Impaginato 141 1. Introduction Bermudagrass is a warm-season, C4, perennial grass. It has short, grey-green blades with rough edges, stems of 1 to 30 cm in length and a deep root system that can penetrate 2 m into the ground; how- ever, most of the root mass is less than 60 cm deep (Xu et al., 2011). Among the many advantages of tur- fgrass areas are erosion and dust control, aquifer recharge and protection from pollutants, heat reduc- tion in urban environments, reduction of noise and pollution, and providing human health and aesthetic benefits (Stier et al., 2013). Water scarcity is an increasing challenge to the turfgrass industry and may result in irrigation restrictions being imposed without regard for damage to turfgrass (Beard and Kenna, 2008). For turf managers, thriving in an indus- try where turf quality is of utmost importance is diffi- cult when water is limiting. Therefore, researches investigating turfgrass resistance to drought stress have become increasingly important (Fry and Huang, 2004). Fu and Huang (2001) investigated the effects of drought stress on two cool-season turfgrasses and found that moderate drought stress had not effects on morphological and physiological characteristics, however in intensive drought stress, antioxidant enzyme activities, chlorophyll content, relative water content and shoot dry weight were decreased. In addition to limited amounts of water, turfgrasses are impacted by low-light environments. Shade is more problematic for warm-season turfgrasses to maintain quality given their higher light saturation point com- pared to cool-season turfgrasses (Fry and Huang, 2004). Turfgrasses perform poorly in reduced light environments due to high traffic rate, daily mowing, and reduced photosynthesis. In shade, increased dis- ease presence adversely affects cool-season turfgrass development, while morphological limitations, such Adv. Hort. Sci., 2016 30(3): 141-149 DOI: 10.13128/ahs-20250 Morpho-physiological alteration in common bermudagrass [Cynodon dactylon (L.) Pers.] subjected to limited irrigation and light condition N. Adamipour (*), H. Salehi, M. Khosh-khui Department of Horticulture Science, College of Agriculture, University of Shiraz, Shiraz, Iran. Key words: antioxidative enzymes, irrigation, photoperiod, turfgrass. Abstract: Bermudagrass (Cynodon spp.) is the most popular warm-season turfgrass used in warm climatic regions of the world due to its recuperative ability, high traffic tolerance, heat tolerance, and relative drought and salt tolerance. However, shade is a microenvironment in which bermudagrass performs poorly. In order to evaluate the interaction of photoperiod and irrigation on [Cynodon dactylon (L.) Pers. California Origin], a greenhouse experiment was conducted at the Research Greenhouse of the Department of Horticultural Sciences, College of Agriculture, Shiraz University, Shiraz, Iran. The experiment was conducted with four field capacity regimes (25%, 50%, 75% and 100%) and three light dura- tions (8, 12 and 16 h) in a completely randomized design factorial arrangements with four replications. Results showed that decreasing field capacity and photoperiod decreased fresh and dry weights shoot and root, chlorophyll and starch contents and superoxide dismutase, catalase and ascorbate peroxidase activities. Decreasing the field capacity and light duration increased proline content. Reducing sugars and peroxidase enzyme in leaves increased with decreasing field capacity. Shoot height and leaf area increased by shortening the photoperiod. In overall, results showed that, the increase in irrigation alleviates the destructive effects of reduced day lengths and vice versa. Further studies are needed to clarify more the interaction between irrigation and light treatments at structural and ultrastructural levels, in common bermudagrass. (*) Corresponding author: adamipournader@yahoo.com Received for publication 9 May 2016 Accepted for publication 27 July 2016 Copyright: © 2016 Author(s). This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. http://creativecommons.org/licenses/by/4.0/ http://creativecommons.org/licenses/by/4.0/ Adv. Hort. Sci., 2016 30(3): 141-149 142 as reduced lateral stem growth, inhibits warm-sea- son turfgrass development (Beard, 1972). Variations of shade responses among species and cultivars (Jiang et al., 2004; Trenholm and Nagata, 2005; Sladek et al., 2009) make it possible to select turf- grasses with superior shade tolerance. Identifying morphological characteristics that are associated with superior shade performance based on genetic variation would add value to germplasm screening for shade tolerant species and cultivars. Esmaili and Salehi (2012) noted in bermudagrass that were treat- ed with short photoperiod duration, verdure fresh and dry weight, shoot height, tiller density, leaf area and chlorophyll and relative water contents were decreased, however electrolyte leakage and proline content were increased. Although bermudagrass, the most widely grown C4 turfgrass on an international basis (Shearman, 2006), has been extensively stud- ied, many challenges and questions still remain when light is a limiting growth factor. The main objective of the present study was to investigate the effects of both irrigation interval and light duration on growth and quality of common bermudagrass. 2. Materials and Methods Plant material and experimental conditions This experiment was conducted at the Research Greenhouse of the Department of Horticultural Sciences, College of Agriculture, Shiraz University, Shiraz, Iran (52◦32’E and 29◦36’N, 1810 m asl). Seeds of common bermudagrass (Cynodon dactylon [L.] Pers. California Origin) were weighed and cultured in plastic pots with 19 cm in diameter and 25 cm in height, without drainage (0.25 g pot−1) filled with 4 kg clay-loam soil with permanent wilting point (PWP) of 19% and field capacity (FC) 29%. Watering was car- ried out daily prior to beginning of treatments. Plants were kept in a greenhouse with 31/25°C (day/night) temperature and 35% relative humidity for one m o n t h b e f o r e t h e b e g i n n i n g o f t r e a t m e n t s . Treatments were conducted at four irrigation levels (25%, 50%, 75% and 100% FC) and three photoperiod duration [8, 12 and 16 h as short day length (SDL), intermediate day length (IDL) and long day length (LDL)]. Watering was carried out daily before seed germination and after turf establishment. Then, the t u r v e s w e r e w a t e r e d e q u a l l y w h e n r e q u i r e d . Established turves were clipped from 3 cm above soil by a hand mower and were transferred to a covered frame which temperature, light (intensity and length) and relative humidity were controlled with digital sensors. The environmental condition of covered frame was 31°C, white and creamy fluorescent lamps one m above the pots with a constant light intensity of 3000 lux, and 35% relative humidity for applying simultaneous irrigation and photoperiod treatments. Pots were weighed daily and set to different irriga- tion treatments (25, 50, 75 and 100% FC), during the whole of experiment. After three months, plants were harvested in order to measure morphological and biochemical traits. Growth parameters Growth parameters including, shoot height (cm), leaf area (cm2) and fresh and dry weights of shoot and root (g) were measured. Dry weights were mea- sured when the materials dried at 60°C for 48 h. Chlorophyll content Chlorophyll content was measured according to the method of Saini et al. (2001) using the following formula: Chlorophyll (mg/g f.w.) = [20.2(OD 645 nm) + 8.02(OD 663 nm) × V/ (f.w.×1000)] Where: OD is optical density, V is the final solution volume in ml and f.w. is tissue fresh weight in mg. Proline content Proline was determined according to the method described by Bates et al. (1973). Using spectropho- tometer (Biowave II, England) at 520 nm wavelength, appropriate proline standards were included in calcu- lation of its content in samples. Total soluble sugars and starch analysis The total soluble sugars were measured using the method as previously described by Dubois et al. (1956). The total soluble sugar content of samples was measured at 490 nm of absorbance and glucose solution was used at different concentrations for standard curve drawing. The starch content was quantified using the Bradford method (McCready et al., 1950). The starch content was measured at absorbance of 630 nm and calculated using the stan- dard curve of glucose and multiplying it by 0.92. Antioxidant analysis Fresh samples were homogenized in extraction buffer (0.1 M phosphate buffer pH 6.8) with mortar and pestle on ice. The homogenate was then cen- trifuged at 12,000 g for 15 min at 4°C and the super- natant was used as the crude extract for the superox- Adamipour et al. - Morpho-physiological alteration in common bermudagrass subjected to limited irrigation and light condition 143 ide dismutase (SOD), guaiacol peroxidase (POD), ascorbate peroxidase (APX) and catalase (CAT). The SOD, POD, APX and CAT enzymes were estimated u s i n g t h e m e t h o d s p r e v i o u s l y d e s c r i b e d b y B e a u c h a m p a n d F r i d o v i c h ( 1 9 7 1 ) , C h a n c e a n d M a e h l y ( 1 9 9 5 ) , N a k a n o a n d A s a d a ( 1 9 8 1 ) a n d Dhindsa et al. (1981), respectively. Experimental design and data analysis This study was conducted in a completely ran- domized design with factorial arrangements and two factors: field capacity and photoperiod with four replicates. Data were analyzed using statistical soft- ware (SAS Software) and mean comparisons were performed using LSD test at 5% level. 3. Results and Discussion Results of analysis of variance (Tables 1 and 2) showed that photoperiod (except for soluble sugar) and irrigation had significantly influenced the mea- sured traits and also the interaction of photoperiod and irrigation had a significant effect on fresh and dry weights of shoot, proline content and the level of activity of superoxide dismutase. Shoot height and leaf area Shoot height and leaf area significantly declined by decreasing field capacity from 100% to 25% (Table 3). Shoot height and leaf area decreased (47.09% and 27.77%, respectively) at 25% FC compared to 100% FC (Table 3). Ryan (2011) reported that growth can be reduced through impairment of cell division and cell expansion which occurs at a lower water stress threshold rather than photosynthetic inhibition. Fu and Huang (2001) reported that shoot growth of both Table 1 - Analysis of variance of photoperiod, field capacity and interaction between photoperiod and field capacity measured traits Source of variability df Shoot height (cm) Leaf area (cm2) Shoot fresh weight (g) Shoot dry weight (g) Root fresh weight (g) Root dry weight (g) Sugars of shoot (mg g-1 d.w.) Photoperiod 2 38.45 ** 0.04** 212.24** 20.59** 36.59** 4.87** 5.77 NS Field capacity 3 314.82** 0.33** 141.40** 141.40** 1337.54** 324.86** 79732.82** Photoperiod * field capacity 6 0.0 NS 0.0 NS 1.25** 1.25** 0.0 NS 0.0 NS 0.0 NS Error 33 0.17 0.002 0.24 0.24 0.00 0.08 7.01 CV - 2.18 4.32 3.58 6.57 0.00 2.02 1.96 ** and NS significant at the 0.01 level and not significant respectively. Table 2 - Analysis of variance of photoperiod, field capacity and interaction between photoperiod and field capacity measured traits ** and NS significant at the 0.01 level and not significant respectively. Source of variability df Superoxide dismutase (Ug-1 FW) Catalase (Ug-1 FW) Peroxidase (Ug-1 FW) Ascorbate peroxidase (Ug-1 FW) Chlorophyll (mg g-1 FW) Proline (µmol g-1 FW) Starch content (mg g-1 DW) Photoperiod 2 4044.08** 97.06** 969.12** 28933.33** 0.59** 4.99** 3.33** Field capacity 3 55360.44** 234.71** 5248.15** 257973.85** 1.74** 7238.96** 105235.35** Photoperiod * Field capacity 6 2.52 NS 0.0 NS 0.0 NS 0.0 NS 0.00 NS 2.27** 0.00 NS Error 33 235,20 3.26 27.65 260.03 0.00 0.12 0.00 CV - 9.91 5.65 6.67 1.79 0.0 2.68 0.00 Variables Photo- period Field capacity (%) Mean 100% 75% 50% 25% Shoot length LDL 22.25 d* 21.27 e 16.67 h 11.05 k 17.81 C (cm) IDL 23.75 c 22.77 d 18.17 g 12.55 j 19.31 B SDL 25.35 a 24.37 b 19.77 f 14.15 i 20.91 A Mean 23.78 A 22.80 B 18.20 C 12.58 D Leaf area LDL 1.22 bc 1.22 bc 1.17 c 0.87 e 1.12 B (cm2) IDL 1.23 bc 1.23 bc 1.18 c 0.88 e 1.13 B SDL 1.32 a 1.32 a 1.27 ab 0.97 d 1.22 A Mean 1.26 A 1.25 A 1.20 B 0.91 C Shoot fresh LDL 20.30 a 20.26 a 16.16 b 14.06 c 17.69 A weight (g) IDL 16.32 b 16.26 b 12.81 d 9.13 e 13.63 B SDL 13.85 c 12.83 d 9.69 e 5.34 f 10.42 C Mean 16.82 A 16.45 A 12.88 B 9.51 C Shoot dry LDL 11.30 a 11.26 a 7.16 d 5.06 e 8.69 A weight (g) IDL 10.32 b 10.26 b 6.81 d 3.13 f 7.63 B SDL 9.85 b 8.83 c 5.69 e 1.34 g 6.42 C Mean 10.49 A 10.11 A 6.55 B 3.17 C Root fresh LDL 39.78 a 38.69 b 29.54 g 16.99 j 31.25 A weight (g) IDL 37.80 c 36.71 e 27.56 h 15.01 k 29.27 B SDL 36.81 d 35.72 f 26.57 i 14.02 l 28.28 C Mean 38.13 A 37.04 B 27.89 C 15.34 D Root dry LDL 19.78 a 18.69 b 13.46 d 8.54 f 15.12 A weight (g) IDL 19.69 a 18.60 b 13.37 d 8.45 f 15.03 A SDL 18.82 b 17.73 c 12.50 e 7.43 g 14.12 B Mean 19.43 A 18.34 B 13.11 C 8.14 D Proline content LDL 5.57 i* 7.06 g 14.35 f 23.25 c 12.56 C (mol g−1 f.w.) IDL 5.63 i 6.51 h 15.19 e 24.20 b 12.88 B SDL 5.66 i 6.89 gh 17.29 d 24.74 a 13.65 A Mean 5.62 D 6.82 C 15.61 B 24.07 A *In each variable, data followed by the same letters (small letters for inte- ractions and capital letters for means) are not significantly different using LSD at 5% level. LDL= long day length. IDL= intermediate day length SDL= short day length. Table 3 - Effect of field capacity and photoperiod and their interaction on shoot length, leaf area, shoot fresh and dry weight, root fresh and dry weight and chlorophyll content Adv. Hort. Sci., 2016 30(3): 141-149 144 kentucky bluegrass and tall fescue generally were not affected by surface soil drying but under full drying, shoot growth declined for both species. The reduced leaf area is a modification to avoid evopo-transpira- tion loss and to increase water use efficiency in grass- es which helps to tolerate water stress. Low leaf sur- face area would reduce transpiration rate also by lowering stomatal activity (Riaz et al., 2010). Turf shoot height showed considerable difference in LDL treatments compared to SDL treatments. Reducing photoperiod significantly increased the shoot height and leaf area (Table 3). Shoot height and leaf area increased significantly with shortening day length that it’s maximum and minimum decreased (14.82% and 8.19%, respectively) was observed at SLD com- pared to LDL (Table 3). Similar results have reported on bermudagrass (Tegg and Lane, 2004) and zoysia- grass (Qian and Engelke, 1999). Shoot fresh weight Reducing field capacity from 100% FC to 25% FC significantly decreased the shoot fresh weight to 43.46% at 25% FC compared to 100% FC (Table 3). Riaz et al. (2010) demonstrated that, water deficit conditions had a significant inhibitory effect on shoot fresh and dry weights of three bermudagrass culti- vars. The extended photoperiod (16 h) significantly increased fresh weight compared to shorter pho- t o p e r i o d s ( 1 2 h a n d 8 h ) . S h o o t f r e s h w e i g h t increased 41.09% under LDL compared to the SDL condition (Table 3). Sinclair et al. (2004) demonstrat- ed that the extended photoperiod increased biomass accumulation of four grasses (‘Pensacola’ bahiagrass, Paspalum notatum Flugge var. Saurde Parodi; ‘Tifton 85’ bermudagrass, Cynodon spp. L. Pers.; ‘Florakirk’ bermudagrass; and ‘Florona’ stargrass, Cynodon nlem- fuensis Vanderyst var. nlemfuensis) compared to short day condition. Interaction between field capacity and photoperiod resulted in the highest and lowest fresh weight in 100% FC-LDL and 25% FC-SDL treatments (Table 3). Shoot dry weight Different percentages of field capacity and pho- toperiod had significant effects on dry weight (Table 3). Reducing field capacity and photoperiod signifi- cantly decreased the dry weight. The shoot dry weight in 100% FC conditions decreased 69.78% compared to 25% FC condition (Table 3). Similar results have been reported on creeping bentgrass (Agrostis stolonifera L.), rough bluegrass (Poa trivialis L.), and perennial ryegrass (Lolium perenne L.) ( P e s s a r a k l i a n d K o p e c , 2 0 0 8 ) , b e r m u d a g r a s s (Cynodon dactylon L.) (Riaz et al., 2010). The highest and lowest dry weight was observed in 100% FC-LDL and 25% FC-SDL treatments, respectively (Table 3). Burton et al. (1988) stated that day length was highly correlated with yield of ‘Coastal’ bermudagrass, with yield reduction occurring in day lengths under 13 h. Therefore, photoperiod influenced dry matter pro- duction of forage grasses. Extended photoperiod throughout the cool-season in short-day length con- ditions substantially decreased forage yield (Sinclair et al., 1997, 2001, 2003). Root fresh weight R o o t f r e s h w e i g h t s i g n i f i c a n t l y d e c l i n e d b y decreasing field capacity from 100% to 25%. Root fresh weight decreased (59.76%) at 25% FC com- pared to 100% FC (Table 3). The impact of partially closing stomata limits CO2 availability and reduces photosynthesis, which is vital to produce and translo- cate carbohydrates to roots to explore deeper mois- ture (Huang, 2006). Huang and Gao (2000) found that severe leakage of organic solutes from roots in drying soil gives evidence that root death of tall fescue culti- vars during drought stress may correlate with root desiccation. There was a significant difference between LDL, IDL and SDL treatments and the high- est and lowest root fresh weights were obtained in LDL and SDL treatments, respectively. Root fresh weight decreased 9.50% at SDL compared to LDL (Table 3). This is in agreement with Wang et al. (2004) who reported that an increase in root growth is associated with extended light duration and is related to increase in internal cytokinin concentra- tion and its increased activity in root tips. Root dry weight As shown in Table 3, reduction in field capacity decreased root dry weight of plants. The highest and lowest root dry weights were observed in 100% FC and 25% FC treatments, respectively and in 25% FC decreased 58.10% compared to 100% FC. Pessarakli and Kopec (2008) demonstrated that, water deficit conditions showed a significant decrease in root dry weight of three turfgrass species. The highest and lowest root dry weight was obtained in LDL and SDL treatments, respectively (Table 3). Root dry weight decreased 6.61% at SDL compared to LDL (Table 3). Beard (1972) reviewed the morphological responses of turfgrasses under shade based on the research conducted before 1995, and found alterations such as: reduced tillering and shoot density, longer intern- odes with a reduced stem diameter, increased leaf length, decreased leaf width, thinner leaves, more Adamipour et al. - Morpho-physiological alteration in common bermudagrass subjected to limited irrigation and light condition 145 vertical leaf orientation, and fewer roots (McBee and Holt, 1966; Almodares, 1980; Dudeck and Peacock, 1992). A shift in allocation of dry matter occurs in response to shade, resulting in more dry matter par- titioning into shoots rather than roots (Allard et al., 1991; Dias-Filho, 2000). In response to lower irradi- ance, accelerated leaf elongation and decrease in partitioning to root dry matter are adaptive strate- gies to enhance light capture (Semchenko et al., 2012). Proline content Reducing field capacity and photoperiod signifi- cantly increased proline content in all plants. The highest amount of proline content was obtained in 25% FC and the lowest one was obtained in 100% FC treatment (Table 3). This is in agreement with (Etemadi et al., 2005) who demonstrated that the increase in drought increased proline content in bermudagrass (Cynodon dactylon L.). During drought stress, plants respond to different stresses with changes they create in their physiological features. Accumulation of soluble material in response to drought is a way to maintain turger. It seems that the accumulation of free proline in plants is the general reaction to the stress. However several other amino acids increase under drought and salinity stress. But the degree of changes is not comparable with proline accumulation (Gzik, 1996). In a comparative study between perennial ryegrass and red fescue for the amount of resistance to the drought, it was seen that the amount of proline in red fescue was more than perennial ryegrass (Bandurska and Jozwiak, 2010). The highest and lowest proline content was obtained in SDL and LDL treatments, respectively (Table 3). This is in agreement to the findings reported on the effects of decreased photoperiod on bermodagrass (Esmaili and Salehi, 2012). Interaction between field capacity and photoperiod resulted in the highest and lowest proline content in 25% FC-SDL and 100% FC- LDL treatments (Table 3). Chlorophyll content Field capacity and light durations had significant effects on leaf chlorophyll content. The highest and lowest chlorophyll content, were observed in 100% FC and 25% FC treatments, respectively (Table 4). Induction of drought has caused a reduction of elec- tron carrier in photosynthesis and a reduction in chlorophyll content which has been reported by (Zuily et al., 1990; Moran et al., 1994). Prolonged drought, heat, and the combined stresses could lead to loss of chlorophyll and lipid peroxidation, resulting in further turf quality decline (Jiang and Huang, 2001). Water is required to facilitate photosynthesis in plants. Low energy electrons are extracted from water and are energized through light energy cap- tured by chlorophyll. These energized electrons enable the production of NADPH and ATP which are then used to reduce CO2. CO2 is taken up from the atmosphere through stomata. Stomata are very sen- sitive to external environmental factors such as light, CO2, water status, and temperature (Hopkins and Hüner, 2004). The loss of chlorophyll by the plant in an intense stress can be associated with photo oxida- tion and consequently oxidative stress (Kato and Shimizu, 1985). Kaiser (1987) indicated that an irre- versible decrease in plant photosynthetic capacity occurs as RWC declines below 30%, leading to cell death from membrane damage in chloroplasts. Table 4 - Effect of field capacity and photoperiod and their interaction on proline, sugars and starch contents, a c t i v i t y o f S u p e r o x i d a s e d i s m u t a s e , C a t a l a s e , Peroxidase, and Ascorbate peroxidase enzymes Variables Photo- period Field capacity (%) Mean 100% 75% 50% 25% Chlorophyll LDL 1.81 a 1.78 b 1.48 e 0.99 j 1.52 A content IDL 1.74 c 1.71 d 1.41 h 0.92 k 1.44 B (mg Chl g−1f.w.) SDL 1.45 f 1.42 g 1.12 i 0.63 l 1.15 C Mean 1.67 A 1.64 B 1.33 C 0.85 D Sugars of shoot LDL 62.63 de 67.19 c 199.39 b 212.15 a 135.34 A (mg g−1d.w.) IDL 61.81 e 66.37 cd 198.22 b 211.33 a 134.52 A SDL 61.45 e 66.02 cd 198.22 b 210.98 a 134.17 A Mean 61.96 D 66.53 C 198.72 B 211.49 A Starch content LDL 225.30 a 224.10 d 91.50 g 41.30 j 145.50 A (mg g−1 d.w.) IDL 224.90 b 223.70 e 91.10 h 40.90 k 145.10 B SDL 224.30 c 223.20 f 90.60 i 40.40 l 144.60 C Mean 224.80 A 223.70 B 91.10 C 40.90 D Superoxide LDL 136.00 cd 148.50 c 266.00 a 116.00 def 166.62 A dismutase IDL 128.50 cde 143.50 c 261.50 a 109.50 ef 160.75 A (Ug-1 FW) SDL 106.00 fg 118.50 def 236.00 b 86.00 g 136.62 B Mean 123.50 C 136.83 B 254.50 A 103.83 D Catalase LDL 31.48 ef 34.45 cd 40.71 a 31.18 efg 34.45 A (Ug-1 f.w.) IDL 28.89 fgh 31.86 de 38.12 ab 28.59 ghi 31.86 B SDL 29.53 efg 29.53 efg 35.78 bc 26.25 i 29.53 C Mean 28.98 C 31.95 B 38.20 A 28.67 C Peroxidase LDL 68.40 fg 70.75 f 94.58 c 112.40 a 86.53 A (Ug-1 f.w.) IDL 60.80 hi 63.15 hi 86.99 d 104.81 b 78.94 B SDL 52.83 j 55.18 ji 79.02 e 96.84 c 70.97 C Mean 60.68 C 63.03 C 86.86 B 104.68 A Ascorbate LDL 869.64 de 879.64 d 1160.36 a 854.29 ef 940.98 A peroxidase IDL 829.64 gh 839.64 fg 1120.36 b 814.29 hi 900.98 B (Ug-1 f.w.) SDL 784.64 kj 794.64 ij 1075.36 c 769.29 k 855.98 C Mean 827.97 B 837.97 B 1118.69 A 812.61 C *In each variable, data followed by the same letters (small letters for inte- ractions and capital letters for means) are not significantly different using LSD at 5% level. LDL= long day length. IDL= intermediate day length SDL= short day length. Adv. Hort. Sci., 2016 30(3): 141-149 146 Detrimental effects on chloroplast biochemistry or chlorophyll fluorescence occur when RWC drops below 60% in tall fescue (Huang et al., 1998). Surface drying had no effects on chlorophyll content in ken- tucky bluegrass (Poa pratensis L.) and tall fescue (Festuca arundinacea Schreb.) while under full dry- ing, chlorophyll content decreased in both grasses (Fu and Huang, 2001). Our findings were in agree- ment are (Fu and Huang, 2001) who reported that amount of chlorophyll in bermudagrass under mod- erate stress is not reduced, but it will be reduced in the severe drought. Chlorophyll content decreased with decreasing day length and the highest and low- est ones were observed in LDL and SDL treatments, respectively (Table 4). Shorting photoperiod caused decrease in chlorophyll content. In a research, the resistances to low light stress in both bermudagrass and paspalum have been examined and it was con- cluded that resistance to low light stress in the pas- palum is more than bermudagrass (Jiang et al., 2004). Baldwin et al. (2008) reported that bermudagrass showed significant decrease in chlorophyll content in response to short day length condition. Total soluble sugars and starch content Regardless of photoperiod, decrease in field capacity significantly increased total soluble sugars in the shoot (Table 4). Starch content declined by decreasing field capacity from 100% to 25% (Table 4). Shoot starch content, were highest and lowest in 100% and 25% FC treatments, respectively (Table 4). On the other hand, total soluble sugars during the drought can increase making these compounds non- photosynthetic routes and growth stopping due to the destruction of in soluble sugars and their change to soluble sugars (Hissao, 1973). Although some researchers have suggested that the destruction of starch can also increase monosaccharaides (Düring, 1992). The researchers stated that an increase of amylase in water stress causes starch degradation and the conversion of this large molecule into smaller units (Movahhedi-Dehnavi et al., 2004). Different photoperiod had no significant effects on total solu- ble sugars (Table 4). Shoot starch content decreased by different light durations and the highest and low- est one was observed in LDL and SDL treatments, respectively (Table 4). Starch content decreased in response to shortening the photoperiod. Some researchers have reported that prolonging photope- riod increases carbohydrates (Hay and Pederson, 1986; Solhoug, 1991; Wang et al., 1998). Other researchers reported that the photoperiod had no effect on carbohydrates production (Sicher et al., 1982; Logendra and Janes, 1992). Antioxidant enzyme activities A P X , P O D , C A T a n d S O D e n z y m e s a c t i v i t i e s showed significant differences among field capacity and photoperiod treatments. The activities of APX were not significantly different between 100% FC and 75% FC treatments while were significantly increased in 50% FC treatment and minimum APX activity was observed at 25% FC treatment (Table 4). Bian and Jiang (2009) investigated the accumulation of reac- tive species of oxygen and antioxidants activity and t h e p a t t e r n o f g e n e e x p r e s s i o n o f a n t i o x i d a n t enzymes in the kentuchy bluegrass in the drought c o n d i t i o n . T h e y o b s e r v e d t h a t d r o u g h t s t r e s s increased the activity of APX and CAT and decreased SOD and they stated that antioxidant enzymes and their gene expression might be different or occur in the immune system of kentuchy bluegrass roots and l e a v e s . P O D e n z y m e a c t i v i t i e s i n c r e a s e d w i t h decrease in field capacity levels. Differences in leaf POD enzyme activities were not detected between 100% FC and 75% FC treatments. The maximum and minimum POD activity was obtained in 25% FC and 100% FC treatments, respectively (Table 4). In a research on drought tolerance of three cultivars of creeping bentgrass, it was observed that long-term drought stress reduced the activity of antioxidants such as POD and increased lipid peroxidation and the ‘Greenwich’ showed high resistance to drought (Da- Costa and Huang, 2007). CAT and SOD enzymes activ- ities significantly increased with decreasing field capacity from 100% to 50% then, declined in 25% FC treatment (Table 4). Shao et al. (2005) reported that in the of drought stress, the production amount of three enzymes, CAT, SOD and POD in resistant bermudagrass varieties have been significantly more than drought-sensitive ones. General declines in antioxidants, including CAT were reported in the response of three species of creeping bentgrass to drought stress. Moreover, they found that the species Agrostis canina L. was the most resistant species to drought (DaCosta and Huang, 2007). Liu et al. (2008) in a research, physiologically and morpho- logically investigated the five cultivars of kentuchy b l u e g r a s s u n d e r d r o u g h t a n d h e a t s t r e s s a n d observed that drought and heat stress simultaneous- ly reduces SOD enzyme in all cultivars and stated that an increase of SOD enzyme activity cannot inhibit stress and would only delay free radicals accumula- tion. Results of present study indicated that regard- Adamipour et al. - Morpho-physiological alteration in common bermudagrass subjected to limited irrigation and light condition 147 less of field capacity treatments, APX, POD, CAT and SOD enzymes activities significantly decreased in response to decreasing day length therefore, the m a x i m u m a n d m i n i m u m e n z y m e s a c t i v i t y w a s observed in LDL and SDL treatments (Table 4). Similar findings have been previously reported by (Burritt and Mackenzie, 2003) who stated that when the begonia plant is transferred from low light to bright light, CAT activity increases. Also, they stated that when the (Picea abies L.) seedlings are transferred from low light to high light, the activity of CAT enzyme decreases. Xu et al. (2010) investigated the effect of nitric oxide and sodium nitroprusside in tall fescue under high light stress and concluded that using sodium nitroprusside reduces enzyme activity of SOD, CAT and APX, but using nitric oxide increases the activity of mentioned enzymes. Jiang et al. (2005) demonstrated that, low light conditions showed a significant decrease in activity APX and CAT of bermudagrss and paspalum. Grace and Logan (1996) reported that the CAT enzyme activity varies depend- ing on light intensity. The CAT enzyme activity in Schefflera [Schefflera arboricola (Hayata) Merrill] and Vinca (Vinca major L.) plants did not change with a change in light intensity, but in Mahonia (Mahonia repens (Lindley) Don.), CAT enzyme activity increased with an increase of light intensity. Interaction between field capacity and photoperiod resulted in the highest and lowest SOD enzyme activities in 50% FC-LDL and 25% FC-SDL treatments (Table 4). 4. Conclusions The results proved that the reduction in photope- riod led to a progressive increase in shoot height and leaf area, however, the increase in irrigation inhibit- ed their progressive growths. Additionally, the reduc- tion in photoperiod caused a decrease in fresh and dry weight of root and shoot. However, the increase in irrigation led to alleviation of these negative effects during the day-time and thus increased the fresh and dry weight of root and shoot. Therefore, it appears as though the increased irrigation might have contributed to the enlargement and flexibility of cells, which, in turn helped increasing the dry and fresh weight of root and shoot. The reduced pho- toperiod led to a reduction in chlorophyll and starch contents and enzymes activities, and the increased irrigation compensated this reduction to some extent. This phenomenon might be, at least in part, explained by the fact that irrigation reduced ABA production, inhibited ROS production and thus inhib- ited the closure of stomata. In overall, the increase in irrigation caused the destructive effects of reduced photoperiod to diminish, and vice versa. It seems that the interaction of photoperiod and irrigation treatments has superior effects on alleviating of the symptoms of stressed plants, than their separate. Further studies are needed to clarify more the inter- action between irrigation and light treatments at structural and ultrastructural levels, in common bermudagrass. References ALLARD G., NELSON C.J., PALLARDY S.G., 1991 - Shade effects on growth of tall fescue: I. Leaf anatomy and dry matter partitioning. - Crop Sci., 31: 163-167. ALMODARES A., 1980 - The adaptation of Stenotaphrum secundatun (Walt.) Kuntze and Festuca arundinacea Schreb. to tree shade environments as affected by mowing heights. - Texas A&M University, College Station, Texas, USA. BALDWIN C.M., LIU H., McCARTY L.B., 2008 - Diversity of 42 bermudagrass cultivars in a reduced light environ- ment. II International Conference on Turfgrass Science a n d M a n a g e m e n t f o r S p o r t s F i e l d s . - A c t a Horticulturae, 783: 147-158. BANDURSKA H., JOZWIAK W., 2010 - A comparison of the effects of drought on proline accumulation and peroxi- dases activity in leaves of Festuca rubra L. and Lolium perenne L. - Plant. Physiol., 79: 111-116. BATES L.S., WALDREN R.P., TEARE I.D., 1973 - Rapid deter- mination of free proline for water stress studies. - Plant Soil., 39: 107-205. BEARD J.B., 1972 - Turfgrass: science and culture. - Prentice-Hall, Englewood Cliffs, NJ, USA, pp. 658. BEARD J.B., KENNA M.P., 2008 - Water quality and quanti- ty issues for turfgrasses in urban landscapes. - Council for Agriculture Science and Technology, Ames, IA, USA. BEAUCHAMP C., FRIDOVICH I., 1971 - Superoxide dismu- tases: improved assays and an assay predictable to acrylamide gels. - Anal. Biochem., 44: 276-287. BIAN S., JIANG Y., 2009 - Reactive oxygen species, antioxi- dant enzyme activities and gene expression patterns in leaves and roots of Kentucky bluegrass in response to drought stress and recovery. - Sci. Hort., 120: 264-270. BURRITT D.J., MACKENZIE S., 2003 - Antioxidant metabo- lism during acclimation of Begonia erythrophylla to high light levels. - Ann. Bot., 91: 783-794. BURTON G.W., HOOK J.E., BUTLER J.L., HELLWIG R.E., 1988 - Effect of temperature, daylength, and solar radiation on production of Coastal bermudagrass. - Agron. J., 80: Adv. Hort. Sci., 2016 30(3): 141-149 148 557-560. CHANCE B., MAEHLY A.C., 1955 - Assay of catalase and peroxidase. - Methods Enzymol., 2: 764-775. DACOSTA M., HUANG B., 2007 - Changes in antioxidant enzyme activities and lipid peroxidation for bentgrass species in response to drought stress. - J. Am. Soc. Hort. Sci., 132: 319-326. DHINDSA R.S., PLUMB-DHINDSA P., THORPE T.A., 1981 - Leaf senescence: correlated with increased levels of membrane permeability and lipid peroxidation, and decreased levels of superoxide dismutase and catalase. - J. Exp. Bot., 32: 93-101. DIAS-FILHO M.B., 2000 - Growth and biomass allocation of the C4 grasses Brachiaria brizantha and B. humidicola under shade. - Pesqui. Agropecu. Bras., 35: 2335-2341. DUBOIS M., GILLES K.A., HAMILTON J.K., REBERS P.A., SMITH F., 1956 - Colorimetric method for determina- tion of sugar and related substances. - J. Anal. Chem., 28: 350-356. DUDECK A.E., PEACOCK C.H., 1992 - Shade and turfgrass culture, pp. 269-284. - In: WADDINGTION D.V., R.N. C A R R O W , a n d R . C . S H E A R M A N ( e d s . ) T u r f g r a s s . American Society of Agronomy: Madison, Wisconsin, USA. DüRING H., 1992 - Evidence for osmotic adjustment to drought in grapevines (Vitis vinifera L.). - Vitis, 23: 1-10. ESMAILI S., SALEHI H., 2012 - Effects of temperature and photoperiod on postponing bermudagrass (Cynodon dactylon [L.] Pers.) turf dormancy. - J. Plant. Physiol., 169: 851-858. ETEMADI N., KHALIGHI A., RAZMJOO K.H., LESSANI H., ZAMANI Z., 2005 - Drought resistance of selected bermudagrass [Cynodon dactylon (L.) Pers.] accessions. - Int. J. Agric. Bio., 4: 612-615. FRY J., HUANG B., 2004 - Applied turfgrass science and physiology. - John Wiley & Sons, Inc, Hoboken, NJ, USA, pp. 320. FU J., HUANG B., 2001 - Involvement of antioxidants and lipid peroxidation in the adaptation of two cool-season grasses to localized drought stress. - Environ. Exp. Bot., 45: 105-114. GRACE S.C., LOGAN B.A., 1996 - Acclimation of foliar antioxidant systems to growth irradiance in three broadleaved evergreen species. - J. Plant. Physiol., 112: 1631-1640. GZIK A., 1996 - Accumulation of proline and pattern of α- amino acids in sugar beet plants in response to osmot- ic, water and salt stress. - Environ. Exp. Bot., 36: 29-38. HAY R.K.M., PEDERSON K., 1986 - Influence of long pho- toperiodon the growth of timothy (Phleum pretense L.) varieties from different latitudes in northern Europe. - Grass Forage Sci., 41(4): 311-317. HISSAO T., 1973 - Plant responses to water stress. - Annu. Rev. Plant. Biol., 24: 519-570. HOPKINS W.G., HüNER N.P.A., 2004 - Introduction to plant physiology. - John Wiley & Sons, New Jersey, USA, pp. 528. HUANG B., 2006 - Plant environment interactions.- CRC Press/Taylor and Francis, Boca Raton Fl, USA. HUANG B., FRY J., WANG B., 1998 - Water relations and canopy characteristics of tallfescue cultivars during and after drought stress. - HortScience, 33: 837-840. HUANG B., GAO H., 2000 - Root physiological characteris- tics associated with drought resistance in tall fescue cultivars. - Crop Sci., 40: 196-203. J I A N G Y . , C A R R O W R . N . , D U N C A N R . R . , 2 0 0 5 - Physiological acclimation of seashore paspalum and bermudagrass to low light. - Sci. Hort., 105: 101-115. JIANG Y., DUNCAN R.R., CARROW R.N., 2004 - Assessment of low light tolerance of seashore paspalum and bermudagrass. - Crop Sci., 44: 587-594. JIANG Y., HUANG B., 2001 - Physiological responses to heat stress alone or in combination with drought: a compari- son between tall fescue and perennial ryegrass. - HortScience, 36: 682-686. KAISER W.M., 1987 - Effects of water deficit on photosyn- thetic capacity. - Physiol. Plant., 71: 142-149. KATO M., SHIMIZU S., 1985 - Chlorophyll metabolism in higher plants. VI. Involvement of peroxidase in chloro- phyll degradation. - Plant. Cell Physiol., 26: 1291-1301. LIU J., XIE X., DU J., SUN J., BAI X., 2008 - Effects of simulta- neous drought and heat stress on kentucky bluegrass. - Sci. Hort., 115: 190-195. LOGENDRA S., JANES H.W., 1992 - Light duration effects on carbon partitioning and translocation in tomato. - Sci. Hort., 52: 19-25. McBEE G.G., HOLT E.C., 1966 - Shade tolerance studies on bermudagrass and other turfgrasses. - Agron. J., 58: 523-525. McCREADY R.M., GUGGLOZ J., SILVIERA V., OWENS H.S., 1950 - Determination of starch and amylose in vegeta- bles. - Anal. Chem., 22: 1156-1158. MORAN J.F., BECANA M., ITURBE-ORMAETXE I., FRECHILLA S., KLUCAS R.V., APARICIO-TEJO P., 1994 - Drought induces oxidative stress in pea plants. - Planta, 194: 346-352. MOVAHHEDI-DEHNAVI M., MODARRES A.M., SANAVI- SOROUSH-ZADE A., JALALI M., 2004 - Changes of pro- line, total soluble sugars, chlorophyll (SPAD) content and chlorophyll fluorescence in safflower varieties under drought stress and foliar application of zinc and manganese. - Biaban., 9: 93-110. NAKANO Y., ASADA K., 1981 - Hydrogen peroxide is scav- enged by ascorbate-specific peroxidase in spinach chloroplasts. - Plant Cell Physiol., 22: 867-880. PESSARAKLI M., KOPEC D.M., 2008 - Comparing growth responses of selected cool-season turfgrasses under salinity and drought stresses. - Acta Horticulturae, 783: 169-174. QIAN Y.L., ENGELKE M.C., 1999 - Influence of trinexapac- ethyl on diamond zoysiagrass in a shaded environment. - Crop Sci., 39: 202-208. R I A Z A . , Y O U N I S A . , H A M E E D M . , K I R A N S . , 2 0 1 0 - Morphological and biochemical responses of turfgrass- Adamipour et al. - Morpho-physiological alteration in common bermudagrass subjected to limited irrigation and light condition 149 es to water deficit conditions - Pakistan. J. Bot., 42: 3441-3448. RYAN M.G., 2011 - Tree responses to drought. - Tree Physiol., 31: 237-239. SAINI R.S., SHARME K.D., DHANKHAR O.P., KAUSHIK R.A., 2001 - Laboratory manual of analytical techniques in horticulture. - Agrobios, Jodhpur, India, pp. 49-50. SEMCHENKO M., LEPIK M., GÖTZENBERGER L., ZOBEL K., 2012 - Positive effect of shade on plant growth: amelio- ration of stress or active regulation of growth rate? - J. Ecol., 100: 459-466. SHAO H.B., LIANG Z.S., SHAO M.A., WANG B.C., 2005 - Changes of some physiological and biochemical indices for soil water deficits among 10 wheat genotypes at seedling stage. - Colloids, Surf. B., 42: 107-113. SHEARMAN R.C., 2006 - Fifty years of splendor in the grass. - Crop Sci., 46: 2218-2229. SICHER R.C., KREMER W.G., CHATTERTON N.J., 1982 - Effects of shortened day length upon translocation and starch accumulation by maize, wheat and pamgola grass leaves. - Can. J. Bot., 60: 1304-1309. SINCLAIR T.R., BENNET J.M., RAY J.D., 1997 - Enviromental limitation to potential forage production during the winter in Florida. - Soil. Crop Sci. Soc. Flo. Proc., 56: 58- 63. SINCLAIR T.R., MISLEVY P., RAY J.D., 2001 - Short photope- riod inhibits winter growth of subtropical grasses. - Planta, 213: 488-491. SINCLAIR T.R., RAY J.D., MISLEVY P., PREMAZZI L.M., 2003 - Growth of subtropical forage grasses under extended photoperiod during short day length months. - Crop Sci., 43: 618-623. SINCLAIR T.R., RAY J.D., PERMAZZI L.M., MISLEVY P., 2004 - Photosynthetic photon flux density influences grass responses to extended photoperiod. - Environ. Exp. Bot., 51: 69-74. SLADEK B.S., HENRY G.M., AULD D.L., 2009 - Evaluation of z o y s i a g r a s s g e n o t y p e s f o r s h a d e t o l e r a n c e . - HortScience, 44: 1447-1451. SOLHOUG K.A., 1991 - Effects of photoperiod and temper- ature on sugars and fructans in leaf blades, leaf sheaths and stem, and roots in relation to growth of Poa pratensis. - J. Plant. Physiol., 82: 171-178. STIER J.C., STEINKE K., ERVIN E.H., HIGGINSON F.R., MCMAUGH P.E., 2013 - Turfgrass and Issues, pp. 105- 145. - In: STIER J.C., P.B. HORGAN, and A.S. BONOS (eds.) Turfgrass: Biology, use, and management. American Society of Agronomy, Madison, WI, USA. TEGG R.S., LANE P.A., 2004 - A comparison of the perfor- mance and growth of a range of turfgrass species under shade. - Aust. J. Exp. Agric., 44: 353-358. TRENHOLM L.E., NAGATA R.T., 2005 - Shade tolerance of St. Augustine grass cultivars. - HortTechnology, 15: 267-271. WANG Z., YUAN Z., QUEBEDDEAUX B., 1998 - Photoperiod alters partitioning of newly-fixed 14C and reserve car- bon into sorbitol sucrose and starch in apple leaves, stems and roots. - Aust. J. Plant. Physiol., 25: 503-506. W A N G Z . H . , X U Q . , H U A N G B . , 2 0 0 4 - E n d o g e n o u s cytokinin level and growth responses to extended pho- toperiods for creeping bentgrass under heat stress. - Crop Sci., 44: 209-213. XU J., WANG Z., CHENG J.J., 2011 - Bermudagrass as feed- stock for biofuel production: a review. - Bioresour. Technol., 102: 7613-7620. XU Y.F., SUN X.L., JIN J.W., ZHOU H., 2010 - Protective roles of nitric oxide on antioxidant systems in tall fescue leaves under high-light stress. - Afr. J. Biol., 9: 300-306. ZUILY F.Y., VAZQUEZ T.A., VIEIRA D.J., 1990 - Effect of water deficit on cell permeability and on chloroplast integrity. - Bulletin de la Société Botanique de France, Actual. Bot., 137: 115-123.