Salt stress is increasingly becoming a major problem in turfgrass management, as golf courses and other turf areas are located on salt-laden lands, coastal areas, and areas where fresh water sources are limited. In areas where fresh water is limited or costly, the use of reclaimed water on golf courses and other turfgrass areas is a strategy to maintain sustainability and conserve water or is mandated. Depending on the source of recycled water and the growing environment to which it is applied, there are various potential pros and cons of using reclaimed water. Among other issues, the water often contains high levels of salts, potentially at high-enough concentrations to cause salt-stress damage to plants (Harivandi, 2013). But the pros of using reclaimed water, such as beneficial nutrients and cost efficiency, have allowed the use of reclaimed water to become a desirable turf management practice. Therefore, it is necessary to understand how turfgrass plants will cope with irrigation water containing high salt levels or prolonged salt exposure because of accumulated salts in soils, whether the salt stress is due to natural incidence or from use of reclaimed water.
Creeping bentgrass, the most desirable and predominant C3 species on golf course greens in the United States, is considered moderately sensitive to salt-stress damage (Harivandi, 2013). Salinity tolerance in C4 turfgrasses was reported to be closely associated with restricted Na+ accumulation in shoots (Marcum and Murdoch, 1994). Creeping bentgrass also exhibits considerable susceptibility to other abiotic and biotic stresses such as summer stress and dollar spot [Sclerotinia homoeocarpa (Fry and Huang, 2004)]. How the phytohormone profile of creeping bentgrass changes in response to salt irrigation is not yet fully understood or documented. Determination of hormone changes in response to salt stress will indicate potential salt tolerance attributes as well as potential susceptibility to other abiotic and biotic stresses during salt-stress incidence. Our results will also apply to other widely used grasses such as for forage or bioenergy. In addition, a better understanding of how hormones respond to salt stress may lead to the development of breeding methods to identify salt-tolerant cultivars via the use of these hormones as biomarkers or potential biochemical pathways to genetically alter for improved salt tolerance.
Hormonal changes in response to salt stress have been documented in other species such as wheat (Triticum aestivum) and rice (Oryza sativa) (Javid et al., 2011). However, how hormones and other metabolites respond to stress can be highly species dependent and depend on salt stress severity and duration. In addition, cultural practices such as mowing that could modify hormone contents of turfgrasses are highly different from that in other monocots used as agronomic crops. Significant variation in salt-stress tolerance has been found in creeping bentgrass germplasm (Marcum, 2001); however, the major physiological factors contributing to cultivar differences in salt tolerance have still not been fully elucidated. In addition, how phytohormone profiles change in response to salt stress in turfgrass species is not well investigated. Understanding how hormone profiles change will help to elucidate mechanisms governing salt-stress tolerance in creeping bentgrass.
Visual methods to measure plant cell viability by dye techniques have been used in other crop species, but have not yet been used within turf science. Researchers often encounter difficulties in the triphenyl tetrazolium chloride (TTC) method (Berridge et al., 2005) and find it difficult to isolate specific root classes, as is required in the methods of Ruf and Brunner (2003) from perennial grass species. This is largely because grasses have fine, fibrous, and dynamic root systems. Thus, investigation of additional techniques to evaluate cell viability in turfgrass species is desirable. One method to identify nonviable cells that is primarily used in flow cytometry or in fixed-tissue procedures is the use of SYTOX® dyes (Molecular Probes, Eugene, OR). These dyes are unable to penetrate live cell membranes. The dyes are able to penetrate dead cell membranes and then exhibit fluorescent properties when bound to the DNA within that dead cell. Recently, it was shown that these techniques are effective on living plant tissue of Arabidopsis thaliana (Truernit and Haseloff, 2008). Here, we aim to evaluate SYTOX dye staining procedures and which dye color is most effective to use on creeping bentgrass leaves and roots. We also aimed to determine whether the dye technique coupled with image color analysis is a quick, easy method for investigation of differential salt tolerance among turfgrass cultivars.
The objectives of this study were to understand how the major phytohormones ABA, IAA, ZR, JA, SA, and ethylene responded to salt stress in two creeping bentgrass cultivars contrasting in salt tolerance, Mariner (relatively salt tolerant) and Penncross (salt sensitive), maintained as low-mown turfgrass. In addition, we have evaluated physiological traits, cell viability, and quantified salt uptake and nutrient content of the two cultivars to determine relative tolerance levels.
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