak-Donmez, E. 2000. Pollen Morphology of Wild Vitis sylvestris Gmelin (Vitaceae). Turkish J. Bot., 24:147-150.
16. Javady, T. and Arzani, K. 2001. Pollen Morphology of Five Iranian Olive (Olea europaea L.) Cultivars. J. Agr. Sci. Tech., 3: 37-42.
17. Koubouris, G. C., Metzidakis, I. T. and Vasilakakis, M. D. 2012. Intraspecific Variation in Pollen Viability, Germination and Ultrastructure of Olea europaea L. Afr. J. Biotechnol., 11(70):13442-13446.
18. Lanza, B., Marsilio, V. and Martinelli, N. 1996. Olive Pollen Ultrastructure: Characterization of Exine Pattern through Image Analysis-Scanning Electron Microscopy (IA-SEM). Scien. Hort., 65: 283-294.
19. Lanza, B. and Marsilio, V. 1999. Ultrastructural Image Analysis and Biometric Studies on Pollen Grain to Distinguish Olive cvs. Acta Hort., 474:133-136.
20. Li, X., Yang, J., Li, X. G. and Yang, J. 2002. Application of Numerical Taxonomy of Pollen Morphology on Origination, Evolution and Classification of Pyrus L. in China. J. Fruit Sci., 19(3):145-148.
21. Li, X., Xiang, L., Wang, Y., Luo, J., Wu, Ch., Sun, Ch. and Xie, M. 2014. Genetic Diversity, Population Structure, Pollen Morphology and Cross-compatibility among Chinese Cymbidiums. Plant Breed., 133(1):145–152.
22. Loussert, L. and Brousse, G. 1978. Olive Tree: Mediterranean Agricultural Techniques of Olive Production. Publishing «Neuve and Larose» House, PP. 44-111.
23. Moore, P. D. and Webb, J. A. 1983. An Illustrated Guide to Pollen Analysis. Hodder and Staughton, London.
24. Mulas, M., Agabbio, M. and Nieddou, G. 1987. Etude au Microscope Electronique à Balayage du Pollen de L'amandier pour L'identification des Variétés. Proc. VII Grempa, Reus-Espagne, 17-19 Giugno, PP. 229-247.
25. Muzzalupo, I., Vendramin, G. G. and Chiappetta, A. 2014. Genetic Biodiversity of Italian Olives (Olea europaea) Germplasm Analyzed by SSR Markers. Sci. World J., Article ID 296590: 12 .
26. Nazeri Joneghani, V. 2008. Pollen Morphology of the Genus malus (Rosaceae). Iran. J. of Sci. Technol., Trans. A, 32(2):89-97.
27. Pacini, E. and Vosa, C. G. 1979. Scanning Electron Microscopy Analysis of Exine Patterns in Cultivars of Olive (Olea europaea L.). Ann. Bot., 44: 745-748.
28. Pacini, E., Ciampolini, F. and Crest, M. A. 1980. Technique for Observing the Same Anther or Pollen Grain Both by TEM and SEM. Grana, 19: 193-195.
29. Padula, G., Giordani, E., Bellini, E., Rosati, A., Pandolfi, S., Paoletti, A., Pannelli, G., Ripa, V., De Rose, F., Perri, E., Buccoliero, A. and Mennone, C. 2008. Field Evaluation of New Olive (Olea europaea L.) Selections and Effects of Genotype and Environment on Productivity and Fruit Characteristics. Adv. Hort. Sci., 22(2): 87-94.
30. Petruccelli, R., Giordano, C., Salvatici, MC., Capozzoli, L., Ciaccheri, L., Pazzini, M., Lain, O., Testolin, R., Cimato, A. 2014. Observation of Eight Ancient Olive Trees (Olea europaea L.) Growing in the Garden of Gethsemane. Comptes Rendus Biologies, 337(5):311-317.
31. Radičević, S., Nikolić, D., Cerović, R., Đorđević, M. 2013. In vitro Pollen Germination and Pollen Grain Morphology in Some Sweet Cherry (Prunus avium L.) Cultivars. Roman. Biotechnol. Lett., 18(3): 8341-8349.
32. Ribeiro, H., Cunha, M., Calado, L. and Abreu, I. 2012. Pollen Morphology and Quality of Twenty Olive (Olea europaea L.) Cultivars Grown in Portugal. In: “Proc. VIth IS on Olive Growing”, (Eds.): Sampaio, E. M. and Pinheiro, A. C. Acta Hort., 949: 259-264.
33. Roselli, G. 1979. Identificazione di Cultivar da Alcuni Caratteri del Polline. Rivista di Ortoflorofrutticoltura, 63:435-445.
34. Sorkheh, K., Vezvaei, A., Wirthensohn, M. G. and Martínez-Gómez, P. 2008. Pollen Ultrastructure Characterization in Californian and Australian Almond Cultivars. J. Am. Pomol. Soc., 62(4): 173-177.
35. Stanley, R. G. and Linskens, H. F. 1974. Pollen: Biology, Biochemistry Management. Springer-Verlag, Berlin and New York.
36. Talaie, A. R. and Imani, A. 1998. Morphology of Pollen Grains as an Index for Identification of Local Iranian Almond Varieties. Acta Hort., 470: 280-285.
37. Thakur, D. R. and Thakur, S. S. 1970. Pollen Morphology and Germination in Some Temperate Drupe Plants. J. Palynol., 6: 96-100.
38. Trentacoste, E. R. and Puertas, C. M. 2011. Preliminary Characterization and Morpho-agronomic Evaluation of the Olive Germplasm Collection of the Mendiza Province (Argentina). Euphytica, 177: 99-109.
39. Trujillo, I., Ojeda, M. A., Urdiroz, N. M., Potter, D., Barranco, D., Rallo, L and. Diez, C. M. 2014. Identification of the Worldwide Olive Germplasm Bank of Córdoba (Spain) Using SSR and Morphological Markers. Tree Gene. Genom., 10:141–155.
40. Varasteh, F. and Arzani, K. 2009. Classification of Some Iranian Pomegranate (Punica granatum) Cultivars by Pollen Morphology Using Scanning Electron Microscopy Hort. Environ. Biotechnol. 50(1):24-30.
41. Wrońska-Pilarek, D. 2011. Pollen Morphology of Polish Native Species of the Rosa Genus (Rosaceae) and Its Relation to Systematic. Acta Soc. Bot. Pol., 80(3):221-232.