J Agro Crop Sci (2012) ISSN 0931-2250
DROUGHT STRESS
Effects of Increasing Salinity Stress and Decreasing Water
Availability on Ecophysiological Traits of Quinoa
(Chenopodium quinoa Willd.) Grown in a MediterraneanType Agroecosystem
C. Cocozza1, C. Pulvento2, A. Lavini2, M. Riccardi2, R. d’Andria2 & R. Tognetti1
1 Dipartimento di Bioscienze e Territorio, Universit
a degli Studi del Molise, Pesche, IS, Italy
2 Institute for Agricultural and Forest Mediterranean Systems (ISAFoM-CNR) Ercolano, NA, Italy
Keywords
abscisic acid; Chenopodium quinoa; drought;
salinity; stomatal conductance; water
potential
Correspondence
C. Cocozza
Dipartimento di Bioscienze e Territorio,
Universit
a degli Studi del Molise
Pesche
IS, Italy
Tel.: +39 0874 404735
Fax: +39 0874 404123
Email: [email protected]
Accepted October 30, 2012
doi:10.1111/jac.12012
Abstract
Quinoa is a native Andean crop for domestic consumption and market sale, widely
investigated due to its nutritional composition and gluten-free seeds. Leaf water
potential (Ψleaf) and its components and stomatal conductance (gs) of quinoa, cultivar Titicaca, were investigated in Southern Italy, in field trials (2009 and 2010). This
alternative crop was subjected to irrigation treatments, with the restitution of 100 %,
50 % and 25 % of the water necessary to replenish field capacity, with well water
(100 W, 50 W, 25 W) and saline water (100 WS, 50 WS, 25 WS) with an electrical
conductivity (ECw) of 22 dS m 1. As water and salt stress developed and Ψleaf
decreased, the leaf osmotic potential (Ψp) declined (below 2.05 MPa) to maintain
turgor. Stomatal conductance decreased with the reduction in Ψleaf (with a steep
drop at Ψleaf between 0.8 and 1.2 MPa) and Ψp (with a steep drop at Ψp between
1.2 and 1.4 MPa). Salt and drought stress, in both years, did not affect markedly
the relationship between water potential components, RWC and gs. Leaf water potentials and gs were inversely related to water limitation and soil salinity experimentally
imposed, showing exponential (Ψleaf and turgor pressure, Ψp, vs. gs) or linear (Ψleaf
and Ψp vs. SWC) functions. At the end of the experiment, salt-irrigated plants
showed a severe drop in Ψleaf (below 2 MPa), resulting in stomatal closure through
interactive effects of soil water availability and salt excess to control the loss of turgor
in leaves. The effects of salinity and drought resulted in strict dependencies between
RWC and water potential components, showing that regulating cellular water
deficit and volume is a powerful mechanism for conserving cellular hydration under
stress, resulting in osmotic adjustment at turgor loss. The extent of osmotic adjustment associated with drought was not reflected in Ψp at full turgor. As soil was drying,
the association between Ψleaf and SWC reflected the ability of quinoa to explore soil
volume to continue extracting available water from the soil. However, leaf ABA content did not vary under concomitant salinity and drought stress conditions in
2009, while differing between 100 W and 100 WS in 2010. Quinoa showed good
resistance to water and salt stress through stomatal responses and osmotic adjustments that played a role in the maintenance of a leaf turgor favourable to plant growth
and preserved crop yield in cropping systems similar to those of Southern Italy.
Introduction
Quinoa (Chenopodium quinoa Willd.) is a herbaceous
annual plant of Amaranthaceous family, a main native
© 2012 Blackwell Verlag GmbH
Andean crop for domestic consumption and for market
sale, which has been widely investigated (e.g. Jacobsen
2011) mainly due to its nutritional composition (Ruales
and Nair 1992, 1993, 1994), Repo-Carrasco et al. 2003 and
1
Cocozza et al.
as alternative to typical cereals in coeliac diets, because
seeds are gluten-free (Alvarez-Jubete et al. 2009). In the
Andean highlands, quinoa is subjected to a range of adverse
climatic factors such as drought, frost, wind, hail and soil
salinity, which have been addressed in several studies
(Grace 1985, Jensen et al. 2000, Garcia et al. 2003, Aguilar
and Jacobsen 2003, Jacobsen et al. 2003a,b, Bertero et al.
2004, Jacobsen et al. 2005, Bois et al. 2006, Geerts et al.
2006, Bhargava et al. 2007, Hariadi et al. 2011), and supposedly, quinoa has desirable characteristics including tolerance to adverse growing conditions (Jacobsen et al.
2003a,b). The spread of quinoa outside its native area,
however, might be jeopardized by recurrent stress, including drought and soil salinity (Jacobsen and Stolen 1993,
Bray 1997, Vacher 1998, Zhu 2001, Pulvento et al. 2010),
particularly in the Mediterranean area. Recent discussion
on research cooperation has highlighted the need for further agronomic investigations to be focused on the process
of cultivation of quinoa (Winkel et al. 2012).
In field conditions, food crops encounter a combination
of different abiotic stresses (Essa 2002, Wang et al. 2003).
In arid and semiarid agroecosystems, drought and salinity
are the main abiotic stresses damaging the potential yield
and causing yield instability in quinoa (Pulvento et al.
2010, Fuentes and Bhargava 2011, Razzaghi et al. 2011a,b).
Crop productivity in water-limited environments derives
from mechanisms that promote tolerance to drought episodes and minimize water loss, thereby maintaining a
favourable water status for leaf development. In this sense,
chemical and hydraulic signals are operative and integrated
in regulation of leaf growth and stomatal conductance
when plants experience drought stress (Davies et al. 1994,
Comstock 2002). In particular, abscisic acid (ABA) may
have hormonal activity affecting grain maturation (King
1976) and act as chemical signals during early stages of soil
drying (Davies and Zhang 1991, Bacon et al. 1998). Jacobsen et al. (2009) found that quinoa (cultivar INIAIllpa) has
a responsive stomatal closure, by which the plants are able
to maintain leaf water potential and photosynthetic rate
during soil drying, resulting in an increase in water-use efficiency. During soil drying, root produced ABA, which was
active in stomatal regulation, inducing a decrease in turgor
of stomata guard cells, leading to a reduction in stomatal
conductance (Jacobsen et al. 2009).
Nevertheless, Jensen et al. (2000) found that the stomatal
response of quinoa was insensitive to drought induced
decrease in leaf water status and hypothesized that high net
photosynthesis rates during early growth stages would
result in initial vigour of plants supporting prompt water
uptake and thus tolerance to later drought. Studies on the
effect of salt stress showed that quinoa has a very efficient
system to adjust osmotically also for abrupt increases in
salinity levels (Hariadi et al. 2011). Jensen et al. (2000)
2
observed that the leaf water relations were characterized by
low osmotic potentials during later growth stages sustaining a potential gradient for water uptake and turgor maintenance during soil drying. Turgor maintenance, obtained
by sensitive stomata closure in response to long-distance
chemical signalling generated in roots (Jensen et al. 1998)
and osmotic adjustment at low water potential in leaves
(Ali et al. 1999), would allow metabolic processes to be
preserved and sustain growth and survival of plants, during
reduction in leaf water status (Hsiao et al. 1986, McCree
1986). Karyotis et al. (2003) reported that seed yield was
markedly reduced by soil saline–sodic conditions, and
Razzaghi et al. (2011b) added that the salinity caused a significant decrease in seed production of quinoa (cultivar
Titicaca), but the increasing salinity did not further
decrease crop yield, due to plant acclimation to salinity levels between 20 and 40 dS m 1. In a Danish breed of quinoa
(cultivar Titicaca also cited as Q52 or KVLQ520Y) grown
in greenhouse conditions, Razzaghi et al. (2011a) observed
that a reduction in soil water potential, caused by soil drying and salinity, reduced transpiration and increased
apparent root resistance to water uptake at the end of the
drought period. However, the definition of indicators that
plant breeders might apply in open field to improve quinoa, for its tolerance or adjustment to saline environments,
is still a matter of debate (Razzaghi et al. 2011a). In particular, there is paucity of information on cultivar-specific
ecophysiological response of quinoa to concurrent salinity
and drought stress in Mediterranean-type agroecosystems.
The mechanisms involved in the phenological flexibility
of quinoa in response to a combination of different abiotic
stresses under field conditions are not fully identified (Jacobsen et al. 2005, Geerts et al. 2008, Rosa et al. 2009, Razzaghi et al. 2011a, 2011b). In addition, current climate and
land-use changes expose the acute vulnerability of the Mediterranean region to climatic extremes, and sagacious irrigation practices must be sought to maintain productivity
in chenopodiaceous species (Tognetti et al. 2003). In particular, climate change may add to existing problems of
desertification, water scarcity, water quality and food production. Growing of tolerant genotypes of quinoa might be
one of the cost-effective strategies for coping with growth
constraints, which are significant factors affecting crop production and sustainability in numerous agricultural regions
of the Mediterranean rim. Indeed, Pulvento et al. (2012)
observed that salt and drought stress did not cause significant yield reduction in quinoa, in 2 years of investigation.
We hypothesized that quinoa, cultivar Titicaca, would
show drought resistance in agronomic conditions of Southern Italy (Pulvento et al. 2010) and that drought-mediating
mechanisms would add to salinity tolerance (Pulvento
et al. 2012). The seasonal trend of leaf water potential, leaf
ABA content and stomatal conductance of quinoa, cultivar
© 2012 Blackwell Verlag GmbH
Effects of Increasing Salinity Stress
Titicaca, under soil drying and rising salinity in a typical
Mediterranean-type agroecosystem of Southern Italy was
investigated to further predict field crop responses and
implement water productivity models. The specific objectives of this work were (i) to monitor plant water potential
components and their involvement in osmotic adjustments
to increasing salinity stress and decreasing water availability
and (ii) to examine the sensitivity of stomatal response in
experimental field conditions.
Materials and Methods
Site description
The field trial was performed in 2009 and 2010 at the CNR
– Institute for Agricultural and Forest Mediterranean Systems (ISAFoM) research station located in the Volturno
River plain (14°50′E, 40°07′N; 25 m above sea level), an
irrigated area of Southern Italy. The soil of the experimental site was characterized by a clay loam texture and volumetric water content of 0.394 m3 m 3 at field capacity (Ψ
of soil at 0.03 MPa) and of 0.217 m3 m 3 at wilting
point (Ψ of soil at 1.5 MPa).
The climatic conditions were typical of a sub-humid
Mediterranean area with long-term (1976–2010) average
annual precipitation of 805 mm. Potential evapotranspiration, ET0, calculated by Penman–Monteith equation, was
about 1157 mm per year (35- year average value). Vapour
pressure deficit (VPD) was calculated from hourly values of
mean temperature (Tmean) and relative humidity (RH).
Experimental layout
The cultivar Titicaca was provided by UCPH of the University of Copenhagen, and the crop was sown in plots of
36 m2 (6 9 6 m) at a density of 20 plants per m2 in both
years. The experiments lasted 96 (from DOY, day of year,
140 to DOY 236) and 99 (from DOY 111 to DOY 210) days
in 2009 and 2010, respectively. Quinoa was sown on the 20
May and harvested on the 24 August 2009, while it was
sown on the 21 April and harvested on the 29 July 2010. In
a completely randomized block design, with three replicates, three irrigation levels were compared: a control with
the restitution of 100 % of the water necessary to replenish
field capacity, the soil layer explored by roots (0.00–
0.36 m) and two treatments with restitution of 50 % and
25 % of the water volume used for the control treatment.
Three treatments irrigated, with the restitution of 100 %,
50 % and 25 % of the water necessary to replenish field
capacity, with saline water (25 WS, 50 WS and 100 WS)
and three treatments irrigated with well water (25 W,
50 W and 100 W) were performed. For the saline treatments, irrigation water had an electrical conductivity
© 2012 Blackwell Verlag GmbH
(ECw) value of about 22 dS m 1, obtained by adding
13.38 g l 1 of sodium chloride (NaCl), 0.45 g l 1 of calcium chloride (CaCl2), 0.34 g l 1 of potassium chloride
(KCl), 1.14 g l 1 of magnesium chloride (MgCl2) and
1.64 g l 1 of magnesium sulphate (M gs O4); these salts
were added to reproduce the ionic content of water
obtained by mixing well water and sea water in the ratio
1 : 1.
The irrigations were supplied in both years on a weekly
basis. In 2009, four irrigations were supplied starting from
the 14th of July (day of year, DOY, 195, after 55 days from
sowing), with a total amount of applied water of
1864 m3 ha 1 and 2024 m3 ha 1 for 100 W and 100 WS
treatments respectively; in 2010, five irrigations were supplied starting from the 16th of June (DOY 167, after
88 days from sowing), with a total amount of applied water
of 1295 m3 ha 1 and 1286 m3 ha 1 for 100 W and
100 WS treatments, respectively.
The irrigation levels, with and without salt, were
arranged in a factorial combination. The experimental layout was a completely randomized block design with three
replicates. Meteorological data were collected using an
automatic weather station (iMETOS, MMM – Mosler tech
support, Berlin, Germany) located in the experimental
field, and they were compared with long-term average values (1976–2010) (Fig. 1). For further details on cropping
practice, experimental design and irrigation protocol, see
Pulvento et al. (2012).
Soil characteristics
In both years, the electrical conductivity of the soil was
measured during the cultivation season, and samples were
taken between rows at the same soil depth used to measure
moisture (0.00–0.12, 0.12–0.36, 0.36–0.60, 0.60–0.96 m).
The electrical conductivity of the soil (EC) was measured
on an aqueous soil extract (ratio of water/soil = 2.5/1), and
values were related to those of the concentration in
saturated paste (ECe) (see Pulvento et al. 2012).
Soil water content (SWC) was monitored on a weekly
basis, using the gravimetric method, before and 24 h after
each watering via soil sampling by a probe (0.08 m in
diameter and 0.12 m long) in each plot (soil bulk density = 1.28 t m 3). Soil samples were taken in each
replicate at soil depths of 0.0–0.36 and 0.36–0.96 m.
Water relations
The period of monitoring was from DOY 178 to DOY 214,
in 2009, and from DOY 158 to DOY 194, in 2010, and all
plant traits were monitored after the completion of the vegetative growth period. Water potential components and the
relative water content (RWC) were measured on fully
3
Cocozza et al.
(a)
(b)
was calculated as Ψp 9 RWC (according to Jensen et al.
2000).
For relative water content (RWC, %) determination, leaf
samples were weighted (FW) and then floated on distilled
water in the dark overnight, at 4 °C. The turgid weight
(TW) was determined after blotting, and the dry mass
(DW) was measured after the samples had been dried for
24 h at 80 °C. Then, RWC was determined according to
Barrs and Weatherley (1962) following the equation:
RWC = [(FW-DM)/(TW-DW)]*100.
The turgor pressure (Ψp, MPa) was estimated from the
difference between Ψleaf and Ψp, assuming leaf matric
potential to be zero.
The stomatal conductance (gs, mmol m 2 s 1) was measured on fully expanded leaves (the same used for plant
water status determination), every 15 days between 11 : 00
and 13 : 00 h, under saturating light intensity using a
dynamic diffusion porometer (AP4; Delta-T Devices Ltd.,
Cambridge, UK).
ABA determination
Fig. 1 Time course of some climatic parameters in the 2 years (2009–
2010) and the 35-year mean value. Minimum and maximum air temperatures are 10-day means; rainfalls are 10-day sums (in column); the
potential evapotranspiration (ET0) values are 10-day means.
expanded leaves collected from the mid-section of the
shoot in two different plants per each plot. Measurements
were conducted between 11 : 00 and 13 : 00 h (local time)
every 15 days.
The leaf water potential (Ψleaf, MPa) was measured using
a pressure chamber Scholander (SKPM 1400; Skye Instruments Ltd., Llandrindod Wells, UK). Harvested leaves were
immediately put in jars and transferred into liquid nitrogen
and stored at 80 °C until analysis of osmotic potential.
The osmotic potential (Ψp, MPa) was determined after
thawing leaves for 15 min, and sap was extracted by applying hydraulic pressure through a cylinder (generating a
compressive force). Sap samples were collected in glass
tubes placed on ice and their osmolarity determined with a
Roebling 13DR automatic osmometer (D-1000; Messtechnik, Berlin, Germany). The osmotic potential for each measured concentration of cell juice was calculated according
to the van’t Hoff equation at a temperature of 20 °C:
Ψp = MiRT, where M is the concentration in molarity of
the solute, i is the van’t Hoff factor (the ratio of amount of
particles in solution to amount of formula units dissolved),
R is the ideal gas constant, and T is the absolute temperature. The osmotic potential at full hydration (Ψp100, MPa)
4
Leaves were harvested on DOY 203 and 211, in 2009, and
on DOY 190 and 202, in 2010, between 11 : 00 and
13 : 00 h (local time). Three replicates for each crop–treatment combination were used. Leaves were immediately put
in jars and transferred into liquid nitrogen and stored at
80 °C until ABA extraction. Twenty milligrams of leaf
(without midribs) was extracted overnight in 1.5 ml distilled water in the dark at 4 °C on a shaker. The extracts
were centrifuged at 10000 9 g for 25 min, and the ABA
content of the supernatants was quantified in an enzymelinked immunosorbent assay (ELISA) using the Phytodetek-ABA kit (AGDIA, Elkhart, IN, USA), according to the
indications of the manufacturer. All assays were made in
triplicate. The hormone concentrations were calculated
using a standard curve of ABA and the relative optical
density, according to the ELISA technique.
Crop harvest
At maturity, plants were harvested, and grain yield was estimated. For dry mass determination, plants were oven dried
at 70 °C to constant weight (see Pulvento et al. 2012).
Statistical analysis
Data were analysed by analysis of variance (ANOVA), using
the SAS statistical package (SAS Inst. Inc., Cary, NC, USA),
and means were compared using the least significant difference (LSD) test.
Data of ABA were averaged on a plant basis, and the
individual means were used for the analysis. The effects of
© 2012 Blackwell Verlag GmbH
Effects of Increasing Salinity Stress
watering and salt treatments on ABA concentration were
tested with the statistical package Statistica (StatSoft Inc.,
Tulsa, OK, USA), using repeated measures analyses of variance. Statistical comparisons were considered significant at
P 0.05.
(a)
Results
Climatic characteristics of the trial site are typical of a subhumid Mediterranean area. Meteorological conditions in
the two experimental years, 2009 and 2010, and long-term
average values (1976–2010) were reported in Fig. 1.
In 2009, abundant rainfalls were recorded in late spring
and early summer, with scattered precipitation < 5 mm
(per rain event) afterwards (until harvest). In 2010, abundant rainfalls occurred in mid-spring and late July
(Fig. 1a). Potential evapotranspiration (ET0) was generally
higher in 2009 than 2010 (Fig. 1b). In 2009, air temperature and ET0 values were generally higher than long-term
values from the second decade of May to harvest. Instead,
in 2010, air temperature values did not differ from the
long-term means except in July. The vegetative growth period was characterized by 180.0 and 173.4 mm of rainfall
(only rain events above 5 mm were considered sufficiently
large and effective) (Fig. 1a), and 544.1 and 454.38 mm
ET0 (Fig. 1b), as cumulative values in 2009 and 2010,
respectively, and by 30.3 and 26.7 °C maximum temperature, and 18.3 and 15.6 °C minimum temperature, as average values (Fig. 1a). During the growing season, daily VPD
(data not shown) showed maximum values in mid-summer
ranging from about 2 kPa (2009) to 1.8 kPa (2010) and
minimum values in late spring ranging from 0.16 kPa
(2009) to 0.06 kPa (2010).
The water content in the soil layer explored by roots
(SWC, 0.00–0.36 m) (Fig. 2) decreased in each irrigation
treatment during the growing season to a minimum value
of 0.15 m3 m 3 reached on DOY 235 and 214 of 2009, in
25 WS and 25 W, respectively, and 0.21 m3 m 3 in 25 W
on DOY 193 of 2010 (Fig. 2). The water content values in
the soil were generally higher in 2010 than 2009, and WS
treatments dried out more slowly than W treatments
(Fig. 2). The salts supplied with irrigation during the crop
cycle, in WS treatments, determined an increase in the ECe
(Table 1) in the 0.00 to 0.36-m soil layer; ECe reached
values of 15.9 dS m 1 and 10.6 dS m 1 in 100 WS,
respectively, in 2009 and 2010 (Table 1).
The evolution of physiological parameters under water
deficit and salinity stress, during the 38- and 34-day periods in 2009 and 2010, respectively, was monitored. Leaf
water potential ranged from 1.47 MPa in 100 W (DOY
196) to 2.43 MPa in 50 WS (DOY 214), in 2009, and
from 0.79 MPa in 50 WS (DOY 158) to 2.22 MPa in
100 WS (DOY 192), in 2010.
© 2012 Blackwell Verlag GmbH
(b)
Fig. 2 Soil water content (SWC) measured before irrigating plants and
at harvest during the growing season of 2009 (a) and 2010 (b) for 0.00
–0.36-m soil layer (only values for the irrigation period were reported).
Arrows indicate the irrigations events. Phenological stages and length
of the crop cycle in the two experimental years according the indications of Jacobsen and Stolen (1993): LV, late vegetative; F, flowering;
ESF, early seed filling; LSF, late seed filling; R, ripening. Dashed line indicates the wilting point.
Table 1 Electrical conductivity of the soil saturated paste (ECe) during
the growing season of the two experimental years (2009–2010), for the
0.00–0.36-m soil layer. DOY corresponded to the beginning of treatment differentiation, day after three irrigations and day of harvest
Treatments
Year
DOY
25 WS
ECe (dS 1)
50 WS
2009
193
209
235
166
193
209
0.8
5.4
8.7
0.9
1.2
6.1
0.8
7.7
12.5
1.2
3.9
9.0
2010
100 WS
0.8
10.7
15.9
1.6
4.9
10.6
Stomatal conductance decreased as Ψleaf became more
negative with a steep drop at leaf water potential between
0.8- and 1.2 MPa (Fig. 3a). Stomatal conductance
decreased with diminishing Ψp, with a steep drop at Ψp
between 0.6 and 0.3 MPa (Fig. 3b). Turgor pressure and
Ψleaf were exponentially correlated with gs across
treatments.
5
Cocozza et al.
(a)
(b)
Fig. 3 Relationships between leaf water potential (Ψleaf) and stomatal
conductance (gs) (a) and Ψleaf and turgor pressure (Ψp) (b) in quinoa,
cultivar Titicaca, in 2009 and 2010. Open circles refer to W plants and
closed circles to WS plants. Regression parameters of exponential functions fitted to data are reported, as well as significance.
The relationship between Ψp and Ψleaf (Fig. 4a) and
between Ψp, Ψp and Ψleaf, and RWC (Fig. 4c,e) showed a
positive linear relation, whereas Ψp100 was not significantly
related to Ψleaf (Fig. 4b). In particular, for a drop of 1 MPa
in Ψp, a loss of 40 % in RWC was observed. The variation
in Ψp values showed a decreasing trend during the experiment, in both years of monitoring (data not shown). Overall, Ψp was significantly different between treatments, in
2009, ranging from maximum 1.72 MPa in 100 W to
minimum 2.29 MPa in 50 WS. In 2010, there were not
significant differences between treatments; however, Ψp
ranged from 1.12 MPa in 100 W to 2.08 MPa in
50 WS (data not shown). The three WS treatments showed
that salt addition in water deficit treatments resulted in
lower Ψp with respect to 50 W and 100 W (data not
shown). Osmotic potential at full turgor showed only
minor variation during the experiment (Fig. 4b); statistical
6
differences in Ψp100 between treatments in the 2 years of
monitoring were not reported. Furthermore, Ψp100 was not
significantly related to SWC.
Significant linear positive relations were found between
SWC and plant water status (Fig. 5). Decreasing SWC followed declining Ψleaf (Fig. 5a) and Ψp (Fig. 5b) values,
whereas gs was not related to SWC (Fig. 5c). Treatments
were clearly separated, with higher values of Ψleaf, Ψp and
gs in W than WS plants, at corresponding SWC.
The ABA concentration in leaves was significantly
higher in 2010 than in 2009 (P < 0.0001), and marked differences were observed between days of monitoring in
both years (Table 2). In particular, the ABA content was
higher in the second day of monitoring than the first, in
both years. In 2009, mean leaf ABA concentration ranged
between 27.3 pmol ABA g 1 of 50 W (DOY 203) and
58.6 pmol ABA g 1 of 25 W (DOY 211), in W treatments,
whereas it varied between 25.2 pmol ABA g 1 of 100 WS
(DOY 203) and 65.5 pmol ABA g 1 of 25 WS (DOY 211),
in WS treatments (Table 2). In 2010, leaf ABA concentration ranged from 48.4 pmol ABA g 1 of 100 W (DOY 190)
to 250.6 pmol ABA g 1 of 100 W (DOY 202), in W treatments, whereas it varied from 18.3 pmol ABA g 1 of
50 WS (DOY 190) to 270.8 pmol ABA g 1 of 25 WS (DOY
202), in WS treatments (Table 2). Significant differences for
simple saline effect (e.g. 25 W vs. 25 WS, 50 W vs. 50 WS,
etc.) were detected between days of monitoring (Table 3).
The interaction ‘treatment’ 9 ‘day of monitoring’ was significant in 2010 (Table 3). In both years, the interaction
between W and WS treatments did not differ significantly,
in term of crop yield; no significant differences were
obtained within these treatments. Grain yield reached a
value (average value of all treatments) of 2.7 t ha 1 and
2.3 t ha 1, respectively, in 2009 and 2010; a detailed analysis
of yield factors can be found in Pulvento et al. (2012).
Discussion
The study depicted the seasonal course of leaf water potentials, leaf ABA content and stomatal conductance of quinoa, cultivar Titicaca, grown in a Mediterranean area
under field conditions, mimicking increasing salinity stress
and decreasing water availability. Yield responses of the
same crop have been described in a companion paper by
Pulvento et al. (2012). Salt and drought stress did not
affect crop yield negatively, while the highest level of saline
water treatment resulted in higher mean seed weight. These
seeds showed relatively high protein and fibre contents,
which make this relatively stress tolerant species promising
for developing new products with good nutritional properties. Low Ψp towards late season sustained a potential gradient for water uptake and turgor maintenance during the
advancement of stress conditions, showing significant
© 2012 Blackwell Verlag GmbH
Effects of Increasing Salinity Stress
(a)
(c)
(b)
(d)
(e)
Fig. 4 Relationships between water relations and physiological parameters of quinoa, cultivar Titicaca, in 2009 and 2010. (a) osmotic potential, Ψp,
vs. leaf water potential, Ψleaf; (b) osmotic potential at full hydration, Ψp100, vs. Ψleaf; (c) turgor pressure, Ψp, vs. relative water content, RWC; (d) Ψp,
vs. RWC; (e) Ψleaf vs. RWC. Open circles refer to W plants and closed circles to WS plants. Regression parameters of linear functions fitted to data are
reported, as well as significance.
differences between treatments and days, in 2009, which
was relatively drier than 2010. Plant water relations were
relatively more sensitive to salt stress with respect to water
limitation, showing lower values of water potential components, RWC and gs; values of plant water relations were in
agreement with the data reported by Jensen et al. (2000)
for quinoa grown in field conditions. Indeed, the stomatal
response to water limitation and salinity stress showed that
quinoa had rather insensitive stomatal response, as in the
present experiment, stomatal closure did not occur until
© 2012 Blackwell Verlag GmbH
Ψleaf and Ψp values were below approximately 1.2 and
0.3 MPa, respectively (Fig. 3). The existence of negative
pressures, at least in part, could be explained by the method
employed to obtain Ψp, which was estimated from the difference between Ψleaf and Ψp, assuming leaf matric potential to be zero. A dilution effect was probably occurring due
to the decrease in relative apoplastic water content, which
Jensen et al. (2000) found to be significant in quinoa under
water stress, by a fraction of 19–14 %. However, the potential benefits of negative turgor pressures in terms of
7
Cocozza et al.
(a)
(b)
(c)
Fig. 5 Relationships between soil water content (SWC) and leaf water
potential (Ψleaf) (a), turgor pressure (Ψp) and SWC (b), stomatal conductance (gs) and SWC (c) in quinoa, cultivar Titicaca, in 2009 and 2010.
Open circles and dashed lines refer to W plants and closed circles and
filled lines to WS plants. Regression parameters of linear functions fitted
to data are reported, as well as significance.
preventing mechanical and osmotic damage associated with
severe desiccation make this topic worthy of further study.
Thus, quinoa can be defined as a crop tolerant to drought
8
and salinity as compared with other dicots (Jensen et al.
2000, Sanchez et al. 2003, Pulvento et al. 2012). This
behaviour would enable quinoa to maintain cell turgor
under saline conditions, thus avoiding permanent damage
from drought stress, also in this Mediterranean-type agroecosystem.
Crop yield was not affected by water and salt stress,
suggesting that plants activated stress tolerance mechanisms to avoid yield losses. In the present experiment,
the soil moisture level was kept relatively high by precipitation until panicle formation (Pulvento et al. 2012). In
both years, SWC was above 50 % of available water until
pre-flowering for all irrigated treatments. However, a certain amount of water was ensured during flowering and
grain filling, even for the less irrigated W and WS treatments (50 and 25), which was enough to stabilize yield.
This is in agreement with Razzaghi et al. (2012) who
attained no significant differences between yield of quinoa Titicaca for full irrigated and for deficit irrigated
treatments in a clay loam soil. Likewise, Geerts et al.
(2008) suggested that drought stress should be only mitigated during plant establishment and the reproductive
stages to stabilize quinoa yield.
Differences in plants subjected to W and WS treatments
were found in the relationships between SWC and Ψleaf
and Ψp, indicating a slightly higher plant hydration in W
than WS plants at similar SWC. As soil was drying, the
association between leaf water potentials and SWC
reflected the ability of quinoa to explore soil volume to
continue extracting available water from the soil. On the
other hand, Ψp100 was not significantly related to SWC
and Ψleaf. The osmotic potential at full turgor (Ψp100)
showed rather constant values during the present experiment, showing only slight fluctuations. Thus, the ability of
leaves to adjust osmotically and thereby decrease the value
of Ψp at full turgor did not appear to be present in different trends between plants experiencing water deficits and
salinity conditions, probably because the concurrent
increase in tissue elasticity resulted in a larger symplast volume at full turgor. A limited osmotic adjustment Ψp100
between fully watered and water-stressed plants was also
found by Jensen et al. (2000), in quinoa grown in pots and
lysimeters. The low osmotic potential in itself will support
the maintenance of the potential gradient for water uptake
at low soil water potential under high evaporative demands
causing deficits in the plant (Ali et al. 1999), as also envisaged in modelling work (Jensen et al. 1993). Increasing
stress conditions with season progression defined distinctive patterns of ABA concentrations. The main reason of
high ABA contents towards the end of experiment in both
years could be explained by the progression of stress intensity. The 2 days selected for ABA monitoring were defined
in coincidence of important stages of stress progression.
© 2012 Blackwell Verlag GmbH
Effects of Increasing Salinity Stress
Table 2 Leaf ABA content in 2009 (DOY 203 and 211) and 2010 (DOY 190 and 202)
Treatments
Year
DOY
2009
203
211
190
202
2010
25 W
42.18
58.64
77.11
235.18
14.28
4.34
2.28
11.56
50 W
27.26
56.68
70.72
146.99
5.59
2.40
13.22
14.94
100 W
38.09
56.68
48.38
250.60
8.09
1.50
15.50
18.31
ANOVA
25 WS
41.55
65.48
79.84
270.84
5.20
2.68
0.46
26.98
50 WS
28.27
61.72
18.29
144.59
1.74
2.94
7.30
1.93
100 WS
W
WS
ns
ns
ns
*
*
ns
ns
**
25.25
58.93
26.04
104.59
6.79
0.76
21.89
6.26
For each DOY, significant differences between treatments (ANOVA test) are indicated.
ns, not significant.
*P 0.05.
**P 0.01.
Table 3 Repeated measures ANOVA of ABA content, for treatments (W
vs. WS) and day of monitoring in quinoa, cultivar Titicaca
Source
Year
2009
2010
Irrigation level
F-statistics
Treatment (T)
Day (D)
25
50
100
25
50
100
0.300
1.496
1.959
1.619
3.479
7.205 **
12.706 **
162.386 ***
47.775 ***
133.804 ***
47.473 ***
20.039 **
T9D
0.435
0.666
3.977
67.7112 ***
25.476 **
13.622 **
Significance values are indicated as *P 0.05.
**P 0.01.
***P 0.001.
The first day was defined when gs and Ψleaf experienced the
first significant reduction with respect to measurements at
the beginning of experiment; the second day coincided
with the evident reduction in water potential values. In
2010, in the second day of ABA analysis, leaves were not
sampled for other physiological parameters, owing to a
sudden occurrence in severe stress conditions that caused
plant injures, such as browning of leaf edges. The earlier
plant phenological stage could explain the higher leaf ABA
content in 2010 than 2009, because sowing was performed
earlier.
In 2009, quinoa regulated the response to stress conditions adjusting Ψleaf and Ψp, whereas RWC was maintained
constant across treatments (data not shown); nevertheless,
gs and Ψp decreased with marked stress to some extent,
with maintenance of Ψleaf above critic threshold; Ψleaf
dropped to 2.6 MPa under high-salinity stress, in the
study of Razzaghi et al. (2011a). In 2010, Ψleaf showed
lower differences between treatments, with Ψp adjusting to
moderate drought, somewhat controlling turgor during
dehydration. Jacobsen et al. (2009) observed, during soil
drying, that quinoa has a prompt stomatal closure, with
isohydric response to water stress, maintaining Ψleaf, resulting in an increase in plant water-use efficiency. This behav© 2012 Blackwell Verlag GmbH
iour was not clear in the present experiment (shallow
relationship between gs and SWC, regardless of the treatment), although under increasing salinity stress and
decreasing water availability, plants showed lower gs, probably leading to down-regulation of transpiration rate,
which might help quinoa to withstand soil water deficit.
However, despite stomatal response to increasing evaporative demand during the season, this was insufficient to prevent Ψleaf from falling to levels below 2 MPa in the driest
period, which confirm an intermediate behaviour between
isohydric and anisohydric response (Tardieu and Simmonneau 1998). Anisohydric species may behave as droughttolerant plants capable of maintaining higher gs at lower
Ψleaf than drought-intolerant species (Bunce et al. 1977).
Isohydric species, by contrast, may be considered as
drought avoiders, keeping their Ψleaf within narrow limits
(Galmes et al. 2007).
In this experiment, a strong exponential relationship
between gs and Ψleaf and Ψp was found, as observed in
Jacobsen et al. (2009) and Jensen et al. (2000). In all cases,
the effects of salinity and drought resulted in strict dependencies between RWC and water potential components,
showing that regulating cellular water deficit and volume is
a powerful mechanism for conserving cellular hydration
under stress, resulting in osmotic adjustment of quinoa. In
particular, for a drop of 1 MPa in Ψp, a loss of 40 % in
RWC was observed. When stressed plants reached the reference value of 70 % RWC (Babu et al. 1999), the low Ψp
( 1.6 and 1.9 MPa in W and WS, respectively) indicated
that an active osmotic adjustment had occurred. However,
no consistent differences were found between treatments in
the considered relationships. The relationship between gs
and Ψleaf showed a somewhat direct stomatal response to
chemical signals or hydraulic factors, independently of Ψleaf
until a specific Ψleaf is achieved.
Quinoa, cultivar Titicaca, therefore, could be cultivated
in drought and salt stress conditions of Mediterranean-type
agroecosystems, considering that the grain yield was not
compromised by the activation of ecophysiological
9
Cocozza et al.
mechanisms compensating for environmental disturbance.
The grain yields in the 2 years were compatible with average yields obtained in the original cultivation area of quinoa (Mujica et al. 2001, Delgado et al. 2009). These results
suggest a high plasticity of quinoa for tolerance and/or
resistance to increasing salinity stress and decreasing water
availability. Future scenarios for Mediterranean-type agroecosystems need to test the adaptation of different quinoa
varieties to areas characterized by recurrent drought and
chronic salinization. Results of this experiment appear
encouraging for the cultivation of quinoa, cultivar Titicaca,
in Southern Italy and the Mediterranean area.
Acknowledgements
We are grateful to the EU 7th Framework Programme for
the support to the project ‘Sustainable water use securing
food production in dry areas of the Mediterranean
region – SWUP-MED’. The authors thank Balsamo Angela,
Calandrelli Davide and Romano Giovanni for technical
support to field trial.
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