Leaf carbon isotope ratios in three landscape speciesgrowing in an arid environment

D.A. Devitt*, S.D. Smith† & D.S. Neuman†

*Department of Environmental and Resource Science, University ofNevada Reno, Reno, NV 89557, U.S.A.

†Department of Biological Sciences, University of Nevada Las Vegas, LasVegas, NV 89154, U.S.A.

(Received 22 June 1996, accepted 24 July 1996)

The effect of leaching fraction (LF = drainage volume/irrigation volume) andtree planting size on the carbon isotope ratio measured in leaves of live oak(Quercus virginiana), desert willow (Chilopsis linearis) and tall fescue (Festucaarundinacea; LF only) were investigated in an arid climate. Irrigations wereapplied to maintain LFs of + 0·25, 0·00 or –0·25 (theoretical). Live oak anddesert willow were planted as 3·8, 18·9 or 56·8 l nursery container plants inlysimeters where hydrologic balances were maintained weekly. Leaf carbonisotope ratios varied by species across the three LFs (live oak > desertwillow > tall fescue). In live oak, leaf carbon isotope ratios increased as LFdecreased but did not change with planting size. In desert willow the ratiosincreased with decreasing LF and with increasing planting size. In tall fescuethe leaf carbon isotope ratio decreased as the LF increased. Leaf carbonisotope ratios also varied with stomatal conductance, leaf xylem waterpotentials and water-use efficiency (WUE). In live oak, both average middaystomatal conductance and leaf xylem water potential decreased withincreasing leaf carbon isotope ratio. Average stomatal conductance for liveoak was negatively correlated with average leaf WUE. Average leaf WUE inlive oak and tall fescue were correlated with average leaf carbon isotope ratios.However, in tall fescue leaf WUE increased as the leaf carbon isotope ratiobecame more negative. In live oak the ratio became more positive as leafWUE increased. This study has demonstrated that leaf carbon isotope ratioscan be used as a way of screening plant response for low water-uselandscapes, but only after careful correlations for each species have beenestablished between growth form and growing conditions.

©1997 Academic Press Limited

Keywords: Quercus virginiana; Chilopsis linearis; Festuca arundinacea; leachingfraction; tree size

Introduction

Population growth in the arid south-western U.S. continues to place increasingpressure on all available water resources. For example, outdoor water use on urban

Journal of Arid Environments (1997) 36: 249–257

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landscapes has been estimated to be as high as 60% of the total amount of water usedon residential sites in Las Vegas, NV (Las Vegas Valley Water District, pers. comm.1995). Because of a wide range in irrigation management practices, plants growing inthese arid urban landscapes are often subjected to a wide range of water availability.Many of these plants are also not well suited to an arid environment where water islimiting. Although much is known about the water use and WUE of agricultural crops(Green & Read, 1983; Sinclair et al., 1983) and to a lesser extent arid land plants ofecological significance (Toft et al., 1989; Ehleringer et al., 1992; Ehleringer, 1993),little is known about these attributes in plants of horticultural value (Devitt et al.,1994).

Quantifying carbon isotopic composition in plants has been shown to be an effectiveway of evaluating water-use efficiency (WUE) (Farquhar et al., 1982; Farquhar &Richards, 1984). Such a tool might be used to help in the process of selecting species(landscape plantings) better suited to an arid environment where water is limiting.However, the distinction must be made between plants that have a high WUE andlandscapes that are designed to be water efficient. Clearly, one species can use watermore efficiently than another in terms of CO2 fixed per unit of water transpired but stillmay use more water based on differences in growth patterns. Large plants with highWUEs could still contribute to an overall higher water-using landscape (water use perlandscape area) than that manifested through a combination of small plants with lowWUEs. Therefore, knowledge of the variation in WUE of plants of similar anddifferent growth patterns may be useful as a tool in plant selection for mixedlandscapes to achieve both a reduced water-use rate and a desired landscapeappearance.

This research was conducted to evaluate the use of leaf carbon isotope ratios inestimating WUE in three landscape species growing in an arid environment underdifferent irrigation regimes, and to determine if significant correlation’s existedbetween carbon isotope ratios, WUE, stomatal conductance and leaf water potential.We hypothesized that water availability would influence leaf carbon isotope ratios andwater relations parameters, allowing isotope ratios to be reasonable indicators ofWUE.

Materials and methods

Southern live oak (Quercus virginiana Mill.) (nursery seedling selection), desert willow(Chilopsis linearis (Cav.) Sweet var. linearis) and tall fescue (Festuca arundinaceaSchreb.) were grown outdoors (Las Vegas, NV) in non-draining 190 l (0·54 mdiameter) rigid plastic containers (lysimeters). A complete description of theexperimental setup can be found in Devitt et al. (1994). Briefly, trees and grass specieswere grown for a 2-year period under three different irrigation regimes. Trees wereplanted as 3·8, 18·9 or 56·8 l (1, 5, 15 gallon container) nursery stock. Tall fescue wasplanted as sod. The lysimeters were filled with a soil mix composed of 75% blow sandand 25% forest litter–bark mix. All lysimeters were lowered into open-ended concretepipes that had a sand base flooring.

In February 1990, after a 3-month establishment period, irrigation treatments wereimposed for a 2-year period by placing the trees and grass under leaching fractions(drainage volume/irrigation volume) of + 0·25, 0·00 or –0·25. These leaching fractionswere maintained by applying irrigations twice weekly based on the equation I = ETa/(1 – LF), where I is the irrigation volume to apply, ETa is the actual evapo-transpiration and LF is the leaching fraction. Thus, a deficit soil water status wasattained by placing a theoretical negative LF (–0·25) into the equation, resulting ineach week’s total irrigation for the –0·25 LF treatment to be less than the previousweek’s ETa. ETa was measured using the hydrologic balance approach; ETa = (irriga-

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tion + precipitation) – drainage – change in storage, where changes in soil waterstorage were estimated as the difference in lysimeter weighings taken every 7 days witha 2270-kg capacity (0·1% accuracy) load cell (Port-A-Weigh 4260, MeasurementsSystems Int., Seattle, WA). To do this, nylon slings were wrapped around thelysimeters and connected to metal hooks that hung from a rectangular metal frameattached to the load cell. The load cell was attached to an electrical hoist that wasmounted on a large movable frame that was positioned over each lysimeter. Drainagefor each individual lysimeter was collected 4 days per week by placing a vacuum of 17kPa for 1 h on two large ceramic extraction cups buried in 10 cm of diatomaceousearth at the bottom of each lysimeter. At the end of the first year, soil water storage wasre-established to initial values in all lysimeters by applying additional water. The sameprotocol for irrigation treatments were then re-imposed for a second year.

Leaf xylem water potential was measured with a pressure chamber (Soil MoistureInc., Santa Barbara, CA) and stomatal conductance was measured with a steady stateporometer (LI-COR 1600, LI-COR, Lincoln, NE) at solar noon twice per month onleaves sampled from the upper-outer canopy area of both the live oak and desert willowtrees. Fifty-four trees were monitored in this phase of the experiment (72 in the largerexperiment). All measurements were taken from 1130–1330h because this was whenclimatic factors were most stable at the study site. Canopy temperatures of tall fescueplots were measured at solar noon twice per month with an infrared thermometer(Everest Interscience, Tustin, CA).

At the conclusion of the experiment (February 1992), all trees were cut at the soilsurface and immediately weighed on a large top loading balance to the nearest gram.All leaves were removed from the oak trees prior to cutting. For the desert willows,leaves that fell during the winter period were collected from the base of the trees. Finalclippings from the tall fescue lysimeters were also taken at this time. All tissue sampleswere oven-dried at 70°C for 48 h.

Carbon isotope fractionation occurring via CO2 assimilation was used to evaluatepotential differences in water-use efficiency (WUE) among the three species studied.The ratio of carbon isotopes in leaf tissue samples (Rsam) relative to that of the PeeDeeBelemnite standard (Rstd) was used to express carbon isotope compositions (13C/12C)on a parts per thousand basis (‰). Carbon isotopic ratios were determined on oven-dried leaf tissues at the University of Utah Stable Isotope Ratio Facility forEnvironmental Research, where instrument error associated with each observation hasbeen estimated at 0·01% and error between repeated analyses at ≤ 0·14% (Ehleringer,1990). Because the study site was located in an urbanized area (population 1 million)in an enclosed basin, and the free atmospheric CO2 was not measured, no attempt wasmade at estimating discrimination values. Farquhar et al. (1989) has shown thatdiscrimination values can be calculated as ∆ – (δa – δp), where δa is the deviation of theisotopic composition of free atmospheric CO2 and δp is the deviation of the isotopiccomposition in the plant. Inoue & Sugimura (1985) have reported that the carbonisotope ratio of atmospheric carbon dioxide can vary substantially on a yearly basis inmajor metropolitan areas. Thus we chose to report carbon isotopic ratios instead ofdiscrimination values.

The treatments were replicated (N = 3) in a randomized complete block design(species 3 planting container size (trees only) 3 leaching fraction) and data analysedwith descriptive statistics, analysis of variance (ANOVA) and/or linear regressionanalysis. Average treatment values were compared based on an LSD generated from amean square of the error term from the corresponding ANOVA.

Results

An ANOVA revealed that leaf carbon isotope ratios varied by species across the three

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LFs (tall fescue = –28·22, desert willow = –27·28, live oak = –26·61; df. = 2,p = 0·001) and by leaching fraction imposed across the three species (–0·25LF = –26·58, 0·00LF = –27·61 and 0·25LF = –27·93; df. = 2, p = 0·001) but no significantinteractions occurred between LF and species. When ANOVAs were conducted basedon tree species, planting size and LF, live oak leaf carbon isotope ratios varied by LFacross the three planting sizes (–0·25LF = –25·68, 0·00LF = –26·88, 0·25LF = –27·34; df. = 2, p = 0·001) but not by planting size, and no significant interactionoccurred between LF and planting size. However, in desert willow, leaf carbon isotoperatios varied by LF with separation based on the –0·25 LF treatment (–0·25LF = –26·39, 0·00LF = –27·58, 0·25LF = –27·88; df. = 2, p = 0·001) and also by plantingsize with separation based on the 56·8 l planting size (3·8 l = –27·72, 18·9 l = –27·40,56·8 l = –26·72; df. = 2, p = 0·014) but with no significant interactions. In tall fescue,the leaf carbon isotope ratio decreased as the leaching fraction increased (IsotopeRatio = –28· 22– 1·93 (LF), r2 = 0·46, p = 0·05).

Average carbon isotope ratios obtained on leaf tissue at final harvest were correlatedwith average plant water measurements obtained over the last 12 months of the study.For live oak trees, average leaf carbon isotope ratios were found to be significantlycorrelated with average leaf xylem water potentials (Isotope Ratio = –31·11 – 3·59(ΨL, MPa), r2 = 0·62, p = 0·001) and also with average stomatal conductances (gs)(Isotope Ratio = –23·75 – 0·02 (gs, mmol m–2 sec–1), r2 = 0·74, p = 0·001). In desertwillow there was no correlation between average leaf carbon isotope ratios and averageleaf xylem water potentials. However, average stomatal conductances were correlatedwith leaf carbon isotope ratios (Isotope Ratio = –24·52–0·02 (gs, mmol m–2 sec–1),r2 = 0·48, p = 0·05). Canopy temperatures were monitored on the tall fescue plotsand no significant correlation was obtained between the average canopy temperaturesand the average leaf carbon isotope ratios.

Relative growth changes occurring in each planting size of both live oak and desertwillow were assessed by measuring the change in trunk diameter occurring over theexperimental period divided by the initial trunk diameter. In both live oak and willowtrees, a decrease in the carbon isotope composition was associated with an increase inthe relative trunk diameter change (RTDC) (live oak = –26·06–3·13RTDC,r2 = 0·18, p = 0·05; desert willow = –26·13–2·48RTDC, r2 = 0·41, p = 0·001).Smaller trees recorded larger relative trunk diameter changes, with desert willowrecording as much as a two-fold increase over live oak at the 3·8 l planting size (desertwillow: RTDC = 0·7157 – 0·0095X, r2 = 0·72, p = 0·001 and live oak:RTDC = 0·3288 – 0·0422X, r2 = 0·55, p = 0·001, where X is planting size in l).

Leaf water-use efficiencies (WUE) were calculated for the live oak trees during thesecond year of experimentation by dividing the total dry weight of leaves collected atharvest by the corresponding yearly evapo-transpiration totals. First year data were notincluded because collection of leaves that dropped during that period was not made.Leaf WUE was also estimated for the tall fescue plots by dividing the totals of weeklydry weight harvests (clippings) by the evapo-transpiration totals during the secondyear. No WUE estimates were made for the desert willow trees, since the leaves hadfallen during the 3-month period prior to the end of the experiment, allowing for onlyan accurate subsample to be taken. Average stomatal conductance (gs) values werefound to be negatively correlated (r2 = 0·77, p = 0·001) with average leaf WUEs oflive oak trees (Fig. 1, where WUE and gs values were averaged by planting size and LFtreatments). Leaf WUEs for live oak trees were highly correlated (r2 = 0·90,p = 0·001) with the average measured leaf carbon isotope ratios (Fig. 2, where WUEand isotope ratios were averaged by planting size and LF treatments). In tall fescue,average leaf WUEs were also highly correlated with average isotope ratios (WUE = –24·83–17·63 (isotope ratio), r2 = 0·99, p = 0·001). However, leaf WUEs increased asthe average carbon isotope ratios became more negative in tall fescue, whereas in oaksleaf WUEs increased as the average carbon isotope ratios became more positive.

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Average stomatal conductance (mmol m–2 s–1)

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Discussion

In this study we compared the carbon isotope ratio in leaf tissue from live oak (Quercusvirginiana), desert willow (Chilopsis linearis) and tall fescue (Festuca arundinacea). Ourdata suggested that differences in growth patterns and turnover of carbon assimilatedinfluenced the final isotope ratios. Older leaves that remained on the tree from theinitial planting to the final harvest (live oak) might have influenced the isotope ratio.If carbon was assimilated into the cellulose of older leaves prior to stress treatments itis possible they would not reflect the same carbon discrimination that would take placeunder more stressful conditions. Francey et al. (1985) have shown that both age andstress can induce variation in carbon isotope discrimination within plant partsdeveloped at different times. Yakir et al. (1990) suggested that the isotopic signal ofleaf organic matter provides an integrated signal that is accumulated over the entiretime the leaf is developing. In this study, we separated larger, older leaves of live oak(3·7 ± 0·6 cm long 3 1·6 ± 0·3 cm wide) from younger, smaller leaves (1·9 ± 0·2 cmlong 3 0·9 ± 0· cm wide) for carbon isotope analysis. The younger leaves had a 22%wider range in isotope ratios (data not shown). The data reported for live oak in thismanuscript reflects the activity in younger leaves that developed during the irrigationtreatments.

In contrast to live oak, the deciduous desert willow sheds all leaves just prior towinter and tall fescue grows in spurts in response to favorable growing conditions. Inaddition to variation by leaf age, other factors influence carbon isotope ratios. Forexample, Hubick & Farquhar (1989) reported that there were differences in the

Figure 1. Average water-use efficiency of live oak (leaf dry weight(g)/evapo-transpiration (1)) asa function of average stomatal conductance. Separation of data based on leaching fractions (LF)and planting size, where points labeled as 1, 5 or 15 refer to nursery container size in gallons(which equates to 3·8, 18·9 and 56·8 l).

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–29

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discrimination value among plant parts in barley and that these differences variedsignificantly among cultivars. In herbaceous species the discrimination values forwhole plants appear to be better correlated with WUE than the discrimination valuesfor leaves only (Poorter & Farquhar, 1994). In our study, we only measured the carbonisotope ratio in leaves. However, we believe that the growth of leaves as opposed to thegrowth of other plant parts was influenced to a greater extent by the irrigationtreatments imposed during the experimental period (especially for the two treespecies). For prairie graminoids, species identity, year of collection, date within thegrowing season and site all contributed in an additive fashion to variation in the carbonisotope ratio (Mole et al., 1994). Final harvests of both live oak and tall fescue weretaken in February, which represented a period of low evapo-transpirational demand(Devitt et al., 1994). Whether the time of sampling had any influence on the isotoperatios measured can only be speculated on; however, with tall fescue a wider range inyield was clearly observed between treatments during the more stressful summerperiod.

All of the trees in this experiment were grown in separate lysimeters with sufficientspacing to be considered as isolated trees. As such, their canopies were closely coupledwith the environment. Conductance values decreased as stress increased, with higherconductance values associated with the smaller planting sizes. Isotope ratios decreasedas stress decreased, with the most negative values being associated with the smallestplanting size under the highest LF. Such a strategy by small juvenile trees may be anadaptation that maximizes growth at the expense of WUE. Greater relative trunkdiameter changes in the smaller trees would support this conclusion. Sandquist et al.

Figure 2. Average isotope ratio measured in the leaves of oak as a function of the average water-use efficiency (leaf dry weight (g)/evapo-transpiration (1)). Separation of data based on leachingfractions (LF) and planting size, where points labeled as 1, 5 or 15 refer to nursery container sizein gallons (which equates to 3·8, 18·9 and 56·8 l).

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(1993) reported that in four of five species examined, WUE was lower and growthmore rapid during the establishment stage than as adults. Differences in carbonisotope ratios between species also correlated with relative growth rates, with the liveoak having smaller relative trunk diameter changes than desert willow at all plantingsizes and also more positive isotope ratios. Tall fescue had the greatest turnover of newgrowth and lowest isotope ratios among the three species. Ehleringer & Cooper (1988)suggested that plants with high WUE may in fact grow more slowly than plants withlow WUE during periods of high soil water availability.

Leaf carbon isotope ratios proved to be reasonable indicators of leaf WUE in bothlive oak and tall fescue. In live oak a 1·9-fold increase in leaf WUE was observed fora 1·0% increase in the leaf carbon isotope ratio while a 1·4-fold increase in leaf WUEfor a 1·0% decrease in the leaf carbon isotope ratio was observed in tall fescue basedon average values. This response became even clearer when replicates within eachirrigation treatment and planting size were pooled. Error bars associated with theseaverage values clearly indicate that variability existed both within isotope ratios andleaf WUEs. Hubick & Farquhar (1989) reported that despite the potential for variationin WUE independent of discrimination values, whole plant WUE and carbon isotopediscrimination were highly correlated. In a previous study we found a reverse trend inWUE between live oak and desert willow (Devitt et al., 1994). However, those valueswere based on total above-ground dry weight and not just leaf dry weight or carbonisotope ratios. When WUE was based on above-ground dry weight, a lower r2 and levelof significance was observed with the carbon isotope ratios. This poorer correlationwas no doubt due to the fact that the majority of this growth (stem and woodproduction) occurred prior to the stress being imposed and that carbon isotope ratioswere measured only on leaves.

In tall fescue, increased leaf WUE was driven by a larger increase in dry matterproduction than a change in evapo-transpiration, and this was associated with adecrease in the leaf carbon isotope ratio. A 1·5-fold increase in evapo-transpiration per1·0‰ decrease in the leaf carbon isotope ratio was associated with a 2·0-fold increasein dry matter yield. Wright et al. (1988) reported that variability in WUE of peanutswas due more to the variation in total dry matter production than in water use.Decreased stress associated with more available water led to an enhanced photo-synthetic capacity in this C3 grass. Dean et al. (1996) has reported that 75% of thevariability in WUE of tall fescue growing under varied combinations of salinity anddrought could be accounted for if matric and osmotic potentials in the soil wereknown, with higher WUEs also being reported under lower stress conditions. A similarresponse has been reported by Condon et al. (1987) for wheat, where a positivecorrelation between discrimination values and WUE were obtained under well wateredconditions. It has been suggested by Farquhar et al. (1988) that this may be related tochanges in internal resistance (CO2, heat transfer) when comparing individual plantswith plants within full canopies.

Finally, although leaf carbon isotope ratios were more positive (higher WUE) for liveoak than for desert willow at the larger planting size, higher ET values were reportedfor oak at all LF treatments (Devitt et al., 1994). This would suggest that some plantsare in fact more efficient in utilizing water, but still use significantly more water thanother plants because of differences in growth patterns or form. Selection of plants forlow water-using landscapes in arid environments where water is a precious limitingresource should thus be based on established total water use estimates. Leaf carbonisotope ratios are useful as a proxy for WUE provided species, size and growingconditions are considered. These ratios could be used as an additional means ofselecting plants for a low water-use landscape.

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We thank the Las Vegas Valley Water District for financial support of this research project. Wewould also like to thank Jeff Andersen, Linda Verchick and Bob Morris for their able laboratoryand field assistance.

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