Carbon Isotope Discrimination, Gas Exchange, and Growth of Container-grown Conifers Under Cyclic Irrigation

The objective of this study was to quantify the effects of cyclic irrigation on growth and physiology of container-grown conifer species in pot-in-pot (PIP) production in the upper Midwest. Trees of four conifer species (Picea glauca var. densata, Picea pungens, Abies fraseri, and Pinus strobus) were grown in 25-L containers and assigned to one of four combinations of irrigation rate (low or high) and daily irrigation cycle frequency (one or four). Irrigation rates were based on common nursery practice in the region (2 cm rainfall equivalent/day) and one-half the standard rate (1 cm rainfall equivalent/day). Cyclic irrigation increased relative height growth and relative caliper growth of Pinus strobus by over 80% and 35%, respectively, compared with once-daily irrigation. The high-rate irrigation increased relative caliper growth of Picea pungens by 40% compared with the low rate. The effects of irrigation regime on needleor shootlevel gas exchange varied by species and date of measurement. Carbon isotope discrimination (DC) of needle and wood tissue was positively correlated (r ‡ 0.64, P < 0.001) with needle conductance to water vapor (gwv) and negatively correlated (r£L0.60, P< 0.001) with intrinsic water use efficiency (WUEi). Carbon isotope discrimination of wood and needle tissue decreased with the low irrigation rate, indicating increased WUEi associated with reduced gwv. Cyclic irrigation had relatively little effect on D C except for Pinus strobus. Our findings suggest that carbon isotope composition of wood and needle tissue provides a sensitive and accurate representation of plant response to varying moisture availability. From a water management perspective, identifying optimal irrigation rates appears to be more important than number of daily cycles for these crops grown in the midwestern United States. Nursery producers are in need of waterconserving irrigation methods that will allow them to adapt to increased water costs and stricter water withdrawal and discharge regulations (Beeson et al., 2004). Growers often overwater by applying a fixed rate of irrigation that may exceed plant water use (Tyler et al., 1996); this can result in excessive water withdrawals and leaching of nutrients and chemicals into surface and groundwater. The development of irrigation programs that conserve water without reducing tree growth and quality can enable the nursery industry to adapt to increased input costs and stricter water use regulations. One potential method of conserving water is cyclic irrigation, in which a fraction of a plant’s daily water allowance is applied several times a day. Plant responses to cyclic irrigation include higher growth index, root growth, trunk diameter, shoot dry weight, height, and crop yield than plants receiving one irrigation event per day (Beeson and Haydu, 1995; Fain et al., 1999; Ismail et al., 2007; Keever and Cobb, 1985; Ruter, 1998; Witmer, 2000). In some cases, caliper growth was 25% greater using cyclic irrigation compared with the traditional method of applying a single cycle in the morning (Beeson and Haydu, 1995; Fain et al., 1999; Witmer, 2000). These studies, however, were conducted in the southeastern United States or in greenhouses and may not reflect conditions common in nurseries in cooler climates such as the upper midwestern United States. Improving our understanding of the physiological mechanisms behind the growth responses to irrigation is important in developing effective water management strategies. Up to 97% of water used by plants is transpired through stomata, and an initial response to limited water availability is stomatal closure (Halevy, 1972; Hand et al., 1982; Medrano et al., 2002; Taiz and Zeiger, 2006), which ultimately inhibits CO2 uptake and reduces photosynthetic assimilation (A). It has been postulated that cyclic irrigation increases growth by removing or reducing midday water limitations, thereby increasing cumulative photosynthesis (Beeson, 1992; Witmer, 2000). For example, applying daily irrigation in two split applications consistently increased midday stomatal conductance (gS) to water vapor for plants from three Rhododendron cultivars (Scagel et al., 2011). Cyclic irrigation, by maintaining higher substrate moisture content in the midday hours than one single application, delays or prevents stomatal closure, resulting in higher cumulative A (Halevy, 1972; Warren and Bilderback, 2002). Water use efficiency (WUE) is the relationship between assimilation and water loss and can be used to measure how efficiently a plant is using water to produce biomass (Anyia and Herzog, 2004). Several studies have demonstrated increases in WUE when trees are exposed to water-limiting irrigation regimes (Anyia and Herzog, 2004; Ningbo et al., 2009; Warren and Bilderback, 2002). Two methods are commonly used to estimate WUE for trees and shrubs: instantaneous WUE from measurements of leaf gas exchange and integrated WUE based on stable carbon isotope discrimination. Instantaneous WUE measures WUE at the leaf level and is the ratio of the rate of A to the rate of transpiration or the rate of leaf conductance to water vapor conductance. The ratio, A/gwv, is often referred to as WUEi, because it removes the effect of vapor pressure deficit inherent in WUE estimated from transpiration rate (Roussel et al., 2009). Intrinsic water use efficiency typically increases as plants undergo water stress, because gwv often decreases more rapidly than A as water becomes limiting. Estimating WUE from gas exchange can present logistical challenges in measurement and interpretation because gas exchange rates vary diurnally and seasonally. Because of this, stable carbon isotope (C) analyses have become an important method of assessing plant responses to environmental stress, especially water deficits (e.g., Farquhar et al., 1989). Plants discriminate against C during photosynthesis, resulting in a lower C isotopic composition in plant tissue than in the Received for publication 20 Dec. 2012. Accepted for publication 1 May 2013. Funding for this study provided by Michigan State University Project GREEEN. In-kind contributions provided by Nursery Supplies, Inc., Scotts Co., Renewed Earth, Inc., and Peterson’s Review Nursery. We thank Dana Ellison, Brad Rowe, Erik Runkle, and three anonymous reviewers for their helpful reviews of a previous version of this manuscript. Former Graduate Research Assistant. Current address: North Carolina Cooperative Extension, 120 Hospital Drive, Suite 1, Lenoir, NC 28645. Associate Professor. To whom reprint requests should be addressed; e-mail [email protected]. 848 HORTSCIENCE VOL. 48(7) JULY 2013 atmosphere. Empirical and theoretical studies have shown that the degree of discrimination against C is determined largely by intercellular CO2 concentration, which is controlled by the ratio of A to gwv (Farquhar et al., 1989). Stable carbon isotope discrimination may be used to estimate WUE during the time in which plant tissues, usually leaves or wood, are formed. Ecological studies have documented decreases in DC of coniferous species in response to limited water availability (Aranda et al., 2010; Brandes et al., 2007; Olivas-Garcia et al., 2000; Zhang and Cregg, 2005). Because both DC and WUEi are functions of A and gwv, they are often strongly correlated. However, DC may provide some practical advantages in evaluating plant response to water availability because it integrates physiological responses over time rather than presenting a series of ‘‘snapshot’’ measurements. Based on previous studies, we hypothesized that cyclic irrigation would reduce tree water stress and improve growth and plant WUE compared with conventional, once-aday irrigation. The objectives of this study were to 1) determine effects of cyclic irrigation programs on growth of container-grown conifers; and 2) explore underlying physiological mechanisms including various indicators of WUE. Materials and Methods Site description. This study was conducted at the Michigan State University Horticulture Teaching and Research Center (lat. 42.67 N, long. 84.48 E, elevation 264 m) near East Lansing, MI. Trees were grown in 25-L (#7) containers in a PIP production system. The soil on site was a well-drained loamy sand (82.6% sand, 8.3% silt, 9.1% clay). Container spacing was 0.5 m and 1 m on-center within rows and between rows, respectively. Rims of the socket pots were 2.5 cm aboveground level and the ground was covered with landscape fabric to control weeds. Plant materials. In Apr. 2008, 100 trees (25 of four species) were transplanted from 10-L (#3) containers (GL1200; Nursery Supplies, Inc., Chambersburg, PA) into 25-L (#7) containers (GL 2800) using an 80:20 (v:v) mix of pine bark and peatmoss (Renewed Earth, Inc., Kalamazoo, MI). Container capacity of the media was 44.5%. The four species used were Fraser fir [Abies fraseri (Pursh) Poir.], Colorado blue spruce (Picea pungens Engelm. var. glauca Regel), black hills spruce [Picea glauca (Moench) Voss var. densata], and eastern white pine (Pinus strobus L.) (Table 1). All plants were originally obtained from a local nursery (Petersons Riverview Farm, LLC, Allegan, MI) as seedling transplants in 2006 and then grown for 2 years in 10-L containers in an 80% pine bark:20% peatmoss (v:v) substrate. Irrigation treatments. Daily irrigation volumes were based on daily water use estimates of similar species from Warsaw et al. (2009). Four trees of each species were randomly assigned to one of four irrigation regimes. Treatments consisted of a combination of daily irrigation rate (low or high) and daily irrigation cycle frequency (one or four). There were eight rows of trees in the study. Each row was assigned at random to receive once-daily irrigation (1·) or cyclic irrigation applied four times daily (4·). Once-daily irrigation events occurred at 0600 HR. Cyclic irrigation treatments were applied at 0600, 1000, 1400, and 1800 HR daily to apply 25% of the daily total at each watering. Within each row, irrigation volume (high or low) was assigned at random. Low (950 mL) and high (1900 mL) irrigation rates of trees corresponded to 1 cm and 2 cm rainfall equivalent, respectively. Irrigation was applied from 15 May 2008 to 25 Sept. 2008 and from 21 May to 1 Oct. 2009. Irrigation was controlled by two timers (8014 Series Solo Rain; Nelson Irrigation Corp., Walla Walla, WA); one timer ran once daily, and the other ran four times per day. Irrigation rate was controlled by selection of an emitter using pressure-compensating spray stakes (Netafim, Fresno, CA). Containers were top-dressed with 130 g of 15N–4P–10K controlled-release fertilizer (Osmocote Plus 8-9 month release; The Scotts Co., Marysville, OH) in the spring of 2008 and 2009. Weeds within containers were controlled through hand-weeding. Pots were turned periodically during the growing seasons to prevent rooting into the native soil. The experimental treatments were arranged in a four · two · two factorial with four species · two irrigation rates · two irrigation cycle frequencies. There were four experimental units (trees) for each species · irrigation rate · cycle frequency combination. The plot of study trees was surrounded by a guard row of similarly sized trees. Growth. Tree growth (caliper and height) was measured at the beginning and end of each growing season. Initial tree height was measured with a meter stick to the highest live point of the tree. Trunk caliper was assessed using a digital caliper. Two caliper measurements were taken perpendicular to each other, north–south-oriented and east– west-oriented, at a height level even with the rim of the container. Relative growth rates were calculated as height (or caliper) growth during the study divided by the initial height (or caliper). Gas exchange measurements—Picea spp. and Abies fraseri. Photosynthetic gas exchange was measured on three dates in the 2009 growing season using a portable gas exchange system (LI-6400; LI-COR Inc., Lincoln, NE) equipped with a 0.2-L cylindrical conifer chamber (LI-6400-05). Lightsaturated photosynthesis (Amax) and gwv were assessed on all trees between 1100 HR and 1500 HR on 24 July, 6 Aug., and 31 Aug. One south-facing shoot of the current season’s growth was selected from the upper third of each tree and flagged so that it could be used in subsequent gas exchange sampling. Gas exchange was measured on clear days [photosynthetic photon flux (PPF) > 1500 mmol·m·s] using a CO2 concentration at 400 mmol·mol and flow of air at 500 mL·min. To minimize temperature effects during each measurement run, the temperature controller of the photosynthesis system was set at the predicted high temperature for the day. Measurements were recorded after the readings had stabilized on the system’s real-time graphics screen. At the end of each season (15 Oct. 2008 and 28 Oct. 2009), shoots used for gas exchange measurements were harvested and scanned using a leaf area meter (LI-3000; LI-COR Inc.) to obtain the projected shoot area, by which gas exchange measurements were adjusted. Intrinsic water Table 1. Mean (± SE) height and caliper of container-grown trees of four species at the start of cyclic irrigation trial. Species Ht (cm) Stem caliper (mm) Picea glauca 68.8 (3.0) 27.2 (0.6) Picea pungens 68.4 (2.6) 28.3 (0.8) Abies fraseri 69.8 (2.4) 26.0 (0.7) Pinus strobus 111.3 (2.8) 33.9 (1.1) N = 16 Fig. 1. Sample cross-section of conifer stem before sectioning into individual annual rings for C analysis. HORTSCIENCE VOL. 48(7) JULY 2013 849 CROP PRODUCTION