Tree Physiology 34, 1102–1117
doi:10.1093/treephys/tpu081
Research paper
Sodium replacement of potassium in physiological processes
of olive trees (var. Barnea) as affected by drought
Ran Erel1,2, Alon Ben-Gal1, Arnon Dag3, Amnon Schwartz2,† and Uri Yermiyahu1,4,†
1Institute
of Soil, Water and Environmental Sciences, Gilat Research Center, Agricultural Research Organization, Mobile Post Negev 85-280, Israel; 2The Robert H. Smith
Institute of Plant Sciences and Genetics in Agriculture, Faculty of Agriculture, Food and Environment, The Hebrew University of Jerusalem, Rehovot, Israel; 3Institute of Plant
Sciences, Gilat Research Center, Agricultural Research Organization, Mobile Post Negev 85-280, Israel; 4Corresponding author ([email protected])
Received June 23, 2014; accepted August 24, 2014; published online October 3, 2014; handling Editor Roberto Tognetti
Potassium (K) is a macro-nutrient understood to play a role in the physiological performance of plants under drought. In
some plant species, sodium (Na) can partially substitute K. Although a beneficial role of Na is well established, information regarding its nutritional role in trees is scant and its function under conditions of drought is not fully understood. The
objective of the present study was to evaluate the role of K and its possible replacement by Na in olive's (Olea europaea L.)
response to drought. Young and bearing olive trees were grown in soilless culture and exposed to gradual drought. In the
presence of Na, trees were tolerant of extremely low K concentrations. Depletion of K and Na resulted in ∼50% reduction
in CO2 assimilation rate when compared with sufficiently fertilized control plants. Sodium was able to replace K and recover
the assimilation rate to nearly optimum level. The inhibitory effect of K deficiency on photosynthesis was more pronounced
under high stomatal conductance. Potassium was not found to facilitate drought tolerance mechanisms in olives. Moreover,
stomatal control machinery was not significantly impaired by K deficiency, regardless of water availability. Under drought,
leaf water potential was affected by K and Na. High environmental K and Na increased leaf starch content and affected the
soluble carbohydrate profile in a similar manner. These results identify olive as a species capable of partly replacing K by Na.
The nutritional effect of K and Na was shown to be independent of plant water status. The beneficial effect of Na on photosynthesis and carbohydrates under insufficient K indicates a positive role of Na in metabolism and photosynthetic reactions.
Keywords: photosynthetic capacity, soluble carbohydrates, stomatal regulation.
Introduction
Potassium (K) is an essential macro-nutrient found in relatively high concentrations in most terrestrial plants. Potassium
plays a central role in the water economy of plants, especially
via osmoregulation and maintenance of cell turgor. Proper K
nutrition is acknowledged to raise plant tolerance to water
stress (Cakmak 2005, Römheld and Kirkby 2010). Positive
effects of K nutrition on drought resistance have been demonstrated in species including hibiscus (Hibiscus rosa-sinensis)
†These
(Egilla et al. 2001), maize (Zea mays L.) (Premachandra
et al. 1991) and sunflower (Helianthus annuus L.) (Lindhauer
1985). Under drought, adequate K levels are vital for maintaining efficient stomatal control, osmotic adjustment, prevention of oxidative damage and high water-use efficiency (WUE,
Shabala and Pottosin 2014). In fact, the beneficial effects of
high K nutritional levels on plants are often reported exclusively under conditions of water stress (Sen Gupta et al. 1989,
Marschner 1995, Cakmak 2005). Potassium is the main
osmoticum causing stomatal movement (Marschner 1995).
authors contributed equally.
The Author 2014. Published by Oxford University Press. All rights reserved. For Permissions, please email: [email protected]
Sodium replacement of potassium in the olive 1103
Under water stress, K-deficient olive (Olea europaea L.) and
sunflower plants failed to regulate stomatal closure (Arquero
et al. 2006, Benlloch-Gonzalez et al. 2008). In addition to
positively effecting tolerance to abiotic stresses, K was shown
to play a crucial role in signaling-stress-causing factors, particularly drought (Anschütz et al. 2014).
Contrary to K, which is acknowledged as an essential
macro-nutrient, sodium (Na) is generally regarded in plant
nutrition for its adverse toxic effects on plants and its antagonistic relationship with K. Nonetheless, in some plant species, Na is apparently capable of at least partially replacing
K. Beneficial effects of Na on plant growth and yield have
long been acknowledged, specifically under conditions of low
K availability. Marschner (1995) classified plants into groups
according to their response to Na: species which can substitute
some of the K by Na and in which Na can sometimes stimulate
enhanced growth even when K is sufficient, and species which
enjoy no benefit from Na at all. Since, under insufficient K, high
Na-accumulating plant species can better regulate cell osmotic
pressure, the presence of Na is expected to enhance drought
resistance (Wakeel et al. 2011) by improved stomatal regulation, higher turgor and enhanced cell expansion. Unfortunately,
there is little solid foundation backing these postulations. In
sugar beet, probably the most studied Na utilizing plant species, Na addition was found to reduce stomatal conductance
(gs) under stress and thus enhance the relative water content
(Hampe and Marschner 1982). In accordance, the halophyte
Aster tripolium was reported to close its stomata in response
to high Na concentration and avoid Na accumulation in guard
cells (Perera et al. 1997, Véry et al. 1998). In the well-studied
glycoside Commelina communis, stomatal response to environmental changes of light, abscisic acid (ABA) and CO2 were
smaller in the presence of Na compared with equivalent K concentration (Jarvis and Mansfield 1980). In addition to enhanced
drought tolerance, K nutrition has a role in the synthesis, transport and accumulation of various metabolites, mainly due to
enzyme regulation (Armengaud et al. 2009). However, the
question whether Na can partly replace K in carbohydrate
(and amino acid) metabolism remains vague. Comprehensive
reviews regarding the nutritional interaction of Na and K are
available in Subbarao et al. (2003), Wakeel et al. (2011) and,
most recently, in Kronzucker et al. (2013).
Although the possible beneficial effect of Na has long been
known, reports quantifying the positive effect of Na on the physiology of perennial evergreen plants are limited to two species
having fundamental physiological and genomic dissimilarity,
cacao (Theobroma cacao) and eucalyptus (Eucalyptus grandis).
Almeida et al. (2010) demonstrated the positive impact of Na
application to eucalyptus trees in K-deficient soil which led to an
increased above-ground biomass. Recently, augmented assimilation capacity and leaf area index were observed in response
to Na application to K-deficient eucalyptus trees (Battie-Laclau
et al. 2013, 2014a). The enhancement in net photosynthesis
rate (A) was partly attributed to higher leaf nitrogen and chlorophyll concentration. Under water stress, eucalyptus trees supplied with K or Na had a higher water demand and thus were
more sensitive to drought (Battie-Laclau et al. 2014b). In cacao
plants, Gattward et al. (2012) also demonstrated a positive
effect of K substitution by Na on assimilation rate and WUE.
Olive trees are grown extensively in their native Mediter­
ranean region. In spite of its widespread traditional cultivation,
knowledge regarding the role of K nutrition in olive physiology
is scarce. Potassium is considered to be particularly important in olive cultivation (Fernandez-Escobar 2010), partially
because of a high correlation between K and yield (Hartmann
et al. 1966). However, the assumption of high requirement of
K for tree health and productive capability lacks solid scientific evidence. Olive is considered to be salt tolerant, depending on the variety (Gucci and Tattini 1997). Salt tolerance in
general (Marschner 1995) and the ability to compartmentalize
Na in cell vacuoles are crucial if Na will function as a nutrient (Wakeel et al. 2011). Hence, plants such as halophytes
(Flowers and Lauchli 1983), which exhibit tolerance to salinity,
are likely able to use Na as an osmoregulant in vacuoles. The
tolerance mechanisms of the olive make it a promising candidate to utilize Na as a nutrient, however no data are available
regarding potential disparate function of Na in olives in general or specifically when K is deficient. Since K is considered
an enhancer of abiotic stress tolerance, the expression of K
deficiency may not materialize except under water stress. For
example, stomatal control of olives subject to K deficiency was
found to differ from K sufficient plants only under water stress
(Arquero et al. 2006, Benlloch-Gonzalez et al. 2008). Thus,
although olive is an important widespread crop grown in harsh
environments, there remains insufficient knowledge regarding
the role of K in elevating drought tolerance and its possible
replacement by Na. Furthermore, while a general supplementary effect of Na has been established, the literature fails to
provide explanations regarding mechanisms for beneficial roles
of Na under conditions of water stress.
The effect of K on growth and productive traits was investigated in a preliminary study (Erel et al. 2008, 2013b). Olive
trees were grown for 4 years in perlite substrate under six
K levels from 0.20 to 5.20 mM. Surprisingly, no considerable
effect of K nutrition on growth, fruit production (Erel et al.
2008, 2013b) or oil composition (Erel et al. 2013a) was
found. Two possible explanations for the lack of response to
K were raised and are the driving hypotheses of the current
study: (i) the absence of water stress and (ii) Na replacement
of K. The objectives of this work were therefore to investigate
the physiological response of olive trees var Barnea to K status and water stress and to examine the possible role of Na
as a substitute for K. This was accomplished by both further
examination of the chosen trees from the original experiment
Tree Physiology Online at http://www.treephys.oxfordjournals.org
1104 Erel et al.
and through an additional study on young trees. The two trials
are complementary and together allow better understanding of
the role of K and possible substitution of Na in olive's response
to drought.
Materials and methods
The first hypothesis, that the role of K nutrition is contingent
on conditions of water stress, was tested using trees from Erel
et al.’s (2013b) preliminary experiment. Olive (O. europaea L.)
trees that had grown outdoors for 4 years under various K
levels were subjected to imposed drought stress and physiological parameters were monitored (EXP-I). Since EXP-I was
originally designed to study solely the role of K+, Na+ was
abundant in the irrigation solution (i.e., 1.8–4.0 mM). The
source of variability in Na concentration was the use of K or Na
as a counter-cation of NO3, therefore high K concentration was
accompanied by low Na and vice versa. In addition, due to the
high water consumption in EXP-I, the water used to prepare
the irrigation solutions was from a local water supplier which
contained ∼0.13 mM K. In order to eliminate the inherited limitations of EXP-I and investigate the second hypothesis that Na
can at least partially replace K in physiological processes in
olive, an additional experiment was run (EXP-II) in which both
K and Na were studied independently in 1-year-old olive plants
in a greenhouse, allowing accurate evaluation of the role of Na
in the presence or absence of K. In EXP-II the source of water
was a reverse osmosis plant providing practically zero background K and Na concentration. Both trials were conducted at
the Gilat Research Center, located in southern Israel (latitude
31°20′N, longitude 34°39′E).
Experimental design and treatments
EXP-I Olive plants var. Barnea were grown in 500 l containers filled with perlite substrate. The experimental setup was
random block designed with three repetitions per treatment.
For 4 years prior to the current study, trees were irrigated
with a solution containing K concentration ranging from 0.20
to 5.20 mM K (Table 1). The main anion in the solution was
NO3− (62 mg l−1 for all treatments) which was accompanied
with either K or Na according to the K level in the solution.
With the exception of K, Na and SO4, the remaining minerals were provided at identical concentrations to all treatments.
During the experiment, developmental and productive traits
were monitored. For detailed experiment description, please
refer to Erel et al. (2013b). Trees were irrigated daily in excess
from September 2006 to July 2010 using nutrient solutions as
described above. Thereafter, eight trees, two from each treatment: T1, T2, T3 and T5 were placed on large scales connected
to a data-logger (CR-1000, Campbell Scientific, Logen, UT,
USA). Each container (tree) was equipped with a 40-l draining
tank which was automatically emptied each morning at 05:00.
Irrigation was scheduled to start at 24:00 to avoid interference
with water uptake measurements. Diurnal water uptake was
monitored for a month. Average measured evapotranspiration
(ET) in July was 98, 92, 126 and 99 l per tree for treatments
T1, T2, T3 and T5, respectively. Meteorological conditions were
monitored at a station located 300 m from the experimental
site. Diurnal solar radiation, air temperature and vapor pressure
deficit (VPD) are plotted in Figure 1. During the experiment
period (July–August 2010) the conditions were fairly stable,
average class A pan evaporation was 8.8 ± 0.92 mm. Water
stress was realized by gradually decreasing irrigation volume
to 75, 50 and 25% of the initial average water uptake of each
individual tree. Midday gs and stem water potential (ψs) were
measured every 2–3 days in order to decide when to employ
each of the drying steps, i.e., only when ψs and gs stabilized.
Each stress step lasted for ∼10 days.
EXP-II One-year-old olive plants var. Barnea were planted
in 60 l containers filled with perlite. Plants were grown in a
controlled greenhouse equipped with wet pad cooling and a
Table 1. Average ± standard deviation of mineral concentration in the irrigation solution during the 4 years of study in EXP-I and the corresponding
dry biomass, total K and Na uptake and leaf K and Na concentrations at the end of the experiment. Shoot was defined as the total above-ground
biomass (leaves, branches, limbs and stem) and root is the below-ground biomass (root neck, big roots and small roots). The type (linear, quadric,
logarithmic [Log] or exponential [Exp]) and significance of best-fit trend lines of plant biomass and mineral partitioning as a function of K level in
irrigation water are noted. n = 3. *P < 0.05, **<0.001, ***<0.0001. N.S., not significant.
Treatment
T1
T2
T3
T4
T5
T6
Mineral concentration (mM)
Cumulative dry weight
(kg tree−1)
K
Na
Shoot
Root
0.20 ± 0.10
0.47 ± 0.09
0.73 ± 0.09
2.06 ± 0.15
2.80 ± 0.19
5.20 ± 0.12
4.02 ± 0.33
3.85 ± 0.17
3.72 ± 0.13
3.25 ± 0.11
2.71 ± 0.14
1.78 ± 0.22
81.2
75.9
91.4
80.2
89.8
73.7
N.S.
42.9
29.1
37.6
37.4
37.7
41.1
N.S.
Tree Physiology Volume 34, 2014
Cumulative K
(g tree−1)
Cumulative Na
(g tree−1)
Leaf concentration
(%DW)
Fruit
Shoot
Root
Shoot
Root
K
Na
21.2
21.2
21.0
20.6
18.8
18.8
N.S.
147
223
442
571
858
857
**Log
47
82
107
245
550
548
*Log
181
150
117
93
89
48
*Exp
263
182
287
265
191
157
N.S.
0.28
0.58
0.93
1.42
1.71
2.20
***Log
0.37
0.19
0.09
0.06
0.05
0.06
**Exp
Sodium replacement of potassium in the olive 1105
Figure 1. Solar radiation, air temperature and vapor pressure deficit (VPD) on dates when physiological parameters were measured. Open air:
Measurements taken from a nearby meteorological station before (solid lines, 27 July 2010) and during (dashed lines, 28 August 2010) imposed
drought. Greenhouse: Measurements taken from the center of the greenhouse before (solid lines, 22 May 2013) and during (dashed lines, 17 June
2013) imposed drought.
Table 2. Average ± standard deviation of mineral concentrations in irrigation solutions during EXP-II and the corresponding leaf K and Na concentrations, dry biomass and stem circumference. Shoot was defined as the total above-ground biomass (leaves, branches, limbs and stem) and
root is the below-ground biomass (root neck, big roots and small roots). Treatment effect was tested by one-way ANOVA (Tukey–Kramer multiple
comparisons test), different letters indicate statistical significance (n = 6).
Treatment
K0-Na0
K0-Na10
K0-Na58
K15-Na0
K100-Na0
K100-Na58
Mineral in irrigation
solutions (mM)
Leaf concentration
(%DW)
Cumulative DW (g tree−1)
K
Na
K
Na
Shoot
Root
Total
0.09 ± 0.03
0.07 ± 0.04
0.08 ± 0.03
0.39 ± 0.04
2.41 ± 0.17
2.48 ± 0.20
0.13 ± 0.04
0.57 ± 0.07
2.42 ± 0.15
0.15 ± 0.04
0.27 ± 0.04
2.84 ± 0.31
0.19c
0.21c
0.20c
0.47b
1.05a
1.01a
0.03c
0.10b
0.19a
0.02c
0.01c
0.01c
2150c
2304c
2533bc
2789ab
2970a
3159a
320b
296b
512a
332b
362ab
496a
2470d
2599cd
3045bc
3130b
3332ab
3655a
removable thermal screen cover. Diurnal solar radiation, air
temperature and VPD are plotted in Figure 1. After 3 months
of acclimation, trees were pruned to 65-cm single shoots with
20 leaves per plant. Differential nutrient application treatments
were initiated immediately after pruning. Treatment K and Na
concentrations and nomenclature are given in Table 2. Three
treatments contained no added K and increasing levels of Na
with target concentrations of 0 mg l−1 Na (K0-Na0), 10 mg l−1
Na (K0-Na10) and 58 mg l−1 Na (K0-Na58). Three treatments
contained no added Na and increasing levels of K with target
concentrations of 0 mg l−1 K (K0-Na0), 15 mg l−1 K (K15-Na0)
and 100 mg l−1 K (K100-Na0). A control treatment contained
Stem
circumference
(mm)
96c
98bc
105b
116a
123a
121a
high levels of K and Na with target concentrations of 100 mg l−1
K and 58 mg l−1 Na (K100-Na58). Divergence from target concentrations was mostly due to the impurity of the salts and
traces of minerals in the desalinated irrigation water. The experiment used a ‘Latin square’ design (balanced rows and columns)
with six replicates. During the growth period trees were pruned
three times to comparable size and water uptake. Target concentrations were achieved by proportionally dissolving commercial grade salts of Ca(NO3)2, NH4H2PO4, KCl, MgSO4, Mg(Cl)2
and NaCl. Differences between treatments were exclusively as
K, Na and Cl concentrations. Micronutrients (Fe, Mn, Zn, Cu, Mo
and B) were applied at a rate of 0.1 ml l−1 of commercial solution
Tree Physiology Online at http://www.treephys.oxfordjournals.org
1106 Erel et al.
(‘Koratin + B’, Fertilizers and Chemicals Ltd, Haifa, Israel). For
each treatment, salts were dissolved in desalinated water with
electrical conductivity <50 µS m−1. Average measured mineral
concentrations of specific elements were: Ca: 2.0 ± 0.1 mM,
Mg: 1.2 ± 0.2 mM, NH4: 0.27 ± 0.04 mM, NO3: 4.6 ± 0.1 mM
and P: 0.30 ± 0.02 mM. The pH was in the range of 5.7–5.8.
Trees were irrigated daily in excess using the described
solutions for 15 months. In a preliminary trial, irrigation was
discontinued in 10 similar olive pots. The pots were weighed
daily until visual stress symptoms (wilting) appeared. The
amount of water lost during the drought period averaged
12.0 l per pot. This amount of water, between pot capacity
and wilting, was determined to be the ‘plant available water’
for subsequent trials. In May 2013, drought was imposed by
gradually decreasing the plant available water by 4 l pot−1 in
three steps (8, 4 and 0 l pot−1). In order to achieve accurate
and identical water content for each plant, pots were weighted
daily early in the morning and the volume of available water
was determined by comparing with the pot’s weight at capacity accordingly: AW = 12 − (Wpc − Ws) where AW is the available water, Wpc is the pot weight after excesses irrigation and
drainage (weighed at dawn) and Ws is the pot weight early in
the morning during stress. Immediately following daily weighing, water was manually supplied to reach the target weight
for each pot if necessary. Each stress step was carried out
over a week. During the periods of imposed stress, morning
stomatal conductance and leaf water potential were determined twice a week in order to monitor plant water status. The
average daily ET before initiation of drought (26 May 2013)
was 4.8 ± 0.42 l per pot.
Mineral concentration, partition and biomass
Leaf analysis was performed according to the protocol formerly
used for olive as described in Erel et al. (2013b). Upon concluding the trials, plants were removed and divided into leaves,
branches, limbs, stems, root necks, thick roots (>10 mm diameter) and fine roots (<10 mm diameter). Each plant fraction
was sampled, weighed, dried and ground. Mineral concentrations were determined as described for leaves. For simplicity, in Tables 1 and 2 the dry weight (DW) and mineral
uptake rates and concentrations of combined total shoot and
root (total above- and below-ground biomass) are presented.
In EXP-I, fruits were harvested and weighed in October each
year. Fruit DW content was determined by oven drying of fruit
paste. Stem circumference was measured ∼10 cm above the
substrate surface.
Gas exchange, chlorophyll fluorescence and water
potential
Net CO2 assimilation rate (A) and stomatal conductance of H2O
vapor (gs) were measured simultaneously by a portable gas
exchange measuring system (LI-6400–40, LI-COR, Inc., Lincoln,
Tree Physiology Volume 34, 2014
NE, USA). In general, chamber conditions were set to an environmental CO2 concentration of 400 µmol CO2 mol−1, a photosynthetic photon flux density (PPFD) of 1000 µmol mol m−2 s−1,
a humidity of 10 mmol H2O mol−1 and a block temperature of
25 °C. Measurement periods predawn and after sunset were
taken in accordance with the environment natural light with
chamber light turned off (PPFD = 0). Electron transport rate
(ETR) was measured using a portable chlorophyll fluorometer,
Mini-PAM (Walz, Effeltrich, Germany) in EXP-I and OS5p (OptiSciences, Hudson, NH, USA) in EXP-II. For both gas exchange
and chlorophyll a fluorescence (F), measurements were taken
from fully illuminated leaves (except early in the morning and
after sunset). During the day, ETR was measured only if PPFD
≥500. Firstly, F was logged and immediately after saturation a
pulse was illuminated in order to log maximal fluorescence (Fm′).
Electron transport rate was calculated accordingly: (Fm′ − F)/
Fm′ × 0.5 × PPFD × 0.875. Stem and leaf water potential (ψs and
ψl) were measured in a pressure chamber (Arimad-3000, MRC,
Holon, Israel). The chamber flow rate was set to 0.03 MPa s−1.
EXP-I Diurnal measurements were taken twice during the
experiment: a day before imposing drought stress on 27 July
2010 and during the stress on 29 August 2010. Measurements
were taken from sunrise until after sunset at ∼2-h intervals. Gas
exchange measurements were made on leaves on the tree’s
periphery, five leaves per tree and two trees per treatment. ψs
was measured on two shoots per tree as described by Naor
et al. (2006) on shaded shoot endings containing six to seven
leaves which were covered by aluminum bags at least 2 h in
advance.
EXP-II Two diurnal measurements were performed before
and during drought stress (22 May and 17 June 2013, respectively) on five replicate trees of treatments K0-Na0, K0-Na10,
K0-Na58 and K15-Na0, and the control. K100-Na0 was essentially identical to the control in regards to leaf mineral concentrations and was therefore not measured. First measurements
were taken before sunrise and the last set was carried out after
sunset. Gas exchange was measured on three to four leaves
per tree and ψl on one uncovered shoot per tree, each at ∼2-h
intervals.
Starch and soluble carbohydrate determination (EXP-II)
Starch and soluble carbohydrates (SCH) were determined a
day before drought was initiated. Treatment K100-Na0 was
excluded. Leaves were collected and immediately frozen in liquid nitrogen, lyophilized, milled and stored at −20 °C. Soluble
carbohydrates were extracted as follows: 20–25 mg leaves
were extracted three times with 80% ethanol at 80 °C. The
extracts were evaporated to dryness and re-dissolved in 4 ml
of double distilled water from which aliquots of 500 μl were
lyophilized. The dried material was re-dissolved and derivatized
Sodium replacement of potassium in the olive 1107
for 90 min at 37 °C (in 40 μl of 20 mg ml−1 methoxyamine
hydrochloride in pyridine), followed by a 30-min treatment
with 70 μl of N-methyl-N-(trimethylsilyl) trifluoroacetamide at
37 °C and centrifugation. After derivatization, the SCH was
analyzed via Agilent 6850 GC/5795C MS (Agilent Technology,
Palo Alto, CA, USA). One microliter was injected in splitless
mode at 230 °C. Helium carrier gas was used at a flow rate of
1 ml min−1. Chromatography was performed using a HP-5MS
capillary column (30 m × 0.250 mm × 0.25 μm). Mass spectra were collected at 2.4 scans s−1 with an m/z 50–550 scan.
Mass spectrometry (MS) temperature was set to 230 °C. Both
ion chromatograms and mass spectra were evaluated using
MSD ChemStation E.02.00.493 software. Carbohydrates were
identified by comparison of retention time and mass spectra
with those of authentic standards (sigma) and each SCH was
quantified using a calibration curve.
Starch content was determined by digesting ethanol insoluble
residue with amyloglucosidase as described in Vishnevetsky
et al. (2000). Glucose in solution was determined by colorimetric reaction with dinitrosalicylic acid according to Miller (1959).
Data analysis
EXP-I Relationships between plant biomass and mineral partitioning to the K level in irrigation water were tested using
Sigmaplot 10.0 software (Systat Software, San Jose, CA, USA).
Linear, quadric, logarithmic or exponential trend lines were fitted to data. The type of best fit regression and significance are
presented in Table 1.
EXP-II Effect of treatments on plant biomass, stem circumference, Fv/Fm, SCH and starch were analyzed by one-way
analysis of variance (ANOVA, Tukey–Kramer multiple comparisons test) using JMP 7 statistical software (SAS Institute, Cary,
NC, USA). The diurnal measurements were integrated over the
day for each individual tree, assuming continuity between the
measured points. Instead of gs, transpiration rate was used for
evaluation. The sum of daily A, transpiration and e− transport
for each tree was analyzed using ANOVA as described above.
The regression of gs and A was plotted using SigmaPlot (Systat
Software, Inc.) along with 95% confidence levels. The relationship between gs and ψl was plotted using SigmaPlot using data
measured during the stress development stages.
Results
EXP-I
Cumulative biomass, yield and minerals Olive trees were
grown in an inert substrate for 4 years under a wide range of
K in irrigation solutions. Nonetheless, accumulated growth of
all treatments was similar: i.e., 124 and 115 kg DW per tree for
T1 and T6, respectively. Likewise, no effect of K on biomass
partitioning was noted (Table 1). Cumulative dry fruit yield was
also independent of treatments. Potassium in the shoot and
root increased in a logarithmic fashion reaching maximum at
the two highest K levels: T5 and T6. The K content in shoots of
the high K treatment was more than fivefold greater than that
of T1. In the roots, the high K treatments accumulated more
than 11 times greater K. Sodium accumulated in the shoots at
relatively high levels in the lowest K treatment and decreased
exponentially as K in the irrigation solution increased. Sodium
accumulation in roots was variable and was not significantly
affected by K level. As described for shoot K, leaf K concentration increased in a logarithmic fashion with K in the irrigation
solution rising from 0.28% up to 2.20%. Treatment T1 had K
concentration below the commonly accepted K deficiency level
of 0.4% (Fernandez-Escobar 2010). Potassium in T2 leaves fell
between 0.4% and the correspondingly accepted sufficiency
threshold of 0.8%. Sodium leaf concentration was inversely
correlated to leaf K, decreasing exponentially from 0.37% in
T1 down to ∼0.06% for T4–T6. Thus, the T1 treatment had
significantly higher Na content compared with K in leaves. Leaf
Na concentration was almost double the accepted threshold
for toxicity of 0.2% (Fernandez-Escobar 2010). Nonetheless,
this did not reduce growth, even after 4 years.
Diurnal physiological parameters of stressed and nonstressed trees Diurnal measurements of non-stressed
plants were collected on 27 July 2012 and drought stressed
measurements on 29 August following gradual drought. Both
days were clear and hot, typical to the arid climate of the
research center (Figure 1). Both diurnal sets, stressed and
non-stressed, are presented in Figure 2. For non-stressed
trees, regardless of the K level, A rose from nearly null early
in the morning to a maximum of ∼15 µmol m−2 s −1 at 10:30
(Figure 2). During the day, A was quite stable until 17:00
to sunset when it decreased. Stressed trees had considerably lower A during the day. Maximum A was measured at
09:10 followed by a gradual decrease towards the afternoon
and a minor peak around 18:30. Under both stressed and
non-stressed conditions, no significant difference in A due
to K was found. In non-stressed trees, gs increased from
early in the morning until 10:30 and remained fairly stable
(0.26 mol H2O m−2 s −1) until 17:10, followed by a sharp
decline towards sunset (Figure 2). Stressed trees exhibited very low gs which peaked at around 9:00 to a value of
∼0.1 mol H2O m−2 s −1 and later dropped to extremely low
values. Again, K level had no effect on gs in both cases.
Diurnal ETR of well-irrigated trees of T1 and T2 started from
almost zero in the morning to ∼120 µmol e− m2 s −1 compared with ∼140 for T3 and T5. This difference was not
significant. Electron transport rate remained stable most of
the day (Figure 2). Electron transport rate of the stressed
trees peaked at 10:30 (∼76 µmol e− m2 s −1) and gradually
decreased until the end of the day. The ψs was substantially
Tree Physiology Online at http://www.treephys.oxfordjournals.org
1108 Erel et al.
Figure 2. Average (symbols) and standard deviation (error bars) of daily measurements of assimilation rate (A), stomatal conductance (gs),
electron transport rate (ETR) and stem water potential (ψ s) of fully irrigated (continuous lines, empty symbols) or drought stressed trees (dashed
lines, full symbols) of olive trees grown for 4 years under different K levels: 0.2 mM (circles), 0.5 mM (diamonds), 0.7 mM (triangles) and 2.8 mM
(squares) in EXP-I.
affected by irrigation level (Figure 2). In the non-stressed
trees, ψs was −0.7 MPa early in the morning, decreased
to −1.8 by midday and recovered to −1.0 around sunset.
Potassium level did not affect ψs. In the stressed trees, ψs
decreased continuously from −1.7 MPa at dawn to −4.5 in the
afternoon and evening. T2 had the lowest ψs for most of the
day, however, differences were insignificant.
EXP-II
Biomass and mineral content In addition to the difference
in tree size and age, EXP-II differed from Exp-I, in part, due
to the manipulations made on K levels with or without Na.
Leaf K increased as a function of K in irrigation water from
0.19 to 0.20%, when no K was added, to slightly >1% in
response to 2.5 mM K in solution (Table 2). When no K was
added, Na concentration in leaves increased as a function
of Na in solution from 0.03 to 0.19%. In the presence of K,
Na was almost completely excluded from leaves. Treatments
K0-Na58 and K100-Na58 (control) received similar Na levels
Tree Physiology Volume 34, 2014
in their irrigation solution but had entirely different effects on
leaf Na concentration, 0.19% if no K was added compared
with 0.01% at the highest K level. Shoot biomass production
(including pruned branches) tended to increase with Na level
with no statistical difference (Table 2). The three no-K treatments produced significantly lower shoot biomass compared
with K100-Na0 and the control. Root biomass was highest in
Na58-K0 and the control, significantly higher than Na0-K0,
Na15-K0 and K15-Na0. When no K was added, total produced
biomass significantly increased with Na, but did not reach the
levels of the trees which received K. Maximum biomass was
found in the control. Stem circumference was comparable to
plant biomass, showing a significant increase in response to
Na in K-depleted plants and a maximum value for the highest
K levels (Table 2).
Starch and SCHs in leaves Starch and SCHs were determined in May before imposing drought. Of the eight indentified SCHs, glucose was the most dominant followed by
Sodium replacement of potassium in the olive 1109
mannitol and sucrose; the three composed ∼90% of the total
SCH. Potassium and Na nutrition significantly affected six
of the eight SCHs. The main trend of elevated K was found
to be an increase in mannitol at the expense of sucrose.
Galactose, myo-inositol and galactinol significantly decreased
with increased K while raffinose was increased. The total
SCH level, however, was not modified by treatment and fluctuated ∼120 mg g−1. Leaf starch concentration was significantly higher in high Na and high K treatments: 86 mg g−1,
compared with a mere 66 mg g−1 for Na0-K0 (Table 3). The
intermediate K and Na treatments had intermediate starch
concentrations.
Diurnal physiological parameters of stressed and nonstressed trees Diurnal measurements of non-stressed plants
for EXP-II were collected on 22 May 2013 and stressed
measurements on 17 June. Both days were relatively warm
(Figure 1). The diurnal measurements are presented in
Figure 3. In order to improve visualization, results from treatment K0-Na10 are presented in the summary table (Table 4)
but not in Figure 3. Under conditions of high water availability,
A was considerably affected by Na and K levels (Figure 3).
Throughout the day, treatment K0-Na0 had the lowest A,
roughly half of the control. The addition of Na to zero-K trees
(K0-Na58) recovered most of the A, although it remained
lower than the control. The A of treatment K15-Na0 was quite
similar to the control. Stomatal conductance and ETR diurnal
curves generally followed those of A (Figure 3). Predawn gs
was notably high for all treatments even though measurements
were taken in the absence of significant PPFD radiation and
with the chamber internal light turned off. The zero-K treatments, K0-Na0 and K0-Na58, exhibited the lowest gs between
09:00 and 14:00 but not at dawn or later in the afternoon.
After sunset, maximal gs was measured in treatment Na0-K0.
Potassium and Na nutrition had a pronounced effect on ETR.
During daylight hours (09:00–17:00) ETR was mostly stable,
averaging 141, 143, 116 and 79 µmol e− m−2 s−1 for control,
K15-Na0, K0-Na58 and K0-Na0, respectively. All treatments
exhibited identical and typical ψl diurnal curves, starting at
−0.7 MPa at dawn, decreasing to a minimum of −1.5 MPa at
around 14:00 and partly recovering in the evening, post dusk,
hours.
Subsequent to prolonged gradual drought stress, A, gs,
ETR and ψl each declined considerably (Figure 3). A and gs
in stressed trees were highest at 09:00, gradually depleted
as the day advanced and demonstrated occasional minor
peaks in the afternoon between 16:00 and 18:00 (Figure 3).
Except for ψl, the nature of the response to nutrition (but not
the actual values) was similar in irrigated and drought conditions. Assimilation rate, gs and ETR were higher in K15-Na0
and the control in the morning and through most of the day. A,
gs and ETR were generally lowest in K0-Na0 while K0-Na58
had either lower or similar values compared with the control. ψl
was notably affected by K and Na levels under drought stress
(Figure 3). Throughout the day, the lowest ψl was measured
for the control, followed by K15-Na0, K0-Na58 and K0-Na0.
Regardless of the nutritional level, ψl was highest in the morning, lowest around 14:00 and subsequently partially recovered
towards the night.
Cumulative daily A, transpiration, ETR and WUE In order
to interpret the diurnal A, transpiration and ETR, each curve
was integrated to provide accumulated daily values of assimilation, transpiration and e− transport of each individual tree.
Additionally, cumulative A was divided by transpiration to
yield intrinsic WUE (Table 4). Under irrigated conditions, total
daily assimilated CO2 was significantly lower in K0-Na0 and
K0-Na10: 0.40–0.47 mol m−2 day−1, compared with 0.75–
0.78 in the control and K15-Na0. Total daily assimilated
CO2 in treatment K0-Na58 lay roughly in between, significantly higher than K0-Na0 and K0-Na10 but lower than the
K treatments. Accumulated CO2 assimilation decreased more
than three times under drought. However, the relative effect
of treatments remained more or less the same. The diurnal transpired H2O of non-stressed trees was the lowest in
K0-Na0, significantly less than K15-Na0 and the control but
not than K0-Na58 and K0-Na10. Under drought, transpiration
dropped by a magnitude of ∼6 and K0-Na0 was the lowest,
although not significantly different. The effect of treatments
on diurnal e− transport strongly resembled that reported for
Table 3. Average SCHs and starch concentrations in leaves 11 months after treatment initiation in EXP-II. Treatment effect was tested by one-way
ANOVA (Tukey–Kramer multiple comparisons test), different letters indicate statistical significance (n = 5).
SCHs and starch concentration (mg g−1)
K0-Na0
K0-Na10
K0-Na58
K15-Na0
K100-Na58
Treatment effect:
Glucose
Mannitol
Sucrose
Fructose
Galactose
myo-Inositol
Raffinose
Galactinol
Total SCH
Starch
54.0a
57.1a
55.3a
60.7a
51.3a
0.8492
17.4b
18.7b
22.6ab
26.2a
26.1a
0.0005
23.6a
22.0ab
23.4a
15.9ab
15.2b
0.0097
3.7a
3.8a
3.2a
3.4a
3.0a
0.3048
2.9a
3.2a
3.0a
2.5ab
1.3b
0.0009
2.8a
2.2ab
2.4ab
1.8ab
1.6b
0.0099
2.1b
2.3ab
2.8a
2.5ab
2.5ab
0.0165
12.1a
10.2ab
10.9a
7.6ab
5.3b
0.0055
119a
119a
124a
121a
106a
0.6833
66b
69ab
86a
78ab
86a
0.0011
Tree Physiology Online at http://www.treephys.oxfordjournals.org
1110 Erel et al.
Figure 3. Average (symbols) and standard deviation (error bars) of daily measurements of assimilation rate (A), stomatal conductance (gs),
e­ lectron transport rate (ETR) and leaf water potential (ψl) of fully irrigated (left column) or drought stressed (right column) trees with different K
and Na nutrition. No added K and Na (circles); No K and 58 mg l−1 Na (triangles); 15 mg l−1 K and no Na (diamonds); 100 mg l−1 K 58 mg l−1 Na
(squares).
assimilation, i.e., lowest in K0-Na0, higher in K0-Na58 and
highest in K15-Na0 and control. Under drought, daily accumulated e− transport was much lower but the response to
the treatments was similar. Water-use efficiency was much
higher under drought. For example: 5.0 compared with
7.8 mmol CO2 mol−1 H2O in the control. Under irrigation, the
Tree Physiology Volume 34, 2014
low K and Na treatments, K0-Na0 and K0-Na10, had significantly lower WUE. Under drought, all treatments had roughly
similar WUE, an average of 8.0 mmol mol−1.
Maximum quantum yield of PSII (Fv/Fm) Under fully
irrigated conditions, predawn Fv/Fm was high (0.82–0.83)
­
Sodium replacement of potassium in the olive 1111
Table 4. Integrated daily assimilation, transpiration, ETR and intrinsic WUE under irrigation and drought regime. The diurnal measurements were
integrated by summing the additive values over the day for each individual tree (n = 5) assuming continuity between the measured points.
Treatment effect was tested by one-way ANOVA (Tukey–Kramer multiple comparisons test), different letters indicate statistical significance.
Treatment
Irrigation
K0-Na0
K0-Na15
K0-Na58
K15-Na0
K100-Na58
Drought
K0-Na0
K0-Na15
K0-Na58
K15-Na0
K100-Na58
Daily assimilation
(mol CO2 m−2)
Daily transpiration
(mol H2O m−2)
Daily ETR
(mol e− m−2)
WUE
(mmol CO2 mol−1 H2O)
0.40c
0.47c
0.62b
0.78a
0.75a
123b
137ab
134ab
159a
152a
3.2c
3.7cb
4.7b
5.8a
5.8a
3.3b
3.4b
4.6a
4.9a
5.0a
0.13B
0.12B
0.17AB
0.26A
0.22AB
16.8A
17.4A
19.8A
27.7A
28.3A
0.59B
0.63AB
0.79AB
1.01A
0.91AB
7.6A
6.6A
8.8A
9.3A
7.8A
retarded growth, ‘willow-like’ branching and, eventually, typical
necrosis spreading from leaf tips to edges (Figure 5). All symptoms were remarkably reduced by the addition of Na to zero-K
plants.
Discussion
General response of olive to limited K and its
substitution by Na
Figure 4. Predawn maximum PSII quantum yield (Fv/Fm) before (black
bars) and during (gray bars) imposed drought stress. Treatment effect
was tested by one-way ANOVA (Tukey–Kramer multiple comparisons
test), different letters indicate statistical significance (n = 6).
for all treatments except K0-Na0 and K0-Na10 (0.77–0.78)
(Figure 4). Under drought, predawn Fv/Fm was impaired in
all treatments (Fv/Fm < 0.80), indicating photodamage. Yet,
Fv/Fm was lowest for K0-Na0 and was significantly higher in
the high Na treatment.
Visual symptoms For the first 3 years in EXP-I, trees could
not be visually distinguished according to their K level. In the
fourth year, T1 exhibited willow-like branch growth patterns.
Subsequently, by the end of the experiment, visual examination
revealed occasional leaf tip necrosis. The appearance of ‘tip
burn’ was likely due to Na toxicity rather than K deficiency as
leaf Na (Table 1) reached approximately double the accepted
threshold values for olives (Fernandez-Escobar 2010). In
EXP-II, visual evidence of K deficiency, chlorotic leaves in the
K0-Na0 treatment, was first noticed ∼4 months after treatment
initiation. Thereafter, symptoms were accelerated, trees showed
Potassium is understood to play many vital roles in plant physiology, including osmotic regulation, enzyme activation, electrochemical balance, phloem and xylem transport and stress
signaling (Wakeel et al. 2011, Shabala and Pottosin 2014).
Typical K deficiency symptoms are characterized by retarded
cell growth, accumulation of simple carbohydrates, lower
chlorophyll content, photosynthesis inhibition, and eventually,
reduced growth and productivity (Römheld and Kirkby 2010).
In the current study, the physiological response of olive to K
levels and drought was investigated in the presence of ample
Na on bearing trees (EXP-I) or under manipulated levels of
Na on young trees (EXP-II). The data obtained indicates relatively low demand of the olive tree for K, especially in the
presence of Na. In most plants, K is needed in concentrations
comparable with those of Na, typically in the range of 2–5% of
plant DW. In contrast, in species that are capable of replacing
K by Na, adequate leaf K requirement is much lower, 0.5–1.0%
(Marschner 1995). In the olive, in the presence of Na, leaf K
dropped to 0.3% with no appearance of visual symptoms and
with no negative effect on growth and yield as demonstrated
in EXP-I. Under the conditions of EXP-I, leaf Na was more than
double compared with leaf K. Olive is considered to be salt
tolerant, depending on the variety (Gucci and Tattini 1997)
and its tolerance to salinity is accredited mainly to the ability
to exclude and retain salt ions in roots (Chartzoulakis 2005)
and therefore might be considered as an ‘excluder’. However,
when K is limited Na is accumulated in the leaf and augments
Tree Physiology Online at http://www.treephys.oxfordjournals.org
1112 Erel et al.
Figure 5. Typical appearance of trees with no K and no Na (left), no K and 2.5 mM Na (center) and 2.5 mM K and Na (right) in the irrigation solution. Picture was taken a year after treatment initiation.
physiological performance, as seen in both EXP-I and EXP-II.
Hence, the olive may also be considered an ‘includer’, i.e., having the ability to accumulate high amounts of Na ions in shoots
(Marschner 1995). The choice of salt translocation mechanism, ‘includer’ or ‘excluder’, is largely dependent on the olive K
level. In the presence of adequate K, Na is excluded from olive
shoots. Only when K is limited, Na is transported to the shoots
and substitutes for K. The ability of the olive to flourish under
conditions of extremely low environmental K levels indicates an
exceptional ability to utilize K or replace it with Na, probably as
a result of physiological adaptation to shallow, rocky infertile
soils common in its natural habitat (Fernández 2014). It is of
little surprise, therefore, that the olive tree is considered to
have a low nutrient demand. The combination of low soil fertility and close vicinity of the olive habitat to the Mediterranean
sea likely drove adaptation mechanisms enabling the use of Na
when the supply of K is short, as found previously for eucalyptus (Almeida et al. 2010).
Plant water status and gas exchange parameters The olive
is morphologically and physiologically well adapted to long periods of drought (Fernández 2014). The physiological response
patterns of olive to drought in the current experiment were
comparable to those previously reported (Angelopoulos et al.
1996, Moriana et al. 2002, Bacelar et al. 2007, Diaz-Espejo
et al. 2007, Ben-Gal et al. 2010). Subsequent to drought,
the peaks of gs and A are early in the morning when WUE
is highest (Moriana et al. 2002, Testi et al. 2008) followed
by a substantial reduction of all parameters for the remainder of the day. Similar diurnal patterns of gas exchange and
water status parameters were found in the current study’s two
experiments. Nonetheless, absolute values were significantly
affected by tree nutritional status of extreme K deficiency and,
Tree Physiology Volume 34, 2014
even more significantly, by limiting Na. Under low supply of
0.20 mM K and sufficient Na (4.0 mM), as demonstrated in
EXP-I, the measured physiological response to drought was
almost unchanged, only ETR seemed to be impaired by low K.
Most importantly, in contrast to our initial hypothesis, neither
K nor Na nutrition enhanced drought resistance of the olive. In
eucalyptus, K and Na nutrition resulted in more severe drought
stress compared with unfertilized control due to rapid decline
of soil water (Battie-Laclau et al. 2014b). However, the fundamental difference between the current and the Battie-Laclau
studies is the way drought was controlled. In the Battie-Laclau
et al. (2013) study, plants were rainfed whereas in the current
study water availability was maintained similar among plants
by monitoring and supplying water differentially. In any case,
the current study joins the former in challenging the generally
accepted understanding that K elevates drought tolerance in
plants (Cakmak 2005, Römheld and Kirkby 2010). The main
impact of K and Na deficiency found in the current study was
pronounced only at the lowest K treatment level, where A, gs
and ETR were notably impaired, regardless of the water availability conditions. Unlike the gas exchange parameters, ψl was
considerably affected by K and Na nutrition under drought.
Leaf or stem water potential is considered to be an accurate
indicator for water stress in many fruit trees (Naor et al. 2006).
Thus, a possible interaction between ψl and environmental or
intrinsic factors is of high interest. Fruit load has been shown
to affect ψs under both stressed and non-stressed conditions
in grapevines (Naor et al. 1997) and in olives (Ben-Gal et al.
2011, Trentacoste et al. 2011, Naor et al. 2013). The present study indicates that K and Na nutritional levels also influence the absolute values of ψl, but only under drought. Leaf
water potential was highly dependent on the K and Na nutritional status (Figure 3). In general, a higher concentration of
Sodium replacement of potassium in the olive 1113
monovalent cations was correlated with lower ψl. The lower
ψl may have been a result of the higher gs and higher transpiration leading to tissue dehydration. Similar to our findings,
K and Na were found to reduce ψl in eucalyptus trees during the dry season (Battie-Laclau et al. 2013, 2014b). In the
rainy season, predawn leaf water potential was similar for low
K, high K or high Na trees. As mentioned, the lower ψl may
be a result of enhanced water uptake of K sufficient trees.
Additionally, the differences in ψl may be a result of leaf morphology and hydraulic properties and due to the effect of K
and Na on leaf area. In eucalyptus, parenchyma thickness and
total intercellular space increased in response to K and Na
nutrition (Battie-Laclau et al. 2014a). Furthermore, bulk elastic modulus (ε) has a role in plant water relations (Saito et al.
2006). In eucalyptus leaves, ε was significantly affected by K
and Na nutrition (Battie-Laclau et al. 2013). The difference in
ψl between trees with comparable water availability is important for broader understanding of ψl as a stress indicator. As
stated above, fruit load has been shown to affect ψl by general decrease in ψl with elevated fruit load. As a result, olive
trees with high fruit load had higher gs for a given ψl (Naor
et al. 2013). In a similar manner, K and Na nutritional levels
were shown in the present study to impact the gs–ψl interaction (Figure 6). Throughout the range of ψl, gs was lower in
the deficient treatment, higher in response to Na application
and highest in the control. Hence, K and Na nutrition levels
should be taken into account when interpreting ψl.
Discussed widely by Kronzucker et al. (2013), the possible
role of Na in stomatal movement is of high interest, but as
Figure 6. Relationship between leaf water potential (ψl) and stomatal conductance (gs) for the four extreme treatments: K0-Na0 (filled
circles), K0-Na58 (triangles), K15-Na0 (diamonds) and K100-Na58
(open circles). Data were obtained on five selected days during the
period of imposed water stress with the addition of the two diurnal
measurements. All data were collected between 08:30 and 12:00.
Each point represents an average ± standard error of five trees per
treatment. Exponential regression was fitted to each treatment.
yet undetermined. The similar response to drought of olive
trees with various K and Na levels indicates that their ability
to regulate transpiration according to water availability was
not impaired, even under severe K deficiency. The lack of
response of stomatal movement (not conductance) to either
K or Na is surprising as K is considered the main ion accumulating in guard cells (Marschner 1995) and since the K level
has previously been shown to effect stomatal regulation in
olives (Arquero et al. 2006, Benlloch-Gonzalez et al. 2008).
Simple carbohydrates such as sucrose also play a substantial role in stomatal movement (Talbott and Zeiger 1998) and
stomatal regulation through feedback inhibition (Kelly et al.
2013). Sodium was also shown to enable stomatal opening
in detached epidermis of C. communis (Willmer and Mansfield
1969, Jarvis and Mansfield 1980). Thus, in K-deficient olive
trees, either Na or SCHs may be responsible for the adjustment of guard cell osmotic pressure allowing appropriate
stomatal opening. Sodium, however, is expected to be less
efficient in stomatal response to environmental changes, such
as light, CO2 and ABA (Jarvis and Mansfield 1980) since the
presence of Na causes impaired stomatal closing. In the halophyte A. tripolium, even under high leaf Na concentration, Na
was excluded from guard cells and not found to replace the
role of K (Perera et al. 1997). The results presented here for
olives suggest that stomatal response to drought remains
normal even at extremely low K levels with or without Na.
Although stomatal regulation was not impaired in the current
study, the absolute values of gs tended to be lower under K
deficiency, whether under drought or not. Likewise, BattieLaclau et al. (2014a) found greater gs in eucalyptus trees
exposed to high K and high Na. The negative effect of K deficiency on gs may be regulated by the ‘stress hormone’ ABA, a
major regulator of stomatal movement (Blatt 2000). Abscisic
acid is produced in roots in response to drought and increases
exponentially as soil water content declines (Dodd 2007). In
a split root experiment, Dodd demonstrated that total water
availability and heterogeneous water distribution affected ABA
transport and consequential root-to-shoot signaling and plant
gas exchange parameters (Dodd 2007, Dodd et al. 2008).
Hence, variation in root or water distribution within the pots
in the current study might have affected the behavior of gs as
presented in Figure 3, i.e., nutritional level may have affected
root distribution which, in turn, influenced ABA production and
stomatal behavior. Such an indirect effect between phytohormones and K nutrition was reported in sunflower, which failed
to regulate stomatal closure under drought probably due to
the involvement of K in ethylene synthesis (Benlloch-González
et al. 2010). In a similar way, K and Na nutrition may have had
an indirect effect on gs by triggering, encouraging or inhibiting
phytohormone synthesis.
One remarkable effect of nutrition was the non-stressed predawn and post-sunset gs seen in EXP-II. All treatments had
Tree Physiology Online at http://www.treephys.oxfordjournals.org
1114 Erel et al.
approximately double gs at dawn (0.11–0.15 mol H2O m−2 s−1)
compared with 0.07 mol H2O m−2 s−1 for the control. Night-time
stomatal opening has been reported for other plant species,
including those well adapted to dry environments (Caird et al.
2007). Although night transpiration is well documented, the
phenomenon is not yet fully understood (Auchincloss et al.
2014). The slow response of stomata to darkness in the presence of Na, as found in C. communis (Jarvis and Mansfield
1980), may partly explain these results. The literature is not
consistent in regard to a nutritional effect on stomatal movement (Caird et al. 2007). We speculate that night-time transpiration, as encountered in the current study, may be an adaptive
mechanism to enhance mineral uptake and/or redistribution
and utilization under deficit conditions as suggested by Daley
and Phillips (2006).
Nutritional effect on photosynthesis, WUE and
photoinhibition
EXP-II clearly showed inhibition of assimilation caused by K
deficiency as occasionally has been reported in the literature
(Cakmak 2005, Römheld and Kirkby 2010, and references
within). Our results show that, similar to eucalyptus, Na partially
restored A of K-deficient plants (Battie-Laclau et al. 2014a).
Cacao K-deficient plants were also shown to have enhanced
A under the presence of Na (Gattward et al. 2012). Two major
factors may affect net assimilation: (i) stomatal conductance
and (ii) assimilation efficiency. The higher A rate in high K trees
as displayed in Figure 2 is likely to be partly a consequence of
lower stomatal limitation. However, this cannot explain the significant difference between K0-Na58 and K0-Na0 treatments,
which had comparable daytime gs but significantly different
A. The assimilation efficiency of the trees was therefore likely
affected by K and Na nutrition. In order to better evaluate interactions between A, gs and nutrition, we have plotted A response
to gs (Figure 7) revealing an asymptotic curve similar to that
found by Tognetti et al. (2004). Both K levels and the high Na
level had fairly identical response curves, i.e., identical A efficiencies. However, at almost any given gs, treatment K0-Na0
had lower A. Thus, in comparison with K0-Na0, the superior
A of the high K treatments was derived from both greater gs
(Figure 2) and greater assimilation efficiency (Figure 7). The
higher A of treatment K0-Na58 compared with K0-Na0 can
also be credited to better assimilation efficiency. Therefore, at
low environmental K, Na is capable of maintaining proper A by
preserving normal assimilation capacity. The reduced assimilation efficiency of K- and Na-depleted plants is probably the
cause of the lower WUE found in this treatment. This reduction
in WUE was completely recovered in the presence of 2.5 mM
Na in the irrigation solution. Nutritional level affected WUE only
for the irrigated regime and not under drought. The significant
effect of water availability on interaction between WUE and K
nutrition is likely also to be due to the nature of the A response
Tree Physiology Volume 34, 2014
Figure 7. The relationship between stomatal conductance and assimilation rate of the four extreme treatments: K0-Na0 (circles), K0-Na58
(triangles), K15-Na0 (diamonds) and K100-Na58 (squares). For
all treatments exponential rise regression was fitted. The raw data
from the two diurnal measurements were used, excluding measurements taken before 08:30 and after 16:30. Thin lines represent 95%
­confidence levels.
to gs. At gs <0.07 mol m−2 s−1, which is typical for drought conditions, A was nearly identical among treatments (Figure 7).
The difference between K0-Na0 and the remaining treatments
was augmented as gs increased. Thus, when water was highly
available, gs was generally high and the assimilation rate of Kand Na-deficient plants was substantially impaired.
The maximum photosystem II (PSII) quantum yield, Fv/Fm, is
considered a reliable indicator of proper activity of PSII of the
photosynthetic system (Maxwell and Johnson 2000) and has
been found relevant for olive (Angelopoulos et al. 1996). In the
current study, K-deficient plants with no Na suffered damage
to their photosynthetic apparatus regardless of the imposed
water stress. Sodium addition diminished the apparent photoinhibition. Under drought, predawn Fv/Fm indicated photodamage to all plants, in accordance with Angelopoulos et al.
(1996). In spite of this, K0-Na0 had significantly lower Fv/Fm
compared with K0-Na58, implying that photoinhibition, similar
to A, was positively affected by the presence of Na regardless
of plant water status.
Starch and SCHs
The SCH profile of the olive leaf is complex and composed
of several non-structural SCHs. A number of SCHs such as
mannitol are thought to play a role in responses to abiotic
stress (Tattini et al. 1996, Dichio et al. 2009). In the present study, both starch content and SCHs were significantly
affected by K and Na nutrition. Starch concentration was lowest in the absence of K and Na as reported for eucalyptus
trees (Battie-Laclau et al. 2014a), probably a result of impaired
net assimilation (Table 4) combined with inhibition of enzyme
Sodium replacement of potassium in the olive 1115
starch synthase (Nitsos and Evans 1969). In contrast to starch,
total SCH level was not modified. In eucalyptus, Na-, and to a
greater extent, K-fertilized trees had a lower SCH compared
with an unfertilized control (Battie-Laclau et al. 2013, 2014a).
The SCH concentrations found here are comparable to those
reported in the literature for olive (Bustan et al. 2011) but the
relative partition of SCHs was strongly influenced by nutrition
level. Mannitol content increased with K and Na. Increase in
mannitol content was reported previously in olive in response
to drought (Dichio et al. 2009), salinity (Tattini et al. 1996)
and even K (Chartzoulakis et al. 2006). The opposite (i.e., a
decrease) was found for galactinol concentration, which was
highest in the absence of K and Na. Galactinol was found to
provide protection from abiotic stresses and particularly from
oxidative damage (Nishizawa et al. 2008). The higher galactinol concentration may be an indicator for oxidative stress in
the low K and Na plants as also reflected in the maximum PSII
quantum yield.
Conclusions
The physiological response to K and Na nutrition and drought
was studied in two experiments using bearing and young olive
trees. Olive was found to be highly efficient in K utilization,
allowing leaf K to drop to extremely low levels prior to the
appearance of deficiency symptoms. Potassium is considered an enhancer of drought resistance; however, we found
no evidence of such in olive exposed to gradual water stress.
Moreover, stomatal control was not significantly impaired by
K deficiency whether under well watered or drought conditions. The most pronounced effect of K and Na deficiency was
lower stomatal conductance and inferior assimilation capacity. Consequentially, the general assimilation rate if no K and
Na were applied was roughly half that of sufficient fertilized
control. The presence of Na in irrigation solution recovered
the assimilation rate to a nearly optimum level by enhancing
both conductivity and photosynthetic efficiency. The inhibitory
effect of K deficiency on assimilation was more pronounced
when stomatal conductance was high and therefore, under
drought the effect of K declined. Under drought, leaf water
potential decreased with K and Na concentration. High K and
Na nutritional levels caused increased leaf starch concentration. Total SCH concentrations were not modified by nutrition,
however, the relative partitioning of individual sugars was
strongly related to K and Na nutrition. To the best of our knowledge, this is the first documentation that the olive is capable of
partially replacing K with Na. Under conditions of deficit K, the
addition of Na resulted in enhanced growth, higher stomatal
conductivity, superior assimilation capacity, higher leaf starch
concentration and reduced deficiency symptoms. The positive
effect of Na on root growth, assimilation performance, photodamage and carbohydrate profile indicates that Na may play a
metabolic role in addition to its acknowledged osmotic role. The
nutritional effect of K and Na was shown to be independent of
plant water status.
Conflict of interest
None declared.
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