Plant Ecology 149: 131–142, 2000.© 2000Kluwer Academic Publishers. Printed in the Netherlands.

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Ecophysiological differences between male and female plants ofPistacialentiscusL.

O. Correia1 & M. C. Diaz Barradas21Departamento de Biologia Vegetal. Faculdade de Ciências da Universidade de Lisboa, Campo Grande, BlocoC-2, Piso 4, 1700 Lisboa, Portugal (E-mail:otiliacc@fc.ul.pt);2Departamento de Biolog´ıa Vegetal y Ecolog´ıa,Facultad de Biologia, Universidad de Sevilla, Apartado 1095, 41080 Sevilla, Spain

Received 25 June 1999; accepted in revised form 5 May 2000

Key words:Chlorophyll fluorescence, Dioecy, Drought, Gas exchange, Water relations

Abstract

Previous studies in spatial distribution of male and female shrubs ofPistacia lentiscushave demonstrated that lessperturbed areas, older communities with a well developed cover, have male-biased sex ratios, whereas in abandonedold agricultural areas there are no significant differences between the number of male and female plants. In thisstudy, we analyse both sexes in terms of their photosynthetic features that could provide a physiological basis forhabitat partitioning between sexes. Rates of light-saturated assimilation and stomatal conductance were studiedin male and female plants during summer. Assimilation rates were higher in the morning than in the afternoonand mean daily maximum assimilation rates reached 10.9 and 6.6µmol m−2 s−1, for male and female plants,respectively. In the absence of drought stress (laboratory conditions), the measured photosynthetic characteristicsof leaves of male and female plants, provided by fluorescence studies and light and CO2 response curves, weresimilar. Under natural stress conditions however, lower CO2 assimilation rates and stomatal conductances wererecorded in female plants. The differences in the light response curve of effective quantum yield (8II ) recordedunder stress conditions showed also higher quantum yield for male plants under low irradiances. From this studywe suggest that the differences observed between male and females are largely due to different degrees of stomatalcontrol rather than to differences in photosynthetic activity, leading to higher water use efficiency (WUE) in femaleplants. However, despite the higher leaf control of water loss by females, they reduce the water potential to the samevalues as male plants, probably due to specific characteristics of the root system or of the conducting xylem. Theseresults suggest that the ecological advantage of male plants in older communities is due to a higher competitionfor water uptake, while in the youngest open areas is the higher WUE in female plants that confer an ecologicaladvantage.

Introduction

Pistacia lentiscusis a common dioecious evergreenshrub occurring in Mediterranean ecosystems in awide variety of habitats, from open communities ingarrigues to close communities in more mesic andshaded sites. Previous studies of spatial distribution ofmale and female shrubs showed that in less perturbedareas from mature stages of succession, with a well de-veloped vegetation cover, the sex ratio (ratio betweenthe number of male and female individuals) was male-biased (higher number of males), while in abandoned

old agricultural areas there were no significant differ-ences between the number of male and female plants(Diaz Barradas & Correia 1999).

Male-biased sex ratios in low resource environ-ments have been reported by other authors (Dawson& Bliss 1989; Sakai 1990; Sakai & Weller 1991; Sheaet al. 1993). However, Verdú & García Fayos (1998)found that the sex ratio inPistacia lentiscusin stressfulhabitats was female-biased. Moreover, they found asignificant correlation between the population densityand the proportion of male individuals, the sex ratioapproaching 1:1 when density increased. This result

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suggested that males had more ecological performancewith increasing competition.

In dioecious species, females usually develophigher reproductive effort than males because theyproduce fruits in addition to flowers, and thus theyhave to allocate more biomass to reproduction (Wal-lace & Rundel 1979; Antos & Allen 1994; Jonassonet al. 1997; Obeso 1997; Hogan et al. 1998; andCorreia et al. 1992, forP. lentiscus). Although a sig-nificant sexual dimorphism was not found, the oldermale plants tended to be taller, with a larger canopyvolume, a higher leaf area index and a significantlyhigher value in basal stem number (Diaz Barradas &Correia 1999). Such characteristics could confer com-petitive advantage to males. Females would probablyhave lower survival rates than males in more advancedstages of succession, when competition is higher andthe individuals are oldest. Delph (1999) also refersthat sexual dimorphism often does not exist until aftermultiple reproductive bouts have occurred. Periodicdroughts are characteristic of Mediterranean ecosys-tems and may be important as enhancers of competi-tion and environmental stress in more advanced stagesof succession.

Most studies on the maintenance of dioecy havefocused on reproductive development. However, dif-ferences during vegetative development, such as bettercompetitive abilities or more efficient use of limitingresources and thereby higher survival might also beimportant. Some investigations have concentrated onphysiological differences between the sexes and on thelink between physiology and the spatial distributionof the sexes (Freeman & McArthur 1982; Dawson& Bliss 1989; Gehring & Monson 1994; Laporte &Delph 1996; Hogan et al. 1998). Some authors pointedout that sexual differences in resource costs mightlead to greater physiological stress in the sex withthe higher reproductive investment (Dawson & Bliss1989; Gehring & Monson 1994; Laporte & Delph1996). The results of these studies have shown thatdifferences in physiological specialisation may aideach sex in meeting different resource demands asso-ciated with reproduction, but the results depend on thespecies without any general pattern. All indications arethat dimorphism in physiological traits is variable bothin degree and in direction within species from site tosite and also between related species (for a review, seeDawson & Geber, 1999). The functional significanceof dimorphism is difficult to interpret, because of theshort-term nature of the measurements of physiologi-cal traits. Dawson and Geber (1999) refer in a recent

review that there are a number of dioecious speciesthat exhibit spatial segregation of the sexes, often inassociation with habitat quality, but that do not showsexual dimorphism in physiology or morphology.

The objective of the present study was to investi-gate the possibility that differences in photosyntheticcharacteristics, between male and female plants, couldplay a role in spatial sex distribution. Specifically wetested the hypothesis that females have higher pho-tosynthetic rates than males, to meet the additionalenergy requirements to reproduction with a similarplant structure. This study was performed duringsummer under extreme environmental conditions. Toaddress these questions, the dependence of leaf pho-tosynthetic rates on both photon flux density (lightresponse curves) and CO2 partial pressure (CO2 re-sponse curves), chlorophyll a fluorescence, as wellas leaf organic nitrogen and specific leaf weight weremeasured both in the laboratory and in the field.

Materials and methods

Site and species characteristics

The field study was conducted in Portugal, inthe Natural Park of Serra da Arrábida (Convento−38◦28′40′′ N; 8◦59′34′′ W, elevation 280 m) on asouth facing slope with a well developed Mediter-ranean maquis, 20 to 30 years old, after last burn-ing. The detailed descriptions of the vegetation ofthis region are available in Catarino et al. (1982).The plant community at the study site belongs tothe thermophilic plant community known asOleo-Ceratonionwhich can be found in very xeric zonesdominated by evergreen sclerophyllous shrubs, suchas Quercus coccifera, Pistacia lentiscus, Arbutusunedo, Phillyrea latifolia, Phillyrea angustifoliaanddrought-semideciduous shrubs, such asRosmarinusofficinalis and someCistaceae(Cistus salvifolius,C. monspeliensis, C. albidusandC. ladanifer). Pista-cia lentiscusL. (Anacardiaceae) is a dioecious scle-rophyllous evergreen species that forms shrubs up to2 m high, sometimes attaining a tree growth form inthe more humid and protected sites.

Field measurements of gas exchange and waterrelations

Measurements were done in the field (1996, 1997) andunder controlled conditions in the laboratory (1996).

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Measurements of net photosynthetic rates (A), stom-atal conductance (g) and transpiration rates (T r) weremade using a portable compact CO2/H2O porometerfrom Walz (Schulze et al. 1982). Rates of net CO2 andH2O-vapour exchange were calculated from the differ-ence in CO2 and H2O concentrations between sampleand reference air in an open system. The gas analyserwas a BINOS 1 (Leybold Heraeus, D-6450 Hanau,Germany). Sensors in the porometer cuvette allowedreadings of leaf temperature (Tl), air temperature (Ta)and air relative humidity (RH). Air temperature andvapour pressure deficit inside the cuvette were main-tained at values close to those of external air by acontinuous mixing of the air during measurements.The porometer head is also equipped with a LiCorquantum sensor (model Li-190SB), outside the cham-ber, wich provided readings of incident photon fluxdensity of photosynthetically active radiation (PFD).All measurements of CO2 exchange were performedat CO2 concentration of ambient air for which thegas analyser was calibrated, and no corrections wereneeded.

Leaf-to-air water-vapour-concentration difference1W (Pa kPa−1) was obtained from measurements ofleaf and air temperatures inside the chamber and at-mospheric relative humidity. All rates were calculatedon a projected leaf area basis, using equations and wa-ter vapour corrections proposed by Cowan (1977) andvon Caemmerer & Farquhar (1981).

Three mature shrubs of male and female plants ofPistacia lentiscusgrowing adjacent to each other wereselected and measured during two summer days in1996 and one day in 1997. Measurements were madeon three to six well exposed leaves from current-yeartwigs of each shrub. During periods of constant PFDa single leaf from the top of the canopy was sealed inthe porometer cuvette for some minutes. Ten measure-ments were made on each leaf and the mean value wasused.

Parallel to these gas exchange measurements, leafwater potential (9) was also measured from 09:00to 17:00 using a pressure chamber (Schölander et al.1965). Each result is the average of two to four leavestaken from the outer canopy of each shrub.

Laboratory measurements of gas exchange

For measurements of light and CO2-response curves,twigs from the selected shrubs were collected in July1996, recut under water, and left to rehydrate over

night at room temperature. We assume that leaveswere then at full turgor.

Measurements were conducted using a minicuvettesystem (Walz, Effeltrich, Germany) with a Gas Ana-lyzer BINOS-100. Humidity was controlled by a ColdTrap KF-18/2 (Walz) connected to the Minicuvettesystem. For light-response curves, conditions in thecuvette were: 23◦C, leaf to air vapour pressure dif-ference of 11–12 Pa/kPa, and ambient CO2 partialpressure (36 Pa). After acclimation for 30 min at alight intensity of 1800µmol m−2 s−1, CO2 and H2Oexchange rates were measured, starting with this lightintensity and decreasing it in small steps until dark-ness. CO2-response curves were made at high PFDs(1800µmol m−2 s−1) by lowering the CO2 pressurefrom 30 Pa to approximately 3 Pa in small steps andthen increasing it again to 150 Pa. Each responsecurve was obtained from measurement on a singleleaf. Three to five leaves were measured from differentplants of each sex.

Partial pressure of CO2 in the leaf air spaces,Pi, was estimated from A and g using a factor of1.6 for the ratio of the binary diffusivities of watervapour/air and CO2/air, relating the conductance ofCO2 to that of water vapour, according to von Caem-merer & Farquhar (1981) and Sharkey et al. (1982).Partial pressure of CO2 in air was controlled by mix-ing CO2-free air with CO2 by means of a CO2/N2 Gasmixing unit GMA-2 (Walz).

Measurements of chlorophyll fluorescence

Measurements of chlorophyll a fluorescence weredone both in the field on the same leaves of gas ex-change measurements, and in the laboratory in thesame twigs used for photosynthesis studies. Mea-surements were carried out on the adaxial surface ofthe leaf at ambient temperature and CO2 concentra-tion, using the pulse-amplitude-modulation technique(Bilger et al. 1995) with the portable fluorometerPAM-2000 (Walz, Effeltrich, Germany). Standardnomenclature of characteristics fluorescence levelsand expressions are those proposed by Shreiber et al.(1995).

The ratio of variable to maximal fluorescenceFv /Fm, that represents the potential maximal PS IIquantum yield or photochemical efficiency of dark-adapted leaves (optimal quantum yield of PS II)was automatically calculated by the PAM-2000 as:Fv /Fm = (Fm - F0)/Fm, whereF0 is the initial minimal

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Figure 1. Daily variation of air temperature (T ), incident photonflux density of photosynthetic active radiation (PFD) and watervapour pressure deficit (1W ) for 24 July and 1 August 1996.

fluorescence on dark adapted leaves for 15 min andFmis maximal dark-adapted fluorescence.

The effective quantum yield of non-cyclic elec-tron transport through the PS II or photochemicalefficiency of PS II in a light-adapted state (8II )was calculated according to Genty et al. (1989) as8II = (F ′m − F)/F ′m = 1F/F ′m, (F ′m = max-imal and F = steady-state fluorescence under ac-tinic irradiance), from measurements with a leaf clipholder (2030-B Walz). The light-response curves of8II were determined in the field (same days of gasexchange measurements) and under controlled condi-tions (laboratory) in leaves from the same rehydratedtwigs used for CO2 and light response curves. The pre-programmed sequence of the fluorometer was used inconjunction with the Leaf-Clip Holder 2030-B, givingillumination times of 5 minutes with the internal LEDactinic light source (up to 500µmol m−2 s−1).

Recordings were taken at ambient temperaturesin the laboratory and in the field. Leaf temperaturesmeasured with fine wire thermocouples on the abax-ial surface, rose to about (35–37◦C) during periodsof maximum sun exposure in the field, and around25–30◦C under laboratory conditions.

Foliar nitrogen concentration

Total N was determined by a C/N Elemental AnalyserEA 1108 (Fisons Instruments), on all leaves on whichphotosynthetic measurements were done.

Analysis of light response curves

The light response curves used a descriptive func-tion (Harley et al. 1986; Hogan et al. 1996):A =8I/(1 + 82I2/A2

max)1/2 − Rd , whereA is the net

photosynthetic rate,I (PFD) is the incident photonflux density,Amax is the light saturated photosyntheticrate,Rd is the rate of respiration in the light, exclusiveof photorespiration, and8 is the initial slope (appar-ent quantum yield). The parameters,8, Amax andRdwere obtained from 5 individual curves. Average val-ues were calculated for each sex and compared with aStudent’st-test.

Analysis of CO2 response (A/Ci) curves

We used a model developed by Farquhar et al. (1980)and Sharkey (1985) as applied by Harley et al. (1992),in which net photosynthesis (A) is expressed as:A =Vc(1 − 0.5O/τCi) − Rd , whereVc is the rate ofcarboxylation of the enzyme ribulose-1,5-biphosphatecarboxylase/oxygenase (Rubisco),Rd is the rate ofrespiration in the light by processes other than pho-torespiration,Ci andO are the leaf intercellular partialpressures of CO2 and O2, and τ is the specificityfactor for Rubisco (Jordan & Ogren 1984). The fac-tor 0.5 reflects the fact that for each two oxygena-tion of Rubisco, one molecule of CO2 is released inphotorespiration.

The rate at which Rubisco is carboxylated (i.e.,Vc) depends upon (i) the amount, activity and kineticproperties of Rubisco, (ii) the rate of ribulose-1,5-bisphosphate (RuBP) regeneration via electron trans-port, and (iii) the rate of triose phosphate utilisation(Farquhar et al. 1980 and Harley et al. 1992). WhenAis limited by the capacity of ribulose-1,5-bisphosphatecarboxylase/oxygenase (Rubisco) the dependence ofAonCi is:A = Vcmax((Ci-0∗)/Kc(1+O/K0)+Ci)−Rd whereVcmax is the Rubisco activity,0∗ is the CO2compensation point in the absence of dark respiration,O is the partial pressure of oxygen in the intercellularair space,Kc andK0 is the Michaelis–Menten con-stant for carboxylation and oxygenation by RubiscoandRd is the rate of ‘dark’ respiration in the light.

Rubisco capacity (Vcmax) was calculated assumingthat, atCi values below 40 Pa, the assimilation of

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Figure 2. Mean values of net photosynthetic rate (A) and stomatal conductance (g) at light saturation, for male (light bars) and female plants(dark bars), measured in the field, using a portable compact CO2/H2O porometer (Walz), during different time periods in July and August 1996.Each bar represents the mean values (± sd) of 10–20 measurements per leaf in 3–6 leaves. The same leaves were measured throughout the day.

CO2 was limited solely by the amount, activity andkinetic properties of Rubisco. The parameters,Vc andVcmax were obtained from 3 individual curves and theaverage values for each sex were compared with aStudent’st-test.

Data analyses

Simple linear regressions, nonlinear regressions andStudent’st-tests were conducted with a PC-based soft-ware package (STATISTICA version 5 for Windows).A Gauss-Newton nonlinear estimation routine wasused to estimate the photosynthetic parameters derivedfrom the light curves and CO2-curves. Slope (and el-evation) of regression lines were compared betweenmales and females with a Student’st-test in a fashionanalogous to that of testing for differences betweentwo population means (Zar 1984).

Results

Microclimate during sampling

Daily courses of photon flux density of photosyntheticactive radiation (PFD), water vapour pressure deficit(1W ) and air temperature (T ) during the studied daysin 1996 are shown in Figure 1. July 24 was a rela-tively cloudy day. On this day a strong decline in PFD

was observed from midmorning to afternoon, witha consequent decrease in air temperature and evap-orative demand (1W ). Temperature ranged between19.5◦C and 30.6◦C and1W from 9 to 28 Pa kPa−1.August 1st was a relatively clear day during the morn-ing. A maximum1W (32 Pa kPa−1) was observedaround 13:30, with a corresponding air temperature of29.3◦C. Although similar values of temperature oc-curred in July and August, the vapour pressure deficitwas much higher in August.

Gas exchange and water relations

Photosynthetic rates (A) and stomatal conductances(g) measured in the field reached maximum levels inmidmorning in both sexes and declined throughoutboth studied days (Figure 2). With increasing pho-ton flux density (PFD) during the morning, stomataopened and CO2 was assimilated at a rapidly increas-ing rate (not shown), decreasing in the afternoon.

Daily variation in water potential (9) was simi-lar in both sexes, with a minimum around midday(Figure 3). Male plants recovered more rapidly in theafternoon than female plants.

Female plants always presented lower photosyn-thetic rates (p < 0.001) and lower stomatal conduc-

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Table 1. Summary of the parameters studied in male and female shrubs ofPistacia lentiscus. Maximum photosynthetic rates (Amax), stomatalconductance (gmax) and transpiration rates (T rmax) recorded during the field measurements. Water use efficiency (WUE, calculated asA:Tr),photosynthetic nitrogen use efficiency (PNUE calculated as the ratio between photosynthesis and leaf nitrogen content), minimum water potential(9min) and total chlorophyll content (Chl a+b) are also presented. Mean values±SD of 3–6 leaves taken from each of three shrubs per sex. Alldata were obtained during the summers of 1996 and 1997.

Sex Amax gmax T rmax WUE N PNUE 9min Chl a + ba

(µmol m−2 s−1) (mmol m−2 s−1) (mmol m−2 s−1) (mmol mol−1) (mmol m−2) (µmol mol−1 s−1) (MPa) (mg dm−2)

Male 10.9± 2.7 173.3± 84.6 2.27± 0.69 4.96± 1.08 195.1± 36.3 58.1± 18.1 −2.94± 0.36 2.59± 0.56

Female 6.5± 3.2 98.5± 88.4 1.08± 0.72 7.16± 3.33 174.9± 35.9 38.5± 21.3 −2.62± 0.41 2.21± 0.67

t-test p < 0.05 p < 0.05 p < 0.05 p < 0.05 p = 0.075 p < 0.05 p = 0.1159 p < 0.05

aData from Diaz Barradas & Correia (1999).

Figure 3. Daily course of water potential (9) on male and femaleplants during 24 July and 1 August 1996. Mean values± sd (n = 4leaves) in two plants per sex.

tance than male plants, despite similar exposures tosunlight of the studied leaves.

The relationship between A and g was lin-ear, with a deviation from linearity at g above100 mmol m−2 s−1, for male shrubs. For values belowto 100 mmol m−2 s−1 the relationship between A andg during the studied days was different in male and fe-male shrubs (Figure 4). The slope of linear regressionwas significantly higher (p < 0.001), in females, thusin male plants an A comparable to that of female plantswas associated with a higher g. Under the conditionsof our study, we also observed a 30% greater wateruse efficiency (WUE, expressed as the ratio of the rateA:Tr ) in female than in male plants (p < 0.05) (Ta-ble 1). The relationship betweenA andg is consistentwith these calculations (Figure 4).

Light saturation of net photosynthesis occurred be-fore noon (around 11:00 h, solar time) in both sexes,at a photon flux density of 1500µmol m−2 s−1. Maleplants displayed significantly (p < 0.05) higher val-ues ofAmax , T rmax , gmax and total chlorophyll during

Figure 4. Relationship between instantaneous net photosynthesis(A) and stomatal conductance (g) measured in the field (24 Julyand 1 August 1996) in male and female plants. Linear regression:female plants,A = 0.135g + 0.58, r2 = 0.77; male plants,A = 0.074g+2.04,r2 = 0.88. For all relationshipsp < 0.001. Thetwo slopes of linear regressions calculated with the above modelsare significantly different (t0.001(2), 98 = 6.935,p < 0.001).

the study period (Table 1). Although female plantsshowed slightly lower average values than males intotal leaf nitrogen (N) there were no significant dif-ferences (Table 1).

In male plants a high higer maximum photosyn-thesis rate was associated with higher photosyntheticnitrogen use efficiency (PNUE, the ratio of photosyn-thesis rate to leaf nitrogen concentration) and lowerwater use efficiency (WUE) (Table 1).

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Figure 5. Light response curves of leaves of male and female plants.Data were obtained under controlled laboratory conditions, usinga minicuvette system from Walz, at ambient CO2, air temper-ature 23◦C and1W 11 Pa kPa−1. The calculated parameters,using the proposed model (A = 8I/(1+ 82I2/A2

max)1/2 − Rd )

were for males and females respectivelly:Amax = 8.53±1.98and 6.47±1.88µmol CO2 m−2 s−1, 8 (quantum yield efficiency)= 0.022±0.01 and 0.021±0.01 mol CO2 mol−1 andRd (dark res-piration during photosynthesis)= 0.958±0.19 and 0.853±0.338µmol CO2 m−2 s−1. Mean values±sd from different shrubs(n = 4). There were no significant difference between both sexesin any of the calculated parameters (p > 0.05).

Light and CO2-response models

Figure 5 shows the light response curves obtainedunder laboratory conditions for both sexes. All thenonlinear light response models described the ex-perimental data in a satisfactory manner (R2 variedbetween 96 and 99.8%) and were statistically signif-icant (p < 0.001). Male plants showed slightly higherlight saturated photosynthesis (Amax) although therewere no significant differences between sexes (Fig-ure 5). There were also no significant differences inquantum yield (8) and dark respiration in light (Rd ).The estimated maximum assimilation rates (Amax) fallin the same general order as the maximum values mea-sured in the field (Figure 2, Table 1). Photosyntheticresponses to light suggest that both male and femaleplants only saturated in full sunlight.

The relationship between the rate of photosyn-thesis (A) and the intercellular CO2 partial pressure(Pi) (Figure 6) was used to analyse the photosyn-thetic characteristics of the leaves for both sexes. Theinitial slope of theA vs Pi represents the region ofRuBP carboxylation limitation, whereas at a highPi ,RuBP regeneration limitation occurs. At lowPi values(< 40 Pa), the data were fitted with a Rubisco-limitedequation of photosynthesis to calculateVcmax, whereasat high Pi there were not sufficient data for calcu-

Figure 6. Dependence of the rate of photosynthesis (A), on the in-tercellular CO2 partial pressure (Pi ), of male and female plants.Measured under controlled laboratory conditions, using a minicu-vette system from Walz, at PFD= 1800 µmol m−2 s−1,T = 23◦C, 1W = 11 Pa kPa−1. The calculated parame-ters, using the proposed models(A = Vc(1 − 0.5O/τCi) − Rd,and A = Vcmax((Ci − 0∗)/Kc(1 + O/K0) + Ci) − Rd)

were for males and females respectivelly:Vc = 28.82±1.36and 31.56±7.11 µmol m−2 s−1, and Vcmax = 34±1.9 and40±10.6 µmol m−2 s−1. Data are means±sd of 3 leaves fromdifferent shrubs.

lations. The calculated values ofVc andVcmax weresimilar for both sexes (Figure 6).

Chlorophyll fluorescence

Under laboratory conditions in twigs rewatered overnight the ratio of variable fluorescence to maximal flu-orescence after dark adaptation (Fv/Fm) was around0.80 for both sexes (Table 2) which is a typical valuein the absence of photoinhibition.

The increasing irradiance levels led to a declinein the middayFv/Fm values measured in the field,as compared to the laboratory values. In Au-gust, when water stress was progressively increasing(9min=−3.2 MPa), a moderate decline inFv /Fm wasobserved. No significant differences were observed inFv /Fm between male and female plants. Female plantstended to present lowerFv /Fm.

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Figure 7. The effective quantum yield of PS II (8II, yield) at steady state, in relation to the incident PFD on the leaf, measured with a portablefluorometer PAM-2000 (Walz). Response of female and male leaves were determined on: Laboratory conditions – rewatered twigs from 4 malesand 4 females shrubs, in July 1996 (12/7, 18/7, 25/7, 31/7); Field conditions – during 24 July and 1 August 1996, measurements were made in2 plants per sex in 4–5 leaves per plant. Linear regression of effective quantum yield of PS II (8II, yield) in relation to incident PFD lower than500µmol m−2 s−1are shown in the plot. The slopes are significantly different (p < 0.001) between females and males in laboratory and fieldconditions.

Table 2. Maximum photochemical effi-ciency of PS II (Fv /Fm) of dark-adaptedleaves for 20 min, measured at midday(12:00–15:00 solar time) for male and fe-male plants of Pistacia lentiscusduringthe study period in 1996, using a portablefluorometer PAM-2000 (Walz). Values aremeans of 10 measurements per shrub orper twig± standard deviation (sd). Therewere no significant difference between sexes(p > 0.05).

Day Male Female

12 July 0.79±0.05 0.81±0.01

(laboratory)

25 July 0.80±0.01 0.80±0.01

(laboratory)

31 July 0.81±0.05 0.80±0.02

(laboratory)

24 July 0.78±0.01 0.78±0.02

(field)

1 August 0.69±0.05 0.66±0.07

(field)

The PFD dependence of PS II yield at steady state,as determined for both sexes by chlorophyll fluores-cence measurements in the laboratory and in the field,is shown in Figure 7. No differences between sexeswere detected in the efficiency of PS II electron flow8II (Yield) when the measurements were done in thelaboratory, under optimal conditions. The courses ofthis efficiency display the same shape for both plants.The8II determined in the absence of excess light washigh for both plants, and showed a strong decline atmoderate PFDs. When PFD< 500 µmol m−2 s−1

this decline was more pronounced in male plants (p <

0.001) (Figure 7).Under field conditions (Figure 7), the decline in

efficiency was more pronounced in female plants.The slopes of the linear regressions were significantlyhigher (p < 0.001) in female than in male plants,contrary to what was observed in the laboratory.

Discussion

The physiological differences between sexes wereonly evident in the field studies. For similar cli-matic conditions of PFD,1W , andT , female plants

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exhibited lower values ofA than male plants (Fig-ures 1 and 2). The low photosynthesis in July andAugust, restricted to the morning period reflects thehigh temperatures and water stress characterising thesummer season (Figures 1 and 2). These patterns arein accordance with the behaviour exhibited by severalMediterranean sclerophyllous species during summer(Harley et al. 1987; Tenhunen et al. 1987). Increasing1W in August tended to reduce stomatal conductance,which in turn should also reduceA. This reductionwas greater in male than in female plants, since in Julyfemale plants already showed lower values.

Despite their lower photosynthetic rates, femaleplants had a higher water use efficiency (WUE), sincethey exhibited lower values of stomatal conductanceand transpiration (Table 1). The ratio of CO2 as-similation rate to stomatal conductance (A:g) is amajor determinant of water use efficiency of plants.The higher slopes ofA vs g found in female plants(Figure 4) agree with the calculated WUE and areconsistent with the calculatedPi values. Females ex-hibited lowerPi values than males (18± 7 Pa and23± 6 Pa for females and males respectively) whichwas a direct consequence of lower leaf conductancewith similar rates ofA. According to Schulze & Hall(1982) these differences may be associated with dif-ferential drought adaptations. Consequently, they mayhave great ecological significance for female shrubs.

TheAmax and quantum yield (8II ) measured underlaboratory conditions, in the absence of stress, do notsignificantly differ between the two sexes (Figure 5).These results support the conclusion that differencesin WUE can result only from differences in stomatalconductance (g), with a higher stomatal control forfemale plants. The assimilation values measured in ourstudy,Amax, and8II , are within the range of those ob-served for other Mediterranean species in other studies(Tenhunen et al. 1987; Sullivan et al. 1996).

To achieve the same rate of photosynthesis at lowerstomatal conductance, a high Rubisco activity and ca-pacity for electron transport, and a large concentrationof nitrogen in the leaf may be required. Nevertheless, ahigh WUE in female plants was associated with a lowPNUE and low nitrogen content (Table 1). Hence, ahigher rate of photosynthesis per unit rate of transpira-tion was associated with a lower rate of photosynthesisper unit nitrogen. A trade-off between the efficiencyof the use of water and the use of nitrogen in pho-tosynthesis has also been found in a comparison ofCalifornia evergreen species (Field et al. 1983). This isprobably associated with a greater investment in cell-

wall components of thicker leaves, collenchyma andsclerenchyma elements (Lambers & Poorter 1992),which is in agreement with the higher leaf longevityobserved for female plants ofPistacia lentiscusbyJonasson et al. (1997).

The observed values ofVc andVcmax (Figure 6)are similar to those referred for other sclerophyllousspecies (53± 15µmol m−2 s−1) (Wullschleger 1993).The Rubisco activity (Vcmax) and the rate of carboxy-lation of Rubisco (Vc) was slightly higher in femaleplants, and the rate of electron transport observedunder controlled conditions were similar (data notshown) between sexes, due to similar8II .

Observed values ofFv /Fm for both plants arewithin the range of 0.75–0.85 given by Björkman& Demmig (1987), Demmig & Björkman (1987)and Schreiber et al. (1995) for various species un-der favourable conditions. TheFv /Fm ratio measuredin the field around midday declines relatively to theoptimal values, probably reflecting the influence ofhigh radiation and drought. A diurnal change in theFv /Fm ratio on sunny days has been described forother species, reaching minimum values at noon whenradiation is maximal, with a sustained recovery inlate afternoon (Bjorkman & Powles 1984; Demmiget al. 1989; Long et al. 1994; Werner & Correia1996; Werner et al. 1999). A reduction of the pho-tosynthetic reactions under summer drought stress andhigh photon flux density is a widespread phenomenon,which can be a consequence of both stomatal closure,causing increased constraint on CO2 diffusion, andnon-stomatal limitation such as decreased chloroplastactivity. It has been also pointed out that photoinhi-bition may play a role in regulating photosyntheticbehaviour during summer stress conditions (Demmig1992; Krause & Weis 1991; Long et al. 1994). Underlaboratory conditions, photoinhibition was not foundin either sex (Table 2).

Under optimal conditions female plants ofPistacialentiscushave a similar or even a higher photosyn-thetic capacity than males. But under stress conditions,their photosynthetic capacity declines more than thatof male plants. Their adaptive strategies to limit waterloss during periods of drought result in a restrictionof photosynthetic CO2 uptake and could give rise tothe problem of effective dissipation of excessive radi-ation. The differences in the light response curves ofeffective quantum yield (8II ), recorded under stressconditions with male plants being photosyntheticallythe most active (lower slopes) (Figure 7), are con-

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sistent with the values of A obtained during the fieldstudies (Figure 2).

These data suggest that male plants are rela-tively more tolerant to summer conditions than fe-male plants, maintaining a substantial positive carbonbalance under conditions of severe soil water stressand evaporative demand. In using the Farquhar et al.(1980) model of CO2 assimilation it is important to re-alize that this model only describes the initial fixationof CO2 in the chloroplast by Rubisco. The subsequenttranslocation and allocation of this carbon to differ-ent plant organs and the translation of this to growthare very much separate questions. This is particu-larly important in the case of female shrubs since fruitproduction (maturation) takes place in summer, whenwater stress is greatest. Female plants, with a slightlylower photosynthetic capacity than males, must allo-cate a great percentage of carbon to fruit formation.Jonasson et al. (1997) found that leaf formation wasstrongly reduced by fruiting in female plants ofPista-cia lentiscusand that they compensated the reducedphotosynthetic capacity by retaining older leaves for alonger time than male plants.

We found few consistent differences in the eco-physiological responses of male and female plants atthe leaf-level, when environmental conditions wereoptimal, like under laboratory conditions. We did notfind differences between sexes in models of net as-similation versus light andPi , particularly when themeasurements were done under controlled favourablesituations. However, under field conditions we ob-served significant differences inA andg that can berelated with low water availability during summer. Theleaf intercellular CO2 (Pi ) during field measurements(data not shown) was always lower in female plantsthan in males. BecauseA andg usually decrease si-multaneously during drought, stomatal closure is oftenconsidered the primary physiological response whichresults in decreasedA. From this study, we suggestthat the differences observed between male and femaleplants were due to a stomatal control rather than to dif-ferences in the photosynthetic apparatus activity. Whyis stomatal conductance much lower in female plants?This could be due to higher internal resistances to wa-ter flow or, more probably, to the relatively reduceddimension of the root system, consequence of a re-duced allocation of energy and resources to the rootsystem in female plants. In fact, despite the higherleaf control of water loss by female plants (Figure 2),they reduce their water potential to the same values asmale plants (Figure 3), probably as a result of lower

water availability, caused by specific characteristics ofthe root system or of the conducting xylem. A betterdeveloped root system in male plants seems to be ev-ident from data of Diaz Barradas & Correia (1999),who found a higher number of basal trunks per plantin male individuals, which suggests a higher sproutingcapacity of males after disturbances, probably due toa more extensive root system. This statement couldexplain the male biased sex ratio in closed vegetation,because male plants would be more competitive forwater uptake. These results also show that males havesome advantages under low irradiance, with a lowerslope of8II vs PFD (Figure 7). Male plants have ahigher8II than females at lower PFD. In open vegeta-tion, corresponding to younger communities, probablythe differences between the root system is not verylarge and a higher WUE in female plants might con-tribute to a better competitive ability than for males,and could represent an ecological advantage explain-ing the sex ratio close to 1:1 in this situation. Theseresults are consistent with the observations obtained inothers dioecious species, where the male bias tendedto increase with the population age, only after differ-ences in reproductive has been expressed repeatedly(Delph 1999).

Functional analysis of physiological dimorphismfor this species will have to take the age of the plantsinto account that could give rise to differences ingrowth related to cumulative differences in allocationpatterns.

Future studies should investigate other periods ofthe year, when no environmental stress is imposedupon the plants or experimentally by removing thestress through watering the plants artificially.

Acknowledgements

This research was supported by a TMR grant (Training& Mobility of Researchers, from European Comis-sion) provided to Mary Cruz Dias Barradas. We aregrateful to Graça Oliveira for the stimulating discus-sion and valuable comments in manuscript as well asfor the English correction.

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