sign in sign up
<<
Home , Abies sibirica

World Applied Sciences Journal 28 (7): 909-913, 2013 ISSN 1818-4952 © IDOSI Publications, 2013 DOI: 10.5829/idosi.wasj.2013.28.07.13834

Needle Anatomy of Mutational Witches' Brooms of Siberian

Mikhail Sergeevich Yamburov and Kseniya Gennadievna Titova

Siberian Botanical Garden of Tomsk State University, Tomsk, Russia

Abstract: Our research has shown that the mutational witches' brooms of Siberian fir, as compared with a normal part of the crown, have higher rates of many morphological and anatomical features of needles, such as needles width, thickness and dry weight, the areas of assimilation, transporting and resiniferous tissues. All these features indicate a more intensive metabolism and can suggest that the needles of witches' brooms can be more productive by accumulation of secondary metabolic products, which are used in the chemical and pharmaceutical industries. The perspectives of studied witches' brooms for the use in selective breeding are discussing now. These unique genotypes of Siberian fir can be used for creating dense branching and highly abundant cultivars, suitable for plantation grown.

Key words: Siberian fir Abies sibirica Mutational witches' brooms Mutational variability Needle anatomy

INTRODUCTION could be caused by different reasons: 1) the development of the parasitic -mistletoe (genus Viscum), 2) Siberian fir (Abies sibirica Ledeb., family ) is infection by parasitic agents-fungi, phytoplasmas, viruses an Asian species, widespread in Siberian dark [1]. [13-15], 3) spontaneous somatic mutation [16-22]. Siberian fir has a high economic value to the Siberian Nowadays, the term is also used in plant pathology region. Its non-resin ducts wood is widely used in the and distinguishes two types of witches' brooms: woodworking industry and Siberian fir oil is extracted from parasitical and mutational those are different in origin the needles of young branches, the components of which [23, 24]. are used in the chemical industry and medicine [2-4]. Parasitical witches' brooms are disease and have no Despite widespread in Siberia and great economic breeding values, they influenced by metabolic products importance, Siberian fir almost is not involved in the of various parasitical agents. In the genus Abies selective breeding in order to obtain the most productive parasitical witches' brooms formed during infection of forms with high content of substances for chemical and rust fungi-Melampsorella caryophyllacerum G. Schrot. medical industries. One of the main reasons for this is the Parasitical witches' brooms of Siberian fir are characterized low level of morphological polymorphism in Siberian fir by patchy distribution, painful sight and premature death [5-7], also low level of karyotype variability [8] and small of needles and oppression of reproductive function [24]. genetic differences between populations [9-12]. Mutational witches' brooms are formed by mutation A source of valuable features for selective breeding of meristematic cells in the buds. Because are of Siberian fir can be spontaneous somatic mutations modular organisms, the whole system of branching that induce marked morphological changes. One of these developed from such buds have mutational features. rare and little-known mutations is a mutation that causes The mutation leads to the formation of witches' brooms increased branching and the formation of so-called is extremely rare in wild populations of and has a high witches' brooms. value for selective breeding. The term "witch's broom" came from the Middle Previously conducted a comparative study of Ages. That time every fragment in the tree crown that parasitical and mutational witches' brooms shoot have dense branching was called so. Dense branching structure showed that they differ by the formation of

Corresponding Author: Yamburov, Siberian Botanical Garden of Tomsk State University, Lenina Prospect 36, 634050, Tomsk, Russia.

909 World Appl. Sci. J., 28 (7): 909-913, 2013 branching [24]. Formation of parasitical witch’s broom branching caused by development of fungi Melampsorella caryophyllacerum-aecium with spores formed only at the annual needles, which leads to disruption of anatomy and death of needles. Fungal metabolites affect on shoots organogenesis. Metabolites cause formation of numerous latent buds on shoots. Latent buds lay along the entire length of branch at the base of needles. Latent buds proliferate and form new annual shoots, needles that compensate deceased needles of last year. Unlike parasitical witches' brooms, branching of mutational witches' brooms formed only by laying an increasing number of lateral buds at the top of shoots. Needles of mutational witches' brooms as needles from normal part of the tree crown live for few years [24]. Mutational witche’s brooms possess valuable features for selective breeding. Currently, many dwarf and abundantly branching cultivars of were obtained from vegetative and seed progeny of mutational witches' Fig. 1: Witches' broom # 1 brooms [17-21]. Also based on the witches' brooms were obtained and studied cultivars of Siberian stone pine (Pinus sibirica Du Tour) which forms a numerous cones at a young age [25-27]. The study of mutational witches' brooms allows us to estimate the mutational variability of Siberian fir and understand the character of changes in morphological and anatomical features caused the emergence of mutations. The aim of the present investigation was to describe the morphological and anatomical features of mutational witches' brooms needles and to evaluate their potential as initial material for selective breeding of Siberian fir.

MATERIALS AND METHODS

The study was carried out on material collected from two witches' brooms (Fig. 1, Fig. 2). Witches' broom # 1 aged about 9 years has been found in the mixed forest, near Kurlek village (Russia, West Siberia, Tomskaiay Oblast, 30 km south from Tomsk, 80 m above sea level). Fig. 2: Witches' broom # 2 Witches broom # 2 aged about 6 years was discovered during an expedition in the dark needle- taiga of 30 units were fixed and stored in 70% ethanol. The number Kuznetsk Alatau Mounties (Russia, South-Central Siberia, of stomata counted on a binocular under 4x magnification Kemerovo Oblast, Tiger-Tish range, near Vysokogornyy in the abaxial side of the needles. The dry weight of 30- creek, 800 m above sea level). needles samples in 10 replicates was determined and then The study was conducted on one-year-old needles of dry weight per one needle was calculated. witches' brooms. For comparison used normal needles Cross sections of the needles were performed with from the same part of the tree crown. Normal needles were a freezing rotary microtome MZ-2. Anatomical collected from shoots with same exposure that housed the observations and micrographs were performed with witches' brooms. Needles were collected in September, microscope Axio Lab.A1 (Carl Zeiss), using Axio Vision after the completion of growth. Needles in an amount of software for receiving, processing and image analysis.

910 World Appl. Sci. J., 28 (7): 909-913, 2013

RESULTS

The studied witches' brooms have characteristic features of the branching and normal development of the needles without traces of rust hyphae. It is the evidence of mutational origin of these witche’s brooms. A needle of witches' brooms for many anatomical and morphological features differ from the needles of the Fig. 3: Abies sibirica needle cross section. 1-epidermis, normal part of the crown (Table 1). 2-palisade mesophyll, 3-spongy mesophyll, 4-vein Needles of both witches' brooms have length less (central cylinder), 5-endodermis, 6-xylem, 7- than normal needles than 16% (witches' broom # 1) and phloem, 8-resin duct, 9-epithelial cells of resin than 7% (witches' broom # 2), but needles of witches' duct. brooms are wider than 3% and 10% and also thicker than 20% and 10%. The needle dry weight of witches' brooms Morphological features of needles were measured was more than 20% and 16%. The cross-sectional area of under 50x magnification, more detailed anatomical the needles of witches' brooms is more than 14% and 18%. features were measured under 400x magnification. In the case of the witches' broom # 1, reducing of needle Following parameters were measured: length, width and length also reduce surface area of needles at 11% thickness of the needles; areas of mesophyll, vein, compared with normal needles (despite the increase of phloem, xylem, resin ducts; thickness of epidermal cells, needles width and thickness). Needles of witches' broom length and width (thickness) of palisade and spongy # 2 are equal to normal by surface area of needles, mesophyll cells, width and thickness of endoderm cells, because the decrease of needles length compensated width and thickness of epithelial cells of resin ducts by significant increase of needles width and thickness. (Fig. 3.). The significance of differences was determined The stomatal density in compared variants has no using the Student's t-test. significant differences, as well epidermal cell thickness.

Table 1: Morphological and anatomical features of needles from witches' brooms and a normal part of the tree crown. Features Witches' broom # 1 Normal needles # 1 Witches' broom # 2 Normal needles # 2 Needle dry weight, mg 4.8±0.02** 4.0±0.01 3.6±0.01** 3.1±0.01 Needle length, mm 23.1±0.5** 27.6±0.7 19.4±0.4** 20.9±0.3 Needle width, µm 1334.7±4.5** 1298.9±5.0 1346.4±13.4** 1224.2±10.5 Needle thickness, µm 489.6±6.6** 408.1±5.3 431±50.5** 388.7±3.7 Needle surface area of, mm² 58.7±0.7** 65.7±2.4 47.0±1.1 46.6±0.9 Stomata density per 1 mm2 of needle surface area 43.0±0.8 42.3±0.7 45.8±0.8 43.9±0.6 Epidermis thickness, µm 24.3±0.3 24.3±0.6 20.0±0.4 19.9±0.7 Needle cross-section area × 10³, µm² 517.2±7.8** 453.1±5.5 470.3±10.1** 397.9±7.0 Mesophyll area × 10³, µm² 354.9±7.7** 320.3±6.3 331.5±8.4* 308.6±6.5 Palisade mesophyll cell length, µm 38±0.4** 41.6±0.5 37.6±0.5** 45.6±0.7 Palisade mesophyll cell thickness, µm 23.5±0.4 24±0.3 25.1±0.3 26.4±0.3 Spongy mesophyll cell length, µm 55.6±0.7** 60.9±1.0 64.1±1.0* 66.4±0.9 Spongy mesophyll cell thickness, µm 37.3±0.5** 39.9±0.7 37.3±0.6* 39.0±0.7 Vein area × 10³, µm² 68.6±2.0** 48.5±0.9 61.4±2.2** 42±1.4 Total phloem area × 103 µm² 4.9±0.1** 4.5±0.2 4.8±0.2** 4.5±0.1 Total xylem area × 103 µm² 6.1±0.2** 5.2±0.2 6.1±0.1** 4.3±0.2 Number of endoderm cells 27.4±0.3** 24.1±0.3 26.7±0.3** 21.8±0.3 Endoderm cell width, µm 44.7±1.0 47±0.7 44.3±0.7 42.6±0.7 Endoderm cell thickness, µm 29.5±0.8 32.3±0.5 32.3±0.6 31.7±0.7 Resin duct diameter, µm 146±1.9** 133.1±2.8 138±2.2** 128.2±2.7 Total resin ducts area × 10³, µm² 33.8±1.5** 28.2±1.0 30.2±0.8** 26.2±0.8 Number of resin duct epithelium cells 16.2±0.2** 14.2±0.2 15.6±0.2** 13.7±0.2 Resin duct epithelium cell width, µm 27.5±0.4** 30.5±0.5 27.9±0.4** 33.4±0.6 Resin duct epithelium cell thickness, µm 13.8±0.3 13.9±0.2 13.7±0.2 12.6±0.3 * Significant difference, p-value = 0.05; ** p-value = 0.01.

911 World Appl. Sci. J., 28 (7): 909-913, 2013

Assimilation tissue of the needles is composed of secondary metabolic products, which are used in the palisade and spongy mesophyll. Spongy mesophyll cells chemical and pharmaceutical industries. In the future, it are larger than palisade mesophyll cells in all variants. will be necessary to study the qualitative composition of Mesophyll area in the needles of witches' brooms more fir oil extracted from witches' brooms. Siberian fir oil is rich than 7% and 11%. The length/thickness ratio of palisade in triterpene acids [28]. Triterpenoids are used in the cells in the mesophyll of witches' brooms is-1.6, while in chemical industry and have a wide range of biological the normal part of the crown is 1.8. Palisade mesophyll activities [29]. cells of witches' brooms have length less than 9% and Thus, we can assume that these unique genotypes 18%, but do not have difference in thickness. Spongy of Siberian fir can be used to create dense branching and mesophyll cells have the same length/width ratio in highly abundant cultivars, suitable for plantation grown. compared variants, but in the needles of witches' brooms Plantation grown of Siberian fir will reduce the pressure they have a length less than 4% and 9% and a thickness on natural populations, where the collectors of raw less than 4% and 7%, respectively. material often make complete defoliation of young Needles of witches' brooms have vein area more than Siberian fir . In addition, dense branching cultivars 41% and 46%. Transporting system is represented by two derived from witches' brooms will be tested in urban open collateral bundles with phloem on abaxial side of greenery. More intensive metabolism of needles may bundle and xylem on adaxial side. The total phloem area in promote a higher resistance in an urban environment. needles of witches' brooms is a little more than in normal needles-at 9% and 7% and the total xylem area is much ACKNOWLEDGEMENTS more than 17% and 42%. Endoderm performing the function of the border between the inner tissue and This work was supported by the Russian Foundation peripheral parts of the needles and providing selective for Basic Research, project No. 12-04-31698 and partially transport of substances to the conducting tissues has no by the Ministry of Education and Science of the Russian statistically significant differences between compared Federation. State registration number of the theme: variants by the cells size, but the number of cells which 01201256296. cover vein in the needles of witches' brooms is more than 14% and 23%. REFERENCES The resin ducts in fir needles contain a mixture of terpenes and serve a protective function. The diameter of 1. Krylov, G.V., I.I. Maradudin, N.I. Miheev and the resin ducts in the needles of witches' brooms is more N.F. Kozakova, 1986. Fir, pp: 239. than 8% and 10% and the total area of resiniferous 2. Atlas of areas and resources of medicinal plants in tissue is more than 15% and 20%. This may indicate that USSR, 1976, pp: 340. the resin ducts of witches' brooms contain more secret. 3. Minaeva, V.G., 1991. Medicinal plants of Siberia, The number of epithelial cells in the resin duct is also pp: 432. more than 14%, but their width is less than 10% and 17% 4. Ciesla, W.M., 1998. Non-wood forest products from and do not have difference in thickness. conifers, pp: 124. 5. Mamaev, S.A., 1968. The problems and methods of CONCLUSION intraspecific of woody plants. Forms of variability. Inst Plant and Animal Ecology. UF AS Siberian fir in wild populations has low variability of USSR, 60: 3-43. needles features [5], as well as many other morphological 6. Kokorin, D.V. and L.I. Milyutin, 2003. Diversity of features. This makes it difficult to work on selective Siberian Fir (Abies sibirica Ledeb.) forms in southern breeding of this species [6]. Our study showed that regions of Central Sibiria. Lesovedenie, 4: 32-35. witches' brooms have higher rates of many morphological 7. Kokorin, D.V., 2004. Variability of Morphological and anatomical features of needles: needles width, Features of Siberian Fir in Southern Siberia, PhD thickness and dry weight, parameters of assimilation, thesis, Krasnoyarsk. transporting and resiniferous tissues. 8. Kvitko, O.V. and E.N. Muratova, 2010. Karyological All these features indicate more intensive metabolism Characteristics of Siberian Fir (Abies sibirica Ledeb.) of needles and suggest that the needles of witches' in Central Siberia. Cell and tissue biology, brooms can be more productive by accumulation of 4(2): 215-222.

912 World Appl. Sci. J., 28 (7): 909-913, 2013

9. Ekart, A.K. and A.Y. Larionova, 2006. Genetic 21. Vrgoc, P., 2002. Witches’ broom of Alepo pine (Pinus diversity and structure of the plain and mountain halepensis Mill.) and its use for new ornamentals. populations of Siberian fir (Abies sibirica Ledeb.). Acta Horticulturae: XX International Eucarpia Vestnik NESC FEB RAS, 4: 72-76. Symposium,572: 199-205. 10. Larionova, A.Y., A.K. Ekart and A.N. Kravchenko, 22. Yamburov, M.S., 2010. Mutational witches' brooms of 2007. Genetic diversity and population structure of some species in the family Pinaceae. PhD thesis, Siberian fir (Abies sibirica Ledeb.) in Middle Siberia. Tomsk State University, Tomsk. Eurasian J. For. Res., 10(2): 185-192. 23. Brown, C.L., H.E. Sommer and H. Wetzstein, 1994. 11. Semerikova, S.A. and V.L. Semerikov, 2006. Genetic Morphological and histological differences in variation and population differentiation in Siberian development of dwarf mutants of sexual and somatic fir Abies sibirica Ledeb. inferred from allozyme origin in diverse woody taxa. Trees: structure and markers. Russian Journal of Genetics, function, 9: 61-66. 42(6): 636-644. 24. Yamburov, M.S., 2009. Morphological features of 12. Semerikova, S.A. and V.L. Semerikov, 2011. Genetic mutational and parasitical "witches' brooms" of variability of Siberian fir Abies sibirica Ledeb. Siberian fir. Vestnik of Tomsk State University, inferred from AFLP markers. Russian Journal of 329: 246-250. Genetics, 47(2): 241-246. 25. Yamburov, M.S. and S.N. Goroshkevich, 2006. 13. Bos, L.A., 1960. Witches' broom virus disease of Witches` brooms in Siberian stone pine as somatic Vaccinium myrtillus in the Netherlands. T. Pl. Ziekten, mutations and initial genetic material for breeding of 66: 259-263. nut-bearing and ornamental cultivars. The breeding 14. Kaminska, M., H. Sliwa and A. Rudzinska-Langwald, and genetic resources of five-needle pines. 2001. Association of Phytoplasma with Stunting, Leaf Conference in Southern Carpathians. Romania. Necrosis and Witches' Broom Symptoms in Magnolia Valiug, pp: 26-27. Plants. J. Phytopathology, 149: 719-724. 26. Yamburov, M.S. and S.N. Goroshkevich, 2007. 15. Kuz'michev, E.P., E.S. Sokolova and E.G. Kulikova, Witches' brooms of Siberian stone pine as 2001. Common Fungai Diseases of Russian Forests, spontaneous somatic mutations: occurrence, pp: 137. properties and possible use in breeding programs. 16. Liese, I., 1933. Inheritance of witches' broom Conifers of boreal zone, 24(2-3): 317-324. formation in the jaw. Magazine. for. Forestry. and. 27. Yamburov, M.S., S.N. Goroshkevich and O.V. Jagdzcesen. Berlin, pp: 17. Hihlova, 2011. The yield of cones of "witches’- 17. Fordham, A.J., 1967. Dwarf conifers from witches- brooms" vegetative progeny of Siberian stone pine brooms. Arnoldia, 27(4-5): 29-50. (Pinus sibirica Du Tour). UIFRO conference 18. Johnson, A.G., S.S. Pauley and W.H. Cromell, 1968. “Breeding and genetic resources of five-needle Pine Dwarf segregates from Witches’ brooms. Proc. Pine. Tomsk, Russia, pp: 84-85. Int. Plant prop. Soc., 18: 265-270. 28. Raldugin, V.A., 2004. Triterpenoids of fir and highly 19. Waxman, S., 1975. Witches’ brooms sources of new efficient plant growth regulator based on them. and dwarf forms of Picea, Pinus and Tsuga species. Russian chemical journal, 48(3): 84-88. Acta Hort. Symposium on propagation in 29. Petrichenko, V.M., E.I. Molokhova and Arboriculture, 54: 25-32. T.V. Sukhinina, 2011. Chemical composition of

20. Waxman, S., 1987. Dwarf conifers from witches’ volatile components and fatty acids in a CO2 extract brooms. Comb. Proc. Intem. Plant Propagators Soc., and the of the Siberian fir. 36: 131-136. Pharmaceutical chemistry journal, 45(7): 412-414.

913