CSAWAC 43 (8) 1115-1266 (2015) Vol. 43 No.

8 August 2015

CLEAN
Soil Air Water
Renewables
Sustainability
Environmental Monitoring

8 | 2015

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1248
Abdul Khaliq1
Muhammad Zia-ul-Haq1
Faizan Ali1
Farhena Aslam1
Amar Matloob1,2
Abubakr Navab1
Saddam Hussain1,3
1

Department of Agronomy, University

of Agriculture, Faisalabad, Pakistan

Department of Agronomy,
Muhammad Nawaz Shareef
University of Agriculture, Multan,
Pakistan

College of Resources and

Environment, Huazhong Agricultural
University, Wuhan, Hubei, P. R.
China

Research Article
Salinity Tolerance in Wheat Cultivars Is Related to
Enhanced Activities of Enzymatic Antioxidants and
Reduced Lipid Peroxidation
Soil salinity is a stringent abiotic constraint limiting crop productivity. Studies were
undertaken to appraise the performance of six diverse wheat cultivars (LU-26S, MH-97,
AARI-2011, Millat-2011, Lasani-2008, and FSD-2008) under non-saline (control) and
saline soil (8 and 16 dS m
1) conditions. Increasing the salinity level severely
diminished emergence and seedling growth attributes in all wheat cultivars.
Nevertheless, cultivar specic responses were evident and a signicant salinity 
cultivar was observed in most cases. The lowest (23%) reduction in nal emergence was
observed for AARI-2011 as compared with the maximum recorded for LU-26S (56%).
AARI-2011 and Millat-2011 recorded the highest seedling dry biomass at a salinity level
of 16 dS m
1. The highest salinity level resulted in the maximum inhibition of
chlorophyll content in Millat-2011 and MH-97 by 59 and 68%, respectively, as compared
with their respective control. With an increase in the salinity level, enhanced activities
of antioxidants like superoxide dismutase and catalase were observed in AARI-2011
compared with all other wheat cultivars. Lasani-2008 showed a maximum (70%)
increase in malondialdehyde content compared with control. AARI-2011 and FSD-2008
appeared as promising wheat cultivars manifesting salt tolerance that was related to
enhanced activities of enzymatic antioxidants and reduced lipid peroxidation,
respectively.
Keywords: Emergence; Salt stress; Seedling growth; Wheat cultivars
Received: November 19, 2014; revised: February 7, 2015; accepted: March 12, 2015
DOI: 10.1002/clen.201400854

1 Introduction
Worldwide, soil salinity has adversely affected about 30% of the
irrigated, and 6% of the total land area [1]. In Pakistan, nearly 6.3
million hectare, comprising 14% of the irrigated land, is believed
to be salt-affected [2]. Crop production on saline soils remains a
challenging task. Salinity poses deleterious effects on growth and
the development of plants by inducing various biochemical and
physiological changes [3]. Wheat is a moderately salt-tolerant
crop [4, 5] and higher salinity levels severely impede its
emergence and seedling growth leading toward poor stand
establishment and consequently reduction in nal yield [6].
Salinity causes reduced emergence and impaired growth
presumably due to salt-induced osmotic effects, specic ion
toxicity, nutrient imbalance, oxidative damage, and alterations
Correspondence: S. Hussain, College of Resources and Environment,
Huazhong Agricultural University, Wuhan, Hubei 430070, P. R. China
E-mail: sadamhussainuaf@gmail.com, shussain@webmail.hzau.edu.cn
Abbreviations: CAT, catalase; chl, chlorophyll; E50, time taken to 50%
emergence; EC, electrical conductivity; EDTA, ethylenediaminetetraacetic acid; EI, emergence index; FEP, nal emergence percentage; MDA,
malondialdehyde; MET, mean emergence time; NBT, nitro-blue
tetrazolium; POX, peroxidase; RL, root length; ROS, reactive oxygen
species; SL, shoot length; SOD, superoxide dismutase; TDB, total dry
biomass; TSS, total soluble salt
2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim

in endogenous levels of hormones [3, 7, 8]. It is reported that both

physiological and biochemical components of the photosynthesis
are affected under salinity [9]. Toxic levels of ions such as Na and
Cl in saline soils result in the impairment of the chlorophyll (chl)
synthesis and various mechanisms such as gas exchange and
chlorophyll uorescence attributes [10, 11]. Another profound
effect of salinity is the increase in malondialdehyde (MDA)
content in response to the stress-induced oxidation of membrane
lipids [12]. An increase in MDA content under salinity suggests
that besides ionic and osmotic effects, oxidative stress is also a
crucial factor, and evaluating the cell membrane stability can be
exploited as a screening tool during salinity. Reactive oxygen
species (ROS) are abundantly produced during salinity, and attack
nucleic acids, proteins, and membrane lipids. Nevertheless, the
extent of such damage largely depends on the balance between
the accumulation of reactive oxygen species and their removal.
Plants usually employ enzymes of antioxidative defence to cope
with salt stress-induced oxidative cross-stress. Stress tolerance is
directly related to an efcient enzymatic defense system [13] and
their enhanced expression and activities are regarded as stressmarkers under salinity [8]. Besides antioxidative enzymes, phenolics
can also scavenge a considerable fraction of free radicals. A greater
H-donating capacity and radical stabilization make phenolics
suitable for this purpose [12].

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Breeding crops for salt-tolerance remains a long-term, cumbersome, and complex phenomenon. Hence, the quest to cope with crop
production under saline conditions remains a challenging task for
crop scientists. Crops and their various cultivars exhibit differential
response to salinity [6] and are characterized as salt-tolerant and
sensitive. Under the present scenario, a better approach may be the
screening of crop cultivars for salt tolerance. Wheat (Triticum
aestivum L.) is the prominent cereal food grain and major staple food
worldwide [14]. Food security in Pakistan is directly related to wheat
production and consumption trends. Over years, wheat productivity
is threatened by looming water crises, uncertainty of climatic
optima and increased incidence of salinity. With dwindling land
resources and little scope for vertical expansion, food requirement
of a burgeoning population need to be met from the available
natural resources. This warrants the need to improve the yield per
unit area and also to cope with crop production even on saline soils.
Comparing the cultivar response of one species to salinity
provides a convenient and useful tool for elucidating fundamental
mechanisms responsible for salt-tolerance. Such information is
crucial to suggest a suitable wheat cultivar for salt-affected soils. The
present study tends to assess the emergence, seedling growth, and
biochemical responses of some wheat cultivars toward salinity.
Underlying biochemical bases of differential salt-tolerance among
wheat cultivars are also discussed.

2 Materials and methods

2.1 Seed procurement
Seeds of six wheat cultivars (hexaploid) viz., LU-26S (salt tolerant) [15],
MH-97 (salt sensitive) [12], AARI-2011, Millat-2011, Lasani-2008, and
FSD-2008 were obtained from Ayub Agriculture Research Institute,
Faisalabad. The salinity tolerance of AARI-2011, Millat-2011, Lasani2008, and FSD-2008 cultivars was not known previously. Seeds were
surface sterilized with 30% ethanol for 3 min, washed thrice with
deionized water to remove traces of ethanol, and were subsequently
dried between layers of lter paper [16].

2.2 Soil salinization

Field soil collected from a depth of 015 cm was dried and wellmixed. A sodium hexametaphosphate based dispersion of soil
particles was produced to assess particle size distribution. A
proportion of sand, silt, and clay was determined based on the
rate of their settlement using a hydrometer [17]. The soil was a sandy
clay loam with sand, silt, and clay of 48.20, 23.47, and 28.33%,
respectively. A saturated soil paste was made by adding distilled
water to 250 g soil and stirring with a spatula and was allowed to
stand for 2 h to attain equilibrium. The extract of the saturated soil
paste was obtained by ltering through a lter press assembly using
a vacuum pump (N820.3FT18, KNF Neuberger, Germany). The pH (7.6)
of the saturated soil paste and electrical conductivity (EC) of the
saturation extract (ECe, 0.97 dS m
1) were determined as per Ryan
et al. [18] using a digital conductivity meter (HI-9811, Hanna
Instruments, USA). The soil saturation percentage based on water
loss due to oven drying at 105C was 24.78%, and was calculated:
Saturation%

Loss in soil weight on drying

 100
Weight of soil after drying

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1249

The amount of NaCl required for attaining EC values of 8 and

16 dS m
1 in soil (as per treatment) was calculated based on the EC
difference (required soil ECinitial soil EC) as follows:
NaCl requiredg=kg

TSS  58:5  Saturation%

100  1000

where TSS denotes total soluble salts (mEq L
1) and was calculated by
multiplying the EC difference with a factor of 10 and 58.5 (molecular
weight of NaCl). To obtain EC values of 8 and 16 dS m
1 in soil, the
amount of NaCl salt was calculated as 1.019 and 2.178 g kg
1 soil,
respectively. To achieve a homogeneous salt distribution, NaCl
(Analytical grade, Merck, Darmstadt, Germany) was thoroughly
mixed with a given amount of eld soil using a small scale cement
mixer.

2.3 Seed bioassay

Thermocol trays (15  9 cm2) were lled with soil (1000 g) of
respective EC (8 and 16 dS m
1). Un-amended (without NaCl) eld
soil with an original EC value of 0.97 dS m
1 was used in control
trays. Fifteen seeds of respective wheat cultivars were uniformly
sown in each tray at eld capacity. The trays were placed in a screen
house under natural solar conditions with a 10/14 h light/dark
photoperiod. Trays were uniformly irrigated with distilled water as
per requirement to avoid water decit. Emergence counts were
made on a daily bases according to AOSA [19] until a constant count
was achieved. Seeds were considered as emerged when the hypocotyl
length was 2 mm. The time taken to 50% emergence of seedlings
(E50) was calculated according to the modied formula of Farooq
et al. [20]:
E50 ti

N
2



ni tj
ti
nj
ni

where N is the nal number of emerged seeds, and ni and nj are the
cumulative number of seeds emerged by adjacent counts at times ti
and tj where ni < N/2 < nj. The mean emergence time (MET) was
calculated according to Ellis and Robert [21]:
P
Dn
MET P
n

where n is the number of seeds, which were emerged on day D, and

D is the number of days counted from the beginning of emergence.
The emergence index (EI) was calculated as described by AOSA [22]:
EI

No: of emerged seeds

No: of emerged seeds
:::::::::
Days of first count
Days of final count

The root and shoot length of ve randomly selected plants from

each replicate was measured at day 35 after sowing with a
measuring tape. Seedling roots and shoots of ve randomly selected
seedlings were oven dried separately at 70C for 48 h and weighed
thereafter using an electric balance (TX323L, Shimadzu, Japan). Total
seedling biomass was calculated as the sum of biomass of roots and
shoots. The numbers of leaves and secondary roots were manually
counted. Seedling thickness (cm) was recorded using a digital
Vernier Caliper.

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A. Khaliq et al.

2.4 Biochemical analyses

Lipid peroxidation in leaves of wheat seedling was determined as
MDA content following the thiobarbituric acid method [23]. The
absorbance of the supernatant was determined at 532 nm on a UVspectrophotometer (UV-4000, ORI, Germany) and corrected for nonspecic absorbance at 600 nm. The MDA content was calculated
using an extinction coefcient of 155 mM
1 cm
1. For the extraction
of antioxidant enzymes, 0.5 g leaf sample was homogenized in 10 mL
of chilled potassium phosphate buffer (pH 7) containing 1 mM EDTA
and 1% (w/v) polyvinyl pyrrolidone at 4C. The homogenate was
squeezed through four layers of cheese cloth. The extract was
centrifuged at 15 000 rpm for 15 min and the supernatant was stored
at 4C for further determination of antioxidants. Soluble proteins
were quantied as per Bradford [24]. Superoxide dismutase (SOD)
activity was determined as described by Giannopotitis and Ries [25].
One unit of SOD activity was dened as the amount of enzyme
inhibiting the photochemical reduction of nitro-blue tetrazolium
by 50%/min at 560 nm. CAT activity based on the consumption of
H2O2 was determined using the method of Dhindsa et al.[26]. The
consumption of H2O2 was measured at 240 nm and one unit of CAT
was dened as the amount of enzyme required to oxidize 1 mM
H2O2 min
1. Peroxidase (POX) activity was recorded as described by
Egley et al. [27]. An increase in the absorbance due to guaiacol oxidation
was measured at 470 nm. One unit of enzyme activity was dened

as the amount of enzyme required to oxidize 1 mM guaiacol min
1.

Total soluble phenolics in leaves and roots of wheat seedlings were
determined as described by Randhir and Shetty [28]. Photosynthetic
pigments (chl a and chl b) in leaves were extracted in 80% ice cold
acetone and measured at 663 and 645 nm [29].

2.5 Experimental design and statistical analyses

Studies were carried out following a completely randomized design
under factorial arrangement with seven replications, and repeated
once in the same season. Since the results of two runs of the
experiment were statistically similar (p  0.05), data were pooled for
further statistical analyses [30]. Least signicance difference test at
p  0.05 was employed to compare treatments means. Correlation
analyses were done to ascertain relationship among different
variables.

3 Results
3.1 Emergence attributes
Salinity had a deleterious effect on emergence attributes of wheat
(Figs. 1 and 2) that varied signicantly (p  0.05) among salinity
levels, wheat genotypes, and their interaction. The time to start

Figure 1. Inuence of salinity levels on (a, b) time to start emergence, and (c, d) E50 of six wheat cultivars. Vertical bars above mean denote standard error
of seven replicates. Means with different letters differ signicantly at 0.05 probability level by LSD test. LSD for salinity levels and cultivars (a) 0.764, (b)
1.081, (c) 0.705, and (d) 0.996.
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Figure 2. Inuence of different salinity levels on (a) nal emergence, (b) MET, and (c) emergence index of six wheat cultivars. Vertical bars above mean
denote standard error of seven replicates. Means with different letters differ signicantly at 0.05 probability level by LSD test. LSD for salinity level  wheat
cultivar interaction is (a) 2.197, (b) 13.02, and (c) 1.546, respectively.

emergence and E50 values were signicantly (p  0.05) delayed at

higher salinity levels (16 dS m
1) over control (Fig. 1a and b). The
time to start emergence was delayed by more than two days at upper
limits of salinity (Fig. 1a). Likewise, salinity levels of 8 and 16 dS m
1
also resulted in 21 and 80% delay in E50 (Fig. 1c) over control. A
cultivar-specic response was also evident (p  0.05) and the wheat
cultivar Lasani-2008 needed less time to complete 50% emergence
(Fig. 1d). The wheat cultivar Lasani-2008 recorded signicantly low
MET under control compared to all other wheat cultivars (Fig. 2a).
Nevertheless, AARI-2011, Lasani-2008, and FSD-2008 recorded
signicantly lower but similar MET under 16 dS m
1 salinity. A
maximum MET was taken by the wheat cultivars LU-26S, MH-97, and
Millat-2011 at the salinity level of 16 dS m
1. Increasing salinity
levels lowered nal emergence percentage and emergence index of
wheat cultivars and interactive effects were signicant for these two
emergence attributes (Fig. 2b and c). MH-97 appeared quite salinity 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim

sensitive at 16 dS m
1, and its nal emergence percentage dropped

by 56% over control which was 23% for AARI-2011. Cultivar
differences were more pronounced at high salinity levels and a
lowest emergence index (1.12) was recorded for MH-97 at the salinity
level of 16 dS m
1 whereas all other wheat cultivars were statistically
alike (p  0.05).

3.2 Seedling growth

Salinity levels, wheat genotypes, and their interaction depicted
signicant (p  0.05) variations in seedling growth attributes (Fig. 3).
Maximum reductions in seedlings growth attributes were observed
at higher salinity levels (16 ds m
1) in all cultivars, however, the
response of the cultivars was different to varying levels of salinity.
Under control treatment, MH-97 and Millat-2011 recorded the
maximum root lengths (13.87 and 13.10 cm), while the minimum

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A. Khaliq et al.

Figure 3. Inuence of salinity levels on seedling growth of six wheat cultivars. Vertical bars above mean denote standard error of seven replicates. Means
with different letters differ signicantly at 0.05 probability level by LSD test. RL: root length, SL: shoot length, RDB: root dry biomass, SDB: shoot dry
biomass, TDB: total dry biomass. LSD (p  0.05) for salinity level  wheat cultivar interaction is (a) 1.025, (b) 1.0301 (c) 2.929, (d) 4.617, (e) 5.574, and (f)
0.316, respectively.

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(9.51 cm) was recorded for LU-26 and Lasani-2008 (Fig. 3a). Interestingly, all wheat cultivars recorded a similar shoot length under
control conditions (Fig. 3b). At a higher salinity (16 dS m
1) level,
AARI-2011, Lasani-2008, and FSD-2008 recorded the highest and
similar root length compared with other wheat cultivars. AARI-2011
and FSD-2008 also recorded higher shoot lengths when compared
with other cultivars. Both root and shoot length of the cultivar
LU-26S (Fig. 3a and b) was most suppressed at higher a salinity level
(16 dS m
1) when compared with all other cultivars. The biomass
accumulation in seedling roots and shoots was also restricted
(Fig. 3c and d) as a result of the exposure to salinity in soil.
Wheat cultivars behaved statistically alike for root dry biomass
under controlled conditions. The salinity  cultivar effect was
more pronounced under saline conditions. Lasani-2008 yielded
the maximum root dry biomass at salinity levels of 8 dS m
1 while
AARI-2011 and Millat-2011 had a similar root dry weight at
16 dS m
1. Despite the highest shoot dry biomass under control
conditions, LU-26S revealed a drastic reduction for a salinity level
of 16 dS m
1. AARI-2011, Millat-2011, FSD-2008, and Lasani-2008
were statistically similar with each other regarding shoot dry
biomass at higher salinity levels (16 dS m
1). Total seedling
dry biomass also followed the same trend compared with root
dry biomass. Lasani-2008 and AARI-2011 showed the greatest
seedling dry biomass at salinity levels of 8 and 16 dS m
1,
respectively (Fig. 3e). The root number (# seedling
1) was
signicantly affected by salinity levels and wheat cultivars
whereas their interaction was non-signicant. Root score was
reduced by 11 and 31% at salinity levels of 8 and 16 dS m
1,
respectively (Fig. 4a). Among wheat cultivars, AARI-2011 recorded
the maximum root number, while the minimum was observed for
Millat-2011(Fig. 4b). In contrast to root score, a signicant
interactive effect of wheat cultivars and salinity levels was
observed for leaf score. The lowest leaf number (# seedling
1)
was recorded for LU-26S and MH-97 at the salinity level of
16 dS m
1 (Fig. 3f). Nonetheless, MH-97, AARI-2011, Lasani-2008,
and FSD-2008 recorded higher leaf score than other cultivars and
were statistically alike (p  0.05). The seedling thickness remained
similar irrespective of wheat cultivars and only the salinity stress
showed a signicant (p  0.05) effect (Fig. 5). The average across
different cultivars, seedling thickness was reduced by 8 and 22%
at salinity levels of 8 and 16 dS m
1, respectively.

1253

Figure 5. Inuence of salinity levels on seedling thickness. Vertical bars

above mean denote standard error of seven replicates. Mean with different
letters differ signicantly at 0.05 probability level by LSD test. LSD for
salinity level is 0.0176.

3.3 Biochemical attributes

Wheat seedlings growing under saline conditions manifested
reduced chlorophyll content although the cultivar specic response
was pronounced due to their signicant (p  0.05) interaction with
salinity levels (Fig. 6a). The chlorophyll contents of MH-97, Millat2011, and Lasani-2008 were reduced over control only at 16 dS m
1 in
contrast to AARI-2011 and FSD-2008 where total chlorophyll
declined even at the lower salinity level of 8 dS m
1 continuing
further at 16 dS m
1. The highest reduction in chlorophyll content
was recorded at 16 dS m
1 for MH-97 and Millat-2011. The root
phenolic content depicted an increase under saline conditions
compared with control with the exception of the salinity-susceptible
MH-97, for which root phenolic content declined with the increase
in salinity (Fig. 6b). At the salinity level of 16 dS m
1, Millat-2011
expressed signicantly higher root phenolics than all other wheat
cultivars. Leaf phenolic content of LU-26S, MH-97, and AARI-2011
showed a considerable increase over control under salinity,
contrarily, these were either dropped or remained at par (p  0.05)
with control for certain cultivars, viz., Millat-2011, Lasani-2008, and

Figure 4. Inuence of salinity levels and cultivars on number of roots. Vertical bars above mean denote standard error of seven replicates. Means with
different letters differ signicantly at 0.05 probability level by LSD test. LSD for salinity levels and cultivars (a) 0.380 and (b) 0.537, respectively.
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Figure 6. Inuence of different salinity levels on (a) total chl content, (b) root phenolic content, (c) and shoot phenolic content, and (d) total soluble
proteins of six wheat cultivars. Vertical bars above mean denote standard error of seven replicates. Means with different letters differ signicantly at 0.05
probability level by LSD test. LSD for salinity level  wheat cultivar interaction is (a) 0.279, (b) 5.967, (c) 7.455, and (d) 9.285, respectively.

FSD-2008 (Fig. 6c). Soluble proteins in leaves varied greatly as a

function of both wheat cultivars and the respective salinity levels
(Fig. 6d). There was a diminishing trend of soluble proteins in all
wheat cultivars at increasing levels of salinity except AARI-2011, for
which soluble proteins were higher than control at both salinity
levels.
Activities of antioxidants (enzymatic) varied signicantly
amongst cultivars, salinity levels, and their interactive effect
(Fig. 7). Regarding SOD, the interactive effect was signicant
(p  0.05) (Fig. 7a). At 16 dS m
1, the SOD activity of the wheat
cultivars AARI-2011 and FSD-2008 was 56 and 15% higher than
control. Contrarily, the SOD activity at this particular salinity was
far less than control (9, 48, 43, and 33%) for the cultivars LU-26S, MH97, Millat-2011, and Lasani-2008, respectively. The interactive effect
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of cultivars and salinity levels was also signicant for CAT. An

increasing salinity level (8 dS m
1) increased the CAT activity that
declined as the salinity level elevated further (16 dS m
1). However,
AARI-2011 depicted still a higher activity of CAT despite the increase
in the salinity level (Fig. 7b). LU-26S exhibited maximum CAT activity
at 8 dS m
1 followed by AARI-2011 and Millat-2011. Nonetheless at
the upper salinity level (16 dS m
1), AARI-2011 and Millat-2011
showed a higher CAT activity and were statistically similar (p  0.05).
Salt stress generally aggravated lipid peroxidation so that the MDA
content was considerably higher in wheat seedlings growing under
NaCl salinity (Fig. 7c). Although the MDA content depicted a gradual
increase with increasing salinity level yet cultivar-specic response
was evident because of signicant interactions. At a higher salinity
(16 dS m
1) level, Lasani-2008 showed the highest MDA content.

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Figure 7. Inuence of different salinity levels on the activity of (a) SOD, (b) CAT, and (c) MDA content of six wheat cultivars. Vertical bars above mean
denote standard error of seven replicates. Means with different letters differ signicantly at 0.05 probability level by LSD test. LSD (p  0.05) for salinity
level  wheat cultivar interaction is (a) 12.341, (b) 0.091, and (c) 8.861, respectively.

FSD-2008 exhibited a lower MDA content and was at par with MH-97
at the same salinity level. Regarding POX, only the salinity had a
signicant effect and the POX activity increased at the salinity level
of 8 dS m
1 over control (Fig. 8). However, it was at par (p  0.05) with
control at higher salinity levels (16 dS m
1).

3.4 Correlation between variables

MDA was negatively correlated with the nal emergence percentage
in wheat of all cultivars except for FSD-2008 (Table 1). SOD was
positively correlated with root length and shoot length in MH-97 and
Lasani-2008. Contrarily, these were negatively correlated for AARI2011 and FSD-2008, however, no correlation was observed in LU-26S
and Millat-2011 for root and shoot length. The correlation between
SOD and seedling dry biomass was positive for MH-97, Millat-2011,
and Lasani-2008, but negative for AARI-2011 and non-signicant for
LU-26S and FSD-2008. POX was positively correlated with root length
and shoot length in Lasani-2008. In Millat-2011 and Lasani-2008, a
positive correlation was observed for POX and seedling dry biomass.
The wheat cultivars MH-97 and Millat-2011 showed a positive
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Figure 8. Inuence of salinity levels on POX activity. Vertical bars above

mean denote standard error of seven replicates. Mean with different letters
differ signicantly at 0.05 probability level by LSD test. LSD for salinity level
is 0.0636.

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Table 1. Correlation analyses showing strength of association between different variables

X-Variable
MDA
SOD
POX
CAT
Phenolics

Y-Variable

LU-26S

FEP
RL
SL
SDB
RL
SL
SDB
RL
SL
SDB
Chlorophyll

0.923
ns

0.278
0.292ns
0.187ns
0.144ns
0.159ns
0.051ns
0.291ns
0.306ns
0.200ns
0.999

MH-97

AARI-2011

0.741
0.997
0.997
0.999
0.240ns
0.243ns
0.358ns
0.999
0.999
0.988
0.956

0.812
0.938
0.844
0.916
0.051ns
0.259ns
0.109ns
0.411ns
0.594ns
0.464ns
0.774

Millat-2011
0.994
ns
0.590
0.488ns
0.839
0.554ns
0.449ns
0.814
0.728
0.806
0.435ns
0.999

Lasani-2008


0.907
0.978
0.997
0.963
0.932
0.972
0.907
0.352ns
0.224ns
0.410ns
0.228ns

FSD-2008
0.689ns
0.829
0.777
0.633ns
0.542ns
0.468ns
0.279ns
0.515ns
0.439ns
0.247ns
0.533ns

FEP, nal emergence percentage; ns, non-signicant; RL, root length; SDB, shoot dry biomass; SL, shoot length.

p < 0.05,  p < 0.01,  p < 0.001.

correlation for CAT with root length and shoot length. CAT and
seedling dry biomass were strongly correlated in MH-97. Phenolics
had positive correlation with chl in Millat-2011, while negative in
LU-26S, MH-97, and AARI-2011.

4 Discussion
Salinity had an overall negative implication for emergence
attributes of wheat cultivars. Such negative consequences of salinity
on wheat emergence might have arisen from the reduction in water
uptake (osmotic effect) and specic ion-toxicity [3]. Salinity either
delayed the emergence commencement or inhibited the same and
the cultivar-specic responses were evident in some instances (Figs. 1
and 2). Time to start emergence, E50 and MET increased, whereas
nal emergence percentage and emergence index decreased with
the increase in salinity. Salinity led to a considerable inhibition or
delay in germination and seedling establishment in durum
wheat [31]. Seedling emergence of a plant is considered as the
most critical stage of a plant under salinity stress [31]. Variation was
also observed among wheat cultivars for various emergence
attributes and differences were more pronounced at higher salinity
levels. The wheat cultivar AARI-2011 performed better under salinity
as was depicted by least reduction in nal emergence than other
cultivars. Better performance of this cultivar under salinity might be
related to differential regulation of emergence processes at
molecular and physiological levels [32]. Wheat has been reported
as a moderately salt-tolerant plant [34] with a threshold salinity level
of 6 dS m
1 [33]; however, it can thrive even at 10 dS m
1, but with a
signicant reduction in yield [34]. Here, the upper limit of salinity
was quite high (16 dS m
1). The emergence and seedling survival by
wheat cultivars AARI-2011 and FSD-2008 at this salinity level seems
quite interesting and eye-catching.
Suppressive effects of salinity on wheat seedling growth were
manifested in the form of reduced elongation of roots and shoots, as
well as less biomass accumulation (Figs. 35). The interaction
between salinity and wheat cultivars was also signicant (p  0.05)
for various attributes. Differential response of wheat cultivars
toward salinity may be due to variable bio-chemical and physiological efciencies pertaining to stress tolerance. A diminished seedling
growth of wheat under salinity may be due to osmotic effects as well
as specic ion toxicity presumably because of salt accumulation in
the intercellular spaces [35]. Disturbances in seedling morphology
under salinity are a secondary expression of salt-induced damage to
2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim

cell organelles and its interference with vital physiological

processes. Genotypes within a species manifest immense variation
for salt tolerance and same was observed during the present study.
Recently, Hussain et al. [6] reported signicant differences among
wheat cultivars regarding their ability to accumulate shoot dry
biomass in response to increasing NaCl salinity. Such differences
seem to have originated in part due to variable leaf emergences,
their elongation and expansion rates [36]. Another possible
explanation of a reduced biomass under salinity could be the loss
of cell turgor due to massive exclusion of Na ions. Interestingly,
during the present study, the seedling growth of LU-26S (previously
recognized as salt-tolerant cultivar) was severely affected even then
MH-97 (moderately salt sensitive) at 16 dS m
1, although these two
cultivars were statistically similar for biomass accumulation at
8 dS m
1. Such a contrasting result might be due to differences in
salinity levels maintained as well as initial seed vigor of two wheat
cultivars. Nevertheless, this aspect needs to be further claried in
future studies.
Increasing salinity levels recorded an increase in MDA content
suggesting higher lipid peroxidation (Fig. 7c). Nevertheless, salttolerant wheat cultivars were better able to survive lipid peroxidation as evident by lower values of MDA in these cultivars. Reduced
emergence and impaired growth of salt-sensitive wheat cultivars
coupled with their enhanced MDA contents suggest more lipid
peroxidation, electrolyte leakage, and compliments the ndings of
Ashraf et al. [12]. The signicance of lipid peroxidation as a salttolerant trait has also been reported for other cereals like barely [37]
and sorghum [38]. A decrease in chl content was observed for wheat
cultivars under salinity and the magnitude of reduction varied
among the cultivars. Salt stress deteriorates many potential
biomolecules such as chl [10, 39]. A variable reduction in chl
content of wheat seedlings under salinity is in line with the nding
of Saqib et al. [40]. Ashraf and Ashraf [41] also documented lower chl
content in the salt-sensitive MH-97 cultivar compared with the salttolerant S-24. A reduction in chl may be an outcome of the salinityinduced degradation or impaired chl biosynthesis due to the decline
in the endogenous content of 5-aminolevulinic acid, a precursor for
protochlorophyllide [42]. A reduced chl content observed in this
study might also be a cross-stress response to salinity induced lipid
peroxidation (Fig. 6a) and ROS related damage to the photosynthetic
system. A reduction in chl content lowers the photosynthesis
rate [43] and explains lower seedling biomass accumulation under
salinity in salt-sensitive wheat cultivars in the present study.

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Soil

Root and leaf phenolic content also varied signicantly with

increasing salinity levels and cultivars (Fig 6b and c). The involvement
of phenolics in salinity tolerance in wheat and brassica has been
reported earlier [12, 16]. Soluble proteins decreased in four out of six
cultivars in response to salinity and are supported by the ndings of
Parida et al. [44]. Under salinity, ROS (superoxides, hydroxyl, and
peroxy radicals) tend to accumulate due to hyper-osmotic and ionic
effects. Plants usually employ enzymatic and non-enzymatic antioxidants to detoxify ROS. Our data depicted a considerable increase in
the activities/levels of various antioxidant enzymes (Fig. 7). Asharf [8]
suggested that production and expression of these enzymes is
enhanced primarily to counteract salinity-induced ROS. Higher
activities of these enzymes in wheat cultivars exhibiting better
emergence and seedling growth suggested their relevance to salinity
tolerance in such cultivars. Sairam et al. [45] reported greater increase
in the activities of SOD, glutathione reductase, and ascorbate
peroxidase in salt-tolerant wheat cultivars as compared with sensitive
ones. Salt-stress generally increased the activities of antioxidants and
such an increase was more pronounced in salt-tolerant wheat cultivars
(Fig. 7). Neto et al. [46] also supported higher activities of antioxidants
in salt tolerant species. A positive correlation of activities of
antioxidant enzymes with stress tolerance has been documented [32,
47], and their enhanced expression has been suggested as a stress
marker. The decline observed in the activities of some antioxidants for
several salt-sensitive wheat cultivars at upper limits of salinity might
be due to their inability to cope with stress induced oxidative damage.

5 Conclusions
In crux, the present study revealed the variable salinity tolerance
among wheat cultivars as evident by their emergence, seedling
growth, and biochemical attributes recorded under salinity.
Regarding nal emergence percentage and dry biomass, AARI2011, Millat-2011, and FSD-2008 performed better. Better performance of AARI-2011 could be explained in terms of its efcient
antioxidant system while that of the FSD-2008 is due to less lipid
peroxidation even at upper limits of salinity (16 dS m
1). Field
studies are, however, needed to validate these results. Moreover,
future studies should assess the accumulation of compatible solutes,
cationic homeostasis, and physiological processes in these wheat
cultivars under salinity.
The authors have declared no conicts of interests.

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