Document Type : Original Article

Authors

1 M.Sc. in Agronomy, Department of Plant Production and Genetics, Faculty of Sciences and Agricultural Engineering, Campus of Agriculture and Natural Resources, Razi University, Kermanshah, Iran

2 Associate Prof., Department of Plant Production and Genetics, Faculty of Sciences and Agricultural Engineering, Campus of Agriculture and Natural Resources, Razi University, Kermanshah, Iran

3 Professor, Department of Water Engineering, Faculty of Agriculture, Campus of Agriculture and Natural Resources, Razi University, Kermanshah, Iran

Abstract

Introduction
Stevia is a perennial herb with sweet leaves, about 300 times sweeter than sucrose and today cultivated in many parts of the world. Drought stress and nitrogen deficiency are the most important environmental stresses that limit the production of this plant and have adverse effects on plant growth and development and other metabolic processes. The effect of drought stress on the plant depends on the type of plant, species, intensity, duration and growth stage of the plant. With the onset of drought decrease leaf water and stomatal conduction, and as it intensifies, loses permeability of the leaf cell membrane and damages. On the other hand, with the closure of the stomata due to lack of water is difficult and the entry of carbon dioxide into the leaves and in general, photosynthesis is reducing. Plant nutrition in drought conditions is also of special importance and proper nutrition in this situation can increase growth and stress tolerance. This element is the most important component of photosynthetic pigment proteins and has a great effect on leaf size and area.
Materials and methods
Therefore, in order to investigate the effect of drought stress and nitrogen fertilizer on the growth and yield of stevia, an experiment was conducted in greenhouse of Campus of Agricultural and Natural Resources, Razi University, Kermanshah, during 2016. Factorial experiment was conducted in a Completely Randomized Design (CRD) with three replications. Treatments included drought stress at three levels (no stress, mild stress and severe stress) and nitrogen fertilizer at five levels (0, 50, 100, 150 and 200 kg.ha-1) from urea source. Irrigation time for non-stress, mid and severe stress were 50, 65 and 80%, respectively, respectively from drainage of usable moisture in the soil. The size of the Cultivation space was 50 × 50 × 50 cm (length, width and height) and they were made of cement. The irrigation system was drip irrigation. the first and second harvest were on the May 23 and September 6, 2016, respectively. Traits were examined including yield and yield components, photosynthetic pigments, relative water content (RWC), water use efficiency (WUE), soluble sugars, proline, superoxide dismutase (SOD), peroxidase (POD) and catalase (CAT). Data were analyzed using SAS and MSTAT-C software and the means were compared using the LSD test at a probability level of 5%.
Results and discussion
The results showed that the drought stress and nitrogen had a significant effect on biomass, leaf dry matter, shoot dry matter, chlorophyll a and b, carotenoids, relative water content, water use efficiency, soluble sugars, proline and enzyme Superoxide dismutase, peroxidase and catalase. The highest amount of leaf dry matter in non-stressed treatments and applying 400 kg.ha-1 N in the first and second cutting were 20.4 and 17.1 g.pl-1, respectively, and the lowest amount of leaf dry matter in severe stress treatments and applying 400 kg N ha-1 the first and second cutting were 6.81 and 5.85 g.pl-1, respectively. With increasing amounts of nitrogen increased chlorophyll a, b, a + b and carotenoids in the first and second cutting. The amount of chlorophyll a, b, a + b and carotenoids was higher in the second cutting, but in carotenoids the values were equal in both cutting. Also, with increasing drought stress decreased the RWC in the first and second cutting and in the non-drought stress in the first and second cutting were 68.4% and 70.4%, and in the severe stress were 55.0% and 58.3%, respectively. Mean comparison of the interactions (stress × nitrogen) on WUE and proline showed that in conditions non-stress and using 200 kg ha-1 nitrogen had the highest WUE. WUE in the first and second cutting were 1.94 and 0.65 g.kg-1, respectively. In this experiment, the amount of superoxide dismutase in the first and second cutting increased with increasing drought stress, but on the contrary, the amount of peroxidase in the second cutting decreased with increasing drought stress. In addition, with the increase of N, the superoxide dismutase in the first and second cutting and also the amount of peroxidase increased in the second cutting.
Conclusion
In general, it can be concluded that stevia production in the second year has a good yield under non-drought stress. In addition, the use of nitrogen in conditions non-stress increased leaf yield. Leaf yield under mid stress with nitrogen application was not very significant. Under drought stress reduced leaf yield and was more severely with nitrogen utilization. Stevia is a summer plant with high water uses. It is recommended to pay attention to the amount of water available in summer and usually have a shorter irrigation cycle than other summer crops in the region, which is economically viable.

Keywords

Main Subjects

 Abdullateef, R.A., Osman, M., 2012. Studies on effects of pruning on vegetative traits in Stevia rebaudiana Bertoni (Compositae). International Journal of Biology. 4, 146-153. https://dx.doi.org/10.5539/ijb.v4n1p146
Abou-Arab, A.E., Abou-Arab, A.A., Abu-Salem, M.F., 2010. Physico-chemical assessment of natural sweeteners steviosides produced from Stevia rebaudiana Bertoni plant. African Journal of Food Science. 4, 269-281. https://dx.doi.org/10.5897/AJFS.9000226
Akbar Khalil, S., Zamir, R., Ahmad, N., 2014. Selection of suitable propagation method for consistent plantlets production in Stevia rebaudiana (Bertoni). Saudi Journal of Biological Sciences. 21, 566-573. https://dx.doi.org/10.1016/j.sjbs.2014.02.005
Amini, S., Ghobadi, C., Yamchi, A., 2015. Proline accumulation and osmotic stress: an overview of P5CS gene in plants. Journal of Plant Molecular Breeding. 3, 44-55. https://dx.doi.org/10.22058/jpmb.2015.17022
Anbazhagan, M., Kalpana, M., Rajendran, R., Natarajan, V., Dhanavel, D., 2010. In vitro production of Stevia rebaudiana Bertoni. Emirates Journal of Food and Agriculture. 216-222. https://dx.doi.org/10.9755/ejfa.v22i3.4891
Anjum, S.A., Farooq, M., Xie, X.Y., Liu, X.J., Ijaz, M.F., 2012. Antioxidant defense system and proline accumulation enables hot pepper to perform better under drought. Scientia Horticulturae. 140, 66-73. https://dx.doi.org/10.1016/j.scienta.2012.03.028
Aranjuelo, I., Molero, G., Erice, G., Christophe Avice, J., Nogues, S., 2011. Plant physiology and proteomics reveals the leaf response to drought in alfalfa (Medicago sativa L.). The Journal of Experimental Botany. 62, 111-123. https://dx.doi.org/10.1093/jxb/erq249
Armand, N., Amiri, H., Ismaili, A., 2016. Interaction of methanol spray and water‐deficit stress on photosynthesis and biochemical characteristics of Phaseolus vulgaris L. cv. Sadry. Photochemistry and Photobiology. 92, 102-110. https://dx.doi.org/10.1111/php.12548
Arnon, A.N., 1967. Method of extraction of chlorophyll in the plants. Agronomy Journal. 23, 112-121.
Atteya, A.M., 2003. Alteration of water relations and yield of corn genotypes in response to drought stress. Bulgarian Journal of Plant Physiology. 29, 63-76.
Barr, H.D., Weatherley, P.E., 1962. A re-examination of the relative turgidity technique for estimating water deficit in leaves. Australian Journal of Biological Sciences. 15, 413-428. https://dx.doi.org/10.1071/BI9620413
Bates, L.S., Waldern, R.P., Tear, I.D., 1973. Rapid determination of free proline for water- stress studies. Plant and Soil. 39, 205-207. https://dx.doi.org/10.1007/BF00018060
Beauchamp, C., Fridovich, I., 1971. Superoxide dismutase: improved assays and an assay applicable to acrylamide gels. Analytical Biochemistry. 44, 276-287. https://dx.doi.org/10.1016/0003-2697(71)90370-8
Bindi, M., Hacour, A., Vandermeiren, K., Craigon, J., Ojanperä, K., Selldén, G., Hogy, P., Fibbi, L., 2002. Chlorophyll concentration of potatoes grown under elevated carbon dioxide and/or ozone concentrations. European Journal of Agronomy. 17, 319-335. https://dx.doi.org/10.1016/S1161-0301(02)00069-2
Ceunen, S., Geuns, J.M., 2013. Steviol glycosides: Chemical diversity, metabolism, and function. Journal of Natural Products. 76, 1201-1228. https://dx.doi.org/10.1021/np400203b
Chaturvedula, V.S., Rhea, J., Milanowski, D., Mocek, U., Prakash, I., 2011. Two minor diterpene glycosides from the leaves of Stevia rebaudiana. Natural Product Communications. 6, 175-178. https://dx.doi.org/10.1177/1934578X1100600205  
Chrysargyris, A., Laoutari, S., Litskas, V.D., Stavrinides, M.C., Tzortzakis, N., 2016. Effects of water stress on lavender and sage biomass production, essential oil composition and biocidal properties against Tetranychus urticae (Koch). Scietia Horticulture. 213, 96-103. https://dx.doi.org/10.1016/j.scienta.2016.10.024
Daniels, L., Hanson, R., Philips, J., Gerhardt, P., Murray, R.G.E., Wood, W., Krieg, N.R., 1994. Chemical Analysis Methods for General and Molecular Bacteriology. Washington, DC: American Society of Microbiology. Chap. 22.
Durán, A.S., Rodríguez, N.M.P., Cordón, A.K., Record, C.J., 2013. Stevia (Stevia rebaudiana), non-caloric natural sweetener. Revista Chilena de Nutrición. 39, 203–206. https://dx.doi.org/10.4067/S0717-75182012000400015
El-Wahab, A., Mohamed, A., 2007. Effect of nitrogen and magnesium fertilization on the production of Trachyspermum ammi L. (Ajowan) plants under Sinai conditions. Journal of Applied Sciences Research. 3, 781-786.
Farooq, M., Wahid, A., Kobayashi, N., Fujita, D., Basra, S.M.A., 2009. Plant drought stress: effects, mechanisms and management. Agronomy for Sustainable Development. 29, 185–212. https://dx.doi.org/10.1007/978-90-481-2666-8_12
Gardner, F.P., Pearce R.B., Mitchell, R.L., 1984. Physiology of Crop Plants. 1st Edition. ‎Iowa State Press. 327p.
Ghaffari, G., Toorchi, M., Aharizad, S., Shakiba, M. R., 2011. Evaluation of traits related to water deficit stress in winter Rapeseed cultivars. Universal Journal of Environmental Research and Technology. 1, 338-350.
Ghorbanli, M., Gafarabad, M., Amirkian, T., Allahverdi Mamaghani, B., 2013. Investigation of proline, total protein, chlorophyll, ascorbate and dehydroascorbate changes under drought stress in Akria and Mobil tomato cultivars. Iranian Journal of Plant Physiology. 3, 651-658. [In Persian]
Gonzalez-Dugo, V., Durand, J. L., Gastal, F., 2010. Water deficit and nitrogen nutrition of crops. A review. Agronomy for Sustainable Development. 30, 529-544. https://dx.doi.org/10.1051/agro/2009059
Hajihashemi, S., Ehsanpour, A., 2013. Influence of exogenously applied paclobutrazol on some physiological traits and growth of Stevia rebaudiana under in vitro drought stress. Biologia. 68, 414-420. https://dx.doi.org/10.2478/s11756-013-0165-7
Hasheminasab, H., Assad, M.T., Aliakbari, A., Sahhafi, S.R., 2012. Influence of drought stress on oxidative damage and antioxidant defense systems in tolerant and susceptible wheat genotypes. Journal of Agricultural Science. 4, 20-30. [In Persian]
Hassegawa, R.H., Fonseca, H., Fancelli, A.L., da Silva, V.N., Schammass, E.A., Reis, T.A., Corrêa, B., 2008. Influence of macro-and micronutrient fertilization on fungal contamination and fumonisin production in corn grains. Food Control. 19, 36-43. https://dx.doi.org/10.1016/j.foodcont.2007.01.006
Huseynova, I.M., 2012. Photosynthetic characteristics and enzymatic antioxidant capacity of leaves from wheat cultivars exposed to drought. Biochimica et Biophysica Acta (BBA)-Bioenergetics. 1817, 1516-1523. https://dx.doi.org/10.1016/j.bbabio.2012.02.037
Kumar, R., Sharma, S., Ramesh, K., Singh, B., 2013. Effects of shade regimes and planting geometry on growth, yield and quality of the natural sweetener plant stevia (Stevia rebaudiana Bertoni) in north-western Himalaya. Archives of Agronomy and Soil Science. 59, 963-979. https://dx.doi.org/10.1080/03650340.2012.699676
Lawlor, D.W., Cornic, G., 2002. Photosynthetic carbon assimilation and associated metabolism in relation to water deficits in higher plants. Plant, Cell and Environment. 25, 275-294. https://dx.doi.org/10.1046/j.0016-8025.2001.00814.x
Muanda, F.N., Soulimani, R., Diop, B., Dicko, A., 2011. Study on chemical composition and biological activities of essential oil and extracts from Stevia rebaudiana Bertoni leaves. LWT - Food Science and Technology. 44, 1865– 1872. https://dx.doi.org/10.1016/j.lwt.2010.12.002
Naureen, G., Naqvi, F.N., 2010. Salt tolerance classification in wheat genotypes using reducing sugar accumulation and growth characteristics. Emirates Journal of Food and Agriculture. 22, 308-317. https://dx.doi.org/10.9755/ejfa.v22i4.4878
Odlare, M., Pell, M., Svensson, K., 2008. Changes in soil chemical and microbiological properties during 4 years of application of various organic residues. Waste Management. 28, 1246-1253. https://dx.doi.org/10.1016/j.wasman.2007.06.005 
Pal, P.K., Kumar, R., Guleria, V., Mahajan, M., Prasad, R., Pathania, V., Singh, R.D., 2015. Crop-ecology and nutritional variability influence growth and secondary metabolites of Stevia rebaudiana Bertoni. BMC Plant Biology. 15, 1-16. https://dx.doi.org/10.1186/s12870-015-0457-x
Parida, A.K., Dagaonkar, V.S., Phalak, M.S., Aurangabadkar, L.P., 2008. Differential responses of the enzymes involved in proline biosynthesis and degradation in drought tolerant and sensitive cotton genotypes during drought stress and recovery. Acta Physiologiae Plantarum. 30, 619-627. https://dx.doi.org/10.1007/s11738-008-0157-3
Pierret, A., Moran, C.J., Doussan, C., 2005. Conventional detection methodology is limiting our ability to understand the roles and functions of fine roots. New Phytologist. 166, 967-980. https://dx.doi.org/10.1111/j.1469-8137.2005.01389.x
Porporato, A., D’odorico, P., Laio, F., Rodriguez-Iturbe, I., 2003. Hydrologic controls on soil carbon and nitrogen cycles. I. Modeling scheme. Advances in Water Resources. 26, 45-58. https://dx.doi.org/10.1016/S0309-1708(02)00094-5
Ramesh, K., Singh, V., Megeji, N.W., 2006. Cultivation of Stevia [Stevia rebaudiana (Bert.) Bertoni]: A comprehensive review. Advances in Agronomy. 89, 137-177. https://dx.doi.org/10.1016/S0065-2113(05)89003-0
Reis, M., Coelho, L., Santos, G., Kienle, U., Beltrão, J., 2015. Yield response of stevia (Stevia rebaudiana Bertoni) to the salinity of irrigation water. Agricultural Water Management. 152, 217-221. https://dx.doi.org/10.1016/j.agwat.2015.01.017
Rivelli, A.R., De Maria, S., Pizza, S., Gherbin, P., 2010. Growth and physiological response of hydroponically-grown sunflower as affected by salinity and magnesium levels. Journal of Plant Nutrition. 33, 1307-1323. https://dx.doi.org/10.1080/01904167.2010.484092
Sairkar, P., Shukla, N.P., Mehrotra, N.N., 2009. Mass production of an economically important medicinal plant Stevia rebaudiana using in vitro propagation techniques. Journal of Medicinal Plants Research. 3, 266-270. https://dx.doi.org/10.5897/JMPR.9000862
Saxton, K.E., Rawls, W.J., 2006. Soil water characteristic estimates by texture and organic matter for hydrologic solutions. Soil Science Society of America Journal. 70, 1569–1578. https://dx.doi.org/10.2136/sssaj2005.0117
Setter, T.L., Flannigan, B.A., Melkonian, J., 2001. Loss of kernel set due to water deficit and shade in maize. Crop Science. 41, 1530-1540. https://dx.doi.org/10.2135/cropsci2001.4151530x 
Sinha, A.K., 1972. Colorimetric assay of catalase. Analytical biochemistry. 47, 389-394. https://dx.doi.org/10.1016/0003-2697(72)90132-7
Taiz, L. Zeiger, E., Moller, I.M., Murphy, A. 2015. Plant Physiology and Development. 6th Edition, Sinauer Associates, Sunderland, CT. p: 761.
Turtola, S., Manninen, A.M., Rikala, R., Kainulainen, P., 2003. Drought stress alters the concentration of wood terpenoids in Scots pine and Norway spruce seedlings. Journal of Chemical Ecology. 29, 1981-1995. https://dx.doi.org/10.1023/A:1025674116183
Vasilakoglou, I., Kalfountzos, D., Gougoulias, N., Reppas, C., 2016. Productivity of two stevia varieties under reduced irrigation and fertilization inputs. Archives of Agronomy and Soil Science. 62, 457-472. https://dx.doi.org/10.1080/03650340.2015.1060554
Woelwer-Rieck, U., Lankes, C., Wawrzun, A., Wüst, M., 2010. Improved HPLC method for the evaluation of the major steviol glycosides in leaves of Stevia rebaudiana. European Food Research and Technology. 231, 581-588. https://dx.doi.org/10.1007/s00217-010-1309-4
Wu, F., Bao, W., Li, F., Wu, N., 2008. Effects of drought stress and N supply on the growth, biomass partitioning and water-use efficiency of Sophora davidii seedlings. Environmental and Experimental Botany. 63, 248-255. https://dx.doi.org/10.1016/j.envexpbot.2007.11.002
Yadav, A.K., Singh, S., Dhyani, D., Ahuja, P.S., 2011. A review on the improvement of stevia [Stevia rebaudiana (Bertoni)]. Canadian Journal of Plant Science. 91, 1-27. https://dx.doi.org/10.4141/cjps10086
Yadav, R.S., Bhushan, C., 2001. Effect of moisture stress on growth and yield in rice genotypes. Indian Journal of Agricultural Research. 35, 104-107.
Yousfi, S., Márquez, A.J., Betti, M., Araus, J.L., Serret, M.D., 2016. Gene expression and physiological responses to salinity and water stress of contrasting durum wheat genotypes. Journal of Integrative Plant Biology. 58, 48-66. https://dx.doi.org/10.1111/jipb.12359