Creeping bentgrass has been widely used as a cool-season turfgrass species for golf course greens, tees, and fairways in cool–humid and transitional climate zones due to its high density and tolerance to short mowing (Wang et al., 2012). It has excellent cold resistance and could tolerate temperatures as low as −35 °C (Gusta et al., 1980). However, creeping bentgrass has suffered winter-kill in some temperate regions such as in northern China. Winter-kill is a general term that is used to define turf loss during the winter and can be caused by a combination factors including desiccation, crown hydration, low temperatures, ice cover, and snow mold (Beard, 1973; Bhowmik et al., 2008). Winter-kill significantly damages the quality of sports surface, especially in golf greens, tees, and fairways.
In northern China such as Beijing area, the daily low temperatures are 8, 0, −6, −9, −6, and 0 °C and the monthly precipitations are 2.3, 0.8, 0.2, 0.3, 0.6, and 0.9 cm in October, November, December, January, February, and March, respectively, based on 30-year average. It appears that limited precipitation in late fall and winter may cause severe drought stress and could be a major factor causing winter-kill because the temperatures are well above killing temperature for creeping bentgrass in this area. Because of severe shortage of water resource and a high cost of irrigation water, golf course managers in this region are reluctant to water the grass in late fall through winter. Previous studies have shown that deficit irrigation could not only improve creeping bentgrass tolerance to stress but also significantly reduce cost of irrigation water (Ervin et al., 2009; Huang et al., 2014). However, little study has been reported on effect of deficit irrigation or mild drought stress before and during cold acclimation impacts on defensive metabolism and freezing tolerance of creeping bentgrass.
Creeping bentgrass undergoes cold acclimation, which is induced by a combination of reduced photoperiod and temperatures in late fall (Dionne et al., 2001a, 2001b; Hoffman et al., 2010). Cool-season turfgrasses require temperatures between 0 and 7 °C for 2–3 weeks to initiate the cold acclimation process. Second stage of cold acclimation is induced by subzero temperatures (Beard, 1973). Soil moisture status affects turfgrass cold acclimation process and thus cold tolerance (Beard, 1973). Previous studies have shown that mild drought stress can induce osmotic adjustment and antioxidant defenses in creeping bentgrass under normal temperature conditions (DaCosta and Huang, 2006, 2007; Fu and Dernoeden, 2009; McCann and Huang, 2007). Reduced soil water content may facilitate cold hardiness in st. augustinegrass [Stenotaphrum secundatum (Walt) Kuntze (Maier et al., 1994)] and arabidopsis [Arabidopsis thaliana (L.) Heynh (Mäntylä et al., 1995)]. This suggests preconditioning of the grass under mild drought stress before cold acclimation may improve plant defensive metabolism and freezing tolerance.
Environmental stress may damage plant cells by the accumulation of toxic reactive oxygen species (ROS), including O2−, H2O2, hydroxyl radicals (OH·), and singlet oxygen (1O2) (Apel and Hirt, 2004; Møller et al., 2007). The overproduction of ROS may damage lipids, proteins, and nuclei acids, resulting in plant senescence and death (Smirnoff, 1993). Plants have developed antioxidant defense mechanisms to eliminate ROS and prevent oxidative damage. Antioxidant enzymes, such as SOD (EC 184.108.40.206), CAT (EC 220.127.116.11), POD (EC 18.104.22.168), and ascorbate peroxidase [APX (EC 22.214.171.124)] protect plants against oxidative stress (Blokhina et al., 2003). SOD constitutes the first line of defense against ROS by dismutating the O2− to H2O2 (Apel and Hirt, 2004), which is finely regulated by CAT, POD, and APX (Wang et al., 2012). Various antioxidant metabolites and enzymes may work coordinately in suppressing ROS toxicity under stressful environments.
The level of antioxidant activity and gene expression is regulated in response to abiotic stresses such as drought and low temperature stresses (Jiang et al., 2010; Menezes-Benavente et al., 2004; Zhou et al., 2012). Catalase is induced by low temperature and an essential enzyme to protect mitochondria against chilling stress. The regulation in antioxidant enzyme activity and isozyme expression under adverse environments is closely related to plant tolerance to abiotic stresses (Allen, 1995; Mittler, 2002).
Plant may undergo osmotic adjustment in response to drought and cold stress (Hoffman et al., 2014; Huang et al., 2014). Accumulation of various osmoprotectants or compatible solutes, such as soluble proteins, simple sugars, and amino acids, in the cell during drought stress and cold acclimation improve plant tolerance to stresses (Ball et al., 2002; Blokhina et al., 2003; Hoffman et al., 2014; Shi et al., 2012).
Deficit or water-saving irrigation cannot only save water and reduce cost but also create mild drought stress which may improve defensive metabolism (Ervin et al., 2009; Huang et al., 2014). Severe drought stress occurred without irrigation in late fall in the region with limited precipitation may weaken defensive metabolism before and during cold acclimation and reduces winter survival of creeping bentgrass. Our hypothesis was that mild drought stress induced by deficit irrigation before and during cold acclimation would improve levels of osmoprotectants, antioxidant metabolism, and freezing tolerance when compared with severe drought stress (occurring in the region with limited precipitation and without irrigation in late fall and winter). In addition, mild drought stress treatment may improve osmorprectants, antioxidant metabolism, and freezing tolerance when compared with well-watered treatment.
The objectives of this study were to examine effects of three soil moisture levels (well-watered, mild drought stress, and severe drought stress) 14 d before and during 21 d of cold acclimation [2 °C (days 14 to 35)] on osmoprotectants, antioxidant metabolism, and freezing tolerance, and investigate if mild drought stress induced by deficit irrigation could improve plant freezing tolerance when compared with well-watered and severe drought stress regimes.
Abedi, T. & Pakniyat, H. 2010 Antioxidant enzyme changes in response to drought stress in ten cultivars of oilseed rape Czech J. Genet. Plant Breed. 46 27 34
Apel, K. & Hirt, H. 2004 Reactive oxygen species: Metabolism, oxidative stress, and signal transduction Annu. Rev. Plant Biol. 55 373 399
Ball, S., Qian, Y. & Stushnoff, C. 2002 Soluble carbohydrates in two buffalograss cultivars with contrasting freeze tolerance J. Amer. Soc. Hort. Sci. 127 45 49
Bandurska, H. & Jóźwiak, W. 2010 A comparison of the effects of drought on proline accumulation and peroxidases activity in leaves of Festuca rubra and Lolium perenne L Acta Societatis Botanicorum Poloniae 79 111 116
Banowetz, G.M., Dierksen, K.P., Azevedo, M.D. & Stout, R. 2004 Microplate quantification of plant leaf superoxide dismutases Anal. Biochem. 332 314 320
Beard, J.B. 1973 Turfgrass: Science and culture. Prentice Hall, Englewood Cliffs, NJ
Beauchamp, C. & Fridovich, I. 1971 Superoxide dismutase: Improved assays and an assay applicable to acrylamide gels Anal. Biochem. 44 276 287
Bhowmik, P.C., Shetty, K. & Sarkar, D. 2008 Cold-stress response of cool-season turfgrass: Antioxidant metabolism, p. 507–530. In: M. Pessarakli (ed.). Handbook of turfgrass management and physiology. CRC Press, New York, NY
Bian, S.M. & Jiang, Y.W. 2009 Reactive oxygen species, antioxidant 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
Black, C.A. 1965 Methods of soil analysis: Part I Physical and mineralogical properties. Amer. Soc. Agron., Madison, WI
Blokhina, O., Virolainen, E. & Fagerstedt, K.V. 2003 Antioxidants, oxidative damage and oxygen deprivation stress: A review Ann. Bot. 91 179 194
Bradford, M.M. 1976 A rapid and sensitive method for the quantitative determination of microgram quantities of protein utilizing the principle of protein–dye binding Anal. Biochem. 7 248 254
DaCosta, M. & Huang, B.R. 2006 Osmotic adjustment associated with variation in bentgrass tolerance to drought stress J. Amer. Soc. Hort. Sci. 131 338 344
DaCosta, M. & Huang, B.R. 2007 Changes in antioxidant enzyme activities and lipid peroxidation for bentgrass species in response to drought stress J. Amer. Soc. Hort. Sci. 132 319 326
Dionne, J., Castonguay, Y., Nadeau, P. & Desjardins, Y. 2001a Amino acid and protein changes during cold acclimation of green-type annual bluegrass (Poa annua L.) ecotypes Crop Sci. 41 1862 1870
Dionne, J., Castonguay, Y., Nadeau, P. & Desjardins, Y. 2001b Freezing tolerance and carbohydrate changes during cold acclimation of green-type annual bluegrass (Poa annua L.) ecotypes Crop Sci. 41 443 451
Ervin, E.H., LaBranche, A. & Zhang, X. 2009 Kentucky bluegrass and creeping bentgrass responses to foliar application of glycinebetaine at three ET replacement level Intl. Turfgrass Soc. Res. J. 11 755 764
Fielding, J.L. & Hall, J.L. 1978 A biochemical and cytochemical study of peroxidase activity in roots of Pisum sativum J. Expt. Bot. 29 969 981
Fu, J.M. & Dernoeden, P.H. 2009 Creeping bentgrass putting green turf responses to two summer irrigation practices: Quality, chlorophyll, canopy temperature, and thatch-mat Crop Sci. 49 1071 1078
Hoffman, L., DaCosta, M., Bertrand, A., Costonguay, Y. & Ebdon, S. 2014 Comparative assessment of metabolic responses to cold acclimation and deacclimation in annual bluegrass and creeping bentgrass Environ. Expt. Bot. 106 197 206
Hoffman, L., DaCosta, M., Ebdon, S. & Watkins, E. 2010 Physiological changes during cold acclimation of perennial ryegrass accessions differing in freeze tolerance Crop Sci. 50 1037 1047
Huang, B.R., DaCosta, M. & Jiang, J. 2014 Research advances in mechanisms of turfgrass tolerance to abiotic stresses: From physiology to molecular biology Crit. Rev. Plant Sci. 33 141 189
Jiang, Y.W., Eric, W., Liu, S.W., Yu, X.Q. & Luo, N. 2010 Antioxidative responses and candidate gene expression in prairie junegrass under drought stress J. Amer. Soc. Hort. Sci. 135 303 309
Jiang, M.Y. & Zhang, J.H. 2001 Effect of abscisic acid on active oxygen species, antioxidative defense system and oxidative damage in leaves of maize seedlings Plant Cell Physiol. 42 1265 1273
Khanna-Chopra, R. & Selote, D.S. 2007 Acclimation to drought stress generates oxidative stress tolerance in drought-resistant than -susceptible wheat cultivar under field conditions Environ. Expt. Bot. 60 276 283
Kochhar, S., Kochhar, V.K. & Khanduja, S.D. 1979 Changes in the pattern of isoperoxidases during maturation of grape berries cv Gulabi as affected by ethephon (2-chloroethyl) phosphonic acid Amer. J. Enol. Viticult. 30 275 277
Kuk, Y.I., Shin, J.S., Burgos, N.R., Hwang, T.E., Han, O., Cho, B.H., Jung, S.Y. & Guh, J.O. 2003 Antioxidative enzymes offer protection from chilling damage in rice plants Crop Sci. 43 2109 2117
Maier, F.P., Lang, N.S. & Fry, J.D. 1994 Freezing tolerance of three st. augustinegrass cultivars as affected by stolon carbohydrate and water content J. Amer. Soc. Hort. Sci. 119 473 476
Mäntylä, E., Lång, V. & Palva, E.T. 1995 Role of abscisic acid in drought-induced freezing tolerance, cold acclimation, and accumulation of LT178 and RABI 8 proteins in A. thaliana Plant Physiol. 107 141 148
McCann, S.E. & Huang, B.R. 2007 Effects of trinexapac-ethyl in creeping bentgrass responses to combined drought and heat stress Crop Sci. 47 2121 2128
Menezes-Benavente, L., Teixeira, F.K., Kamei, C.L.A. & Margis-Pinheiro, M. 2004 Salt stress induces altered expression of genes encoding antioxidant enzymes in seedlings of a Brazilian indica rice (Oryza sativa L.) Plant Sci. 166 323 331
Merewitz, E.B., Gianfagna, T. & Huang, B.R. 2011 Protein accumulation on leaves and roots associated with improved drought tolerance in creeping bentgrass expressing an ipt gene for cytokinin synthesis J. Expt. Bot. 62 5311 5333
Møller, I.M., Jensen, P.E. & Hansson, A. 2007 Oxidative modifications to cellular components in plants Annu. Rev. Plant Biol. 58 459 481
Nakano, Y. & Asada, K. 1981 Hydrogen peroxide is scavenged by ascorbate-specific peroxidase in spinach chloroplasts Plant Cell Physiol. 22 867 880
Nemali, K.S., Montesano, F., Dove, S.K. & van Iersel, M.W. 2007 Calibration and performance of moisture sensor in soilless substrates: ECH2O and theta probes Sci. Hort. 112 227 234
Patton, A.J., Cunningham, S.M., Volenec, J.J. & Reicher, Z.J. 2007 Differences in freeze tolerance of zoysiagrasses: II. Carbohydrate and proline accumulation Crop Sci. 47 2170 2181
Prasad, T.K., Anderson, M.D., Martin, B.A. & Stewart, C.R. 1994 Evidence for chilling-induced oxidative stress in maize seedlings and a regulatory role for hydrogen peroxide Plant Cell 6 65 74
Sarkar, D., Bhowmik, P.C., Young-In-Kwon, K. & Shetty, K. 2009 Cold acclimation responses of three cool-season turfgrasses and the role of proline-associated pentose phosphate pathway J. Amer. Soc. Hort. Sci. 134 210 220
Selote, D.S. & Khanna-Chopra, R. 2006 Drought acclimation confers oxidative stress tolerance by inducing coordinated antioxidant defense at cellular and subcellular level in leaves of wheat seedlings Physiol. Plant. 127 494 506
Sergiev, I., Alexieva, V. & Karanov, E. 1997 Effect of spermine, atrazine and combination between them on some endogenous protective systems and stress markers in plant. Comptes Rendus de L Academie Bulgare des Sciences 51:121–124
Shi, H.T., Wang, Y.P., Cheng, Z.M., Ye, T.T. & Chan, Z.L. 2012 Analysis of natural variation in bermudagrass (Cynodon dactylon) reveals physiological responses underlying drought tolerance PLoS ONE 7 1 12
Srivalli, B., Sharma, G. & Khanna-Chopra, R. 2003 Antioxidative defense system in an upland rice cultivar subjected to increasing intensity of water stress followed by recovery Physiol. Plant. 119 503 512
Wang, K.H., Zhang, X.Z. & Ervin, E. 2012 Antioxidative responses in roots and shoots of creeping bentgrass under high temperature: Effects of nitrogen and cytokinin J. Plant Physiol. 169 492 500
Woodbury, W., Spencer, A.K. & Stahmann, M.A. 1971 An improved procedure using ferricyanide for detecting catalase isozymes Anal. Biochem. 44 301 305
Xu, L.X., Han, L.B. & Huang, B.R. 2011 Antioxidant enzyme activities and gene expression patterns in leaves of kentucky bluegrass in response to drought and post-drought recovery J. Amer. Soc. Hort. Sci. 136 247 255
Zhang, X.Z., Wang, K.H. & Ervin, E.H. 2008 Bermudagrass freezing tolerance associated with abscisic acid metabolism and dehydrin expression during cold acclimation J. Amer. Soc. Hort. Sci. 133 542 550
Zhou, J., Wang, J., Shi, K., Xia, X.J., Zhou, Y.H. & Yu, J.Q. 2012 Hydrogen peroxide is involved in the cold acclimation-induced chilling tolerance of tomato plants Plant Physiol. Biochem. 60 141 149
Zhou, S.P., Sauvé, R.J., Mmbaga, M.T. & Frenkel, C. 2005 Cold-induced antioxidant enzymes changes in Leucanthemum maximum ‘Silver Princess’ HortScience 40 546 548