I

RESPONSE OF MAIZE (Zea mays L.) TO SALINITY AND POTASSIUM SUPPLY

By

TAHIR MAQSOOD M. Sc. (Hons.) Soil Science

A thesis submitted in partial fulfillment of requirement for the degree of

DOCTOR OF PHILOSOPHY

IN

SOIL SCIENCE

INSTITUTE OF SOIL & ENVIRONMENTAL SCIENCES UNIVERSITY OF AGRICULTURE, FAISALABAD

PAKISTAN 2009

II

To The Controller of Examination University of Agriculture, Faisalabad. “We, the Supervisory Committee, certify that the contents and form of thesis

submitted by Mr. Tahir Maqsood have been found satisfactory and recommended that it

be processed for evaluation by External Examiner (s) for the award of degree.

Supervisory Committee

________________________ 1. Chairman : (DR. JAVAID AKHTAR) ________________________ 2. Member : (DR. M. ANWAR-UL-HAQ) _______________________ 3. Member : (DR. RASHID AHMAD)

III

DEDICATED

TO

MY FATHER

MUHAMMAD TUFAIL (LATE)

WHO LOVED ME

&

TO WHOM I MISS

IV

ACKNOWLEDGEMENT

Up and above anything else, all praises to The Allah

Almighty alone, the Omnipotent, the Merciful and

Compassionate. Knowledge is limited and time is short to

express His dignity, the Propitious, the Benevolent and

Sovereignty, Whose blessings and glories have flourished

my thoughts and thrived my ambitions.

Trembling lips and wet eyes pray for the Holy Prophet

Hazrat Muhammad (PBUH) for enlightening our conscience

with an essence of faith in Allah, converging all His

kindness and mercy upon him. I offer my thanks from the

core of my heart to The Holy Prophet Hazrat Muhammad

(PBUH) who is forever a torch of knowledge and guideline

for humanity as a whole.

With humble, profound and deep sense of devotion I

wish to record my supervisor, Professor Dr. Javaid Akhtar,

Institute of Soil and Environmental Sciences, for his

reliable comments, dynamic supervision and sincere help

throughout the course of my studies. I am highly indebted

to Professor Dr. Atta Muhammad Ranjha,, Institute of Soil

and Environmental Sciences, Assistant Professor Dr.

Muhammad Anwar-ul-Haq and Professor Dr. Muhammad Rashid,

Department of Crop Physiology for their friendly behavior,

willing cooperation, sincere help and inspiring guidance

throughout the course of this research work.

V

I am indebted to my Parents for their love,

prayers and moral support through out my studies at

University. I extend my thanks especially to my loving

wife, daughters (Fatima Tahir & Fakiha, Tahir) son

(Muhammad.Abdullaha Tahir) for their consistent patience,

sincere moral support and encouragement during whole of

study. They always raised their hands for my success to

achieve this goal. May Almighty Allah bless us the wealth

of “Emaan” and better health (Aameen). I am also thankful

to the Higher Education Commission, Islamabad, for

providing financial assistance to achieve this goal.

Tahir Maqsood

LIST OF CONTENTS

Chapter Contents Page Chapter1 Introduction 1 Chapter2 Review of literature 6

2.1 Extent of salinity problem at national and global levels 6 2.2 Effect of salinity on plant physiology and morphology 7 2.3 Effects of salinity on crop yields 9

2.3.1 Osmotic stress 9 2.3.2 Specific ion toxicity 10 2.3.3 Nutritional imbalance 12 2.4 Effect of salinity on maize crop 14 2.5 Mechanism of crop tolerance to salinity 23

Chapter 3 Materials & Methods 31 3.1 Growth conditions & experimental techniques 31

3.1.1 Experimental site 31 3.1.2 Seed source 31 3.1.3 Solution culture studies 32

VI

3.1.4 Pot culture studies 32 3.1.5 Analysis of soil in the pot experiment 33 3.1.6 Field experiment 33 3.2 Plant analysis 34

3.2.1 Plant sample collection 34 3.2.2 Leaf sap extraction 34 3.2.3 Determination of Na+ and K+ 34 3.2.4 Determination of Cl- 35 3.3 Physiological attributes 35

3.3.1 Determination of water potential 35 3.3.2 Gas exchange characteristics 35 3.3.3 Growth parameters 35 3.4 Biochemical attributes 36

3.4.1 Protein analysis 36 3.4.2 Determination of total soluble sugars 36 3.4.3 Oil contents determination 37 3.4.4 Crude fiber determination 38 3.4.5 Proline determination 38 3.5 Statistical analysis 38

Chapter 4 RESULTS AND DISCUSSIONS Study-1 Screening of maize genotypes for salt tolerance in solution culture 41

4.1 Results and discussions 41 4.1.1 Results 41 4.1.2 Physical Growth Parameters 41

4.1.2.1 Shoot length (cm) 41 4.1.2.2 Root length (cm) 42 4.1.2.3 Shoot fresh weight (g plant-1) 43 4.1.2.4 Shoot dry weight (g plant-1) 45 4.1.2.5 Root fresh weight (g plant-1) 46 4.1.2.6 Root dry weight (g plant-1) 47 4.1.2.7 Sodium concentration in leaf sap of maize genotypes 48 4.1.2.8 Potassium concentration in leaf sap of maize genotypes 49 4.1.2.9 K+: Na+ ratio in leaf sap of maize genotypes 50 4.1.2.10 Chloride concentration in leaf sap of maize genotypes 52

4.1.3 Discussion 53 Study-2 Differences in salt tolerance and potassium requirement of maize

genotypes a hydroponics study 60

4.2 Results and discussion 60 4.2.1 Results 60

4.2.1.1 Shoot length/ plant height (cm) 60 4.2.1.2 Root length 62 4.2.1.3 Shoot fresh weight (g palnt-1) 64 4.2.1.4 Shoot dry weight (g palnt-1) 66 4.2.1.5 Root fresh weight (g palnt-1) 68 4.2.1.6 Root dry weight (g palnt-1) 70 4.2.1.7 Sodium concentration in leaf sap of maize 72 4.2.1.8 Potassium concentration in leaf sap of maize 74 4.2.1.9 K+: Na+ ratio in leaf sap of maize 76

VII

4.2.1.10 Chloride concentration in the leaf sap of maize 76 4.2.3 Discussion 79

Study-3 Growth response of salt tolerant and salt sensitive maize genotypes to salinity and potassium supply

87

4.3 Results and discussion 87 4.3.1 Results 87

4.3.1.1 Plant height 87 4.3.1.2 Shoot fresh and dry weights 87 4.3.1.3 Grain yield (g plant-1) 88 4.3.1.4 1000 seed weight 88 4.3.2 Ionic concentration in leaf sap 89

4.3.2.1 Na+ concentration in expressed leaf sap 89 4.3.2.2 K+ concentration in leaf sap 90 4.3.2.3 K+: Na+ ratio in the leaf sap 90 4.3.2.4 Chloride concentration in leaf sap 90 4.3.3 Gas exchange parameters and water potential 91

4.3.3.1 Stomatal conductance 91 4.3.3.2 Transpiration rate 92 4.3.3.3 Water Potential 92 4.3.4 Response of maize genotypes to salinity stress and K supply

(Biochemical attributes) 93

4.3.4.1 Seed oil contents 93 4.3.4.2 Crude fiber 94 4.3.4.3 Protein contents 94 4.3.4.4 Total soluble sugars 95 4.3.4.5 Proline contents 96 4.3.5 Discussion 96

Study-4 Physiological and biochemical attributes of maize genotypes in saline soil

103

4.4 Results and discussion 103 4.4.1 Results 103

4.4.1.1 Plant height/shoot length 103 4.4.1.2 Total biomass (g plant-1) 103 4.4.1.3 Grain yield 103 4.4.1.4 Na+ concentration in leaf sap of maize genotypes 104 4.4.1.5 K+ concentration in leaf sap of maize genotypes 104 4.4.1.6 K+: Na+ ratio 105 4.4.1.7 Chloride concentration in leaf sap 105 4.4.1.8 Total soluble sugars 106 4.4.1.9 Proteins contents 106 4.4.1.10 Seed oil contents 107 4.4.1.11 Crude fiber 107 4.41.12 Proline contents 107 4.4.2 Discussion 108

Chapter 5 Summary 114 Chapter 6 Literature cited 118 Chapter 7 Appendices 140

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LIST OF TABLES Table TITLE Page 3.1 Properties of soil used in pot the experiment 33

3.2 Properties of the salt affected soil used in the experiment 34 3.3 A brief of all experiments and the parameters monitored in each

experiment 39

4.1 Shoot length (cm) of maize genotypes grown under different salinity levels and harvested at 4 weeks age

42

4.2 Root length (cm) of maize genotypes grown under different salinity levels and harvested at 4 weeks age

43

4.3 Shoot fresh weight (g plant-1) of maize genotypes grown under different salinity levels and harvested at 4 weeks age

44

4.4 Soot dry weight (g plant-1) of maize genotypes grown under different salinity levels and harvested at 4 weeks age

46

4.5 Root fresh weight (g plant-1) of maize genotypes grown under different salinity levels and harvested at 4 weeks age

47

4.6 Root dry weight (g plant-1) of maize genotypes grown under different salinity levels and harvested at 4 weeks age

48

4.7 Na+ concentration (mol m-3) in leaf sap of maize genotypes grown under different salinity levels and harvested at 4 weeks age

49

4.8 K+ concentration (mol m-3) in expressed leaf sap of maize genotypes grown under different salinity levels and harvested at 4 weeks age

50

4.9 K+: Na+ ratio in leaves of maize genotypes under different treatments grown under different salinity levels and harvested at 4 weeks planting

51

4.10 Cl- concentration (mol m-3) in leaf sap of maize genotypes grown under different salinity levels and harvested at 4 weeks age

53

4.11 Shoot length (cm) of maize genotypes grown in nutrient solution under different salinity treatments and harvested at 4 weeks age

61

4.12 Root length (cm) of maize genotypes grown in nutrient solution under different salinity treatments and harvested at 4 weeks age

63

4.13 Shoot fresh weight (g plant-1) of maize genotypes grown in nutrient solution under different salinity treatments and harvested at 4 weeks age

65

4.14 Shoot dry weight (g plant-1) of maize genotypes grown in nutrient solution under different salinity treatments and harvested at 4 weeks age

67

4.15 Root fresh weight (g plant-1) of maize genotypes grown in nutrient solution under different salinity treatments and harvested at 4 weeks age

69

4.16 Root dry weight (g plant-1) of maize genotypes grown in nutrient solution under different salinity treatments and harvested at 4 weeks age

71

4.17 Na+ concentration (mol m-3) in leaf sap of maize genotypes grown in nutrient solution under different salinity treatments and harvested at 4 weeks age

73

4.18 K+ concentration (mol m-3) in the leaf sap of maize genotypes grown in nutrient solution under different salinity treatments and harvested at 4 weeks age

75

4.19 K+: Na+ in leaf sap of maize of maize genotypes grown in nutrient solution under different treatments

77

4.20 Cl- concentration (mol m-3) in the leaf sap of maize genotypes grown in nutrient solution under different salinity treatments and harvested at 4

78

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weeks age 4.21 Growth response of two maize genotypes grown under control

(1.25 dS m-1) and 10 dSm-1 at two levels of potassium 89

4.22 1000 seed weight and leaf ionic concentration of two maize genotypes grown under control (1.25 dS m-1) and 10 dS m-1 at two levels of potasium

91

4.23 Stomatal conductance (mmol.m-2.sec-1), transpiration rate (mmol.m-2.sec-1) and water potential (MPa) of two maize genotypes grown under control (1.25 dS m-1) and 10 dS m-1 at two levels of potassium

93

4.24 Oil contents (%), crude fiber (%) and protein contents (%) of two maize genotypes grown under control (1.25 dS m-1)and 10 dS m-1 at two levels of potassium

95

4.25 Total soluble sugars (%) and proline contents (µ mol g-1) of two maize genotypes grown under control (1.25 dS m-1) and 10 dS m-1 at two levels of potassium

96

4.26 Pearson correlation in shoot of maize of genotypes Akbar and S-2002 102 4.27 Pearson correlation in shoot of maize of genotypes Akbar and S-2002 102 4.28 Response of two maize genotypes to salinity and potassium addition 105 4.29 K+: Na+, Cl-, total soluble sugars, protein, crude fiber and proline

contents in maize genotypes grown in a saline field at two potassium levels

108

4.30 Pearson correlation in maize of genotypes Akbar and S-20002 113 4.31 Pearson correlation in maize of genotypes Akbar and S-20002 113

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LIST OF FIGURES Figure Title page 4.1 Relationship between shoot fresh weight and sodium concentration 57 4.2 Relationship between shoot fresh weight and chloride concentration in

leaves of maize genotypes 59

4.3 Relationship between shoot fresh weight and sodium concentration in leaves of 7 maize genotypes harvested at 4 weeks age

85

4.4 Relationship between shoot fresh weight and chloride concentration in leaves of 7 maize genotypes harvested at 4 weeks age

85

4.5 Relationship between shoot fresh weight and potassium concentration in leaves of 7 maize genotypes harvested at 4 weeks age

86

LIST OF ABBREVIATIONS SFW = Shoot Fresh Weight RFW = Root Fresh Weight SDW = Shoot Dry Weight RDW = Root Dry Weight SOP = Sulphate of Potash MOP = Murate of Potash MPa = MegaPascal Kharif Crop = A crop which is grown in spring season i.e. March-April.

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Chapter 1

INTRODUCTION

Maize (Zea mays L.) occupies a key position as one of the most important

cereals both for human and animal consumption and grown under various conditions

in different parts of the world. Maize grain has high food value and its oil is used for

cooking purposes while green fodder is quite rich in protein (Dowswell et al.,

1996). In Pakistan, maize is the third most important cereal after wheat and rice.

Being an important “Kharif” crop (spring crop), maize is grown on about one million

hectares with a total production of about 3.13 million tons and an average yield of

3264 kg ha-1 (Anonymous, 2008). In view of its increasing importance, improvement

in agronomic characteristics of maize has received considerable attention in Pakistan,

especially in salt affected soils.

The productivity of crops is adversely affected by high salt content in most

of the soils (Alam et al., 2000). Approximately, 7 % of the world’s land area, 20 % of

the world’s cultivated land, and nearly half of the irrigated land is affected with high

salt contents (Szabolcs, 1994; Zhu, 2001). In view of another projection, 2.1% of the

global dry land agriculture is affected by salinity (FAO, 2003). Effects of salinity are

more obvious in arid and semiarid regions where limited rainfall, high evapo-

transpiration, and high temperature associated with poor water and soil management

practices are the major contributing factors (Azevedo Neto et al., 2006). The

evaporation rate is generally high and exceeds that of precipitation in such regions.

Thus, the insufficient rainfall together with high evaporative demand and shallow

ground water in most locations enhances the movement of salts to the soil surface.

Improper irrigation practices and lack of drainage have aggravated the problem

leading to significant reductions in crop productivity (FAO, 2003).

Total area of Pakistan is 80 million hectares, with a good canal irrigated

system (62,400 km long) that covers an area of 19.43 million hectare (m ha) in the

Indus plains. Incidentally, the salt-affected soils are mainly confined to these plains

and are estimated to be 6.30 m ha (Alam et al., 2000). Intensive and continuous use

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of surface irrigation has altered the hydrological balance of the irrigated areas,

especially in the Indus basin. The substantial rise in the water table has caused

salinity and water logging in vast areas of all four provinces of the country, Punjab,

Sindh, NWFP and Balochistan. The magnitude of the problem can be assessed from

the fact that the area of productive land is being damaged by salinity at the rate of

about 40,000 ha annually (Alam et al., 2000).

Plants have developed a wide range of mechanisms to sustain crop

productivity under salt stress. Increasing evidence suggests that plant species and

varieties vary greatly in their resistance to salinity (Wahid et al., 1997; Akhtar et al.,

2003; Ashraf and Foolad, 2007). Selectivity in ion uptake and transport, vacuolar

compartmentation and even more integrated and complex response such as control of

water content and photosynthetic efficiency may determine the differential genotypic

response (Leidi and Saiz, 1997). According to Qadir and Schubert (2002), the degree

to which different plant species/genotypes can resist soil salinity may partly be

related to their abilities to selective absorption of K+ over Na+. Colmer and Flowers

(2006) summarized the characteristics of salt tolerant genotype including Na+

exclusion, K+/Na+ discrimination, retention of ions in leaf sheath, tissue tolerance, ion

partitioning into different aged leaves, osmotic adjustment, transpiration efficiency,

early vigor and early flowering leading to shorter growing season and the increased

water use efficiency.

Salt stress increases the accumulation of toxic ions such as Na+ and Cl- in

different plant parts, tissues, cells and cell organelles, for example, accumulation of

Na+ and/or Cl- takes place in the chloroplasts of higher plants or in the cytoplasm of

cyanobacterial cells, which affects growth rate, and is often associated with a

decrease in photosynthetic electron transport activities in photosynthesis (Kirst,

1989). Accumulation of excess Na+ and Cl- causes ionic imbalances that may impair

the selectivity of root membranes and induce potassium deficiency (Gadallah, 1999).

Furthermore, a decrease in nitrate reductase activity and an inhibition of photosystem

II and chlorophyll breakdown are associated with increased Na+ accumulation in

plant tissues (Krishnamurthy and Bhagwat, 1995). Cell membrane function is

perturbed due to Na+ replacing Ca2+ resulting in increased cell leakiness. Adequate

regulation and supply of mineral nutrition may help to improve plant growth under

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saline conditions (Akhtar et al., 2003). Of the mineral nutrients, K and Si have been

known for their particular role in enhancing salt tolerance of various crops (Mengel

and Kirkby, 2001; Liang et al., 2007). Excess Na+ and Cl- leads to the appearance of

symptoms like those of K+ deficiency. The deficiency of K+ initially leads to chlorosis

and then necrosis (Gopal and Dube, 2003).

Potassium is essential for protein synthesis, glycolytic enzymes and

photosynthesis; an osmoticum mediating cell expansion and turgor driven

movements and competitor of Na+ under salt stress (Hu and Schmidhalter, 2005).

Accumulation of Na+ and impairment of K+ nutrition is a major characteristic of salt

stressed plants. Therefore, K+/Na+ ratio in plants is considered a useful parameter to

assess salt tolerance (Cakmak, 2005b). Application of K fertilizer reduces the adverse

effects of salinity through its role in stomatal regulation, osmoregulation, energy

status, charge balance, protein synthesis and homeostasis (Marschner, 1995;

Sanjakkara et al., 2001).

Potassium is a major osmoticum, which contributes to osmotic adjustment,

stomatal movement and restriction of Na+ uptake under salinity (Mengel and Kirkby,

2001). Potassium content in plant tissues progressively decreases with an increase in

salinity, owing to higher absorption of Na+ (Grattan and Grieve, 1994). Maintenance

of adequate levels of K is important for plant survival in saline conditions (Chow

et al., 1990). Numerous studies have shown that K mitigates the adverse effects of

salinity on plant growth (Marschner, 1995 and Sanjakkara et al., 2001) by regulating

desirable K+/Na+ ratio, a good indicator of salt tolerance (Zhu, 2003).

Plants have developed a wide range of adaptive/resistance mechanisms to

maintain productivity and ensure plant survival under salt stress. Increasing evidence

suggests that mineral nutrient status of plants plays a crucial role in increasing plant

resistance to environmental stresses including salinity. Among various mineral

nutrients, potassium (K) has a particular role in contributing to plant survival in salt

stressed environment (Marschner, 1995; Mengel and Kirkby, 2001). Improvement of

salt tolerance by the addition of K has been reported in wheat (Shirazi et al., 2001),

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rice (Bohra and Doerffling, 1993), corn (Bar-Tal et al., 2004), tomato (Kaya et al.,

2001a) and cucumber and pepper (Kaya et al., 2001b).

Maintenance of adequate potassium levels is essential for plant survival in

saline habitats. Potassium is the most prominent inorganic plant solute, and as such

makes a major contribution to lower the osmotic potential in the stele of roots that is

a prerequisite for turgor-pressure-driven solute transport in xylem and the water

balance of plants (Marschner, 1995). Despite the presence of significant amount of

data showing reduced uptake and translocation of potassium by plants grown in high

Na+ substrates, there is very little information showing that the addition of potassium

to sodium-dominated soils improved plant growth or yield (Grattan and Grieve,

1999). Cerda et al. (1995) concluded that the beneficial effects of K-fertilizers

increased K+/Na+ within the plant. Potassium supplement reduced the concentration

of Na+ and increased the concentration of K+ in leaves (Kostas and Georgios, 2006).

Kaya et al. (2002b) concluded that the application of supplementary K

reduced the adverse effects of high salinity on plant growth and physiological

development in spinach and lettuce. While working on strawberry, Kaya et al.

(2002a) found that the plants grown at 35 mM NaCl produced less dry matter, fruit

yield and chlorophyll concentration than grown under normal solution. They found

that potassium, supplemented (3 mM K2SO4) in nutrient solution, resulted in an

increase in dry matter, chlorophyll concentration and fruit yield, while the

concentration of K was much lower in the plants grown at high NaCl concentration

and supplementary K application enhanced the K concentration within the plants.

They also concluded that supplementary K can improve plant growth and quality

under saline conditions. For the high salt treatment, supplementing the soil with

KNO3 at 1 g kg-1 resulted in K level similar to those of control. Salinity increased

Na+ and Cl- in tissues of salt tolerant and salt sensitive cultivars. Salt-toxicity

symptoms were observed at 100 and 150 mM NaCl, but not in plants receiving extra

K. Salinity increased mannitol contents up to 41.3% in salt tolerant and 15.8% in salt

sensitive but reduced the starch content in leaves.

Plants growing in saline media have another protective mechanism that

allows them to acclimatize to this unfavorable environment for continued survival

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and growth. One such mechanism that is ubiquitous in plants is the accumulation of

certain organic metabolites of low molecular weight that are collectively known as

compatible solutes (Bohnert et al., 1995; Serraj and Sinclair, 2002; Ashraf and

Harris, 2004; Vinocur and Altman, 2005).

The present study was planned to assess the salt tolerance of maize

genotypes and role of potassium in improving the salt tolerance with the following

objectives:

Assessment of salt tolerance of various maize genotypes in solution culture.

Effect of potassium supply in alleviating salt stress in maize genotypes in solution as well as soil culture.

Role of potassium in improving growth, quality, yield and biochemical parameters of maize genotypes grown in a naturally-saline field.

XVI

Chapter 2

REVIEW OF LITERATURE

Earth is a predominantly salty planet, with most of its water containing

about 3% NaCl. This concentration of salt has rendered the land very salty. It is

projected that about 900 m ha land is affected due to salt, which considerably poses a

serious threat to agricultural productivity (Flowers and Yeo, 1995; Munns, 2002)

because most agricultural crops will not grow under conditions of high salt

concentration. Hence, the existing salinity is a great challenge to food security.

Selection and breeding have always been the common practices by man

for the purpose of high yields and better quality of crops. Selection of crops was also

made with reference to environmental conditions and the properties of soil. Historical

record show a shift in agriculture in the Tigris-Euphrates basin of ancient

Mesopotamia from the cultivation of wheat to the more salt tolerant barley as the

fertile but poorly drained soils became increasingly saline (Jacobsen and Adams,

1958). This dynamic problem seems to be more severe when we have a glance at the

increasing population, particularly in the third world countries.

2.1 EXTENT OF SALINITY PROBLEMS AT NATIONAL AND

GLOBAL LEVELS

Naturally occurring salinization is primarily caused by capillary water

level elevation and subsequent evaporation of saline groundwater. However, man-

made salinization is widely spread. Especially, irrigated land in arid regions is highly

susceptible to salinization. Irrigation practices lead to ground water level elevation

and a subsequent increased evaporation. This is particularly true in countries of arid

and semiarid regions of the world (Supper, 2003).

More than 800 million hectares of land throughout the world are salt-

affected, either by salinity (397 million ha) or the associated condition of sodicity

(434 million ha) (FAO, 2005). This is over 6% of the world’s total land area. Most of

this salinity, and all of the sodicity, is natural. However, a significant proportion of

cultivated agricultural land has become saline because of land clearing or irrigation.

Of the 1500 million ha of land farmed by dry land agriculture, 32 million ha (2%) are

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affected by secondary salinity to varying degrees. Of the current 230 million ha of

irrigated land, 45 million ha are salt-affected (FAO, 2005). High amounts of salts in

soils, taking into account both human made and naturally occurring salinization, are

responsible for yield reduction on one third of the global arable land.

In Pakistan, about 6.30 mha of land are salt affected (Alam et al., 2000).

The canal irrigation system extends over about 62,400 km and is mainly confined to

Indus plain. Although irrigation covers only about 15% of the cultivated land in

Pakistan, yet irrigated land has at least twice the productivity of rain-fed land, and

may therefore produce one-third of the world’s food. The reduced productivity of

irrigated lands due to salinity is, therefore, a serious issue (Rahman, 1998). With the

projected increase in populations of 1.5 billion people over the next two decades

coupled with increased urbanization in developing countries, the world’s agriculture

is facing an enormous challenge to maintain, let alone increase, our present level of

food production (Owen, 2001). Ways must be found to achieve this without resorting

to unsustainable farming practices and without major increases in the amount of new

land under cultivation, which would further threaten forests and biodiversity. It is

estimated that agricultural productivity will need to increase by 20% in the developed

countries and by 60% in the developing countries (Gruhn et al., 2000; Cakmak,

2002). In the light of these demographic, agricultural and ecological issues, the threat

and effects of salinity become even more alarming. Reducing the spread of

salinization and increasing the salt tolerance of crops, particularly the high yielding

ones are, therefore, issues of global importance.

2.2 EFFECT OF SALINITY ON PLANT PHYSIOLOGY AND

MORPHOLOGY

Salt accumulation leads to a deterioration of soil structure and hinders

desirable air-water balance essential for biological processes occurring in plant roots.

As a result of detrimental effects of salinization, crop yields decrease, while arable

land is lost irreversibly (Supper, 2003). Salt stress causes various effects on plant

physiology such as increased respiration rate, ion toxicity, changes in plant growth,

mineral distribution, and membrane instability resulting from calcium displacement

by sodium (Marschner, 1986), membrane permeability (Gupta et al., 2002), and

XVIII

decreased photosynthetic rate (Hasegawa et al., 2000; Munns, 2002; Ashraf and

Shahbaz, 2003; Kao et al., 2003; Sayed, 2003).

Salt stress affects plant physiology at whole plant as well as cellular levels

through osmotic and ionic stress (Hasegawa et al., 2000; Muranaka et al., 2002 a, b,

Ranjbarfordoei et al., 2002; Murphy et al., 2003). In addition to causing osmotic and

ionic stress, salinity causes ionic imbalances that may impair the selectivity of root

membranes and induce potassium deficiency (Gadallah, 2000). The accumulation of

high amounts of toxic salts in the leaf apoplasm leads to dehydration and turgor loss,

and eventually death of leaf cells and tissues (Marschner, 1995). As a result of these

changes, the activities of various enzymes and plant metabolism are affected

(Lacerda et al., 2003). At high rates of transpiration, the xylem of all species contains

much lower chloride and sodium concentrations than those in the external saline

medium. Salt stress enhances the accumulation of NaCl in chloroplasts of higher

plants, affects growth rate, and is often associated with decrease in photosynthetic

electron transport activities (Kirst, 1989). In higher plants, salt stress inhibits PS-II

activity (Kao et al., 2003; Parida et al., 2003), although some studies showed that salt

stress had no effect on PS-II (Brugnoli and Björkman, 1992; Morales et al., 1992).

The reduction of plant growth and dry-matter accumulation under saline conditions

has been reported in several important grain legumes (Tejera et al., 2006).

Many arid and semi-arid regions in the world contain soils and water

resources that are too saline for most of the common economic crops (Pitman and

Lauchli, 2002). The majority of crop plants are relatively salt-sensitive and unable to

tolerate high levels of salinity (Levitt, 1980). Salinity affects plants through osmotic

effects, ion specific effects and oxidative stress (Pitman and Lauchli, 2002). Salinity

results in a reduction of K+ and Ca2+ contents and an increased level of Na+ and Cl-,

which forms its ionic effects. Salt stress induces cellular accumulation of damaging

active oxygen species. Active oxygen species can damage membrane lipids, proteins

and nucleic acids (Mittler, 2002).

Osmotic adjustment of both halophytes and glycophytes is achieved

through the accumulation of organic and inorganic solutes (Yeo, 1998). Therefore, a

greater decrease in cell solute potential than in the external salt concentration may

XIX

indicate an osmotic adjustment. Organic solutes are accumulated in the cytosol to

balance the solute potential of the vacuole, which is dominated by ions (Greenway

and Munns, 1980). A large number of plant species accumulate glycinebetaine and

proline in response to salinity stress and their accumulation may play a role in

combating salinity stress (Ashraf, 1994a; Hanson and Burnet, 1994; Mansour, 2000;

Ashraf and Harris, 2004). Glycinebetaine and proline functions under stress

conditions are presented by Ashraf (1994b), Ashraf and Harris (2004), Hanson and

Burnet (1994) and Mansour (2000). However, data do not always indicate a positive

correlation between the osmolyte accumulation and the adaptation to stress (Wyn

Jones et al., 1984; Rains, 1989; McCue and Hanson, 1990; Ashraf, 1994a; Lutts

et al., 1996; Mansour, 2000; Ashraf and Harris, 2004).

Previous studies suggested that plasma membrane might be the primary

site of salt injury (Epstein et al., 1980; Levitt, 1980; Lauchli, 1990; Mansour, 1997).

To test this hypothesis, the response of the plasma membrane to salinity in genotypes

contrasting in salt tolerance was studied by measuring the plasma membrane

permeability. Plasma membrane permeability probes the changes or differences in the

membrane structure/composition (Simon, 1974; Stadelmann and Lee-Stadelmann,

1989; Magin et al., 1990). Plasma membrane permeability is altered markedly in salt

sensitive cultivars whereas the effect is always marginal in salt tolerant cultivars upon

salt exposure (Mansour et al., 1993; Mansour and Stadelmann, 1994; Mansour, 1997;

Mansour and Salama, 2004).

2.3 EFFECT OF SALINITY ON CROP YIELDS

2.3.1 Osmotic stress Salt stress reduces the plant’s ability to take up water, and this leads to

reduction in growth. This is the osmotic or water-deficit effect of salt stress. Both

cellular and metabolic processes involved in osmotic stress due to salinity are

induced by drought. The rate at which new leaves are produced depends largely on

the water potential of the soil solution, in the same way as for a drought-stressed

plant. Salts themselves do not build up in the growing tissues at concentrations that

inhibit growth, as the rapidly elongating cells can accommodate the salt that arrives

XX

in the xylem within their expanding vacuoles. So, the salt taken up by the plant does

not directly inhibit the growth of new leaves (Munns, 2005).

Reductions in the rate of leaf and root growth are probably due to factors

associated with water stress rather than a salt-specific effect (Munns, 2002). This is

supported by the evidence that Na+ and Cl– are below toxic concentrations in the

growing cells. In rice, spikelet and tiller numbers were more affected by salinity than

1000-seed weight (Zeng et al., 2002). For example, in wheat growing in 120 mM

NaCl, Na+ in the growing tissues of leaves was at most only 20 mM, and only 10 mM

in the rapidly expanding zones, and Cl– only about 50 mM (Hu et al., 2005).

Similarly, Neves-Piestun and Bernstein (2005) found that Na+ and Cl– were only 40

mM in the most rapidly growing tissues, and that the degree of inhibition by salt

stress of either the elongation rate or the total volume expansion rate did not correlate

with the Na+ or Cl– in the tissues of maize growing in 80 mM NaCl. Fricke et al.,

(2004) found only 38 and 49 mM Na+ in mesophyll and epidermal cells, respectively,

in the growing cells of barley after 24 h of exposure to 100 mM NaCl. Therefore, Na+

was not inhibitory to growth, but was probably beneficial as it might be taken up into

the expanding vacuole for osmotic adjustment. This was indicated by the fact that the

growth rate increased with time over 24 h (after a temporary decline when the salt

was applied) while the cellular Na+ increased.

The rapid expansion of the growing cells would help to keep the salt from

building up to high concentrations. Results of experimental manipulation of shoot

water relations suggest that hormonal signals, probably induced by the osmotic effect

of the salt outside the roots, are controlling the rate of cell elongation growth (Munns

et al., 2000). Inhibition of plant growth due to salt stress largely depends on the

severity of the stress. Mild osmotic stress leads rapidly to growth inhibition of leaves

and stems, whereas roots may continue to grow and elongate (Hsiao and Xu, 2000).

The degree of growth inhibition due to osmotic stress depends on the time scale of

the response, the particular tissue and species in question, and whether the stress

treatments imposed abruptly or slowly (Ashraf, 1994b; Munns et al., 2000).

2.3.2 Specific ion toxicity

Toxicity occurs as a result of uptake and accumulation of certain toxic ions

from the irrigation water, within a crop itself. It is different from salinity problem and

XXI

may occur even when the salinity is low. These toxic constituents include mainly

sodium, chloride and sulphate. They can reduce crop productivity and eventually

cause crop failures. Not all crops are equally affected but most crops and woody

perennial plants are sensitive (Abrol et al., 1988).

The salt taken up by the plant concentrates in the old leaves; continued

transport of salt into transpiring leaves over a long period of time eventually results in

very high Na+ and Cl– concentrations, and the leaves die. The cause of the injury is

probably due to the salt load exceeding the ability of the cells to compartmentalize

salts in the vacuole. Salts then would rapidly build up in the cytoplasm and inhibit

enzyme activity. Alternatively, they might build up in the cell walls and dehydrate

the cell (Munns, 2005) but Muhling and Läuchli (2002) found no evidence that maize

cultivars differed in salt tolerance.

Mechanisms for tolerance of the salt-specific effects of salinity are of two

main types: those minimizing the entry of salt into the plant; and those minimizing

the concentration of salt in the cytoplasm. Root cytosolic Na+ concentrations are

probably in the order of 10–30 mM (Tester and Davenport, 2003). Leaf Na+ cytosolic

concentrations are unknown, but are considered to be much less than 100 mM (Wyn

Jones and Gorham, 2002). Roots must exclude most of the Na+ and Cl– dissolved in

the soil solution, or the salt in the shoot will gradually build up with time to toxic

levels.

Husain et al. (2004) used two durum wheat genotypes with contrasting

rates of Na+ transport to leaves to assess the effects of the Na+ exclusion trait on

preventing leaf injury and enhancing yield. They found that older leaves of the high-

Na+ lines lost chlorophyll more rapidly and died earlier than the low-Na+ lines. The

low-Na+ trait improved yield by greater than 20% in saline soil at moderate salinity

(75 mM NaCl). However, yield was not improved at high salinity (150 mM NaCl).

This indicates that traits other than Na+ exclusion are important at high salinity,

where the osmotic effect of the NaCl outweighs its salt-specific effect on growth and

yield. Na+ increment inside plants had toxic effects on seed germination, mainly by

affecting the plant water relations or through displacement of Ca2+ by Na+ from

XXII

critical cell wall binding sites, which could disrupt cell wall synthesis and hence

inhibit plant growth.

According to Loreto et al. (2003) Cl- concentration more than 80 mM in

total tissue water alters plant morphology, stomata become less responsive to

environmental changes and leaf thickness is reduced. Chloride is not adsorbed by

soils but moves readily with the soil water. It is taken up by roots and moves upward

to accumulate in the leaves. The toxic level of chloride causes leaf burn or drying of

leaf tissues, which occurs first at extreme leaf then tips of older leaves and progresses

back along the edges as severity increases. Marschner (1995) found that extreme leaf

burn due to toxic level of chloride leading to early leaf drop, because of which the

whole plant became finally defoliated. Specific ion toxicity, the result of excessive

uptake of certain ions (Na+ and Cl-) is the primary cause of growth reduction under

salt stress (Chinnusamy et al., 2005). According to Jacoby (1993) long term

inhibition of growth under salinity stress might be more a consequence of ion toxicity

than negative water status. Although the rate of canopy development and final size

were an outcome of leaf and stem extension-growth. It had been shown that leaf

injury and loss due to excess salt accumulation might be an important factor

controlling the active size of canopy (Francois and Maas, 1993). Accumulation of

Na+ and Cl- to toxic levels in leaves interfered with the metabolic processes going on

in cytoplasm and retarded the growth and development of plants.

Ion cytotoxicity caused by the replacement of K+ with Na+ in biochemical

reactions and conformational changes and loss of function of proteins as Na+ and Cl-

ions penetrated the hydration shells and interfere with non-covalent interaction

between their amino acids. Dionisiosese and Tobita (2000) reported that increased

concentration of Na+ under saline conditions suppressed the leaf gas exchange and PS

II photochemical activity and consequently hampered the growth and development of

plants.

2.3.3 Nutritional imbalance

Excessive amounts of soluble salts in the root environment cause osmotic

stress, which may result in disturbance of the plant water relations, in the uptake and

utilization of essential nutrients, and also in toxic ion accumulation. As a result of

XXIII

these changes, the activities of various enzymes and the plant metabolism are

affected (Munns, 2002; Lacerda et al., 2003). The interactions of salts with mineral

nutrients may result in considerable nutrient imbalances and deficiencies (McCue and

Hanson, 1990). Ionic imbalance occurs in the cells due to excessive accumulation of

Na+ and Cl– and reduces uptake of other mineral nutrients, such as K+, Ca2+, and

Mn2+ (Karimi et al., 2005). High sodium to potassium ratio due to accumulation of

high amounts of sodium ions inactivates enzymes and affects metabolic processes in

plants (Booth and Beardall, 1991).

Excess Na+ and Cl- inhibits the uptake of K+ and leads to the appearance of

symptoms like those in K+ deficiency. The deficiency of K+ initially leads to chlorosis

and then necrosis (Gopal and Dube, 2003). The role of K+ is necessary for

osmoregulation and protein synthesis, maintaining cell turgor and stimulating

photosynthesis (Freitas et al., 2001; Ashraf, 2004). Both K+ and Ca2+ are required to

maintain the integrity and functioning of cell membranes (Wenxue et al., 2003).

Maintenance of adequate K+ in plant tissue under salt stress seems to be dependent

upon selective K+ uptake and selective cellular K+ and Na+ compartmentation and

distribution in the shoots (Munns et al., 2000; Carden et al., 2003). The maintenance

of calcium acquisition and tranport under salt stress is an important determinant of

salinity tolerance (Soussi et al., 2001; Unno et al., 2002).

Salt stress decreases the calcium/sodium ratio in the root zone, which

affects membrane properties, due to displacement of membrane-associated Ca2+ by

Na+, leading to dissolution of membrane integrity and selectivity (Cramer et al.,

1995; Kinraide, 1998). The increased levels of Na+ inside the cells change enzyme

activity resulting in cell metabolic alteration; disturbance in K+ uptake and

partitioning in the cells and throughout the plant that may even affect stomatal

opening, thus diminishing the ability of the plant to grow. Externally supplied Ca2+

has been shown to ameliorate the adverse effects of salinity on plants, presumably by

facilitating higher K+/Na+ selectivity (Hasegawa et al., 2000). Another key role

attributed to supplemental Ca2+ addition is its help in osmotic adjustment and growth

via the enhancement of compatible organic solutes accumulation (Girija et al., 2002).

Ca2+ has also been implicated in stress protection by stabilizing membranes and

reducing the oxidative damage (Larkindale and Knight, 2002). High K+/Na+ ratio was

XXIV

observed due to ABA treatment in bean plant that seems to limit sodium translocation

to shoot (Khadri et al., 2007).

Excessive amounts of soluble salts in the root environment cause an

osmotic stress, which may result in disturbance of the plant water relations, uptake

and utilization of essential nutrients, and also in toxic ion accumulation. As a result of

these changes, the activities of various enzymes and the plant metabolism are

affected (Bernstein et al., 1974; Munns, 2002; Lacerda et al., 2003). The interactions

of salts with mineral nutrients may result in considerable nutrient imbalances and

deficiencies (McCue and Hanson, 1990). Ionic imbalance occurs in the cells due to

excessive accumulation of Na+ and Cl– and reduces uptake of other mineral nutrients,

such as K+, Ca2+, and Mn2+ (Hasegawa et al., 2000). High sodium to potassium ratio

due to accumulation of high amounts of sodium ions inactivates enzymes and affects

metabolic processes in plants (Booth and Beardall, 1991). According to Weimberg

(1987), high levels of Na+ inhibit the K+ uptake by causing an increase in the Na+/K+

ratio. Many of deleterious effects of Na+ seem to be related to the structural and

functional integrity of membranes. Salinity stress has stimulatory as well as

inhibitory effects on the uptake of some micronutrients by plants. It has been reported

that uptake of Fe, Mn, Zn and Cu generally increases in crop plants under salt stress

conditions. High K+/Na+ selectivity in plants under saline conditions has been

suggested an important selection criteria for salt tolerance (Ashraf, 2002; Wenxue et

al., 2003). In Agropyron spp., the high salt tolerance of A. elongatum relative to A.

intermedium, is associated with its higher uptake of K+ under saline conditions

(Elzam and Epstein, 1969). Experimentation with wheat indicates that salt tolerance

is associated with an enhanced K+/Na+ discrimination trait (Gorham, 1994). Recently,

Munns and James (2003) have found that although Na+ exclusion had a positive

relationship with salinity tolerance of different tetraploid wheats. A similar

mechanism of ion uptake has been observed in barley (Wenxue et al., 2003).

2.4 EFFECT OF SALINITY ON MAIZE Many crop species are sensitive to high concentrations of salt with

negative impacts on the agriculture productivity. Maize (Zea mays L.) is a moderately

salt-sensitive crop (Maas and Hoffman, 1977). Crops differ significantly in their

tolerance to the concentration of soluble salts in the root zone. A number of salt

XXV

tolerant crops are available for better use of saline drainage effluents. Raising the

extent of salinity limits through the selecting more salt tolerant crops enables the

great use of saline water and reduced the need for leaching and drainage. Differences

in salt tolerance exist not only among different genera and species, but also within

certain species. In addition to general osmotic stress, high concentrations of Na+ are

toxic to maize (Fortmeier and Schubert, 1995; Munns, 1993). After exposure of

maize plants to 100 mM NaCl decreased 50% root and shoot fresh weights and

considerable morphological changes were observed. In addition to growth reduction,

the root became yellow and leaves appeared dark-green. Leaves wilted at high

temperatures in consequence of the loss of water and osmotic stress and with a

gradual increase of stress (150 mM NaCl) the younger leaves, gradually became

yellow and leaves developed chlorosis at the tips.

Salt treatments affect plants differently in early growth stages, as Nuran

and Cakirlar (2002) observed that seeds could grow at different salt levels, but they

could not continue their development. A salt stress causes a significant decrease in

shoot length, fresh and dry weights of shoot and leaf area with the increase of stress

treatments. Salt stress at different osmotic potential increased the amounts of proline

of leaf tissues in maize genotypes. Zidan et al. (1990) observed a reduction in growth

of primary roots of maize induced by salinization of the nutrient medium with 100

mM NaCl, accompanied by reductions in the length of the root tip elongation zone (7

mm in control seedlings compared with 4 mm when 100 mM NaCl was added), in the

length of the epidermal cells, and in the apparent rate of cell production. Effects of

salt stress on the activity of antioxidative enzymes and lipid peroxidation were

studied in leaves and roots of two maize cultivars by Azevedo Neto et al. (2006).

They found that in leaves of salt stressed plants, superoxide dismutase, ascorbate

peroxidase , guaiacol peroxidase and glutathione reductase activities increased with

time when compared to the controls. The increase in enzyme activities was more

pronounced in the salt tolerant than in the salt sensitive genotypes. Salt stress had no

significant effect on catalase activity in the salt tolerant, but it was reduced

significantly in the salt sensitive genotypes.

Maize has world wide importance with well-documented history of

cultivation and improvement. In a country like Pakistan, the attention towards the use

XXVI

of balanced fertilizers especially at critical stages because of salt affected and

calcareous nature of soils is necessary to increase the efficiency of applied plant

nutrients. Mansour et al. (2005) worked on two maize cultivars, salt sensitive

Trihybrid 321 and salt tolerant Giza 2, were studied, mainly their adaptation to NaCl

imposition at cell and whole plant level. Changes in growth and mineral content of

roots and shoots, glycinebetaine and free proline levels of shoots, plasma membrane

permeability and solute potential (ψs) of leaf sheath subepidermal cells were

measured. NaCl decreased fresh mass, dry mass, relative growth rate of shoots and

roots, and leaf area ratio in both cultivars. Greater decrease (except leaf area ratio)

was obtained in Giza 2 than in Trihybrid 321. NaCl stress resulted in accumulation of

glycinebetaine and free proline in shoots of both cultivars. The magnitude of increase

in both omsolytes was higher in Giza 2 than in Trihybrid 321. Salt stress induced Na+

and Cl- accumulation while it decreased K+ and Ca2+ levels in shoots and roots of

both cultivars. The increase in Na+ and the decrease in K+ and Ca2+ was greater in

Giza2 than in Trihybrid 321, while Cl- was increased more in Trihybrid 321

compared to Giza2. NaCl increased plasma membrane permeability in both cultivars.

Salt stress decreased cell ψs in both cultivars, especially in Giza 2. It was concluded

that Na+ exclusion from the shoot was not correlated with salt tolerance and that

proline and glycinebetaine accumulation in the shoot was a possible indicator for salt

tolerance in the maize genotypes studied (Mansour et al., 2005)

Khan et al. (2003) studied root growth response of 10-days-old seedlings

of 100 maize accessions at 0, 60, 80 and 150 mM NaCl concentration in solution

culture. The non-linear least square method was used to quantify the salt tolerance of

maize accessions. The estimated salinity threshold (Ct), (the NaCl concentration at

which root growth starts to decrease), C0, and C50 (the concentrations at which roots

stop growing and 50% of its control value) revealed considerable differences between

the accessions. No general consistency for tolerance was, however, found between

the estimates of Ct and C50. Different genetic systems appeared to be involved in

controlling the inheritance of Ct and C50. Both Ct and C50 appeared to quantify

accession tolerance, and the expression of root growth as a function of NaCl

concentrations provides a useful guideline for salt tolerance. Estimates of broad sense

XXVII

heritability for relative root length were moderate in size (0.62 to 0.82), suggesting

the scope for enhancing salt tolerance in maize through selection and breeding.

Rozeff (1995) reported that salts interfered with sugar production by two

different fashions; firstly by affecting the growth rate and yield of the cane and

secondly by affecting the sucrose contents of stalk. The study conducted by Akhtar

(2000), revealed that at a salinity level of 12 dS m-1 marked reduction in yield and

yield contributing parameters of sugarcane genotypes could be attributed to reduced

photosynthetic efficiency under salt stress. Deteriorated soil conditions as the result

of salt stress adversely affected sugarcane growth which resulted in decreased cane

weight, reduced cane number and relatively thinner canes with the resultant decrease

in cane yield (Wahid, 2004). An excess of ions adversely affected the elongation and

differentiation of cane stalk internodes and storage of sucrose therein.

The build up of salts adversely affected the cane juice quality, decreased

% commercial cane sugar (CCS) and increased % reducing sugar. The Na+, Cl- and

SO4-2 contents of the juice as well as bagasse percentage were significantly increased

under saline growth conditions (Gomathi and Thandapani, 2005). Lingle and

Weigand (1997) reported an increase in juice osmolality and a decrease in total

soluble solids and sucrose per unit increase in salinity. This may be due to salt

induced stimulation of sucrolytic activities of acid and neutral invertases. Salinity

reduced both fresh weight of stalks and their sugar content. The link of decreasing

sucrose content in cane stalks with increasing salinity could be that Cl- together with

K+ accumulated in storage tissues where it raises the osmotic potential. This impedes

the sucrose transport into storage cells and competed with Cl- for storage space.

Furthermore, Cl- in storage tissues decreased the pool of plant K+ available for other

metabolic functions in the plants (Wahid, 2004).

Many researchers have reported the salinity response of different varieties

of wheat (Rashid et al., 1999; Shirazi et al., 2001; Pervaize et al., 2002); chickpea

(Katerji et al., 2001b; Gandour, 2002); barley (Touchan and Coons, 1991; Flowers

and Hajibagheri, 2001), lentil (Katerji et al., 2001a), and maize (Khan et al., 2003).

In a gravel-culture study, Akhtar et al. (1998) observed that salinity reduced the

growth of five wheat genotypes (Pato, LU-26S, Pb-85, Tehere and Pak-81). Due to

XXVIII

salinity, shoot fresh weight was reduced from 82% in Pb-85 to 29% in Pak-81.

Amongst wheat genotypes, maximum Na+ concentration was in the oldest leaves in

Pak-81 and minimum in Tehere. Addition of salts decreased the K+ concentration

significantly in all genotypes and the lowest concentration was observed in Pato.

Tehere had the lowest and Pb-85 the highest K+ concentration in the oldest leaf sap,

the youngest leaf maintained high mean K+ concentration.

Inhibition of the uptake of essential nutrients by salinity may result in

severe reductions in yield, depending on the plant species. Supplemental fertilization,

particularly of K+, Ca2+, NO3- and in some cases micronutrients, leads to a recovery

of physiological parameters and stimulates growth (Cramer and Nowak, 1992;

Marschner, 1995; Zhu, 2001). In addition, application of K+ and Ca2+ may also

improve plant performance by reducing the uptake of salts (Romero-Aranda et al.,

1998). The selection of salt tolerant plant species or genotypes is a common practice

to reduce yield losses under saline conditions. Declines in water quality below the

threshold values do not preclude their potential use for irrigation of the considered

crops, however the adoption of both intense management practices and the use of salt

tolerant genotypes are recommended to maintain crop productivity at acceptable

levels and to ensure land sustainability. It should be stressed, however, that the

threshold values might considerably vary among different cultivars or rootstocks

(Storey and Walker, 1999; Chartzoulakis et al., 2002). Furthermore, in woody plants

salt tolerance of a given genotype can display significant variations from one area to

another. Such variations have been reported for citrus (Maas, 1993) and grapevines

(Arbabzadeh and Dutt, 1987), which may be due to differences in environmental

factors (soil fertility, soil physical conditions and climatic factors) and also plant

genetic diversity. It was demonstrated again that maize is relatively tolerant to

salinity during the germination stage and this behavior is considered to be crop

specific (Pablo and Peinemann, 1998). The precise mechanism by which salt affects

carbon partitioning remains unclear. However, the higher retranslocation of nitrogen

compounds via the phloem (Gilbert et al., 1998) may be taken as an indication of a

relative increase of the root sink strength, although a reduced shoot sink strength due

to decreasing growth rates cannot be excluded as an alternative explanation. It is well

XXIX

established that carbon partitioning between shoots and roots is flexible and highly

responsive to the environment (Dalton et al., 1997; Fageria et al., 1997).

Earlier research showed that Na+ concentration in the apoplastic fluid of maize leaves increased up to 4 mol m-3 and that of Cl– to 5 mol m-3 with 100 mol m-3

external NaCl (Wang and Zhao, 1997). Katerji et al. (2000) conducted a long term experiment on the use of saline water and concluded that fresh and dry matter production of maize plants were significantly reduced by increasing NaCl concentration in irrigation water. Greater availability of nutrient elements such as N, P and K could not counteract the ionic absorption effect. Moreover, the response of young maize plants to nutrient applications tended to diminish with increasing salinity. A high negative correlation (r=-0.8488) between NO3

- and Cl- concentrations in shoots was determined. However, phosphorus and potassium concentrations remained constant. Growth was retarded by exposure of maize plants to salinized nutrient solution. The growth reduction increased with the increase of salt concentration. Roots were less affected by salt treatment (100 mM NaCl) although the reduction of fresh biomass was up to 30%. A pot experiment was conducted (Abou El-Nour, 2002) to investigate the influence of salinity on maize plants grown on two soils (sandy and clayey). Treatments were: tap water; high EC water (EC 5.6 dS m-1) and high EC water + micronutrients. Results showed a highly marked significant effect on plant growth due to the tested soil types. Root and shoot dry weight decreased by 52.5 and 60.6%, respectively for plants grown on the sandy soil as compared with those grown on the clayey soil. On the other hand, root and shoot dry weight decreased by about 24 and 21%, respectively due to salinity treatment as compared with control treatment. Such reduction may be due to the inadequacy of nutrients present in the growing medium. The upper row of corn ears was produced with non-saline irrigation water. Irrigation water of 8 dS m-1 was applied to grow ears. Salinity not only reduced the size of the ears but also reduced the number of ears. The total yield with irrigation water having an ECiw of 8 dS m-1 was less than half of that without salt. Similarly, Irshad et al. (2002) reported that soil salinity reduced the plant height, shoot and root dry weight of maize. Adverse effects of salinity on sorghum plant growth have been reported by Khan et al. (1995). Plant growth may be adversely affected by a salinity-induced nutritional disorder as well as osmotic and specific ion effects (Pessarakli., 1991; Ehret et al., 1990). Cicek and Cakirlar (2002) investigated the effect of salinity with different osmotic potentials on shoot length, total fresh and dry weight, amount of organic (proline) and inorganic (K+ and Na+) substance of leaf tissue, the Na+/K+ ratio and leaf area in two maize (Zea mays L.) cultivars (Intendata, C.6127 and DK.623). Plants were grown for 30 days in controlled conditions growth room. Salinized culture solutions at different osmotic potentials (0, -0.1, -0.3 and -0.5 M Pa) prepared by adding varying amounts of NaCl and CaCl2 to the main culture solution were applied to plants from the beginning of the germination. It was observed that shoot length, total fresh and dry weight and leaf area decreased, amount of proline, Na+ and Na+/K+ ratio increased. However the amounts of K+ did not change significantly with increasing stress, and salt stress caused a similar decrease in leaf relative water content in both maize cultivars.

XXX

Through seed selection a variety of sweet corn (Zea mays L. amylacea)

was tested by Bastias et al. (2004) and showed an extreme tolerance to high salt and

boron levels. Concentrations of 100 and 430 mol m-3 NaCl and 20 and 40 mg kg−1

boron were imposed as treatments. The plants did not exhibit symptoms of toxicity to

either NaCl or boron during the 20 days of treatment. Na+ accumulation was

substantial in roots, while boron was translocated to leaves. Despite the fact that

boron increased slightly the effect of salinity on CO2 assimilation, no effect on

photochemical parameters was observed in this ecotype. Changes in dry or fresh

weight are widely used parameters to determine plant sensitivity to NaCl. Plant

growth, expressed in terms of dry weight, plant height and leaf area decreased in the

presence of NaCl. The difference in growth between plants grown at 100 and 430

mol m-3 NaCl was statistically significant and the magnitude of the responses of the

different parameters varied. Thus, plant height was only slightly reduced under saline

conditions (about 15%), while a larger reduction was measured in dry matter

production (40%) and leaf area (50%). In contrast, application of boron levels

considered to be toxic for most crops (20 to 40 mg kg−1) did not significantly alter the

growth of the maize cultivar. Thus, for the control treatment (no NaCl), dry weight

and leaf area were not reduced with extra boron while plant height was slightly

stimulated.

Eker et al. (2006) tested the salt tolerance of 19 hybrid maize varieties grown in nutrient solution during the early growth stages under controlled environmental conditions. Results showed that varieties differed greatly in their response to the NaCl treatment. The development time and severity of leaf symptoms caused by 250 mM NaCl varied markedly among the varieties. The decrease in the shoot dry matter production as a consequence of the NaCl treatment was higher than the decrease in root growth. There was also a marked genotypic variation in concentrations of K+, Ca2+ and Na+ in roots and particularly in shoots. The K+/Na+ and Ca2+/Na+ ratios were significantly greater in most of the tolerant varieties. The most sensitive variety contained a 4-fold greater Na+ concentration in shoots than the most tolerant variety. Patel et al. (2000) studied salt buildup due to irrigation water salinity and fertilizer application in field lysimeters planted with green peppers (Capsicum annuum). Water was applied by sub-irrigation, and the fertilizers were incorporated at the soil surface. Three sub-irrigation water salinities, 1, 5 and 9 dS m-

1 and two water table depths; 0.4 and 0.8 m, were used. The soil salinity was determined by first measuring the bulk soil salinity by time domain reflectometry (TDR) and then converting it to soil solution salinity (ECsw). It was found that the salinity of the sub-irrigation water affected ECsw in the upper soil profile when the water table was maintained at 0.4 m depth. The sub-irrigation water also affected the

XXXI

lower half of the soil profile when the water table was maintained at 0.8 m depth, however, it did not affect any salt buildup in the upper half. Also, the addition of N, P, and K fertilizers did not contribute to the salt buildup in the soil. Although water table depth and sub-irrigation water salinity affected ECsw, they did not affect the green pepper yield. Yadav et al. (2004) conducted an experiment using saline water on five Rabi forage crops. Results indicated that irrigation with fresh canal water resulted in a 115% increase in Rabi forage yield compared with the saline drainage water. It was suggested that alternating fresh water with saline drainage water irrigation increased the yields of all the forage crops compared with using saline drainage water only (EC=3.6 –7.4). In comparison with canal irrigation water, there was a 36%, 42%, 54%, 68%, and 85% yield reduction in rye grass, oat, Persian clover, Egyptian clover and senji, respectively when only saline drainage water (EC=7.4) was used for irrigation, reflecting their relative tolerances of salinity. Yields declined linearly for all crops with increases in the quantity of salt applied (Yadav et al., 2004). Azevedo Neto et al. (2004) concluded that salinity (25 mmol L-1 per day

NaCl salt for 15 days) reduced the dry mass of maize shoot and root. The shoot dry

weight reduced from 33.8% to 66.5% while root dry weight decreased by up to

61.4%. The different genotypes response was due to their variable tolerance.

Similarly Mansour et al. (2005) reported that when maize seedling were exposed to

150 mmol NaCl for 15 days, fresh matter and dry matter of root and shoot were

reduced significantly. Salinity induced growth reduction in maize species has been

previously reported (Alberico and Cramer 1993; Cramer 1993; Cramer et al.,

1994a,b). Dordipour et al. (2004) reported that the effect of salinity depends on the

stage at which the plant is exposed to this stress. Addition of seawater to barley plants

at stem elongation (T1) decreased yield by 62%, but when applied at ear formation it

did not significantly affect the yield compared to the control. Growth and yield of

barley were best in treatment T0 (control). This is in accordance with the finding of

Francois et al. (1994) who demonstrated that the time or stage of salinity stress had a

significant effect on grain weight of wheat. Visual observations of growth indicated

that chlorosis, necrosis and margin/tip burns of leaves appeared at early growth

stages and developed as growth progressed. The physiological mechanisms

underlying leaf growth inhibition under salt stress are not fully understood.

Apoplastic pH is considered to play an important role in cell wall loosening and tissue

growth and was demonstrated to be altered by several growth-limiting environmental

conditions. In the same study, they have evaluated the possibility that inhibition of

maize leaf elongation by salinity is mediated by changes in growing cell wall

XXXII

acidification capacity. The kinetics of extended apoplast pH changes by leaf tissue of

known expansion rates and extent of growth reduction under stress were investigated

and found to be similar for non-stressed and salt-stressed tissues at all examined

apoplast salinity levels (0.1, 5, 10, or 25 mM NaCl). Beatriz et al. (2001) showed the

results of non-destructive daily leaf length measurements. Salinity (80 mol m-3 NaCl)

reduced leaf growth and shoot development. The effect of salt stress on leaf growth

was started from leaf 3. Maximal leaf length attained was reduced by 20% and 30% in

the third and fourth leaves, respectively, and maximal elongation rate of the fourth

leaf was decreased by 47% due to the stress. Since net CO2 assimilation on a fresh

weight basis was unaffected by salt treatment or sprinkling, it may be concluded that

shoot growth rate is not limited by net photosynthesis. The lower carbon assimilation

per plant observed in later growing stages is a reflection of a reduced leaf area and is

in agreement with earlier results (Cramer et al., 1994a; Lewis et al., 1989), which

showed that salinity affects maize leaf growth rate more strongly and earlier than net

photosynthesis. The reduction of shoot growth upon exposure to salinity cannot be

explained by a shortage of nutrient supply since the concentrations of K+ and amino-

N in the xylem sap were increased by salt treatment. This is in general agreement

with the findings of Munns (1985) who showed that K+ concentration in the xylem

sap of barley did not decline until the external NaCl concentration exceeded 100 mol

m-3. Cicek and Cakirlar (2008) concluded that the application of NaCl strongly

decreased plant growth (plant height, fresh and dry biomass of the shoot). The

decrease in plant height at –0.7 MPa NaCl ranged between 21-46%.

2.5 MECHANISM OF CROP TOLERANCE TO SALINITY

Potassium is essential for protein synthesis, glycolytic enzymes and

photosynthesis; an osmoticum mediating cell expansion and turgor driven

movements and competitor of Na+ under salt stress (Hu and Schmidhalter, 2005).

Accumulation of Na+ and impairment of K+ nutrition is a major characteristic of salt

stressed plants. Therefore K+/Na+ ratio in plants is considered a useful parameter to

assess salt tolerance (Cakmak, 2005a, b). Numerous studies have shown that

application of K fertilizer mitigates the adverse effects of salinity through its role in

stomatal regulation, osmoregulation, energy status, charge balance, protein synthesis

and homeostasis (Marschner, 1995; Sanjakkara et al., 2001).

XXXIII

Potassium is often considered to be a nutrient of primary importance for

cereal and oil seed crops. Potassium plays a major part in the enzyme system that

controls the metabolism of photosynthesis (Cakmak, 1994). Plants exposed to

environmental stress factors, such as drought, chilling, high light intensity, salinity

and nutrient limitations, suffer from oxidative damage catalyzed by reactive oxygen

species (ROS), e.g. super oxide, hydrogen peroxide and hydroxyl radical. Increasing

evidence suggests that improvement of potassium (K) nutritional status of plants can

greatly lower the ROS production (Cakmak, 2005b).

Soil salinity is an increasing constraint threatening crop production

globally. Around 30% of cultivated soils are affected by accumulation of salts

(Epstein et al., 1980; Zhu et al., 1997). Soil salinity generally results from excess

accumulation of NaCl and exerts detrimental effects on crop production by causing

ion toxicity, inducing osmotic stress (water deficiency) in root environment and in

plants (Zhu et al., 1997; Zhu, 2001).

Like most of other environmental stresses, salt stress also strongly affects

photosynthesis and cause an oxidative stress by inducing water deficiency (stomatal

closure), ion toxicity and K-deficiency. Consistent with this result, most of salt-

tolerant genotypes respond to salinity by increasing anti-oxidative defense systems

for detoxification of ROS (Rodriguez-Rosales et al., 1999; Sudhakar et al., 2001;

Zhu, 2001). Accumulation of Na+ and impairment of K nutrition is major

characteristic of salt-stressed plants. Therefore, K+: Na+ ratio in plants is considered

a useful guide to assess salt tolerance. Selection or breeding genotypes with high K+:

Na+ ratio is an important strategy to minimize growth deceases in saline soils (Deal

et al., 1999; Santa-Maria and Epstein, 2001). Rascio et al. (2001) identified a wheat

mutant with a high ability to accumulate K+ in the shoot and showed that this mutant

compared to other wheat genotypes greatly improved tissue hydration, seed

germination and seedling growth under increasing concentration of NaCl.

Saline soils generally have higher concentrations of Na+ than K+ and Ca2+

which may result in passive accumulation of Na+ in root and shoot (Bohra and

Doerffling, 1993). High levels of Na+ can displace Ca2+ from root membranes,

changing their integrity and thus affecting the selectivity for K+ uptake (Cramer et al.,

XXXIV

1987). Xylem loading of K+ is regulated by K+ uptake from external solution (Engels

and Marschner, 1992). This indicates that Na+ salinity besides reducing the K+ uptake

rate; also interfere to a greater extent in K+ translocation from root to shoot, which

results in a lower K+ shoot content and a higher K+ root content. The inhibitory effect

of salinity on K+ translocation was stronger with low K+ concentration in the nutrient

solution, when compared at two levels of K+ supply in maize seedlings i.e. 0.1 and 1

mmol L-1 (Botella et al., 1997). Thereby, salinity did not affect root dry weight, but

low levels of K+ in the nutrient solution significantly reduced shoot dry weight.

Similar responses have been found in spinach plants, which responded to an

increasing K+ concentration, reducing the differences in shoot growth between plants

grown under low salinity and those grown under high salinity (Chow et al., 1990).

The salinity-induced inhibition of shoot growth at low levels of K+ in the root

medium was attributed to the effect of K+ deficiency and/or Na+ toxicity on the

plants.

Any stress that is causing K+ leakage out of the cell will eventually lead to

a reduction in cell growth. Ben-Hayyim et al. (1987) have shown that growth was

linearly correlated with K+ content in callus cells of citrus roots. Increasing levels of

Na+ in the external medium reduced K+ in the cell. Salt tolerant cells were able to

hold the K+ in the vacuole against leakage when Na+ was increased in the external

medium. Termaat and Munus (1986) also suggested that salt stress might result in

limited transport of essential nutrients to the shoot. They have shown that the net

transport of K+, Ca2+, Mg2+ and total nitrogen to the shoot was lower in NaCl-exposed

plants. Salt tolerance has been partially linked to the regulation of leaf Na+

concentration (Taleisnik and Grunberg, 1994) and to selectivity for K+ over Na+

(Cuartero and Fernandez-Munoz, 1999). The plants have different pathways to avoid

Na+ from reaching to the leaves; by controlling Na+ influx at the plasmalemma of

root cells (Jacoby, 1993), by removing Na+ from the xylem stream and sequestering

Na+ in parenchyma cells of roots and the lower part of stem (Johanson and

Cheesman, 1983), by retranslocating Na+ from shoots to roots via phloem. In

suspension cells of Brassica napus, increased tolerance to NaCl arises by alteration

of K+ uptake system (Lefebvre, 1989), and tobacco cell cultures show enhanced K+

uptake capacity when adapted to NaCl (Watad et al., 1991). Salt tolerant cultivars of

XXXV

wheat translocate less Na+ from roots to shoots than salt sensitive genotypes

(Schachtman et al., 1989).

The differences within plant species in their capacity to satisfy metabolic

requirements for K+, higher K+ fluxes and lower Na+ fluxes in the presence of salinity

are related to differences in salt tolerance (Cerda et al., 1995). Taleisnik and

Grunberg, (1994) observed that, K+/ Na+ selectivity ratio was higher in tomato

cultivar ‘Edkawi’ than in ‘Ace’. The difference between the cultivars was due to

greater replacement of K+ contents by Na+ in all the plant parts of cultivars Ace. This

indicates that Edkawi has a higher capacity to retain K+ under salinity, a feature that

may contribute to its salt tolerance. Photosynthetic capacity and quantum yield of

oxygen evolution were sharply reduced under high salinity conditions with

decreasing K+ supply in spinach plants due to malfunctioning of photosystem II

(Chow et al., 1990). Their results suggest that there were higher K+ requirements for

shoot growth under high salinity (250 mM NaCl) than low salinity (50 mM NaCl)

conditions. By increasing total salt content with addition of K+ to roots, we can

alleviate reductions in plant and shoot biomass imposed by an increase in salinity and

overcome Na+ toxicity. The higher Na+ accumulation in leaves may help in turgor

maintenance, but cannot substitute for adequate K+ levels in leaves, because K+ is

specifically required for protein synthesis and enzyme activation (Marschner, 1995).

Therefore, maintenance of adequate cytoplasmic levels of K+ and K+/Na+ ratios in the

cell is essential for normal functioning under saline conditions (Greenway and

Munns, 1980; Chow et al., 1990). Addition of K+ to a saline culture solution has been

found to increase the plant dry weight and K+ content with a corresponding decrease

in Na+ content in rice and bean plants (Muhammed et al., 1987; Benlloch et al.,

1994). Since Na+ in most of the natural salinity cases is accompanied with chloride,

competition with nitrate was suggested as a practical agricultural method to prevent

salt damage to tomato (Kafkafi et al., 1982) and avocado (Bar et al., 1997).

In addition to reducing water availability and producing toxic ion effects

in saline condition, high concentrations of Na+ and Cl- ions will normally upset or

inhibit the cotton plant nutrition. Therefore, soil test interpretation in measuring

nutrient availability and recommending fertilizer levels may be different in saline and

non-saline soils. Keshavarz et al. (2004) performed an experiment in order to

estimate K critical level, 15 different main cotton fields under saline conditions and

XXXVI

10 fields under non-saline conditions were selected in Khorasan province (Iran) in

2001-2002. Then, a field experiment with a completely randomized block design

with two-potassium rates of 0 (K0) and 200 (K1) kg ha-1 K2SO4 and three replications

was carried out. Secondly, to determine the effects of sources and rates of K on the

yield of cotton bolls, a completely-randomized-block factorial experiment was

conducted with two sources of K [K2SO4 (SOP) and KCl (MOP)] and five rates of K

[0, 50, 100, 150 and 200 kg ha-1 K2O] at two locations, with saline (EC = 17 dS m-1,

Kava= 200 mg kg-1) and non-saline conditions (EC = 2.1 dS m-1, Kava=180 mg kg-1).

The results showed that, the use of K increases cotton bolls yield significantly (13%

and 6% for saline and non-saline soils, respectively). The type of fertilizers used had

a significant effect on the cotton yield. The maximum yield of cotton bolls in saline

condition was obtained using SOP (25% yield increase in comparison with MOP) and

in nonsaline condition by MOP (10% yield increase in comparison with SOP). The

relation between the potassium application and the yield of cotton bolls (response

curve) showed that the maximum yield was obtained with the use of 125 and 100 kg

ha-1 K2O under saline and nonsaline conditions, respectively. With respect to the

positive effects of K in saline soils, the application of K fertilizer is highly

recommended under those conditions.

Botella et al. (1997) observed that salinity significantly reduced shoot

growth when the level of K+ in the solution was 0.1 mmol L-1, probably due to

salinity induced K deficiency. NaCl reduced K+ net uptake rates and to a greater

extent K+ translocation from root to shoot that resulted in a lower shoot content and

higher concentration in the nutrient solution. Addition of K to nutrient solution

inhibited the uptake of Na+ and improved plant growth under salt stress. Chow et al.

(1990) also observed the effect of K on growth and photosynthetic efficiency under

saline conditions. They found that plant and shoot growth under salinity were related

to K+ uptake into leaves. There was greater decrease in photosynthetic capacity with

decreasing K+ supply to roots under high salinity. A reduction in yield occurred under

high salinity and low K supply which partly accounted for the lower plant and shoot

biomass at high salinity and low nutrient KCl concentrations. Response of spinach

and lettuce grown at higher salinity to supplementary K and P. High salinity reduced

germination percentage, root elongation and increased membrane permeability

compared to control. Supplementary K and P produced fresh weight, chlorophyll

XXXVII

content, water usage and membrane permeability values similar to or slightly lower

than control. Added K and P reduced leaf and root Na+ levels that were much higher

than control values but significantly lower than in saline treatment in most cases.

Salt stress adversely affects the crop performance due to nutritional

disorders. These disorders may result from the effect of salinity on nutrient

availability, competitive uptake, transport or partitioning within a plant (Ashraf,

1994a, b; Marschner, 1995). All these factors cause adverse affects on plant growth

and development at physiological and biochemical levels (Levitt, 1980; Gorham et

al., 1985; Munns, 2002; Munns and James, 2003). For example, salinity reduces

phosphate uptake and accumulation in crops grown in soils, primarily by reducing

phosphate availability but in solution cultures ion imbalances may primarily result

from competitive interactions. Salinity dominated by Na+ salts not only reduces Ca2+

availability, but also reduces Ca2+ transport and mobility to the growing regions of

the plant, which affects the quality of both vegetative and fruit parts. For example,

Na+ reduces K+ uptake and Cl- reduces NO3- uptake. Nutrient additions, on the other

hand, have been more successful in improving crop quality. Exogenous application of

nutrients may also reduce the incidences of injury as has been observed in the

reduction of Cl- toxicity symptoms in certain crops by nitrate applications (Grattan

and Grieve, 1999). While working with maize, Irshad et al. (2002) found that

application of urea caused increased nutrient uptake that resulted in enhanced plant

growth under salt stress. Similarly, additions of N to the growth medium also

improved salt tolerance in Daucus carota and Vigna unguiculata (Ravikovitch and

Porath, 1967), Zea mays (Khalil et al., 1967; Ravikovitch, 1973), Vitis vinifera

(Taylor et al., 1987), Malus pumila (El-Siddig and Ludders, 1994). The addition of

KNO3 can also overcome the effects of high salinity on fruit yield and whole plant

biomass in pepper plants (Kaya and Higgs, 2003). While assessing the role of KNO3

in ameliorating the adverse effects of salt on melon, Kaya et al. (2007) found that

addition of supplementary KNO3 to the rooting medium helped the melon plants to

avoid Na+ toxicity, improved cell membrane stability and Ca2+, K+ and N uptake.

From these reports, it is clear that addition of nutrients to salinity-affected soils may

improve the plant mineral nutrient status, which results in growth enhancement under

saline conditions. However, plants have to utilize sufficient amount of metabolic

energy for uptake of nutrients. To reduce energy losses and increase nutrient

XXXVIII

utilization efficiency, nutrients can be applied foliarly (Mengel and Kirkby, 2001). In

a number of studies, it has been found that foliar application of nutrients improved

the growth by increasing nutrient utilization efficiency under normal or stress

conditions. For example, foliar application of P fertilizer counteracted the salt

induced reduction in P, K and Ca2+ which resulted in increased dry weight, leaf area,

stomatal frequency, and yield (Malakondaiah and Rajeswararao, 1978). In an

extensive review, Grattan and Grieve (1999) were of the view that foliar applications

of those nutrients that become deficient under salt stress conditions could maintain

nutrient status of plants thereby leading to enhanced salt tolerance. However, of the

mineral nutrients, different salts of potassium have been extensively used as a foliar

spray to maintain ion homeostasis and hence improved growth under saline

conditions. For example, the foliar application of KNO3 improved the K nutrition and

growth of Legineria spp. under saline conditions. While working with sunflower,

Akram et al. (2007) reported that foliar sprays of different K inorganic salts

considerably improved the ion homeostatic conditions. They also found that

improved K nutrition due to foliar spray of K salts protected the cell membrane as

estimated by extent of ion leakage. Improved tissue K status of sunflower plants

regulated the plant photosynthetic activity through stomatal movement. They

concluded that all K salts improve the growth of sunflower plants however, the

effectiveness of K salts in improving growth depends upon a number of factors

including concentration of salt, plant developmental stage at which applied, and

accompanying anions in the specific salt. Thus, foliar application of inorganic salts

can also be beneficial to improve crop salt tolerance. Of the mineral nutrients, K

plays a particular role in contributing to the survival of crop plants under

environmental stress conditions. Potassium is essential for many physiological

processes (Mengel and Kirkby, 2001).

Potassium increases root growth and improves drought tolerance, builds

cellulose and reduces lodging, enhances many enzyme actions, aids in photosynthesis

and food formation, helps translocation of sugars and starches, produces grains rich

in starch, increases protein content of plants, maintains turgor, reduces water loss and

wilting and increase tolerance to crop diseases and nematodes. Plants are generally

classified as being either halophytes or glycophytes based mainly on their ability to

grow in saline environments. According to Flowers and Yeo (1986), the most useful

XXXIX

criteria for separating these two groups in the ability of halophytes to complete their

life cycle at excess concentrations of 100 to 200 mM NaCl. In contrast, glycophytes

are plant species characterized by relative low salt tolerance (Miller and Doescher,

1995). Halophytes usually achieve high salt tolerance by inclusion of salts while

glycophytes exhibit relatively limited salt tolerance by excluding salt from leaf cells.

Glycophytes may osmotically adjust by producing biochemical compatible organic

solutes within the cytoplasm. Notwithstanding, in many cases an important factor in

salt damage is dehydration due to the intracellular accumulation of salt as suggested

in the Oertli hypothesis (Flowers et al., 1991). It is well known that response of

glycophytes to salinity may be influenced by the plant species and environmental

conditions, and also varies at different stages of development within the same

species.

The plant may show more tolerance to salinity under one set of conditions

than the other. Despite the environmental factors, it is well documented that some

crops are more tolerant to salinity than others. From the extensive review, it is

evident that plants respond differently at different growth stages to saline growth

conditions. Generally, the plants are more sensitive at germination and seedling stage

than that at adult stage. Secondly, the selection criteria for discrimination between

salt tolerant and salt sensitive species can not be determined on a single attribute of

the plant. Thirdly, there are contrasting reports regarding the magnitude of salinity

effect on different parameters and plant response to the applied nutrient to combat the

salinity problem. The environmental, atmospheric and soil conditions of Pakistan are

quite different from other palace of world where the different studies were carried out

on maize with different aspects of salinity. The present study on this aspect of salinity

and potassium nutrition and their interaction has not been carried out so remarkably.

Therefore, different studies were planned in various growth conditions to find out the

magnitude of salinity effects and maize tolerance to applied salinity and its response

to the potassium addition. Different parameters were selected to find out the salt

tolerance of the maize crop and its effect on crop quality and growth parameters.

Thus the present study emphasized on these aspects of salt tolerance of maize crop

and its response to the applied potassium.

XL

Chapter 3

MATERIALS AND METHODS

The research work presented in this manuscript was conducted to evaluate the performance of maize genotypes under the different levels of salinity (NaCl) in the wire house of Saline Agriculture Research Center (SARC), Institute of Soil & Environmental Sciences, University of Agriculture, Faisalabad. These studies were carried out in solution culture, pot culture and in field conditions to find out the performance of maize genotypes in the said environment. The details of experimentation are given as under.

3.1 GROWTH CONDITIONS AND EXPERIMENATL TECHNIQUES

3.1.1 Experimental site

Solution culture and pot culture studies were conducted in the wire house

of Saline Agriculture Research Center (SARC), Institute of Soil & Environmental

Sciences, University of Agriculture, Faisalabad, during the period 2004-2006. The

wire house has a glass roof with no control over temperature, humidity and light as

the sides are open having only a wire net to control birds. While the field experiment

was conducted at a farmer’s field, Chak No. 86/G.B. district Toba Tek Singh, Punjab,

Pakistan.

3.1.2 Seed source

Synthetic (lines and approved) maize genotype seeds were collected from

Maize and Millet Research Institute (MMRI), Yousafwala, Sahiwal, Pakistan, while

the hybrid genotypes were collected from Styre Company which were available in

market easily and were being used by the farmers.

Improved genotypes Status Hybrid genotypes1. SL 2002 (Sahiwal-2002) Approved variety 2. Ev 1098 Line 3. Ev 6098 Line 4. S-2002 (Sargodha-2002) Approved variety 5. Agati 2000 Approved variety 6. Akbar Approved variety 7. Ev 5098 Line

1. Q 2139 2. Q2094 3. Q2100 4. Q2109 5. Q9515 6. Q0806 7. Q2414 8. Q8915

3.1.3 Solution culture studies (Study 1 & 2)

XLI

Seeds of maize genotypes were germinated in polythene-coated iron trays

containing acid washed gravel. Distilled water was used for marinating optimum

moisture for germination and seedling establishment. At two-leaf stage, seedlings of

uniform size were transplanted in foam-plugged holes of polystyrene sheets floating

over 1/2 strength Hoagland’s nutrient solution (Hoagland and Arnon, 1950). Salinity

was developed in respective treatments by adding NaCl in three/four applications,

starting two days after transplanting. Proper aeration of the culture solution was

provided for 8 hours daily by an aeration pump. The pH of the solution was

monitored daily and adjusted at 6.0±0.5, when needed. The substrate solutions were

changed fortnightly. After four weeks plants were harvested manually and different

parameters (plant height, root length (total root length), and shoot and root fresh

weights) were recorded, plant leaves were stored in eppendorf tubes in the

refrigerator for the determination of Na+, K+ and Cl-. The experiment was laid out in

a Completely Randomized Design (CRD) in factorial arrangements with three

replications.

3.1.4 Pot culture studies (Study 3)

The soil was collected from an agricultural field of research area of

Institute of Soil & Environmental Sciences, University of Agriculture, Faisalabad,

passed through 2mm sieve and filled in glazed earthen pots @ 12 kg per pot. The K

concentration of soil used in pot was determined and the original level of the soil K

was kept as control and was considered as the 1st K level. The 2nd level of K was

kept @ 200 mg kg-1 of soil (200 kg ha-1) by adding [potassium sulphate]. The desired

salinity in pots of saline treatments was developed by mixing required amount of

NaCl in the soil before filling the pots. The recommended doses of nitrogen and

phosphorus @ 200 and 150 kg ha-1 respectively, were applied in the form of urea and

Single Super Phosphate, respectively. Potassium fertilizer was not added in the

control while half of N and whole of P doses were applied to all pots before sowing

while remaining half of N was applied at the tillering stage. The soil was pulverized

and seed were sown at depth of 2cm. After germination, three uniform seedlings were

selected and remaining plants were uprooted. The pots were irrigated with tap water

(EC 0.98 dS m-1). At maturity, plants were harvested manually and different yield

parameters (plant height, shoot fresh weight, 1000 seed weight and grain yield plant-

XLII

1) were recorded, plant leaves were stored in eppendorf tubes for the determination of

Na+, K+ and Cl-. Maize grains were stored for the determination of oil contents, crude

fiber, total soluble sugars and protein contents. Leaf water potential and proline

contents were also determined. The experiment was laid out in a Completely

Randomized Design (CRD) in factorial arrangements with three replications.

3.1.5 Analysis of soil in the pot experiment

Soil samples collected were analysed in laboratory according to USDA

system. The initial soil (pre-sowing) analysis used in pots is as under;

Soil Texture: CLAY LOAM Sand % = 48, Silt % = 28, Clay % = 24

Table 3.1 Properties of soil used in the pot culture

Parameters Units

ECe 1.25 dS m-1

pHs 7.6

SAR 2.2 (mmol L-1)1/2

Soil K 100 mg kg-1 Soil texture Clay loam

Saturation percentage 30%

3.1.6 Field experiment (Study 4)

A field experiment was conducted in the farmer’s field at Chak. No.

86/G.B Toba Tek Singh, Pakistan, in a naturally salt affected soil to assess the

performance of salt tolerant and salt-sensitive genotypes of maize, previously tested

in solution and pot culture studies under greenhouse conditions. The properties of soil

of the site are presented in Table 3.2. A Randomized Complete Block Design

(RCBD) was employed for analysis of variance and Duncan’s Multiple Range Test

was used to separate treatment means (Fisher, 1925). At maturity, plants were

harvested manually and different yield parameters (plant height, total biomass and

grain yield) were recorded. Fresh plant leaves were placed in eppendorf tubes and

was kept in the refrigerator for the determination of Na+, K+ and Cl- later. Maize

grains were used for the determination of oil, crude fiber, total soluble sugar and

XLIII

protein contents. Proline content was also determined using the method of Bates et al.

(1973).

Table 3.2 Properties of the salt affected soil used in the field experiment.

Parameters Unit Soil depth 0-15 cm 15-30 cm ECe dS m

-1 9.95 10.60

pHs --- 8.31 8.20 SAR (mmol L

-1)

1/2 18.65 17.10

K mg kg-1

70.0 85.50 Texture ---- Clay loam

3.2 PLANT ANALYSIS

3.2.1 Plant sample collection

In the solution culture experiments, the youngest fully expanded leaves

were detached at harvesting, rinsed in distilled water, blotted with tissue paper and

stored in separate eppendorf tubes at freezing temperature for leaf sap extraction to

determine Na+, K+ and Cl-. In the soil culture experiments, the leaf next to the flag

leaf was sampled and stored in the same way as described above for determination of

Na+, K+ and Cl- (Gorham et al., 1984).

3.2.2 Leaf sap extraction

Frozen leaf samples were thawed and crushed using a stainless steel rod

with tapered end. The sap was collected in other eppendorf tubes by Gilson pipette

and centrifuged at 6500 rpm for 10 minutes. The supernatant sap was taken in new

eppendorf tubes and was stored in the refrigerator for the determination of Na+, K+

and Cl- (Gorham et al., 1984).

3.2.3 Determination of Na+ and K+

The leaf sap was diluted as required by adding distilled water. The dilution

factor was correlated with the original value and sodium and potassium

concentrations were by using Sherwood 410 Flame photometer with the help of self

prepared standard solutions using reagent grade salts of NaCl and KCl.

3.2.4 Determination of Cl-

XLIV

Chloride in the diluted leaf sap was determined using Sherwood 926

Chloride analyzer.

3.3 PHYSIOLOGICAL ATTRIBUTES

3.3.1 Determination of water potential

The third leaf from top (fully expanded youngest leaf) was excised to

determine the leaf water potential (pot culture study) with Scholander type pressure

chamber (Scholander et al., 1965).

3.3.2 Gas exchange characteristics

Measurement of transpiration (E) and stomatal conductance (gs) were

made on fully expanded youngest leaf (pot culture study) of each plant using an open

system LCA-4 ADC portable infrared gas analyzer (Analytical Development

Company, Hoddesdon, England). Measurements were performed from 9 to 11 a.m.

with the following specifications/ adjustments: molar flow of air per unit leaf area

403.3 mmol m-2 s-1, atmospheric pressure 99.9 KPa, water vapour pressure in the

chamber ranged from 6.0 to 8.9 mbar, PAR at leaf surface was ranged from 1500 to

1711 μ mol m-2 s-1, temperature of leaf ranged from 28.4 to 32.7oC, ambient

temperature ranged from 22.7 to 29.9oC, ambient CO2 concentration 352 μ mol mol-1.

3.3.3 Growth parameters

The following morphological and growth parameters were recorded from

different studies that are usually affected by salinity.

1. Plant height (Total length of stem to highest leaf) (cm) 2. Root length (Total summation of primary root length) (cm) 3. Shoot fresh weight (g plant-1) 4. Root fresh weight (g plant-1) 5. Shoot dry weight (g plant-1) 6. Root dry weight (g plant-1) 7. 1000 seed weight (g) 8. Grain yield (Mg ha-1)

3.4 BIOCHEMICAL ATTRIBUTES

XLV

Biochemical parameters were determined in pot culture study and field

experiment which are usually affected by salinity and they are also the indicator of

plant tolerance against salinity.

3.4.1 Protein analysis

Total soluble proteins were determined using the method of Lowery et al.

(1951). A sample of 0.2 g seed material (grain) was taken and chopped in 5 ml

phosphate buffer 0.2 M (pH 7.0). Two tubes containing 0.5 ml and 1.0 ml of seed

extract were prepared for protein estimation. Solution of 0.5, 0.1, 0.2, 0.4, 0.6 and

1.0 ml of standard Bovine Serum Albumin (BSA) were simultaneously used in the

experiment. The volume of each tube was topped to 1.0 ml with distilled water. The

blank contained only 1.0 ml distilled water. One ml of solution (copper reagents) was

added to each test tube. The reagents in the test tube were thoroughly mixed and

allowed to stand for 10 minutes at room temperature. Then 0.5 ml of Folin-phenol

reagent (1:1 diluted) was added, mixed well and kept for 30 minutes at room

temperature. The optical density (O.D) was read at 620 nm on spectrophotometer

(Hitachi-220).

3.4.2 Determination of total soluble sugars

Total soluble sugars were determined according to the method of Yemm and

Willis (1954). Well-grinned seed material (0.1g) was extracted in 80% ethanol

solution. Dried material was ground so as to pass through 1mm sieve of millimicro

mill (Model Culatti, DFH-48) and it was shaken for 6h at 600C. This extract was used

for the estimation of total soluble sugars. Plant extract (100 μ l) was taken in 25 ml

test tubes and 6 ml anthrone reagent was added, and then heated in boiling water bath

for 10 minutes. The test tubes were incubated for 20 minutes at room temperature

(25oC). Optical density was read at 625 nm on spectrophotometer (Hitachi 220).

Blank was also run in the same way. The soluble sugars were calculated from a

standard curve developed using glucose.

3.4.3 Oil content determination

XLVI

For oil content, seed samples of each treatment were dried for 16 h at 50oC

and then ground into a powder. The procedure adopted was the official method of the

Association of Official Analytical Chemists (AOAC) (1984).

Procedure 1. Weighed accurately 4 to 5 g. of the ground sample into a filter paper and

enclosed in a second filter paper folded in such a fashion as to prevent

escape of meal. The second filter paper was left open at the top like

thimble. A piece of absorbent cotton was placed in the top of thimble to

distribute the solvent as it dropped on the sample.

2. Wrapped sample was placed in the Butt extraction tube and assembled the

apparatus. 25 ml of petroleum ether was added to the tarred extraction

flask before attaching to the tube.

3. Extraction tube was heated on the water bath at such a rate that the solvent

would drop from the condenser on the center of the thimble at the rate of

150 drops per minute.

4. Extraction was continued for 4 hours.

5. The extraction flask cooled and disconnected. Evaporated the ether on a

water bath until no odor of ether remained. A gentle stream of clean, dry

air was used to facilitate removal of the solvent. It was cooled to room

temperature, carefully removed the moisture outside the flask and then

weighed it. Repeated the heating until constant weight was obtained.

6. Moisture in the ground sample was determined as follows:

a. Weighed 5 g. into a tarred dish. b. Slipped the cover on the bottom of the dish and placed the dish in a

forced draft oven. It was dried at 101oC for 2 hours. c. Then removed the dish from oven and covered immediately. Cooled in

desiccator containing an efficient desiccant for 30 minutes and weighed.

Loss in weight Moisture in ground sample, % == ---------------------- x 100 Weight of sample

Calculation

XLVII

Weight of oil Oil in ground sample, % == ----------------------- x 100 Weight of sample

3.4.4 Crude fiber determination

Crude fiber (%) was determined by the method of AOAC, 1995.

Procedure

Two grams of grain material were extracted with petroleum ether. The

material was transferred to 600 ml beaker, avoiding fiber contamination from paper

or brush. Then 1.0 g of freshly prepared asbestos, 200 ml boiling 1.25% H2SO4, and

one drop of diluted antifoam were added. Bumping chips were also added. The

beaker was placed on the digestion apparatus with pre-adjusted hot plate and boiled it

exactly for 30 min, rotated the beaker periodically to keet the solids from adhering to

sides. The beaker was removed and the contents were filtered. The mat and residues

were dried for 2 hr at 130±2oC, then cooled in desiccator and weighed, ignited for 30

min at 600±15oC, cooled in desiccator and reweighed.

% Crude fiber in ground sample = (Loss in wt. on ignition – loss in wt. of asbestos

blank) × 100/wt. of sample.

3.4.5 Proline determination

Extraction and determination of proline (pot culture study and field

experiment) were performed according to the method of Bates et al. (1973). Leaf

samples (1.0 g) were extracted with 3% sulphosalicylic acid. Extracts (2 ml) were

held for 1.0 h in boiling water by adding 2.0 ml ninhydrin and 2.0 ml glacial acetic

acid, after which cold toluene (4.0 ml) was added. Proline contents were determined

by spectrophotometer at 520 nm and calculated as μmol g-1 DW against standard

proline.

3.5 STATISTICAL ANALYSIS

The data were subjected to a statistical analysis using Steel and Torrie,

1980. The standard errors (±) were used to compare means (Fisher, 1925).

Table 3.3 A brief of all experiments and the parameters monitored in each

XLVIII

experiment.

Experiments conducted Parameters monitored

Study 1 Solution culture study

This study was carried out in a wire

house during spring season. Fifteen

maize genotypes (synthetic and hybrid)

were grown in nutrient solution. Three

salinity levels viz., control/natural

salinity of water (12.5 mol m-3 NaCl), 70

and 100 mol m-3 were developed with

NaCl. The pH of the solution was

maintained. At the age of 4 weeks plant

were harvested.

Study 2 Solution culture study

This study was also carried out in a wire

house. The prominent group of genotypes

(synthetic) from the 1st study on the basis

of plant growth parameters and leaf ionic

concentrations were selected and used in

this study. The same three salinity levels

as used in 1st study with three levels of

potassium (1.0, 5.5 and 8.0 mM K) were

used with three replications. Plants were

transplanted in the nutrient solution and

salinity was developed up to desired

levels. Plants were harvested at the age of

28 days after the imposition of salinity.

Study 3 Pot culture study

This study was also carried out in a wire

house. The soil was collected from

Plant height, total root length, shoot fresh

weight, root fresh weight and their dry

weight were recorded. Leaf ion contents

Na+, K+, Na+: K+ ratio and Cl-1 were

determined by the described methods.

The above growth parameters were

studied because plant growth and their

related parameters are usually effected by

the salinity and the magnitude of salinity

is determined from these parameters.

Leaf ionic concentrations also revealed

the extent of salinity and its tolerance.

The following parameters were studied to

find out the effect of salinity within and

among the genotypes and plant response

to the added potassium. Root length,

shoot length, shoot fresh and root fresh

weights, shoot and root dry weight, Leaf

ionic concentrations like Na+, K+, Na+:

K+ ratio and Cl-1 were determined.

Transpiration rate, stomatal conductance

were measured with IRGA. At maturity

plants were harvested and different yield

XLIX

agriculture field from research area,

University of Agriculture, Faisalabad.

Glazed earthen pots were filled with soil

at the rate of 12 kg per pot. The original

level of soil K was considered as control

(1st level). The 2nd level of K was kept @

200 mg kg of soil by adding potassium

sulphate. The desired level of salinity (10

dS m-1) was developed by mixing NaCl

salt in the soil. Nitrogen and phosphorus

were applied at 200 and 150 kg ha-1. The

seeds of two maize genotypes (salt

tolerant and salt sensitive) were sown in

these pots in the spring season. The pots

were irrigated with tap water.

Study 4 Field experiment

A field experiment was conducted in a

saline located near Faisalabad (latitude

31° 25/North and longitude 73° 90/East).

The climate was arid with annual rain fall

250-300 m, 1625 mm annual evaporation

and 35-70% humidity. Before laying

experiment soil sample collected and

analyzed. The filed was properly laid out

according to treatments i.e. control (no

added K and K@ 200kg ha-1. The

sterilized seed of genotypes S-2002 and

Akbar were sown in plot size 4×4m. SOP

was used as potassium source.

parameters (plant height, shoot fresh

weight, 100 seed weight and grain yield

plant-1 were recorded. After 65 days of

germination plant leaves were stored for

the determination of Na+, K+, Na+: K+

ratio and Cl-1 so that the effect of salinity

and the extent of salinity can be

determined. Maize grains were stored for

the determination of oil contents, crude

fiber, total soluble sugars and protein

contents. Fresh leaves were used for the

determination of water potential and

proline contents. So that the effect of

salinity on crop quality and crop response

to potassium applied determined.

After 65 days of germination plant leaves

were stored for the determination of Na+,

K+, Na+: K+ ratio and Cl-1. The plants

were harvested at maturity and plant

height, total biomass were recorded.

Cobs were separated for grain yield. 0.5 g

of plant tissue was stored in toluene for

proline determination. Grains were also

used for the determination of oil contents,

crude fiber, total soluble sugars and

protein contents. These parameters were

studied to find the effect of salinity on

crop quality.

L

STUDY-1: SCREENING OF MAIZE GENOTYPES FOR SALT TOLERANCE IN A NUTRIENT SOLUTION CULTURE

4.1 RESULTS AND DISCUSSION

4.1.1 Results

4.1.2 Physical growth parameters

4.1.2.1 Shoot length (cm)

The data regarding the shoot length (SL) of maize genotypes are presented

in Table 4.1. S-2002 produced the maximum shoot length while Akbar produced the

shortest plants among the synthetic genotypes and all the genotypes differed

significantly in all the treatments (Appendix 1). In case of hybrid genotypes, Q 2139

and Q 2094 produced the highest plants and varied non-significantly, while Q 0806

had the shortest plants among all the hybrid genotypes. In non-saline (control)

conditions S-2002 produced significantly taller plants (92.6 cm) compared to other

genotypes followed by SL 2002 (80.6 cm), Ev 6098 (78.3 cm) and Agati 2000 (72.6

cm), among the synthetic genotypes. Among the hybrid genotypes, Q 2139 and Q

2094 produced the highest shoot length while Q 0806 had the minimum shoot length

at all the salinity levels.

The shoot length of maize genotypes dropped significantly at 70 mol m-3

NaCl and S-2002 produced higher shoot length as compared to other genotypes on

absolute or relative basis. Among the synthetic/recommended genotypes, Akbar

remained sensitive and produced minimum shoot length, however, hybrid genotypes

produced far less shoot length. A similar trend was also observed at 100 mol m-3

NaCl where S-2002 and Akbar remained tolerant and sensitive, respectively among

the synthetic/recommended genotypes. The reduction was more pronounced in all

hybrid genotypes and the maximum reduction was observed in the case of Q 0806.

On the basis of their respective value in terms of % of respective control more

reduction was observed in the case of genotypes prefixed with Q as compared to the

synthetic lines/varieties, with the exception of genotype Q 2109.

Table 4.1 Shoot length (cm) of maize genotypes grown under different

LI

salinity levels and harvested at 4 weeks age. Genotypes Control (12.5 mol m-3

NaCl)/natural salinity70 mol m-3 NaCl 100 mol m-3 NaCl

Synthetic SL 2002 Ev 1098 Ev 6098 S-2002

Agati 2000 Akbar

Ev 5098 Hybrid Q 2139 Q 2094 Q 2100 Q 2109 Q 9515 Q 0806 Q 2414 Q 8915

80.6±1.2070.3±1.2078.3±1.4592.6±1.76 72.6±1.20 61.6±1.7670.6±0.67

72.3±1.20 71.3±1.45 61.0±1.15 53.0±1.1559.0±1.1551.3±0.88 68.3±1.76 66.6±1.45

63.0±1.15 (78)55.6±0.88 (79)64.6±0.88 (83)83.3±0.88 (90) 59.0±0.58 (81) 51.3±0.88 (83)53.6±0.88 (76)

51.3±0.88 (71) 51.0±1.15 (71) 47.6±1.20 (78) 50.0±1.53 (94)41.3±1.45 (70)34.6±0.88 (68) 46.3±1.45 (68) 47.0±1.73 (71)

51.6±0.67 (64)44.3±0.67 (63)50.0±0.58 (64)76.3±0.88 (82) 46.3±0.88 (64) 39.6±0.33 (64)42.0±0.58 (59)

36.6±0.88 (51) 36.0±1.53 (50) 32.3±1.33 (53) 35.3±0.67 (67)28.6±0.88 (49)22.3±0.88 (44) 33.6±0.67 (49) 32.6±1.45 (49)

Each value is an average of 3 replications ± S.E. Values in ( ) are % of respective control. 4.1.2.2 Root length (cm)

Data regarding the root length of maize genotypes are presented in Table

4.2. The maximum root length (RL) was observed in S-2002 while Akbar produced

the shortest roots among the synthetic genotypes under all treatments (table 4.2). For

hybrid genotypes, genotype Q 2094 produced the maximum root length while Q

0806, Q 2414 and Q 8915 produced the minimum root length in all treatments and

varied non-significantly (Apendix-2). In non-saline (control) conditions, S-2002

produced significantly higher root length (83 cm) compared to other genotypes

followed by SL 2002 (75.3 cm), Agati 2000 (69.6 cm), Ev 1098 and Ev 5098 (63.6

and 63.6 cm plant-1), among the synthetic genotypes. The hybrid genotypes produced

much lower root length as compared to the synthetic genotypes.

The root length of maize genotypes was reduced significantly at 70 mol m-3

NaCl and S-2002 produced significantly higher root length as compared to other

genotypes. Among the synthetic/recommended genotypes, Akbar remained sensitive

and produced minimum root length, however, hybrid genotypes had far less root

length. A similar trend was also observed at 100 mol m-3 NaCl where S-2002 and

Akbar remained tolerant and sensitive, respectively among the synthetic/

LII

recommended genotypes, however, the reduction was more pronounced in case of all

hybrid genotypes on the basis of their absolute and relative values as compared to the

synthetic/recommended genotypes. On the basis of their value in terms of % of

respective control, more reduction was observed in case of genotypes prefixed with Q

as compared to the synthetic/recommended lines/varieties. The genotypes Q 9515

and Q 0806 had almost the same values at 70 mol m-3 NaCl stress. Genotypes Q

9515 and Q 2414 had the similar % reduction in root length at 70 mol m-3 NaCl

stress.

Table: 4.2 Root length (cm) of maize genotypes grown under different salinity levels and harvested at 4 weeks age.

Genotypes Control (12.5 mol m-3

NaCl)/natural salinity 70 mol m-3 NaCl 100 mol m-3 NaCl

Synthetic SL 2002 Ev 1098 Ev 6098 S-2002

Agati 2000Akbar

Ev 5098 Hybrid Q 2139 Q 2094 Q 2100 Q 2109 Q 9515 Q 0806 Q 2414 Q 8915

75.3 ±1.33 63.6 ±1.33 62.6 ±0.88 83.0 ±1.53 69.6 ±0.33 61.6 ±0.67 63.6 ±0.67 39.0 ±1.15 45.6 ±1.20 37.0 ±1.15 36.0 ±1.53 34.3 ±2.33 32.6 ±1.76 31.3 ±1.20 30.6 ±1.20

64.3±1.20 (85)61.6±3.71 (97)57.3±1.45 (91)72.3±0.67 (87)60.3±0.33 (87)53.3±1.76 (86)56.6±1.67 (89)

32.6±1.45 (84)33.6±1.20 (74)30.3±0.88 (82)29.6±1.33 (82)26.6±2.03 (78)26.3±1.20 (81)24.3±1.20 (78)22.3±0.88 (73)

60.6±1.20 (81) 51.6±0.67 (81))51.3±1.33 (82) 67.6±1.20 (82) 54.3±0.33 (78) 45.3±1.76 (74) 52.6±1.33 (83)

27.0±1.15 (69) 28.3±1.33 (62) 25.0±1.15 (68) 24.3±1.20 (68) 19.3±1.45 (56) 20.3±1.20 (62) 18.0±0.58 (57) 16.6±0.67 (54)

Each value is an average of 3 replications ± S.E. Values in ( ) are % of respective control.

4.1.2.3 Shoot fresh weight (g plant-1) (The genotypes prefixed as Q were hybrid while the remaining were

synthetic and recommended (varieties/lines) genotypes). In non-saline (control)

conditions, S-2002 produced significantly highest shoot fresh weight (31.6 g plant-1)

compared to other genotypes followed by SL 2002 (22.6), Agati 2000 (20.8) and Ev

6098 (20.0 g plant-1), among the synthetic genotypes. The hybrid genotypes produced

much lower shoot fresh weight as compared to the synthetic/recommended

genotypes.

LIII

Appendix3 showed significant variation among genotypes regarding

hybrid and synthetic. The interactions between and within the synthetic and hybrid

genotypes were also significant. The shoot fresh weight of maize genotypes was

reduced significantly at 70 mol m-3 NaCl and S-2002 produced significantly highest

shoot fresh weight as compared to other genotypes. Among the synthetic/

recommended genotypes, Akbar remained sensitive and produced lowest shoot fresh

weight, however hybrid genotypes produced far less shoot fresh weight. A similar

trend was also observed at 100 mol m-3 NaCl where S-2002 and Akbar remained

tolerant and sensitive, respectively among the synthetic/recommended genotypes,

however the reduction was more pronounced in case of all hybrid genotypes on the

basis of their absolute and relative value as compared to the synthetic/recommended

genotypes. Overall, the synthetic genotypes produced more shoot fresh weight as

compared to the hybrid genotypes on both salinity levels as compared to the control.

Table 4.3 Shoot fresh weight (g plant-1) of maize genotypes grown under different salinity levels and harvested at 4 weeks age. Genotypes Control (12.5 mol m-3

NaCl)/natural salinity 70 mol m-3 NaCl 100 mol m-3 NaCl

Synthetic SL 2002 Ev 1098 Ev 6098 S-2002

Agati 2000 Akbar

Ev 5098 Hybrid Q 2139 Q 2094 Q 2100 Q 2109 Q 9515 Q 0806 Q 2414 Q 8915

22.6 ±0.8116.4 ±0.6820.0 ±0.7231.6 ±1.1620.8 ±0.3515.1 ±0.9618.7 ±0.85

17.9 ±0.3415.9 ±0.7715.7 ±0.4512.6 ±0.8812.5 ±0.697.0 ±0.205.2 ±0.415.2 ±0.21

19.2±0.58 (85) 13.6±0.92 (83) 15.4±0.68 (77) 25.6±1.29 (81) 16.1±0.58 (78) 11.0±0.42 (73) 14.9±0.32 (80)

15.4±0.29 (86) 13.6±0.66 (86) 13.5±0.37 (86) 10.8±0.75 (86) 10.7±0.59 (86) 6.0±0.17 (86) 4.5±0.34 (86) 4.4±0.18 (86)

16.0±0.13 (71) 10.5±0.44 (64) 12.1±0.10 (60) 23.1±1.20 (73) 12.8±0.26 (62) 9.0±0.61 (60)

12.1±0.62 (65)

13.2±0.25 (73) 11.7±0.56 (73) 11.5±0.32 (74) 9.2±0.64 (73) 9.2±0.50 (73) 5.1±0.15 (73) 3.8±0.28 (74) 3.8±0.15 (74)

Each value is an average of 3 replications ± S.E. Values in ( ) are % of respective control.

4.1.2.4 Shoot dry weight (g plant-1)

In non-saline (control) conditions S-2002 produced maximum shoot dry

weight (5.7 g plant-1) compared to other genotypes followed by SL 2002 (4.4 g plant-

LIV

1), and Ev 6098 (3.7 g plant-1). Among the hybrid genotypes Q 2139 and Q 2094

produced highest shoot dry weight and varied non-significantly while Q 8915 and Q

2414 had the minimum shoot dry weights with non- significant difference.

The Anova table showed that there was a significant variation or

differences at both salinity levels (Appendix4). The interactions were also significant

within and among the hybrid and synthetic/recommended lines/varieties. The shoot

dry weight of maize genotypes was reduced significantly at 70 mol m-3 NaCl and S-

2002 produced highest root dry weight as compared to other genotypes. Among the

synthetic/recommended genotypes, Akbar remained sensitive and produced lower

shoot dry weight, while; the hybrid genotypes produced far less shoot dry weight. A

similar trend was also observed at 100 mol m-3 NaCl, where S-2002 and Akbar

remained tolerant and sensitive, respectively among the synthetic/recommended

genotypes. The reduction was more pronounced in case of all hybrid genotypes on

the basis of their absolute and relative value as compared to the

synthetic/recommended genotypes and more reduction was observed in case of Q

9515. On the basis of their respective value in terms of % of control, more reduction

was observed in case of genotypes prefixed with Q as compared to the

synthetic/recommended lines/varieties.

Table 4.4 Shoot dry weight (g plant-1) of maize genotypes grown under different salinity levels and harvested at 4 weeks age.

Genotypes Control (12.5 mol m-3

NaCl)/natural salinity70 mol m-3 NaCl 100 mol m-3 NaCl

LV

Synthetic SL 2002 Ev 1098 Ev 6098 S-2002

Agati 2000 Akbar

Ev 5098 Hybrid Q 2139 Q 2094 Q 2100 Q 2109 Q 9515 Q 0806 Q 2414 Q 8915

4.4±0.09 3.1±0.04 3.7±0.10 5.7±0.07 3.4±0.07 2.8±0.06 3.1±0.08

3.1±0.09 2.8±0.07 2.7±0.04 2.4±0.18 2.2±0.07 1.5±0.03 1.2±0.08 1.0±0.07

3.9±0.03 (89) 2.7±0.04 (87) 3.1±0.05 (85) 5.2±0.04 (92) 3.0±0.03 (89) 2.3±0.04 (82) 2.7±0.08 (87)

2.8±0.04 (89) 2.7±0.16 (94) 2.6±0.08 (96) 2.1±0.12 (91) 2.1±0.10 (93) 1.2±0.04 (79) 1.2±0.06 (96) 1.1±0.05 (108)

3.6±0.07 (82) 2.3±0.05 (75) 2.9±0.04 (79) 4.9±0.05 (87) 2.7±0.03 (80) 1.9±0.04 (68) 2.2±0.04 (72)

2.6±0.05 (84) 2.3±0.10 (82) 2.2±0.06 (82) 1.8±0.09 (77) 1.0±0.03 (46) 0.8±0.03 (52) 0.7±0.06 (63) 0.8±0.04 (77)

Each value is an average of 3 replications ± S.E. Values in ( ) are % of respective control.

4.1.2.5 Root fresh weight (g plant-1) In non-saline (control) conditions, root fresh weight produced by ‘S-2002’

was significantly higher than other genotypes compared to other genotypes followed

by SL 2002, and Ev 6098 among the synthetic genotypes. Among the hybrid

genotypes, Q 2139 and Q 2094 produced highest root fresh weight and varied

significantly while Q 8915 had the minimum root fresh weight. Hybrid and synthetic

genotypes showed significant variations under different salinity levels. On the basis

of their respective values in terms of % of control more reduction was observed in

case of genotypes prefixed with Q as compared to the synthetic/recommended

lines/varieties. The root fresh weight of maize genotypes was reduced significantly at

70 mol m-3 NaCl, with S-2002 being the most tolerant genotype. Among the synthetic

genotypes, Akbar remained sensitive and produced lesser root fresh weight, while,

hybrid genotypes produced far less root fresh weight. A similar trend was also

observed at 100 mol m-3 NaCl where S-2002 and Akbar remained tolerant and

sensitive, respectively among the synthetic genotypes, however, the reduction was

more pronounced in case of all hybrid genotypes on the basis of their absolute and

relative values as compared to the synthetic/recommended genotypes and greater

relative reduction was observed in case of Q 8915.

Table 4.5 Root fresh weight (g plant-1) of maize genotypes grown under

LVI

different salinity levels and harvested at 4 weeks age. Genotypes Control (12.5 mol m-3

NaCl)/natural salinity 70 mol m-3 NaCl 100 mol m-3 NaCl

Synthetic SL 2002 Ev 1098 Ev 6098 S-2002

Agati 2000 Akbar

Ev 5098 Hybrid Q 2139 Q 2094 Q 2100 Q 2109 Q 9515 Q 0806 Q 2414 Q 8915

7.37±0.18 6.13±0.09 7.22±0.07

11.27±0.09 7.15±0.08 5.87±0.08 6.50±0.15

5.76±0.33 5.39±0.32 4.97±0.12 4.31±0.24 3.66±0.15 2.21±0.07 1.84±0.17 1.81±0.07

6.90±0.12 (94) 5.22±0.06 (85) 6.25±0.13 (87)

10.73±0.09 (95) 6.47±0.07 (90) 5.08±0.04 (87) 5.91±0.07 (91)

4.67±0.14 (81) 4.45±0.11 (83) 4.36±0.20 (88) 3.49±0.20 (81) 2.96±0.03 (81) 1.78±0.05 (81) 1.44±0.11 (78) 1.39±0.04 (77)

5.97±0.20 (81)4.23±0.09 (69)5.39±0.15 (75)9.97±0.07 (88)5.43±0.13 (76)4.16±0.14 (71)4.93±0.04 (76)

3.90±0.06 (68)3.72±0.13 (69)3.61±0.15 (73)2.91±0.16 (67)2.37±0.04 (65)1.48±0.06 (67)1.14±0.11 (62)1.11±0.04 (61)

Each value is an average of 3 replications ± S.E. Values in ( ) are % of respective control.

4.1.2.6 Root dry weight (g plant-1)

On the basis of their respective values in terms of % of control more

reduction was observed in case of genotypes prefixed with Q as compared to the

synthetic lines/varieties. The effect of salinity and the variation in genotypes was

significant (Appendix 6). In non-saline (control) conditions, S-2002 produced

maximum root dry weight compared to other genotypes followed by Agati 2000, and

SL 2002 among the synthetic genotypes (Table 4.6). Among the hybrid genotypes, Q

2139 and Q 2100 had the highest root dry weight while Q 8915 and Q 2414 had the

minimum root dry weights with non- significant difference.

The root dry weight of maize genotypes was reduced significantly at 70

mol m-3 NaCl, and S-2002 produced maximum root dry weight as compared to other

genotypes. Among the synthetic/recommended genotypes, Akbar remained sensitive

and produced lower root dry weight, however, hybrid genotypes produced far less

root dry weight. A similar trend was also observed at 100 mol m-3 NaCl where S-

2002 and Akbar remained tolerant and sensitive, respectively among the

synthetic/recommended genotypes, however, the reduction was more pronounced in

case of all hybrid genotypes on the basis of their absolute and relative value as

LVII

compared to the synthetic genotypes and more % reduction was observed in case of

Q 9515.

Table 4.6 Root dry weight (g plant-1) of maize genotypes grown under different salinity levels and harvested at 4 weeks age. Genotypes Control (12.5 mol m-3

NaCl)/natural salinity70 mol m-3 NaCl 100 mol m-3 NaCl

Synthetic 2002

Ev 1098 Ev 6098 S-2002

Agati 2000 Akbar

Ev 5098 Hybrid Q 2139 Q 2094 Q 2100 Q 2109 Q 9515 Q 0806 Q 2414 Q 8915

1.11±0.02 0.99±0.01 1.02±0.02 1.79±0.10 1.15±0.03 1.02±0.02 1.03±0.03

0.88±0.06 0.90±0.06 0.82±0.03 0.69±0.04 0.57±0.05 0.36±0.01 0.30±0.03 0.30±0.01

0.94±0.02 (85)0.73±0.02 (73)0.55±0.02 (83)1.53±0.06 (85)0.93±0.04 (81)0.72±0.02 (71)0.68±0.02 (66)

0.71±0.03 (81)0.63±0.02 (70)0.68±0.06 (83)0.56±0.05 (81)0.45±0.01 (79)0.30±0.01 (82)0.24±0.02 (78)0.22±0.01 (73)

0.88±0.01 (80) 0.63±0.01 (64) 0.78±0.01 (77) 1.44±0.05 (80) 0.79±0.01 (68) 0.58±0.05 (57) 0.59±0.02 (57)

0.60±0.01 ((68) 0.58±0.02 (64) 0.46±0.03 (56) 0.35±0.01 (51) 0.23±0.01 (41) 0.18±0.02 (50) 0.18±0.01 (59) 0.14±0.01 (48)

Each value is an average of 3 replications ± S.E. Values in ( ) are % of respective control.

4.1.2.7 Sodium concentration in leaf sap of maize genotypes

The data regarding the Na+ concentration (mol m-3) in leaf sap of maize genotypes are

presented in Table 4.7. The results showed a significant variation among hybrid and

synthetic genotypes under different salinity levels (70 and 100 mol m-3 NaCl) for Na+

concentration (mol m-3) in leaf sap of maize genotypes. The interactions between and

within synthetic and hybrid genotypes were significant (Appendix 7).

Maize genotypes differed significantly with regard to Na+ concentration in

their youngest fully expanded flag leaf when compared within a particular treatment.

In case of synthetic genotypes, Akbar had the maximum Na+ concentration in the leaf

sap and minimum Na+ concentration was observed in S-2002 as compared to control

at both the salinity levels. In case of hybrid genotypes Q 8915 had the highest Na+

concentration in leaf sap but Q 9515 and Q 0806 differed non-significantly at 70 mol

m-3 NaCl level. The minimum concentration in this treatment was observed in Q

2139. The comparison between synthetic and hybrid genotypes showed that synthetic

LVIII

genotypes had lesser concentration of Na+ in leaf sap as compared to the hybrid

genotypes at 70 or 100 mol m-3 NaCl as compared to the control. On the basis of their

respective values in terms of % of control, more Na+ concentration was observed in

case of genotypes prefixed with Q as compared to the synthetic. The performance of

synthetic genotypes is better as compared to the hybrid genotypes in saline and non-

saline conditions. On relative basis as compared to respective control, synthetic

genotypes had lesser Na+ concentration while hybrid genotypes had about 4.0 times

higher Na+ concentration in leaf sap at 70 and 100 mol m-3 NaCl.

Table 4.7 Na+ concentration (mol m-3) in leaf sap of maize genotypes grown under different salinity levels and harvested at 4 weeks age. Genotypes Control (12.5 mol m-3

NaCl)/natural salinity70 mol m-3 NaCl 100 mol m-3 NaCl

Synthetic SL 2002 Ev 1098 Ev6098 S-2002 Agati 2000 Akbar Ev 5098 Hybrid Q 2139 Q 2094 Q 2100 Q 2109 Q 9515 Q 0806 Q 2414 Q 8915

42.3±2.83 44.2±3.32 43.1±3.01 35.1±1.59 43.5±3.12 48.1±3.19 41.0±2.14 57.3±3.66 49.2±2.87 55.9±3.94 49.8±3.28 50.4±2.35 55.5±3.69 56.8±4.58 53.5±1.95

83.7±7.39 124±9.27 104±9.35 64.5±4.87 91.2±7.11 139±9.40 113±7.80

143±11.43154±13.31147±13.40162±13.91159±14.22159±15.18171±14.61180±13.40

93.6±1.58 1256±1.60 111±0.47 76.1±1.23 101±0.80 146±2.50 118±0.99

145±2.23 154±1.66 162±1.57 158±1.93 163±1.78 168±2.36 160±1.39 183±3.35

Each value is an average of 3 replications ± S.E.

4.1.2.8 Potassium concentration in leaf sap of maize genotypes

Salinity reduced K+ concentration in leaf sap of all maize (15 genotypes)

by 20% and 35% at 70 and 100 mol m-3 NaCl, respectively (Table 4.8). Both salinity

levels differed significantly with regard to their effect on potassium concentration in

maize leaves (Appendix 8). In the control treatment, S-2002 had the highest

concentration of K+ in leaf sap and varied significantly with the other genotypes. The

comparison between synthetic and hybrid genotypes showed that S-2002

accumulated the highest concentration of K+, while, Akbar accumulated the lowest

concentration of K+ in leaf sap. In case of hybrid genotypes all the genotypes had less

LIX

K+ concentration in leaf sap than that of Akbar and lowest concentration of K+ was

observed as compared to the synthetic genotypes in all the treatments. On the basis of

their respective values in terms of % of control, less K+ accumulation was observed

in case of genotypes prefixed with Q as compared to the synthetic/recommended

lines/varieties. The performance of synthetic/recommended genotypes is better as

compared to the hybrid genotypes in saline and non-saline conditions. On an average

basis, with respect to control, the decrease in K+ concentration of leaf sap of

synthetic/recommended genotypes/lines was about 20% at 70 mol m-3 NaCl and

about 28% at 100 mol m-3 NaCl.

Table 4.8 K+ concentration (mol m-3) in leaf sap of maize genotypes grown under different salinity levels and harvested at 4 weeks age. Genotypes Control (12.5 mol m-3

NaCl)/natural salinity 70 mol m-3 NaCl 100 mol m-3 NaCl

Synthetic SL 2002 Ev 1098 Ev6098 S-2002 Agati 2000 Akbar Ev 5098 Hybrid Q 2139 Q 2094 Q 2100 Q 2109 Q 9515 Q 0806 Q 2414 Q 8915

135±7.67 118±6.91 148±7.60

189.2±12.29 124±10.20 98.8±10.03 113±8.14

71.4±5.37 70.0±4.39 60.4±3.42 54.1±4.88 57.9±3.93 51.3±2.50 66.7±4.59 64.5±2.36

111±0.93 97.4±1.91 120±0.32 158±1.07 94.5±0.65 69.6±3.07 90.2±2.94

47.5±1.28 46.3±1.62 43.5±1.25 46.3±1.70 37.8±1.04 32.4±1.37 42.3±0.98 42.3±2.04

97.8±4.59 87.7±2.89 106±4.56 147±5.52 82.4±3.06 62.9±1.60 80.1±1.65

36.4±2.29 34.6±3.34 31.3±2.32 30.5±1.38 33.5±2.27 30.0±1.49 31.1±2.07 33.6±2.25

Each value is an average of 3 replications ± S.E.

4.1.2.9 K+: Na+ ratio in leaf sap of maize genotypes

K+: Na+ ratio dropped significantly due to salinity and reduction was more

with the increase in salinity level (Table 4.9). The maximum reduction was found at

100 mol m-3 NaCl treatment. A higher reduction was observed in case of hybrid

genotypes as compared to the synthetic ones due to the applied stress.

The genetic variation under different treatments was also significant with

reference to K+: Na+ ratio of leaf sap (Appendix 9). Synthetic genotype S-2002

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maintained the maximum K+: Na+ ratio in control (5.4), 70 mol m-3 (2.5) and at 100

mol m-3 NaCl (2.4) treatments. The minimum K+: Na+ ratio, on the other hand, was

observed in Akbar. On the other hand, the hybrid genotypes had lower K+: Na+ ratios

than Akbar. Q 2094 had a significantly higher K+: Na+ ratio than all other hybrid

genotypes. The hybrid genotype Q 0806 had the minimum K+: Na+ ratio among all

the hybrid genotypes in control treatment. On the basis of their respective value in

terms of % of control less K+: Na+ ratio was observed in case of genotypes prefixed

with Q as compared to the synthetic/recommended lines/varieties. The performance

of synthetic/recommended genotypes is better as compared to the hybrid genotypes in

saline and non-saline conditions. On an average basis with respect to control, in

Table 4.9 K+: Na+ ratio in leaf sap of maize genotypes grown under different salinity levels and harvested at 4 weeks after planting.

Genotypes Control (12.5 mol m-3

NaCl)/natural salinity 70 mol m-3 NaCl 100 mol m-3 NaCl

Synthetic SL 2002 Ev 1098 Ev6098 S-2002 Agati 2000 Akbar Ev 5098 Hybrid Q 2139 Q 2094 Q 2100 Q 2109 Q 9515 Q 0806 Q 2414 Q 8915

3.24±0.09 2.63±0.05 3.43±0.25 5.41±0.43 2.84±0.09 2.04±0.13 2.70±0.12

1.24±0.08 1.44±0.09 1.13±0.03 1.12±0.03 1.12±0.05 0.93±0.03 1.21±0.04 1.23±0.03

1.33±0.13 0.74±0.06 1.13±0.10 2.50±0.18 1.14±0.08 0.54±0.03 0.81±0.08

0.33±0.03 0.34±0.02 0.32±0.04 0.31±0.02 0.23±0.02 0.21±0.03 0.32±0.03 0.24±0.02

1.05±0.03 0.74±0.03 0.91±0.04 2.43±0.10 0.84±0.02 0.44±0.01 0.70±0.02

0.24±0.02 0.23±0.02 0.24±0.01 0.23±0.01 0.23±0.02 0.20±0.01 0.21±0.01 0.22±0.01

Each value is an average of 3 replications ± S.E.

synthetic/recommended genotypes/lines K+: Na+ ratio was about 13% of control at 70

mol m-3 NaCl and about 9.28% of control in their respective leaves at 100 mol m-3

NaCl. In hybrid genotypes K+: Na+ ratio was about 10.6% & 5.25% of control at 70

and 100 mol m-3 NaCl, respectively.

4.1.2.10 Chloride concentration in leaf sap of maize genotypes

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On an average basis of 15 maize genotypes used in this study, the

concentration of Cl- observed in leaf sap was 43.5 mol m-3, 63.1 mol m-3 and 74.9 mol

m-3 in control, 70 mol m-3 NaCl and 100 mol m-3 NaCl saline treatments, respectively

(Table 4.10). The Cl- concentration was significantly increased under 70 mol m-3

NaCl (1.5 times) and 100 mol m-3 NaCl (1.7 times) conditions (average of 15

genotypes). Increasing NaCl concentration in the culture solution had significantly

increased Cl- concentration in leaf sap (Table 4.10). However, the different genotypes

varied significantly Cl- contents in leaves. Q 8915 accumulated the highest

concentration of Cl- of all the genotypes under 100 mol m-3 NaCl as compared to the

control. All eight hybrid genotypes accumulated higher concentrations of Cl- as

compared to the synthetic genotypes except Akbar under both saline treatments as

compared to the control. Among the synthetic genotypes S-2002 had the lowest

concentration of Cl- while Akbar had the highest concentration of Cl-. On the basis of

their respective value in terms of % of control more % Cl- accumulation was

observed in case of hybrid genotypes as compared to the synthetic genotypes, which

accumulated less % of Cl- concentration. The performance of synthetic genotypes is

better as compared to the hybrid genotypes in saline and non-saline conditions.

Table 4.10 Cl- concentration (mol m-3) in leaf sap of maize genotypes grown under different salinity levels and harvested at 4 weeks age. Genotypes Control (12.5 mol m-3

NaCl)/natural salinity70 mol m-3 NaCl 100 mol m-3 NaCl

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Synthetic SL 2002 Ev 1098 Ev6098 S-2002 Agati 2000 Akbar Ev 5098 Hybrid Q 2139 Q 2094 Q 2100 Q 2109 Q 9515 Q 0806 Q 2414 Q 8915

41.2±2.88 39.3±1.08 39.4±2.60 27.5±0.08 35.2±1.75 43.7±2.93 35.5±4.70

44.4±2.76 48.6±2.09 51.9±3.73 45.9±1.80 49.4±2.22 48.5±4.25 46.7±1.38 55.4±2.56

44.3±2.24 41.9±2.20 43.1±2.82 30.4±2.48 38.0±1.75 54.6±3.70 38.7±2.21

77.7±2.63 78.4±4.69 72.6±1.65 81.4±2.14 74.2±5.00 79.0±4.75 92.4±4.40 99.4±5.08

55.3±2.56 56.0±3.01 58.1±5.92 40.6±2.90 51.6±3.24 73.8±3.55 53.5±0.93

83.4±7.79 91.1±7.78 93.2±5.86 87.0±6.59 91.6±7.05 92.1±7.95 92.0±12.54104±8.97

Each value is an average of 3 replications ± S.E.

4.1.3 Discussion

The primary objective of this study was to observe the pattern of gain in

growth parameters (shoot length/plant height, root length, shoot/root fresh and dry

weights) of maize genotypes belonging to two groups (synthetic/recommended and

hybrid line/varieties) of maize genotypes under saline conditions.

Our results show that the saline growth medium had an adverse effect on

growth parameters of both groups of maize genotypes but the growth inhibitory effect

of NaCl was more pronounced in case of hybrid genotypes as compared to the

synthetic/recommended genotypes. Application of NaCl significantly decreased plant

growth. The decrease in plant height was much more in the case of hybrid genotypes

as compared to the synthetic genotypes at 100 mol m-3 NaCl. Better plant height and

root length were observed for the genotype S-2002 as compared to the control (Table

4.1 & 4.2), among all the 15 maize genotypes.

The effect of high salinity (100 mol m-3 NaCl) was more pronounced than

the effect of low salinity (70 mol m-3 NaCl). The concentration of Na+ and Cl- in the

leaves increased significantly under both salinity levels while leaf K+ concentration

and K+: Na+ ratio decreased significantly and the decrease was more in hybrid

genotypes as compared to the synthetic genotypes. Higher concentrations of external

Na+ might have disrupted the osmotic balance of the cells thus inducing the water

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deficit. The higher concentrations of Na+ in leaves also become toxic and lead to salt

injury (Serrano et al., 1999).

The response of individual genotypes also varied significantly, when

compared within a treatment. Genotypic variability was also conspicuous under

different treatments. Varietal comparison further showed that the genotypes

accumulating more Na+ in their leaves showed poor growth performance. The

maximum shoot fresh and dry weight was produced by S-2002 while the minimum

shoot fresh and dry weight was observed in case of Akbar from synthetic genotypes

(Table 4.3 & 4.4). Hybrid genotypes produced less shoot fresh/ dry weights due to

higher accumulation of Na+ in leaves and had lower K+ concentration and K+: Na+

ratio due to which their performance was very poor. Maize is as a salt-sensitive plant

(Fortmeier and Schubert, 1995), although this sensitivity is dependent on the cultivar

(Cramer et al., 1994a). Various approaches like engineering techniques and the use of

amendments as well as mineral nutrients are advocated to improve plant survival

under salt stress (Marschner, 1995). Nevertheless, plant species and their genotypes

differ genetically in their ability to adapt to salt stress environment (Rozeff, 1995 ;

Wahid et al., 1997). Characteristics like dry matter production, Na+ accumulation,

K+/Na+ ratio and Ca2+/Na+ ratio have been considered a useful guide to assess salt

tolerance and selection of genotypes on this basis is an important strategy to

minimize growth reduction in saline soils (Santa-Maria and Epstein, 2001). The

decrease in dry matter production of genotypes in the presence of NaCl was due the

ion toxicity as Na+ displaced K+ and resulted in metabolic imbalances which reduced

growth and yields (Zhu, 2002). Chinnusamy et al. (2005) also reported that under salt

stress, the predominant cause of reduced plant growth appeared to be ion toxicity

rather than osmotic stress. Ion cytotoxicity was caused by the displacement of K+ by

Na+ in biochemical reactions and conformational changes and the loss of functions of

proteins as Na+ ions penetrated the hydration shells and interfered with non-covalent

interactions between their aminoacids. The magnitude of decline in dry matter

production among genotypes varied possibly because of their differential selectivity

for K+ over Na+ (Ashraf, 2002; Curtain and Naidu, 1998).

Bastias et al. (2004) measured a large reduction in dry weight production

(40%) of maize local cultivars when grown in 100 and 430 mM NaCl. The ecotype of

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maize Zea mays L. amylacea appeared to be more tolerant than that of the Pioneer

hybrid 3578 (Cramer et al., 1994a) and of the line G2 and SRO73 cultivars (Abd-El

Baki et al., 2000; Shabala et al., 1998), which have been reported to be as most salt

tolerant maize cultivars known after a long-term experiment.

It has been reported that maize cultivars with low concentrations of Na+ in

the shoot are more salt tolerant, suggesting that Na+ exclusion may be positively

correlated with salt tolerance (Fortmeier and Schubert, 1995). A number of screening

techniques/criteria have been proposed for screening of genotypes against salinity but

no one seems to be flawless or universally applicable. Shoot fresh/ dry weight and

Na+, K+ and K+: Na+ ratios are mostly considered as selection criteria. Different

physiological traits such as potassium selectivity, exclusion and/or compartmentation

of sodium, osmotic adjustment and the accumulation of organic solutes have all been

related to salt tolerance of cultivars of different species (Kingsbury and Epstein,

1986; Weimberg, 1987; Yeo et al., 1990 and Barrett-Lennard et al., 1999). Jamil et

al. (2005) found a considerable effect of salinity (0, 4.7, 9.4 and 14.1 dS m-1) on

germination, shoot length, shoot and root fresh weight of canola, cabbage and

cauliflower. It has been reported that the reductions in the rate of leaf and root growth

are probably due to factors associated with water stress rather than a salt-specific

effect (Munns, 2002). This is supported by the evidence that Na+ and Cl– are below

toxic concentrations in the growing cells. For example, in wheat growing in 120 mM

NaCl, Na+ in the growing tissues of leaves was at most only 20 mM, and only 10 mM

in the rapidly expanding zones, and Cl– only about 50 mM (Hu et al., 2005).

Similarly, Neves-Piestun and Bernstein (2005) found that Na+ and Cl– were, only 40

mM in the most rapidly growing tissues, and that the degree of inhibition by salt

stress of either the elongation rate or the total volume expansion rate did not correlate

with the Na+ or Cl– in the tissues of maize growing in 80 mM NaCl. The reduction of

plant growth and dry-matter accumulation under saline conditions has been reported

in several important grain legumes (Tejera et al., 2006). In rice, spikelet and tiller

numbers were more affected by salinity than 1000-seed weight (Zeng et al., 2002).

LXV

Sodium in higher amounts in plant tissues significantly reduced growth,

which was evident from our results where the hybrid genotypes had maximum Na+

concentration in the leaf sap and produced minimum dry matter, characteristics of

salt sensitive genotypes. In contrast, the genotype S-2002 had a lower shoot Na+

concentration and produced maximum shoot fresh weight. Qadir and Schubert (2002)

reported that plant species/genotypes varied not only in their rates at which they

absorbed Na+ but also in the manner by which they translocated Na+ to their shoots.

Under salinity stress the shoot fresh weight production by all genotypes negatively

correlated (r=0.74) with Na+ concentration (Fig. 4.1). The higher concentrations of

Na+ and Cl- in leaves become toxic and lead to salt injury (Serrano et al., 1999; Saqib

et al., 2005). These results were in agreement with Munns et al. (2006) who reported

that the salt tolerance in wheat was associated with low shoot Na+ concentration.

A high K+: Na+ ratio as another attribute of salinity-tolerant S-2002

genotype. Reduction in K+/Na+ ratio in the presence of salinity could be due to the

antagonism of Na+ and K+. Similarly, Hu and Schmidhalter, (1997) attributed the

differences among sugarcane genotypes in K+: Na+ ratio to their ability to restrict

both the uptake of Na+ by root cells from soil and also the movement of Na+ to the

shoots by controlling their influx into the root xylem from root cells. A high K+: Na+

ratio maintained by roots shows the selectivity for K+ over Na+ and the preferential

loading of K+ rather than Na+ into xylem (Carden et al., 2003). Cicek and Cakirlar

(2008) found a considerable decline caused by salt stress treatments in K+: Na+ ratio,

plant height, fresh and dry biomass of the shoot in soybeans cultivars.

Fig. 4.1 Relationship between shoot fresh weight and Na + concentration

LXVI

y = -5.9946x + 202.27R2 = 0.7486

0

50

100

150

200

0 5 10 15 20 25

Sodium concentration (mol m-3)

Shoo

t fre

sh w

eigh

t (g

plan

t-1)

In the present study, the high yielding synthetic genotypes had lowest

shoot Na+ and Cl- concentration and low yielding hybrid genotypes were higher in

Na+ and Cl- concentration in leaf sap. Earlier findings of Ashraf and Khanum (1997)

also showed that a salt tolerant line possessed lower Na+ or Cl- concentrations in

shoot than a salt sensitive line. Similar findings were reported by Ashraf and O’Leary

(1994) in two lines of alfalfa.

The concentrations of Na+ and Cl- were increased significantly under

saline conditions while the concentration of K+: Na+ ratio was decreased

significantly. The trend of effects was also similar in case of synthetic genotypes, but

not to the extent that was found in case of hybrid genotypes. Higher concentrations of

external Na+ and Cl- might have disrupted the osmotic balance of the cells thus

inducing a water deficit effect. The higher concentrations of Na+ and Cl- in leaves

also become toxic and leads to salt injury (Serrano et al., 1999). Salt injury in plant

leaves and stems is due to higher influx of Na+ and Cl- accompanying a reduction in

K+ uptake (Sharma, 1995). The response of individual genotypes also varied

significantly, when compared within a particular treatment. Genetic variability was

also very conspicuous with regard to leaf ionic composition. Varietal comparison

further showed that the genotypes accumulating more Na+ and Cl- in their leaves

showed poor growth performance. Saqib et al. (1999) observed similar findings in

wheat.

A significant negative correlation was found between shoot fresh weight

and chloride concentration (Fig 4.2). Under salinity stress the shoot fresh weight

LXVII

production by all genotypes significantly correlated with Cl- concentration (r=0.56).

The figure showed that the shoot fresh weight of hybrid genotypes decreased

relatively faster with the insrease in Cl- concentration as compared to the synthetic

genotypes. Among the synthetic genotypes, Akbar had the higher Cl- concentration,

while S-2002 had the lower Cl- conentration in its leaf sap. Amor et al. (2004)

concluded that due to highest salinity (200 mM NaCl) Na+ and Cl- were largely

accumulated in shoots and induced mineral nutrition disturbance within the plant

shoot, as their Ca2+, Mg2+, and K+ concentrations significantly declined in the

perennial halophyte C. maritimum. Reductions in the rate of leaf and root growth are

probably due to factors associated with water stress rather than a salt-specific effect

(Munns, 2002). This is supported by the evidence that Na+ and Cl– are below toxic

concentrations in the growing cells. In rice, spikelet and tiller number were more

affected by salinity than 1000-seed weight (Zeng et al., 2002). For example, in wheat

growing in 120 mM NaCl, Na+ in the growing tissues of leaves was at most only 20

mM, and only 10 mM in the rapidly expanding zones, and Cl– only about 50 mM (Hu

et al., 2005). Similarly, Neves-Piestun and Bernstein (2005) found that Na+ and Cl–

were, only 40 mM in the most rapidly growing tissues, and that the degree of

inhibition by salt stress of either the elongation rate or the total volume expansion

rate did not correlate with the Na+ or Cl– in the tissues of maize growing in 80 mM

NaCl. Fricke et al. (2004) found only 38 and 49 mM Na+ in mesophyll and epidermal

cells, respectively, in the growing cells of barley after 24 h of exposure to 100 mM

NaCl. Mansour et al. (2005) worked on two maize cultivars, salt sensitive Trihybrid

321 and salt tolerant Giza 2, were studied, mainly their adaptation to NaCl imposition

at cell and whole plant level. Changes in growth and mineral content of roots and

shoots were measured. NaCl decreased fresh mass, dry mass, relative growth rate

(RGR) of shoots and roots, and leaf area ratio (LAR) in both cultivars. Salt stress

induced Na+ and Cl- accumulation while it decreased K+ and Ca2+ levels in shoots and

roots of both cultivars. The increase in Na+ and the decrease in K+ and Ca2+ were

greater in Giza2 than in Trihybrid 321. Cl- was increased more in Trihybrid 321

compared to Giza2. On the basis of shoot fresh/dry weights and Na+, K+ and Cl-

concentrations and K+: Na+ ratio, the synthetic genotypes were selected for further

LXVIII

studies and the hybrid genotypes were excluded due to their poor performance under

the given treatments.

Fig. 4.2 Relationship between shoot fresh weight and chloride concentration in leaves of maize genotypes

y = -3.1137x + 108.96R2 = 0.5626

0

20

40

60

80

100

120

0 5 10 15 20 25

Chloride concentration (mol m-3)

Sho

ot fr

esh

wei

ght (

g pl

ant-1

)

LXIX

STUDY-2 DIFFERENCES IN SALT TOLERANCE AND POTASSIUM

REQUIREMENT OF MAIZE GENOTYPES A

HYDROPONICS STUDY

4.2 RESULTS & DISCUSSION

4.2.1 Results

4.2.1.1 Shoot length/plant height (cm)

The data regarding the shoot length/plant height are presented in Table

4.11. Shoot length of seven maize genotypes at three levels of salinity including

control presented individually at each salt treatment, showed a decrease in shoot

length with the addition of NaCl. The effect of salinity was more pronounced in all

maize genotypes especially in Akbar showing significant difference or strong impact

on the plant height of all genotypes. Analysis of variance showed that increasing

supply of NaCl had significant effect on plant height (Appendix11). The interactions

were also significant at all salinity levels. The genotype S-2002 produced higher

shoot length than Akbar, which remained low at control. At 70 mol m-3 NaCl stress

Akbar and Ev 5098 had smaller shoot length while at 100 mol m-3 NaCl both

genotypes (Akbar and Ev 5098) followed the same trend. Wide variation was

observed among maize genotypes, with highest level of NaCl salinity stress in the

solution culture and a decrease in shoot length was observed showing same trend

with 1.0 mM K level as compared to 5.5 mM K treatment. However, with the

addition of 8.0 mM K in the solution culture a significant increase was found in most

of the genotypes but the most efficient response was observed in S-2002 as compared

to other genotypes of the same treatment. There was a substantial increase in plant

height with the addition of 5.5 mM K and there was a higher increase when 8.0 mM

K was added. The genotypes Ev 5098 and Ev 1098 varied non-significantly at 100

mol m-3 NaCl concentration with 8.0 mM K supply while S-2002 produced higher

shoot length/plant height at all K s under saline treatments.

Relative performance of different genotypes under 70 and 100 mol m-3

NaCl concentration varied significantly at 1.0, 5.5 and 8.0 mM K level. It also

LXX

showed that with the increased in potassium level in the external medium, plant

height of all genotypes increased and among all genotypes S-2002 performed better.

LXXI

4.2.1.2 Root length (cm)

The data regarding the root length are presented in Table 4.12. Root length

of all maize genotypes decreased with increasing concentration of NaCl in the growth

medium, but the major reduction was observed in the case of Akbar, Agati 2000 and

Ev 5098, these genotypes also varied non-significantly at 70 mol m-3 NaCl. Ev 1098

and Ev 6098 produced the smaller root length at 70 mol m-3 NaCl. While, uder 100

mol m-3 NaCl Ev 1098 produced the least root length, while Agati 2000 and Akbar

varied non-significantly at this salinity+1.0 mM K treatment. The application of 5.5

mol m-3 K nutrition increased the root length in all the genotypes but the maximum

increase was observed in S-2002. Analysis of variance of the data recorded show that

increasing supply of NaCl had significant effect on root length of each genotype

(Appendix 12). There was considerable variation among the genotypes with respect

to root length under saline conditions. Comparison of different genotypes showed

that EV 1098, Ev 6098, Agati 2000, Ev 5098 and Akbar had the lowest root length

among all the genotypes under different salt treatments. S-2002 was the highest under

different salinity stress and at different potassium treatments, whereas the remaining

genotypes were intermediate in root length. The data for root length also showed that

the addition of 5.5 mM K caused the maximum increase in the root length of all the

genotypes as compared to the 8.0 mM K level under all the NaCl treatment. The

differences among the different genotypes were not consistent at all the NaCl and K-

levels. The interaction between salinity and potassium was significant (Appendix 12).

It was observed that most of the reduction was observed at 100 mol m-3 NaCl, while

maximum increase in root length was found with the addition of 5.5 mM K.

Where as, with the addition of 5.5 and 8.0 mM K their relative

performance of all genotypes increased, which are shown in % of respective control

of each genotype. The magnitude of salinity effect increased with the addition of

NaCl stress and the addition of K levels alleviated the effect of salinity. The

alleviation of salinity due to K application varied among the genotypes.

LXXII

LXXIII

4.2.1.3 Shoot fresh weight (g plant-1)

The data regarding shoot fresh weight of genotypes are presented in Table

4.13 Shoot fresh weight of maize genotypes decreased consistently with increasing

concentration of NaCl in growth medium. There was a considerable variation among

the genotypes with respect to shoot fresh weight under saline conditions. Under

control conditions, S-2002 produced the maximum shoot fresh weight while Akbar

produced the minimum. With the increase of NaCl stress (70 mol m-3), the shoot

fresh weight of all genotypes decreased but the maximum decrease was observed in

case of Akbar. The interactions were also significant at all salinity levels. The

comparison of two levels (1 and 5.5 mM K) at 70 and 100 mol m-3 NaCl showed

variation in the shoot fresh weight of genotypes and S-2002 showed a better

performance while Akbar had lowest shoot fresh weight. Comparison of saline

treatment alone to the saline treatments with 5.0 mM potassium level.

On an over all basis (7 genotypes), the shoot fresh weight showed a

significant decrease under saline (70 and 100 mol m-3 NaCl) with the addition of 1.0

mM K. The shoot fresh weight was 24.9, 21.2 and 18.2 g under control, 70 and 100

mol m-3 NaCl respectively at 1.0 mM K level while with 5.5 mM K level with the

same saline treatments the average response was 60.6, 47.8 and 37.9 g, respectively

and 62.9, 49.8 and 40.2 g, respectively with the addition of 8.0 mM K. With the

addition of 5.5 mM K at 70 and 100 mol m-3 NaCl, a significant increase was

observed in all the genotypes but this increase was more prominent at 5.5 mM K

level as compared to the 8.0 mM K-level. S-2002 produced the maximum while

Akbar produced the minimum shoot fresh weight at all salinity and K-level treatment.

Both the treatments had a significant effect on all the genotypes. The magnitude of

salinity was more in case of Akbar, Ev 5098, Ev 1098 and Ev 6098. Salinity

decreased shoot fresh weight, with varietal differences. K alleviated the effect of

salinity but it was subjected to the response of each genotypes.

LXXIV

LXXV

However, the inhibitory effect of 100 mol m-3 NaCl was more significant

due to considerable decrease in the shoot fresh weight of all the genotypes. With the

increased in NaCl stress the relative performance of all the genotypes decreased at 70

and 100 mol m-3 NaCl, while, with the addition of 5.5 and 8.0 mM K their relative

performance increased and S-2002 performed better among all genotypes while

Akbar had the lowest values.

4.2.1.4 Shoot dry weight (g plant-1)

The data regarding shoot dry weight of seven maize genotypes are

presented in Table 4.14. Shoot dry weight of all maize genotypes declined

consistently with the increasing concentration of NaCl in the growth medium. At 70

mol m-3 NaCl stress, genotypes S-2002 produced the maximum shoot dry weight

followed by SL 2002 and Agati 2000, while, the minimum shoot dry weight was

produced by Akbar. With the increase in NaCl stress (100 mol m-3) the shoot dry

weight of maize genotypes decreased but the minimum shoot dry was observed in

Akbar while maximum was produced by S-2002. Analysis of variance (Appendix14)

showed that with increasing concentration of NaCl significantly affected the shoot

dry weight of genotypes. The interactions between genotypes, salinity and potassium

were also significant. Shoot dry weights of all the genotypes increased progressively

with the addition of potassium (1.0 to 8.0 mM) but more significant and prominent

increase was observed with the addition of 5.5 mM K in all the genotypes. The

genotype S-2002 produced the maximum shoot dry weight while Akbar had the

minimum increase in shoot dry weight in response to potassium addition. The shoot

dry weight was increased under 70 and 100 mol m-3 NaCl with the application of 5.5

and 8.0 mM K, however this increase varied within and among the genotypes

according to their responsiveness, efficiency and utilization of the potassium addition

but the reduction due to salinity was less.

On an over all average basis, a considerable variation among the

genotypes with respect to SDW under saline conditions and K application was

observed. At 1.0 mM K-level under control, 70 and 100 mol m-3 NaCl concentration,

the SDW was observed as 4.5, 4.0 and 3.6 g plant-1 respectively, while at 5.5 mM K-

level, the SDW was observed as 9.4, 7.6 and 6.1 g plant-1 respectively at control, 70

LXXVI

and 100 mol m-3 NaCl concentration. With the application of 8.0 mM K, the shoot

dry weight was 9.61, 7.8 and 6.3 g plant-1 at control, 70 & 100 mol m-3 NaCl

concentration. At 1.0 mM K level, under 100 mol m-3 NaCl stress, S-2002 showed

the maximum shoot dry weight and Akbar had the lowest. Similarly, under 70 mM

NaCl stress the same trend was observed. With the increased in the rate of potassium

application, shoot dry weight increased although the increase was less with the

increase in NaCl concentration in the external medium.. On relative basis with

increasing level of salinity i.e. 70 to 100 mol m-3 NaCl, shoot dry weight of all

genotypes decreased however reduction was more in Akbar and less in S-2002. With

the addition of potassium (5.5 and 8.0 mM K) shoot dry weight of all genotypes

increased with respective to control.

4.2.1.5 Root fresh weight (g plant-1)

Root fresh weight of all maize genotypes as affected by salinity levels and

addition of potassium showed a considerable variation among the genotypes (Table

4.15). Comparison of different genotypes showed that Akbar had the lowest root

fresh weight, while S-2002 had higher value at 70 and 100 mol m-3 NaCl stress even

with 1.0 mM K-level followed by SL 2002. Analysis of variance (Appendix15)

showed significant effect on root fresh weight of all the genotypes, with increasing

concentration of NaCl and interaction of genotypes and salinity was also significant,

however, the interaction for genotypes × salinity × potassium was found non-

significant. Addition of potassium to the saline medium showed a significant effect

on all genotypes while it was also very prominent in case of S-2002, as more root

fresh weight was observed, while Akbar remained sensitive to salinity even with the

addition of potassium. With salinity + 1.0 mM K in the rooting medium a substantial

decrease was obvious in mean values of root fresh weight as compared to the

treatment salinity + 5.5 and 8.0 mM K addition, but the effects of K and NaCl varied

among the genotypes. On over all basis (7 genotypes), the root fresh weight was

decreased due to highest saline treatment (100 mol m-3 NaCl) concentration and the

over all root fresh weight production under (control, 70 and 100

LXXVII

mol m-3 NaCl) concentration at 1.0 mM K level was 8.3, 6.8 and 5.6 g plant-1 but

under same saline conditions with 5.5 mM K application, the observed root fresh

LXXVIII

weight was 22.9, 19.4 and 162 g, respectively. Under (control, 70 & 100 mol m-3

NaCl) concentration at 8.0 mM K addition, the root fresh weight was 24.7, 20.9 and

16.2 g plant-1, the genotype S-2002 produced the maximum root fresh weight while

Agati-2000 and Ev-6098 produced the same weight and Akbar produced the

minimum root fresh weight as compared to the same salinity level with 5.5 mM K

and 8.0 mM K addition.

Relative root fresh weight of different genotypes under 70 and 100 mol m-

3 NaCl concentrations showed that S-2002 had ahigher tolerance to salinity followed

by SL 2002, Agati 2000 and Ev 6098, while Akbar had the lowest root fresh weight.

With increasing level of potassium from 1.0 to 5.5 mM under 100 mol m-3 NaCl

concentration, root fresh weight of all genotypes increased. Relatively higher increase

was observed in S-2002 followed by SL 2002 and Agati 2000. The addition of 8.0

mM K did not have a remarkable effect as compared to 5.5 mM K, especially under

100 mol m-3 NaCl concentration.

4.2.1.6 Root dry weight (g plant-1)

With the increase in NaCl concentration root dry weight of all genotypes

decreased progressively (Table 4.16). At 70 mol m-3 NaCl concentration with low

(1.0 mM) K level, genotype S-2002 produced maximum root dry weight, whereas

Akbar had lower root dry weight. At 100 mol m-3 NaCl stress, the same trend was

observed for all the genotypes. The genotype S-2002 remained higher, while, Akbar

remained low in producing root dry weight. Analysis of variance (Appendix16)

showed a significant effect on root dry weight of all genotypes with increasing

concentration of NaCl. All the possible interactions of genotypes with salinity and

potassium were significant except for genotypes × salinity × potassium where it was

found non-significant. The root dry weight of maize on an overall average basis with

1.0 mM K under control, 70 and 100 mol m-3 NaCl was 1.3, 0.9 and 0.8 g plant-1,

respectively while at the same NaCl stress with the addition of 5.5 mM potassium

the average root weight was 3.4, 2.8 and 2.3 g plant-1 respectively. Maize

genotype S-2002 produced highest root dry weight (1.9 g plant-1) in control followed

by Agati 2000 (1.3 g plant-1 SL 2002 (1.2 g plant-1) and Ev 5098 (1.2 g plant-1), and

the former three genotypes varied non-significantly. By the addition of highest

LXXIX

potassium level (8.0 mM K) the overall average root dry weight was 3.6, 2.9 and 2.3

g plant-1 under control, 70 and 100 mol m-3 NaCl concentration, respectively. At 100

mol m-3 NaCl concentration with 1.0 mM K level, the maximum root dry weight was

observed in case of S-2002 genotype as compared with 5.5 and 8.0 mM K level. The

root dry weight of all maize genotypes dropped significantly under 70 mol m-3 NaCl

concentration and S-2002 produced significantly higher root dry weight as compared

to other genotypes. Among the synthetic/ recommended genotypes, Akbar remained

sensitive and produced the lowest root dry weight. Relative performance of different

genotypes under 70 and 100 mol m-3 NaCl concentration showed that S-2002 had the

highest tolerance followed by SL 2002 while Ev 1098 was the most sensitive in terms

of root dry weight. With increasing level of potassium in the growth medium from

5.5 to 8.0 mM under 100 mol m-3 NaCl concentration, root dry weight of all

genotypes increased as compared to 1.0 mM K. A higher increase was observed in S-

2002 followed by SL 2002 and Agati 2000 by the addition of 5.5 and 8.0 mM

potassium.

4.2.1.7 Sodium concentration in leaf sap of maize genotypes

The data regarding sodium concentration in the leaf sap of seven maize

genotypes grown under different levels of salinity and potassium are presented in

Table 4.17. Data revealed that increasing concentration of NaCl had a significant

effect on the leaf sodium concentration under low (1.0 mM) K level. Different

genotypes had different concentrations of sodium at varying levels of NaCl. The

highest concentration of sodium was observed in Akbar followed by Ev 5098, Ev

1098 and Ev 6098, whereas the lowest sodium concentration was observed in S-2002

at 70 mol m-3 NaCl concentration. At higher level of NaCl (100 mol m-3)

concentration the same trend was observed in all the genotypes. Analysis of variance

(Appendix 17) showed that increasing NaCl concentration had a significant effect on

leaf sodium concentration. All interactions for genotypes × salinity × potassium were

significant.

Maize genotypes significantly differed regarding Na+ concentration in their youngest fully expanded leaf among the treatments. In the control treatment, Na+ concentration was low, however increased significantly under 70 mol m-3 NaCl concentration while addition of potassium decreased Na+ concentration in the leaf sap of maize

LXXX

genotypes, for instance, in S-2002, Na+ concentration was 72.0, 62.1 and 56.3 mol m-

3 at 70 mol m-3 NaCl with 1.0, 5.5 and 8.0 mM K concentration respectively and remained 81.1, 56.8 and 50.1 mol m-3 respectively under three K-levels (1.0, 5.5 & 8.0) at 100 mol m-3 NaCl concentration. The highest Na+ concentration was observed in genotype Akbar while the remaining genotypes had intermediate Na+ concentrations in their leaves. The addition of 8.0 mM K treatment decreased the Na+ concentration in leaf sap of maize genotypes, especially in S-2002.

4.2.1.8 Potassium concentration in leaf sap of maize

The increasing concentration of NaCl in the growth medium had a significant effect on K+ concentrations in leaf sap of maize genotypes (Table 4.18). Salinity at moderate and higher level of external supply of NaCl reduced the K+ concentration in the leaves of all maize genotypes. The genotype S-2002 had highest K+ concentration, while, Ev 5098 and Akbar had the lower concentration of K+ in the leaf sap. On the other hand, with the increase in external level of NaCl (100 mol m-3) concentration in the external solution with the 1.0 mM K level, the same trend was observed in all the genotypes. Analysis of variance (Appendix18) showed that increasing NaCl concentration had a significant effect in decreasing leaf K+ concentration of all maize genotypes.

Genotypes also varied significantly under different treatments with regards to K+ concentration in leaf sap. Genotypes grown with external supply of 8.0 mM K had more K+ in their leaves as compared to those grown with 1.0 mM K and 5.5 mM K under the given salinity levels. S-2002 had the maximum concentration of K+ while Akbar had the lower value whereas the remaining genotypes had the intermediate K+ concentration in their respective leaves. Comparison of leaf K+ concentration of each genotype versus different salinity and K-levels showed that the K+ concentration reduced considerably at 100 mol m-3 NaCl and increased significantly with the addition of 5.5 and 8.0 mM K in all the genotypes. Again, K+ concentration was lowest in Akbar and highest in genotype S-2002.

4.2.1.9 K+: Na+ ratio in leaf sap of maize genotypes

Leaf K+: Na+ ratio decreased consistently with the increase in NaCl level.

More reduction was observed at 100 mol m-3 NaCl as compared to 70 mol m-3 NaCl

concentration. The genotype S-2002 maintained better K+: Na+ ratio at lower (70 mol

m-3) and higher (100 mol m-3) NaCl concentrations, compared to other genotypes.

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With the addition of 5.5 and 8.0 mM K K+: Na+ compared at all levels of NaCl stress.

Akbar had lower K+: Na+ ratio and genotype Ev 1098 and Ev 6098 varied non-

significantly.

The genetic variation under different treatments was also significant with

reference to leaf K+: Na+ ratios. S-2002 maintained better K+: Na+ in salt treatment

with the supply of K except the control, whereas EV-6098, Ev 1098, Ev 5098 and

Agati 2000 had non- significant K+: Na+ ratios at 70 and 100 mol m-3 NaCl

concentration under both K levels (5.5 and 8.0 mM) as compared to the control. The

minimum K+: Na+ ratios were observed in Akbar in all the saline treatments at 1.0,

5.5 and 8.0 mM K treatments.

4.2.1.10 Chloride concentration in leaf sap of maize

Increasing supply of NaCl in the external medium increased the Cl-

concentration in leaves of all the maize genotypes significantly as compared to

control (Table 4.20).

Data showed that the highest Cl- concentration was observed in case of

Akbar and the lowest value was observed in S-2002 under the treatment (70 mol m-3

NaCl + 8.0 mM K) while, all other genotypes had intermediate value. At 100 mol m-3

NaCl + 8.0 mM K, SL 2002, Ev 6098 and Agati 2000 varied significantly while S-

2002 had the lowest Cl- concentration. Akbar accumulated the maximum

concentration of Cl- at 70 mol m-3 NaCl + 5.5 mM K. Saline treatments caused a

significant increase in Cl- concentration in leaf sap of different maize genotypes as

compared to the control, but this increase was not significant. Potassium application

had no significant effects on Cl- accumulation in all genotypes. In case of highest

saline + K treatment, Akbar again accumulated the highest chloride concentration

while S-2002 had the lowest value. Although the application of a higher level of

potassium (8.0 mM K) decreased the accumulation of Cl- in the leaves of maize, the

application of this highest level of K had no significant effect on almost all the

genotypes.

4.2.2 Discussion

The primary objective of the hydroponic experiment was to observe the

pattern of gain in fresh and dry weights of seven maize genotypes under different

LXXXII

saline conditions with three K levels supply. In addition, the relationships between

growth and ion accumulation in these genotypes were also determined. It is well

documented that general plant health can be guessed on the basis of a number of

growth parameters. Although salinity can induce a rapid reduction in root growth

(Noaman and El-Haddad, 2000; Munns et al., 2006), shoot growth decreased

proportionally more than root growth, causing an increase in the root/shoot ratio. In

addition, salinity significantly decreased tiller number and their appearance (Mass et

al., 1983; Mass and Grattan 1999). Salinity significantly reduces the total dry matter,

yield, and the degree of reduction in total dry matter depending on genotypes and salt

concentrations. Salt stress also leads to deterioration of soil structure and hinders

desirable air-water balance essential for biological processes occurring in plant roots.

As a result of the detrimental effects of salinization, crop yields decrease, while

arable land is lost irreversibly (Supper, 2003). Salt stress causes various effects on

plant physiology such as increased respiration rate, ion toxicity, changes in plant

growth, mineral distribution, and membrane instability resulting from calcium

displacement by sodium (Marschner, 1986), membrane permeability (Gupta et al.,

2002), and decreased photosynthetic rate (Hasegawa et al., 2000; Munns, 2002;

Ashraf and Shahbaz, 2003; Kao et al., 2003; Sayed, 2003).

Salt stress affects plant physiology at whole plant as well as cellular levels

through osmotic and ionic stress (Hasegawa et al., 2000; Muranaka et al., 2002a, b,

Ranjbarfordoei et al., 2002; Murphy et al., 2003). Despite causing osmotic and ionic

stress, salinity causes ionic imbalances that may impair the selectivity of root

membranes and induce potassium deficiency (Gadallah, 2000). The accumulation of

high amounts of toxic salts in the leaf apoplasm leads to dehydration and turgor loss,

and eventually death of leaf cells and tissues (Marschner, 1995). Ion cytotoxicity

caused by the replacement of K+ with Na+ in biochemical reactions and

conformational changes and loss of function of proteins as Na+ and Cl- ions

penetrated the hydration shells and interfere with non-covalent interaction between

their amino acids. Dionisiosese and Tobita (2000) reported that increased

concentration of Na+ under saline conditions suppressed the leaf gas exchange and PS

II photochemical activity and consequently hampered the growth and development of

plants.

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An ionic imbalance occurs in the cells due to excessive accumulation of

Na+ and Cl– and reduced uptake of other mineral nutrients, such as K+, Ca2+, and

Mn2+ (Karimi et al., 2005). Excess Na+ and Cl- concentrations inhibit the uptake of K+

and leads to the appearance of symptoms like those in K+ deficiency. The deficiency

of K+ initially leads to chlorosis and then necrosis (Gopal and Dube, 2003). The role

of K+ is necessary for osmoregulation and protein synthesis, maintaining cell turgor

and stimulating photosynthesis (Freitas et al., 2001; Ashraf, 2004). Both K+ and Ca2+

are required to maintain the integrity and functioning of cell membranes (Wenxue et

al., 2003). Maintenance of adequate K+ in plant tissue under salt stress seems to be

dependent upon selective K+ uptake and selective cellular K+ and Na+

compartmentation and distribution in the shoots (Munns et al., 2000; Carden et al.,

2003).

Potassium is essential for protein synthesis, glycolytic enzymes and

photosynthesis, an osmoticum mediating cell expansion and turgor driven movements

and competitor of Na+ under salt stress (Hu and Schmidhalter, 2005). Accumulation

of Na+ and impairment of K+ nutrition is a major characteristic of salt stressed plants.

Therefore K+/Na+ ratio in plants is considered a useful parameter to assess salt

tolerance (Cakmak, 2005b). Numerous studies have shown that application of K

fertilizer mitigates the adverse effects of salinity through its role in stomatal

regulation, osmoregulation, energy status, charge balance, protein synthesis and

homeostasis (Marschner, 1995; Sanjakkara et al., 2001). Improvement of potassium

nutritional status of plants can greatly lower the ROS production (Cakmak, 2005b).

Kaya et al. (2002b) discussed the response of spinach and lettuce grown at higher

salinity to supplementary K and P. High salinity reduced germination percentage,

root elongation and increased membrane permeability compared to control.

Supplementary K and P produced fresh weight, chlorophyll content, water usage and

membrane permeability values similar to or slightly lower than control. Added K and

P reduced leaf and root Na+ levels that were much higher than control values but

significantly lower than in saline treatment in most cases.

The application of KNO3 improved the K nutrition and growth of

Legineria spp. under saline conditions. While working with sunflower, Akram et al.

(2007) reported that foliar spray of different K inorganic salts considerably improved

LXXXIV

the ion homeostatic conditions. They also found that improved K nutrition due to

foliar spray of K salts protected the cell membrane as estimated by extent of ion

leakage. Improved tissue K status of sunflower plants regulated the plant

photosynthetic activity through stomatal movement. They concluded that all K salts

improve the growth of sunflower plants however, the effectiveness of K salts in

improving growth depends upon a number of factors including concentration of salt,

plant developmental stage at which applied, and accompanying anion in specific salt.

Thus, foliar application of inorganic salts can also be beneficial to improve crop salt

tolerance. Of the mineral nutrients, K plays a particular role in contributing to the

survival of crop plants under environmental stress conditions. Potassium is essential

for many physiological processes (Mengel and Kirkby, 2001).

Among different parameters shoot dry weight (SDW) is considered to be

the most sensitive plant response parameter to nutrient deficiency and is given as

pivotal phase in screening experiments. It is therefore, generally used as a selection

criterion for evaluating genotypes for salinity and nutrient efficiency at seedling

stage. Our results revealed root length, shoot length, SFW, SDW, RFW, and RDW

are presented in (Table: 4.11 to 4.16). There was significant effect of K-levels,

salinity and their interaction regarding SDW especially of these genotypes on which

basis the selection was done. These significant interactions clearly indicated that

variability exist among the maize genotypes for SFW and SDW production when

grown with different salinity and K levels. These results are in accordance with Gill

et al. (1997) who also reported significant differences among the genotypes for dry

matter accumulation at different K levels. These results are in agreement with the

findings of Ashraf and Tufail (1995) who observed that a salt tolerant sunflower line

produced significantly greater fresh and dry biomass of both shoots and roots than the

sensitive accessions at different salinity levels of the growth medium e.g. corn is

tolerant at germination but is more sensitive at seedling than adult stage (Mass et al.,

1983). Undoubtedly, plant breeders have made significant achievements in few years

improving salinity tolerance in a number of potential crops using artificial selection.

However, most of the selection procedures used so far were based merely on

differences in agronomic characters and these findings were in agreement with

Ashraf (2004). Although the screening and selection under the appropriate field

LXXXV

conditions might be the ideal, one, there are many compelling reasons that have led

investigators to choose artificial systems. These include year to year variability,

spatial variability of soil within the field; the need to keep cost down and the need to

select a character difficult to study in field e.g. root morphology, rate of root growth,

shoot length etc (Vose, 1963).

The concentration of Na+ was increased significantly under saline

conditions especially at 100 mol m-3 NaCl stress while the concentration of K+ and

K+/Na+ ratio was decreased. Higher concentrations of external Na+ might have

disrupted the osmotic balance of the cells thus inducing the water deficit. The higher

concentrations of Na+ in leaves also become toxic and lead to salt injury (Serrano et

al., 1999). Salt injury in plant leaves and stem is due to higher influx of Na+

accompanying a reduction in K+ uptake (Sharma, 1995) that are in accordance with

our results where more growth reductions under saline conditions than at 8.0 mM K-

level. The response of individual genotypes also varied significantly when compared

within a particular treatment. Genetic variability was also very conspicuous with

regard to leaf ionic composition under different treatments. Varietal comparison

further showed that the genotypes accumulating more Na+ in their leaves showed

poor growth performance as compared with the application of 5.5 and 8.0 mM K.

The maximum shoot fresh weight under 100 mol m-3 NaCl stress was found in S-

2002 followed by SL-2002 while minimum shoot fresh weight was observed in

Akbar. Similarly the genotypes varied significantly under different treatments.

Similarly, genotypic variation in Na+ exclusion may result in K+/Na+ ratio

discrimination (Cuin et al., 2003). The net ionic concentration in plants is the result

of ion uptake through selective and non-selective channels in the plant cell membrane

and subsequent loading in the xylem. In plants, a large number of K+ selective

membrane channels and non-selective cation channels permeable to both K+ and Na+

have been identified in different plant species (Amtmann and Sanders, 1999). The

results in Table 4.17, 4.18 and 4.19 demonstrate a significant genotypic difference

in K+ (Fig 4.5) and Na+ contents in plant and in the discrimination of K+/Na+ ratio,

which is in consistance with earlier reports in rice (Asch et al., 2000; Zhu et al.,

2001).

LXXXVI

Similarly, Cramer et al. (1994a) also found that for maize genotypes, the

concentration of K+ and its ratio over Na+ were also more related with salt tolerance

in two hybrids. Because of higher selectivity of K+ in most tolerant genotypes, higher

K+ over Na+ contents in leaves appear to protect the plant from the effect of toxic ions

(Rangel, 1992). In fact, it is possible that a high K+/Na+ ratio is more important for

many species than simply maintaining a low concentration of Na+ (Cuin et al., 2003;

Mark and Romola, 2003). Thus the ratio of K+/Na+ is an important factor to be

considered as a selection criterion. Therefore, the selection of S-2002 as having

highest K+/Na+ ratio is in accordance with the above-mentioned findings. Reductions

in the rate of leaf and root growth are probably due to factors associated with water

stress rather than a salt-specific effect (Munns, 2002). This is supported by the

evidence that Na+ and Cl– are below toxic concentrations in the growing cells

themselves. In rice, spikelet and tiller number were more affected by salinity than

1000-seed weight (Zeng et al., 2002).

` The concentration of Na+ and Cl– increased linearly with increased salinity,

but the extent of increase was greater in case of genotype Akbar. Contrarily, the

concentrations of nutrients (K+ and K+/Na+) studied here were substantially higher in

genotype S-2002 as compared to Akbar (Table 4.18 and 4.19). This indicated that

tolerant genotype managed to restrict the entry of Na+ and Cl– in to the roots and

maintain higher contents of K+ and K+/Na+ in the leaf sap. The enhanced contents of

these nutrients appeared to buffer the toxicity of Na+ and Cl–, and enabled the tolerant

genotype to exhibit better growth in terms of plant height and shoot fresh weight. For

example; in wheat growing in 120 mM NaCl, Na+ in the growing tissues of leaves

was at most only 20 mM, and only 10 mM in the rapidly expanding zones, and Cl–

only about 50 mM (Hu et al., 2005). Similarly, Neves-Piestun and Bernstein (2005)

found that Na+ and Cl– were only 40 mM in the most rapidly growing tissues, and that

the degree of inhibition by salt stress of either the elongation rate or the total volume

expansion rate did not correlate with the Na+ or Cl– in the tissues of maize growing in

80 mM NaCl. A significant negative correlation (r= 0.88) was found between leaf

sodium concentration and shoot fresh weight (Fig: 4.3). A significant negative

correlation (r= 0.86) between the leaf chloride and shoot fresh weight was observed

LXXXVII

(Fig. 4.4). Excess Na+ and Cl- inhibits the uptake of K+ and leads to the appearance of

symptoms like those in K+ deficiency.

The concentration at which Cl– becomes toxic is even less defined. Roots

must exclude most of the Na+ and Cl– dissolved in the soil solution, or the salt in the

shoot will gradually build up with time to toxic levels. Plants transpire about 50 times

more water than they retain in their leaves (Munns, 2005). In conclusion, root

growth was relatively more sensitive to NaCl stress than shoot growth, as genotype

S-2002 with greater root mass exhibited greater NaCl tolerance. The Na+

concentration in the leaves was more inhibitory to plant growth than Cl–. The

relatively lower concentration of Na+ and Cl–, and the higher concentration of K+ (Fig

4.5) and K+/Na+ in leaves of genotype S-2002 reflected a better ability to withstand

salinity. Such pattern of growth and of distribution of nutrients is a plausible strategy

of salinity tolerance at this stage of maize growth, and it may be accorded due

consideration while selecting maize genotypes for growing in naturally saline field

conditions. On the other hand Akbar showed a high sensitivity to saline conditions.

Fig 4.3 Relationship between shoot fresh weight and sodium concentration in leaves of 7 maize genotypes harvested at 4 weeks age.

y = 258.35e-0.0444x

R2 = 0.8884

0

20

40

60

80

100

120

140

160

0 10 20 30 40 50 Sodium concentration (mol m-3)

Shoo

t fre

sh w

eigh

t (g

plan

t-1)

Fig 4.4 Relationship between shoot fresh weight and chloride concentration in leaves of 7 maize genotypes harvested at 4 weeks age.

LXXXVIII

y = 259.39e-0.0619x

R2 = 0.8632

0

20

40

60

80

100

120

140

0 10 20 30 40 50

Chloride concentration (mol m-3)

Shoo

t fre

sh w

eigh

t (g

plan

t-1)

LXXXIX

Fig: 4.5 Relationship between shoot fresh weight and potassium concentration in leaves of 7 maize genotypes harvested at 4 weeks age.

y = 11.484e0.0074x

R2 = 0.8484

05

1015202530354045

0 50 100 150 200

Potassium concentration (mol m-3).

shoo

t fre

sh w

eigh

t (g

plan

t-1

)

XC

STUDY-3 GROWTH RESPONSE OF SALT TOLERANT AND SALT SENSITIVE MAIZE GENOTYPES TO SALINITY AND POTASSIUM SUPPLY

4.3 RESULTS AND DISCUSSION

4.3.1 Results

4.3.1.1 Plant height

Plant heights of two maize genotypes with and without the application of potassium at 200 mg kg-1 soil are given in Table 4.21. Soil salinity (10 dS m-1) significantly reduced plant height in both genotypes but the reduction was significantly higher in Akbar compared to S-2002 (Appendix 21). Plant height was higher in case of S-2002 in absolute and relative terms. The application of potassium significantly improved plant height in S-2002 while in Akbar there was non significant differences. The plant height of salt tolerant and salt sensitive maize genotypes differed significantly between saline and saline + 200 mg kg-1 soil K level. On relative basis, plant height for S-2002 and Akbar was 93% and 69% of their controls respectively at 10 dS m-1 + 100 mg kg-1 soil K-level, while at 200 mg kg-1 soil potassium level, S-2002 had 91% while Akbar had 68% of their respective controls.

4.3.1.2 Shoot fresh and dry weights

The shoot fresh and dry weight of salt tolerant and salt sensitive genotypes as observed under different treatments are given in Table 4.21. Addition of salts (ECe

10 dS m-1) significantly reduced the shoot fresh weight of both genotypes but this reduction was significantly greater in the salt-sensitive than in the salt-tolerant genotype on an absolute or relative basis. The addition of 200 mg kg-1 soil K significantly improved the shoot fresh weight in both genotypes, although their values still lower in the in the control.

A similar trend was observed in for shoot dry weight and S-2002 produced more weight in control and also with the addition of K and remained significantly better than Akbar. There was a significant difference under both K levels in salt tolerant genotype in non-saline (control) and saline conditions. Although salinity reduced shoot dry matter but the application of potassium decreased the effect of salinity on both genotypes. The comparison on a relative basis indicated that shoot fresh weight of S-2002 was 76% and 80% at 100 and 200 mg kg-1 soil K while, Akbar had 70% and 75% respectively. Comparison of shoot dry weight on a relative

XCI

basis showed that S-2002 and Akbar had 75% and 74% of control respectively at 100 mg kg-1 soil K level while at 200 mg kg-1 soil potassium level, S-2002 had 79% while Akbar had 65% of their respective controls.

4.3.1.3 Grain yield (g plant-1)

Data regarding grain yield of two maize genotypes are presented in Table

4.21. Grain yield per plant of two maize genotypes decreased significantly with the

increase in salt concentration (ECe 10 dS m-1) to the soil. The decrease in grain yield

of salt tolerant genotype was less than as compared to salt sensitive genotype Akbar.

The addition of K at 200 mg kg-1 to the growth medium increased grain yield

significantly in maize genotype S-2002 than Akbar. Both the genotypes varied

significantly. There was a significant difference under both K-level (100 and 200 mg

kg-1 soil) in salt tolerant genotype in non-saline (control) and saline condition,

although the salinity reduced the grain yield but with the application of potassium,

salt tolerant genotype produced more as compared to the salt sensitive one. The

comparison on relative basis indicated that grain yield of S-2002 was 90% and 84%

of control at 100 and 200 mg kg-1 soil, K while Akbar had 72% and 71%

respectively, for the two K levels.

4.3.1.4 1000 seed weight

Table 4.22 show 1000 seed weight of both genotypes at two K levels (100

and 200 mg kg-1 soil) in saline and non-saline treatments. Addition of salts (ECe 10

dS m-1) decreased the 1000 seed weight of both genotypes, but the application of K

significantly increased the 1000 seed weight in S-2002 while the increase was non-

significant in Akbar (Appendix23). The performance of S-2002 was more prominent

and significant than that of salt sensitive genotype Akbar with and without the

application of K, as well as with and with out the imposition of NaCl stress in soil

cultured pots.

Table 4.21. Growth response of two maize genotypes grown under control (1.25 dS m-1) and 10 dS m-1 at two levels of potassium

Plant height

XCII

Genotypes ECe dS m-1 K1 (100 mg kg-1 soil) K2 (200 mg kg-1 soil) S-2002 1.25

10.0 85.8±1.76 80.0±1.53

98.3±1.45 89.67±2.33

Akbar 1.25 10.0

69.0±1.73 48.0±3.21

74.3±0.88 50.7±1.45

Shoot fresh weight (g plant-1)

S-2002 1.25 10.0

163.3±6.55 125.3±4.71

268.8±9.59 216.4±3.93

Akbar 1.25 10.0

100.9±0.71 70.7±5.21

137.6±1.33 104.4±2.43

Shoot dry weight (g plant-1) S-2002 1.25

10.0 32.5±1.71 24.6±0.35

52.4±0.10 41.6±1.91

Akbar 1.25 10.0

17.3±0.35 12.9±0.76

22.4±1.10 14.5±0.47

Grain yield (g plant-1) S-2002 1.25

10.0 415±2.10 376±1.70

518±2.37 436±1.61

Akbar 1.25 10.0

270±1.35 195±1.15

295±1.75 210±2.10

Each value is an average of 3 replications ± S.E. .

There was a significant increase of plant growth by application of K (200

mg kg-1 soil) in salt tolerant genotype in non-saline (control) while at (ECe 10 dS m-1),

although the salinity decreased 1000 seed weight but with the application of

potassium, salt tolerant genotype produced significantly more as compared to the salt

sensitive one. The comparison on relative basis indicated that 1000 seed weight of S-

2002 and Akbar was 79% and 72% at 100 mg kg-1 soil K-level while, both genotypes

had 78% and 71% respectively, at 200 mg kg-1 K level with respect to control.

4.3.2 Ionic concentrations in leaf sap

4.3.2.1 Na+ concentration in leaf sap

Maize genotype Akbar had significantly higher Na+ concentration in its

leaf sap than S-2002 (Table 4.22).s The application of potassium decreased the Na+

concentration in leaf sap of both genotypes, however the decrease was more in S-

2002.

The application of salinity resulted a significant increase in sodium

concentrations in both genotypes as compared to control but higher the sodium

XCIII

concentration in Akbar than S-2002. The application of potassium alleviated the

sodium concentration in the salt tolerant genotype compared to salt sensitive one.

4.3.2.2 K+ concentration in leaf sap

The data regarding K+ concentration in leaf sap of two maize genotypes

are presented in Table 4.22. The concentration of K+ in the expressed leaf sap of the

selected maize genotypes decreased significantly with the increase in soil salinity

(ECe 10 dS m-1). The decrease in K+ concentration in the leaf sap of salt tolerant

maize genotype (S-2002) was less than Akbar. The addition of K at 200 mg kg-1 of

soil to the growth medium increased K concentration in leaves of both genotypes but

resulted in significantly higher K+ concentration in genotype S-2002 than in Akbar.

4.3.2.3 K+: Na+ ratio in leaf sap

Addition of salts decreased K+: Na+ ratio significantly in the leaf sap of

both maize genotypes (Table 4.22). The maximum K+: Na+ ratio was observed in the

leaf sap of maize genotype S-2002 and minimum was observed in Akbar at 10 dS m-1

NaCl. With the addition of 200 mg kg-1 K to soil, K+: Na+ ratio was improved in both

genotypes but more improvement was observed in S-2002 as compared to Akbar.

4.3.2.4 Chloride concentration in leaf sap

S-2002 had lower concentration of Cl- in leaf sap while Akbar had higher

concentration with a significant difference between the two genotypes. Salinity

significantly increased the Cl- concentration in both genotypes as compared to

control. The addition of 200 mg kg-1 K to the soil did not affect Cl- concentration in

both genotypes. Both the genotypes varied significantly. Salinity caused a significant

increase in both the genotypes while the application of potassium had non-significant

effect on Cl- concentration in leaf sap (Appendix 27).

Table 4.22. 1000 Seed weight (g) and leaf ionic concentration of two maize

genotypes grown under control (1.25 dS m-1) and 10 dS m-1 at two levels of potassium

XCIV

1000 Seed weight (g) Genotypes ECe dS m-1 K1 (100 mg kg-1 soil) K2 (200 mg kg-1 soil) S-2002 1.25

10.0 278±3.93 221±2.08

324±3.76 253±2.91

Akbar 1.25 10.0

214±2.33 154±3.21

231±3.76 164±4.67

Na+ concentration (mol m-3) in leaf sap of maize S-2002 1.25

10.0 30.6±1.76 63.5±2.51

19.8±2.34 45.3±3.64

Akbar 1.25 10.0

42.3±3.21 95.8±3.43

37.2±2.15 83.8±1.94

K+ concentration (mol m-3) in leaf sap of maize S-2002 1.25

10.0 81.5±1.93 76.4±1.12

115.3±3.15 97.6±2.54

Akbar 1.25 10.0

48.1±1.64 27.8±1.52

61.7±3.51 39.4±2.63

K+: Na+ ratio in leaf sap of maize S-2002 1.25

10.0 2.7±0.17 1.2±0.46

5.8±0.27 2.2±0.26

Akbar 1.25 10.0

1.1±0.25 0.3±0.11

1.7±0.21 0.5±0.14

Cl- concentration (mol m-3) in leaf sap of maize S-2002 1.25

10.0 26.7±1.23 48.4±1.71

24.9±0.85 45.5±0.78

Akbar 1.25 10.0

45.7±0.61 89.4±2.31

44.9±0.82 88.0±1.35

Each value is an average of 3 replications ± S.E.

4.3.3 Gas exchange parameters and water potential

4.3.3.1 Stomatal conductance

Salinity (ECe 10 dS m-1) caused a significant reduction in stomatal

conductance of both the genotypes (S-2002 and Akbar) (Table 4.23). However, the

reduction in stomatal conductance was less pronounced in genotype S-2002. The

addition of 200 mg kg-1 K to soil increased stomatal conductance in both genotypes

and the improvement was more in Akbar as compared to S-2002.

The comparison on relative basis indicated that stomatal conductance in

genotype S-2002 was 95% at 100 mg kg-1 soil K and 94% at 200 mg kg-1 soil K while

Akbar had 89% and 95% in treatment 1.25 dS m-1 at 100 mg kg-1 soil, respectively.

XCV

Maximum value for stomatal conductance was observed in genotype S-2002 while

Akbar had the minimum value for this parameter.

4.3.3.2 Transpiration rate

Salinity (ECe 10 dS m-1) caused a reduction in transpiration rates of maize

genotypes (Table 4.23). Salt tolerant maize genotype S-2002 maintained its

transpiration rate while salt sensitive genotype varied significantly in its transpiration

rate at both potash levels.. The application of 200 mg kg-1 K to soil increased

transpiration rate non-significantly in Akbar and its value remained lower as

compared to the S-2002 that attained higher transpiration rates (Appendix 29). The

transpiration rate of salt tolerant and salt sensitive maize genotypes differed

significantly under saline and saline + 200 mg kg-1 soil K level. On a relative basis,

S-2002 and Akbar had 100% and 85% of control respectively at saline + 100 mg kg-1

soil K level while, at saline + 200 mg kg-1 soil potassium level, S-2002 had 94%

while Akbar had 95% of respective control.

4.3.3.3 Water potential

Water potential of two maize genotypes showed that the salt treatment had

a significant inhibitory effect on the salt sensitive maize genotype Akbar (Table4.23).

Even the addition of 200 mg kg-1 soil K level had no positive effect on water

potential of the sensitive genotype. Leaf water potential of both genotypes decreased

due to salinity but a consistent decrease was observed in Akbar rather than S-2002.

The salt tolerant maize genotype maintained its leaf water potential even at high

salinity (ECe 10 dS m-1).

Water potential of salt tolerant and salt sensitive maize genotypes differed

significantly under saline and saline + 200 mg kg-1 soil K level. On a relative basis,

S-2002 and Akbar had 91% and 55% at 1.25 dS m-1 respectively at 100 mg kg-1 soil

K-level and while, at 200 mg kg-1 soil potassium level, S-2002 had 83% while Akbar

had 60% of their respective controls.

XCVI

Table 4.23. Stomatal conductance (m mol m-2.sec-1), transpiration rate (m mol m-2.sec-1) and water potential (MPa) of two maize genotypes grown under control (1.25 dS m-1) (1.25 dS m-1) and 10 dS m-1 at two levels of potassium

Stomatal conductance (m mol m-2.sec-1)

Genotypes ECe dS m-1 K1 (100 mg kg-1 soil) K2 (200 mg kg-1 soil) S-2002 1.25

10.0 198.2±1.05 187.8±2.10

204.8±2.64 193.7±0.84

Akbar 1.25 10.0

151.5±1.10 134.8±1.84

166.4±1.80 158.7±1.68

Transpiration rate (m mol m-2. sec-1) S-2002 1.25

10.0 3.2±0.04 3.2±0.05

3.6±0.05 3.4±0.04

Akbar 1.25 10.0

2.0±0.03 1.7±0.04

2.2±0.02 2.1±0.05

Water Potential (MPa) S-2002 1.25

10.0 -0.31±0.01 -0.34±0.01

-0.25±0.01 -0.30±0.01

Akbar 1.25 10.0

-0.41±0.02 -0.74±0.03

-0.36±0.02 -0.60±0.02

Each value is an average of 3 replications ± S.E.

4.3.4 Response of maize genotypes to salinity stress and k supply

(biochemical attributes) 4.3.4.1 Seed oil contents

Data regarding seed oil contents are given in Table 4.24. Saline growth

medium caused a significant reduction in the oil contents of two selected genotypes

of maize. Both genotypes differed significantly in their response to salinity (ECe 10

dS m-1) level and in the production of oil contents. Addition of 200 mg kg-1 K to soil

significantly increased the oil contents in the salt tolerant maize genotype; while,

Akbar responded non-significantly (Appendix 31).

Seed oil contents (%) of salt tolerant and salt sensitive maize genotypes

differed significantly under saline and saline + 200 mg kg-1 soil K level. On a relative

basis, S-2002 and Akbar had 88% and 86% of control respectively at 100 mg kg-1 soil

K level. At 200 mg kg-1 soil potassium level, S-2002 had 85% while Akbar had 79%

of respective control.

XCVII

4.3.4.2 Crude fiber

Addition of salts (10 dS m-1 NaCl) in the rooting medium had a significant

decreased crude fiber of both the genotypes (Table 4.24). Both genotypes differed

significantly in crude fiber content production under saline conditions (Appendix 32).

The salt tolerant genotype S-2002 differed non-significantly at both K levels (100 and

200 mg kg-1 soil K) + saline treatment, while it differed significantly with the salt

sensitive genotype. Salinity caused a decrease in crude fiber contents in both

genotypes.

On a relative basis, S-2002 and Akbar had 88% and 89% of the respective

controls, respectively at 100 mg kg-1 soil K-level while at 200 mg kg-1 soil potassium

level, S-2002 had 76% while Akbar had 91% of respective control. Both genotypes

also differed significantly under non-saline treatment at both K levels. Genotype S-

2002 had a higher crude fiber contents on absolute basis than Akbar.

4.3.4.3 Protein contents

Salinity (10 dS m-1 NaCl) in the growth medium significantly reduced

protein contents in maize seed of the two genotypes (Table 4.24). The salt tolerant

genotype S-2002 had maximum protein contents at 10 dS m-1 NaCl (Apendix 33).

Whereas, the opposite was true for the salt sensitive genotype Akbar that had a lower

amount of protein contents in its seeds. The addition of K (200 mg kg-1) in soil

increased the protein contents slightly in S-2002 genotype and Akbar had non-

significant difference.

There was a significant difference between both K levels in salt tolerant

genotype in non-saline (control) while at soil salinity (ECe 10 dS m-1) although the

salinity reduced the protein contents (%) but with the application of potassium, salt

tolerant genotype produced more protein contents as compared to the salt sensitive

one. The comparison on relative basis indicated that protein contents of S-2002 was

91% at 100 mg kg-1 soil K and 90% at 200 mg kg-1 K while Akbar had 80% and 75%

of respective control, respectively.

XCVIII

Table 4.24. Oil contents, crude fiber and protein contents (%) of two maize genotypes grown under control (1.25 dS m-1) and 10 dS m-1 at two levels of potassium

Oil contents (%) Genotypes ECe dS m-1 K1 (100 mg kg-1 soil) K2 (200 mg kg-1 soil) S-2002 1.25

10.0 3.17±0.04 2.79±0.06

3.62±0.07 3.08±0.06

Akbar 1.25 10.0

2.42±0.12 2.10±0.03

2.86±0.05 2.26±0.04

Crude fiber (%) S-2002 1.25

10.0 1.17±0.02 1.03±0.03

1.41±0.01 1.07±0.03

Akbar 1.25 10.0

1.05±0.02 0.94±0.03

1.12±0.02 1.02±0.02

Protein contents (%) S-2002 1.25

10.0 5.16±0.12 4.70±0.11

5.60±0.11 5.08±0.13

Akbar 1.25 10.0

3.21±0.06 2.60±0.11

3.67±0.12 2.78±0.12

Each value is an average of 3 replications ± S.E.

4.3.4.4 Total soluble sugars

Data regarding total soluble sugars are given in Table 4.25. Saline growth

medium significantly decreased total soluble sugars contents in seeds of the two

maize genotypes. The most significant effect of salinity on total soluble sugars was

observed in case of salt sensitive genotype Akbar. The addition of potassium (200 mg

kg-1) to soil significantly increased soluble sugars. While on the other hand, addition

of K (200 mg kg-1) had no significant effect in case of genotype Akbar (Appendix

34).

There was a significant difference under both K-level in salt tolerant

genotype in non-saline (control) while with the application of potassium, salt tolerant

genotype produced more protein contents as compared to the salt sensitive one at soil

salinity (ECe 10 dS m-1). The comparison on relative basis indicated that total soluble

sugar in S-2002 was 91% at 100 mg kg-1 soil K and 81% at 200 mg kg-1 K level,

while; Akbar had 80% and 75% of their respective controls, respectively.

4.3.4.5 Proline Contents

XCIX

Imposition of salts (NaCl) in the growth medium significantly enhanced

the proline concentration in the leaves of both genotypes. A higher concentration of

proline was found in the salt tolerant maize genotype (S-2002). The addition of K had

no significant effect on the proline contents in the leaves of the salt sensitive

genotype (Appendix 35). The salt sensitive genotype had the lowest contents for

proline concentration under saline conditions and saline + K treatments. Both the

genotypes differed significantly in the concentration of proline contents in saline

environment as well as in saline + 200 mg kg-1 K in soil.

The comparison on relative basis indicated that S-2002 and Akbar had

228% and 181% proline concentration at 100 mg kg-1 soil K while, 230% and 152%

of proline concentration of respective control was found at 200 mg kg-1 soil K.

Table 4.25. Total soluble sugars (%) and proline (µ mol g-1) of two maize genotypes grown under control (1.25 dS m-1) and 10 dS m-1 at two levels of potassium

Total soluble sugars (%)

Genotypes ECe dS m-1 K1 (100 mg kg-1 soil) K2 (200 mg kg-1 soil)

S-2002 1.25 10.0

1.14±0.04 1.04±0.02

1.35±0.04 1.10±0.03

Akbar 1.25 10.0

0.85±0.01 0.68±0.02

0.91±0.05 0.69±0.03

Proline (µ mol g-1) S-2002 1.25

10.0 2.1±0.17 4.8±0.41

2.0±0.13 4.6±0.21

Akbar 1.25 10.0

1.6±0.21 2.9±0.31

1.4±0.15 2.7±0.21

Each value is an average of 3 replications ± S.E.

4.3.5 Discussion

The physiological, morphological and biochemical characteristics of the

maize genotypes were significantly affected by salinity, potassium application and

their interactions. The interaction between salinity, K levels and genotypes was also

significant for most of the parameters except water potential and crude fiber contents,

indicating differential response of genotypes to salinity and K levels.

Growth reduction due to salinity is mainly attributed to water deficit due to

lowered water potential in the root zone, nutritional imbalance and specific ion

C

toxicity arising from higher internal concentration of Na+ (Greenway and Munns,

1980). In the present study, salinity caused a significant reduction in growth

parameters of the two maize genotypes while a significant increase in growth was

observed with the addition of K (200 mg kg-1 soil).

The increase in growth is mainly associated with the supply of K nutrition

under saline conditions. It has been reported by Zhao et al. (1991) that K nutrition

significantly promoted the adventitious roots. The tolerant (S-2002) and sensitive

(Akbar) genotypes also varied significantly regarding different growth parameters

under the saline treatment. S-2002 and Akbar were selected from previous studies as

tolerant and sensitive genotypes, respectively. The shoot fresh weight of S-2002 was

about 1.5 times higher than that of Akbar at ECe 10 d Sm-1 and 1.89 times higher at

ECe 10 dS m-1 + K level respectively, and the same trend was observed in case of

shoot dry weight. However, for other growth parameters like, plant height/shoot

length, shoot fresh and dry weight, maize genotype S-2002 performed better than

Akbar at both salinity and salinity + K conditions.

Maize genotype Akbar had higher Na+ concentration in leaf sap in saline

(10 dS m-1) and saline + K (200 mg kg-1 soil) treatments compared to S-2002. On the

other hand, S-2002 had higher K+ concentration and about 2.7 times higher K+: Na+

ratio than Akbar under saline treatment while in saline + K (200 mg kg-1 soil), S-2002

had about 2.5 times higher K+: Na+ ratio than Akbar. A number of studies have

shown that growth performance of plants growing under saline conditions depends on

their ability to minimize the accumulation of Na+ and to have high concentrations of

K+ and K+: Na+ ratio in their actively growing leaves (Rashid et al., 1999; Munns and

James, 2003; Wenxue et al., 2003). Excess Na+ and Cl- inhibits the uptake of K+ and

leads to the appearance of symptoms like those in K+ deficiency. The deficiency of

K+ initially leads to chlorosis and then necrosis (Gopal and Dube, 2003). The role of

K+ is necessary for osmoregulation and protein synthesis, maintaining cell turgor and

stimulating photosynthesis (Freitas et al., 2001; Ashraf, 2004). Both K+ and Ca2+ are

required to maintain the integrity and functioning of cell membranes (Wenxue et al.,

2003). Maintenance of adequate K+ in plant tissue under salt stress seems to be

dependent upon selective K+ uptake and selective cellular K+ and Na+

compartmentation and distribution in the shoots (Munns et al., 2000; Carden et al.,

CI

2003). The maintenance of calcium acquisition and transport under salt stress is an

important determinant of salinity tolerance (Soussi et al., 2001; Unno et al., 2002).

In this study, higher K+ and K+: Na+ ratio and lower Na+ concentration helped maize

genotype, S-2002 in maintaining relatively higher photosynthetic rate and stomatal

conductance and, hence, a relatively higher growth. The better performance of S-

2002 under saline conditions and K addition might be due to restricted uptake and

transport of Na+ and better regulation of K+ uptake. High K+: Na+ ratio was observed

in bean plants that seem to limit sodium translocation to shoot (Khadri et al., 2007).

Asch et al. (2000) and Zhu et al. (2001) reported the same results in rice crop. In fact,

it is possible that high K+: Na+ ratio is more important for many species than simply

maintaining a low concentration of Na+ (Cuin et al., 2003; Mark and Romola, 2003).

Salinity is not only responsible for ion toxicity and imbalance, but also

indirectly leads to low photosynthesis in plants and photosynthesis is directly related

to stomatal conductance, transpiration, chlorophyll contents and water potential. At

low or moderate soil salinity, decreased growth is primarily associated with a

reduction in photosynthetic area rather than a reduction in photosynthesis per unit

leaf area (Munns, 1993). At high salinity, however, leaf photosynthesis can be

reduced by lowered stomatal conductance as result of water imbalance (Brugnoli and

Lauteri, 1991) or by non-stomatal factors that may be caused by toxic ions. Evidence

in support of this comes from strong negative correlations between ions and

photosynthetic activity, where Na+ and Cl- has been implicated primarily in crop

species such as rice (Yeo et al., 1985) and wheat (Rawson, 1986) and Cl- in woody

perennials such as citrus (Walker et al., 1993) and grapevine (Downton, 1977;

Walker et al., 1981). Because transpiration, stomatal conductance and chlorophyll

content in leaves can be measured by a non-destructive, rapid and easy technique

using a porometer, these physiological traits may be important to be used as

screening criteria if they would be closely associated with salt tolerance of genotypes

at a given level of salinity. In this study, significant genotype variation in net

transpiration rate, stomatal conductance and leaf water potential were observed.

However the genotypic variation was greater when K (200 mg kg-1 soil) was added to

the saline growth medium. It is clear that maize genotype S-2002 maintained its

stomatal conductance and transpiration rate as compared to the salt sensitive

CII

genotype Akbar and there was an improvement in photosynthetic parameters due to

the application of K (200 mg kg-1 soil). The effectiveness of screening method has

been examined in a number of studies as an index for response to salinity stress. For

example, this technique was used for screening groundnut genotypes for tolerance to

iron-deficiency chlorosis (Samdur et al., 2000) and it was also used to estimate tissue

tolerance for high Na+ accumulation (Munns and James, 2003). The stomatal

conductance of both genotypes differed significantly with each other at salinity level

(10 dS m-1 NaCl) and with K supply (200 mg kg-1 soil) to saline growth medium and

S-2002 maintained its stomatal conductance as compared to the genotype Akbar.

Previous studies showed that the stomatal conductance was linearly correlated with

maximum net photosynthesis rate in soybean (Ma et al., 1995), in rice (Laza et al.,

1996), and in wheat (Gutierrez-Rodriguez et al., 2000) and similar relationship was

observed in our study (Table 4.23). The data for transpiration showed that the

genotype S-2002 had non-significant values at salinity level while significantly

differed at salinity + K addition (200 mg kg-1 soil) treatment as compared to the

Akbar. According to many workers, transpiration rate is important for controlling the

accumulation of salt ions in shoot (Walker et al., 1990; Storey et al., 1993; Moya et

al., 1999; Storey and Walker, 1999). It seems that transpiration rate as a screening

criterion may be more important for drought stress than salt stress. Because one of

the stresses caused by salinity is osmotic stress or water deficit, the trait of leaf

transpiration in salt tolerant genotypes should be improved in order to further

increase their salt tolerance under saline conditions. On other hand low osmotic

potentials in the soil solution induced water deficit in plant tissues. As a consequence,

the turgor in plants may decrease, resulting from faster decrease in water potential

than in osmotic potential. Salt tolerance of maize genotypes may also vary with their

leaf water relations. The salt tolerant genotype S-2002 and salt sensitive genotype

Akbar varied significantly at salinity level as well as with the addition of K nutrition

to saline condition and S-2002 maintained its leaf water potential and remained better

as compared to Akbar genotype. Munns (1993) proposed that water deficit in plants

occur before the plants suffer from ionic effect (ion toxicity and ion imbalance).

Excess Na+ and Cl- inhibits the uptake of K+ and leads to the appearance of

CIII

symptoms like those in K+ deficiency. The deficiency of K+ initially leads to chlorosis

and then necrosis (Gopal and Dube, 2003).

Osmoregulation (i.e. maintenance of turgor) is considered to be an

important adaptation to drought (Hsiao, 1973; Morgan, 1984) and salinity stress

(Greenway and Munns, 1980; Ashraf and Waheed 1993; Ashraf and Foolad, 2005).

The leaf water potential and stomatal conductance were found lowest in Akbar, while

the better values of these two variables in salt tolerant S-2002 can be explained in

view of some earlier studies. For instance, in cowpea it was found that depression in

stomatal conductance did not affect leaf water potential under water deficit condition.

In fact, salt stress also imposes water deficit conditions to some extent (Greenway

and Munns, 1980; Ashraf 1994a, b).

It is now well evident that photosynthetic activity is one of the major

factors controlling growth (Lawlor, 1987; Shannon, 1998). Photosynthesis leads to

the production of organic osmotica, which play an important role in osmoregulation.

It is thus, expected that the rate of photosynthesis in salt tolerant species is inhibited

less than that in salt sensitive ones (Mansour et al., 2005). The production of organic

solutes like total soluble sugars, proteins and free proline are closely related to the

well functioning of all the activities related with growth under saline medium (Ashraf

and Foolad, 2007; Ashraf and Bashir, 2003). The salt tolerant genotype S-2002 had a

better production of osmoregulators, which maintained its water potential as

compared to Akbar. In genotype S-2002 there was a positive correlation between

growth and photosynthetic activity (Table 4.26). A large number of plant species

accumulate glycinebetaine and proline in response to salinity stress and their

accumulation may play a role in combating salinity stress (Ashraf, 1994b; Hanson

and Burnet, 1994; Mansour, 2000; Ashraf and Harris, 2004).

The oil contents and crude fiber also quantified along with other chemical

and biochemical variables. The oil contents differed significantly among the

genotypes under saline conditions and K application increased this parameter due to

the well maintenance of water potential by S-2002 as compared to Akbar. With the

maintenance of water potential, a plant might have better photosynthetic activities

like stomatal conductance and transpiration rate. These results are in accordance with

CIV

the findings of Francois (1994) who reported that oil and protein content in seed meal

of canola were not affected by salinity. Yermanos et al (1964) and Irving (1988)

reported the same findings in case of safflower. While assessing the role of KNO3 in

ameliorating the adverse effects of salt on melon, Kaya et al., (2007) reported that

addition of supplementary KNO3 to the rooting medium helped the melon plants to

avoid Na+ toxicity, improved cell membrane stability and Ca2+, K+ and nitrogen

uptake. From these reports, it is clear that addition of nutrients in soil may improve

the plant mineral nutrient status, which results in growth enhancement under saline

conditions. However, plants have to utilize sufficient amount of metabolic energy for

uptake of nutrients.

Sodium concentration in leaf sap of the two maize genotypes (S-2002

and Akbar) had significant negative correlations with plant height, potassium

concentration and potassium: sodium ratio while, it was positively correlated

with chloride concentration in shoot.

The oil contents of genotypes S-2002 and Akbar were positively

correlated with their crude fiber contents, total soluble sugars, protein contents

and stomatal conductance. On the other hand crude fiber contents, total soluble

sugars and protein contents were negatively correlated with proline

concentration.

Table 4.26 Pearson correlation in maize genotypes Akbar and S-2002 Plant

height Grain yield

Shoot dry wt

Na+ K+ K+: Na+ Cl-

Plant height

0.927 0.000

0.498 0.099

-0.929 0.000

0.598 0.044

0.852 0.000

-0.942 0.000

CV

Grain yield 0.825 0.001

0.553 0.062

-0.906 0.000

0.589 0.044

0.819 0.001

-0.951 0.000

Shoot dry wt

0.948 0.000

0.906 0.000

-0.595 0.041

0.894 0.000

0.678 0.015

-0.570 0.053

Na+ -0.733 0.007

-0.537 0.072

-0.741 0.006

-0.676 0.016

-0.882 0.000

0.967 0.000

K+ 0.869

0.000

0.929

0.000

0.953

0.000

-0.624

0.030

0.827

0.001

-0.643

0.024

K+: Na+ 0.926 0.000

0.802 0.002

0.943 0.000

-0.873 0.000

0.848 0.000

-0.826 0.001

Cl- -0.522 0.081

-0.182 0.571

-0.481 0.114

0.795 0.002

-0.399 0.198

-0.645 0.024

The values above the diagonal line are for Akbar and those below for S-2002 genotype. Below the Pearson correlation, is the P-value. Significance: P-value = 0.05

Table 4.27 Pearson correlation in maize genotypes Akbar and S-2002 Oil Crude

fiber Protein Total

soluble sugar

Proline Water potential

Stomatal conductance

Oil 0.771 0.003

0.874 0.000

0.795 0.002

-0.613 0.034

-0.829 0.001

0.746 0.005

Crude fiber 0.666 0.018

0.894 0.000

0.646 0.023

-0.547 0.066

-0.892 0.000

0.759 0.004

Protein 0.780 0.003

0.421 0.173

0.702 0.011

-0.547 0.066

-0.919 0.000

0.695 0.012

Total soluble sugar

0.948 0.000

0.673 0.016

0.785 0.002

-0.786 0.002

-0.848 0.000

0.614 0.034

Proline 0.583 0.046

-0.155 0.630

-0.643 0.024

-0.535 0.073

0.665 0.018

-0.665 0.018

Water potential

-0.854 0.000

-0.441 0.151

-0.771 0.003

-0.861 0.000

0.674 0.016

-0.747 0.005

Stomatal conductance

0.634 0.027

0.188 0.559

0.839 0.001

0.645 0.024

-0.857 0.000

-0.755 0.005

The values above the diagonal line are for Akbar and those below for S-2002 genotype. Below the Pearson correlation, is the P-value. Significance: P-value = 0.05

CVI

STUDY-4 PHYSIOLOGICAL AND BIOCHEMICAL ATTRIBUTES OF MAIZE GENOTYPES IN SALINE SOIL

4.4 RESULTS AND DISCUSSION

4.4.1 Results

4.4.1.1 Plant height (cm)

Data regarding plant height are given in Table 4.28. Salinity alone (10 dS

m-1) significantly reduced plant height of both maize genotypes (S-2002 and Akbar).

With the application of potassium (200 kg ha-1) plant height was increased in both

genotypes but this increase was significant in case of S-2002 and non-significant in

case of Akbar when compared with salinity alone (control). Plant was statistically

greater in S-2002 than Akbar. On relative basis, genotype S-2002 and Akbar had

plant height 122.3 % and 103.8% of respective control, at salinity + 200 kg ha-1

respectively.

4.4.1.2 Total biomass (g plant-1)

Data regarding total biomass are given in Table 4.28. Salinity had a

significant effect on the total biomass of both genotypes when grown under natural

field saline conditions. S-2002 produced a significantly greater total biomass than

Akbar. With the addition of potassium (200 kg ha-1) nutrition, the total biomass was

increased in both genotypes, but this increase was significant in case of S-2002 and

non significant in Akbar. On a relative basis, genotype S-2002 and Akbar had total

biomass 111% and 109% of their respective controls at salinity + 200 kg ha-1.

4.4.1.3 Grain yield (Mg ha-1)

Data regarding grain yield are given in Table 4.28. Salinity significantly

decreased the yield in both genotypes. S-2002 produced a significantly higher yield

than Akbar. With the application of potassium nutrition, the yield was increased in

both the genotypes but this increase was significant in case of S-2002 while non-

significant in case of Akbar. On a relative basis, genotype S-2002 and Akbar had

yield 105% and 110% of their respective controls at salinity + 200 kg ha-1.

CVII

4.4.1.4 Na+ concentration in leaf sap of maize genotypes

The concentration of Na+ in the youngest fully expanded leaves of maize

genotypes as affected by salinity (10 dS m-1) and salinity + potassium treatment are

given in Table 4.28. Natural saline conditions caused a significant increase in Na+

concentration in maize leaf sap (Appendix 39). However, under saline filed

conditions, S-2002 and Akbar varied significantly although application of potassium

nutrition (200 kg ha-1) to the salt affected field decreased Na+ concentration in the

leaves of the maize genotypes. S-2002 accumulated the lowest leaf Na+ concentration

under saline and saline + K (200 kg ha-1) application while Akbar had more Na+

concentration in its leaf sap.

4.4.1.5 K+ concentration in leaf sap of maize genotypes

Under the natural salt-affected field conditions, salinity alone and in

combination with potassium application (200 kg ha-1) significantly increased the

concentration of K+ in maize leaves (Table 4.28). Saline field conditions caused a

significantly higher reduction in K+ concentration than the salinity + potassium (200

kg ha-1) treatment for S-2002 and Akbar. However, for Akbar the reduction in leaf K+

concentration under salinity alone was significantly higher than that of saline +

potassium application (Appendix 40).

On the other hand, the application of potassium at the rate of 200 kg ha-1 to

the saline field significantly increased leaf K+ concentration but this increase was

more pronounced and significant in case of S-2002 as compared to Akbar. S-2002

had a higher K+ concentration in both treatments than Akbar.

CVIII

Table 4.28 Response of two maize genotypes to salinity and

potassium addition

Plant Height (cm) Genotypes Saline (ECe 10 dS m-1) Saline (ECe 10 dS m-1) + 200 kg K ha-1

S-2002 62.6±2.03 76.7±1.86 Akbar 43.3±2.60 45.0±2.08

Total biomass (g plant-1)

S-2002 540.3±7.86 601.33±6.33 Akbar 363.0±6.43 399.00±7.98

Grain yield (Mg ha-1) S-2002 2.1±0.04 2.2±0.10 Akbar 1.2±0.15 1.3±0.06

Sodium concentration (mol m-3) in leaf sap S-2002 73.0±2.31 63.7±0.88 Akbar 96.7±3.28 93.4±1.86

Potassium concentration (mol m-3) in leaf sap S-2002 81.0±1.15 112.0±0.60 Akbar 47.3±0.76 55.7±1.76

Each value is an average of 3 replications ± S.E.

4.4.1.6 K+ : Na+ ratio in leaf sap

CIX

A significant reduction in K+: Na+ ratio was observed in both genotypes

due to salinity. The effect of salinity alone was significantly higher than that of

potassium application. The lowest K+: Na+ ratio was observed in Akbar under saline

field conditions, whereas S-2002 had the highest K+: Na+ ratio under the same

treatment. On the other hand, with the application of K (200 kg ha-1), the K+: Na+

ratio was significantly increased. Under the saline field with K application, S-2002

had highest ratio while Akbar had the lowest one. Both genotypes differed

significantly under the both the treatments (Appendix 41).

4.4.1.7 Chloride concentration in fully expanded leaves

Data regarding Cl- concentration in the leaf sap are given in Table 4.29.

The presence of salts in the saline growth medium significantly increased the Cl-

concentration in the leaf sap of maize genotypes. Both the genotypes differed

significantly for the leaf ionic content of Cl- under the naturally salt affected soil. The

addition of potassium treatment (200 kg ha-1) to the saline field had significantly

affected on leaf Cl- concentrations in case of both genotypes S-2002 and Akbar.

Akbar. Genotype S-2002 had lower, while Akbar had higher concentrations of Cl- in

leaf sap.

4.4.1.8 Total soluble sugars

Data regarding total soluble sugars contents are given in Table 4.29. The

presence of salts in the growth medium of naturally salt affected filed significantly

reduced the total soluble sugars in the maize seed of both the genotypes. S-2002 had

highest sugar contents while Akbar had the lowest. The application of K at the rate of

200 kg ha-1 increased the total soluble sugar contents in both the genotypes but S-

2002 had the higher sugar contents as compared to the Akbar. Both the genotypes

differed significantly under both conditions (Appendix 43). On relative basis

genotype S-2002 and Akbar had 114% and 107% of their respective control at

salinity + 200 kg ha-1 K, respectively.

4.4.1.9 Protein contents

CX

Data regarding protein contents are given in Table 4.29. Salinity (ECe 10

dS m-1) present in the growth medium had a significant effect on protein contents.

The salt tolerant genotype S-2002 had the highest amount of soluble proteins under

saline condition, whereas reverse was true for the salt sensitive genotype Akbar.

Addition of K at the rate of 200 kg ha-1 significantly increased the protein contents in

both the genotypes but S-2002 showed significant increase as compared to Akbar

having non-significant difference (Appendix 44). Akbar remained at lowest level in

protein production as compared to S-2002. On relative basis genotype S-2002 and

Akbar had 112.8% and 108.4% values of their respective control at salinity + 200 kg

ha-1 K, respectively.

4.4.1.10 Seed oil contents

Data regarding seed oil contents are given in Table 4.29. Performance of

both genotypes under the natural salt affected field (ECe 10 dS m-1) varied

significantly. Saline growth medium caused a significant reduction in oil contents of

maize genotype Akbar as compared to the S-2002. The addition of K at the rate of

200 kg ha-1 caused a significant increase in oil contents of both genotypes S-2002 and

Akbar (Appendix45). The genotypes Akbar remained at lowest level and S-2002 was

at highest level in the production of oil contents under salt affected field with the

addition of K (200 kg ha-1) and S-2002 had 112.2% while, Akbar had 107.8% relative

value of their respective control (saline).

4.4.1.11 Crude fiber

Data regarding crude fiber contents are given in Table 4.29. The crude

fiber contents of both the maize genotypes under naturally salt affected soil differed

significantly at salinity alone and at salinity + K-application at the rate of 200 kg ha-1.

S-2002 remained better for crude fiber contents and Akbar had the lowest contents at

both the treatments (saline alone and saline + K). K addition had significant effect in

case of S-2002 while non-significant effect in case of Akbar. On relative basis

genotype S-2002 had 110% while, Akbar had 107% of their respective control (saline

treatment).

CXI

4.4.1.12 Proline contents

Data regarding proline contents are given in Table 4.29. The leaf proline

contents of both the genotypes varied significantly under the salt affected field

conditions. S-2002 had maximum concentration of proline against salinity stress

whereas Akbar had significantly less proline contents. Addition of K at the rate of

200 kg ha-1 to the saline field had no significant effects on the proline accumulation

(Apendix 47).

Table 4.29 K+: Na+ ratio, chloride concentration (mol m-3), total soluble sugars

(%), protein contents (%), seed oil contents (%), crude fiber (%) and proline contents (µ mol g-1) in maize genotypes grown in a saline field at two potassium levels.

Genotypes Saline (ECe 10 dS m-1) Saline (ECe 10 dS m-1) + 200 kg K ha-1 K+: Na+ ratio in leaf sap

S-2002 1.1±0.06 1.7±0.22 Akbar 0.49±0.05 0.60±0.04

Chloride concentration (mol m-3) in leaf sap S-2002 31.67±0.88 22.00±1.15 Akbar 57.33±1.45 47.00±1.00

Total soluble sugars (%) S-2002 1.29±0.04 1.48±0.05 Akbar 1.03±0.04 1.11±0.01

Protein contents (%) S-2002 3.43±0.07 3.87±0.12 Akbar 2.13±0.05 2.31±0.07

Seed oil contents (%) S-2002 3.11±0.03 3.51±0.10 Akbar 2.24±0.06 2.41±0.05

Crude fiber (%) S-2002 1.10±0.03 1.21±0.03 Akbar 0.85±0.02 0.91±0.01

Proline contents (µ mol g-1) S-2002 4.1±2.45 4.1±1.94 Akbar 1.7±1.07 1.6±0.77

Each value is an average of 3 replications ± S.E.

CXII

4.4.2 Discussion

The physical, chemical and biochemical characteristics of the two maize

genotypes were significantly affected by salinity and varied with the application of K.

The ionic relations were also disturbed significantly. The effect of salinity alone was

significantly higher than that of saline + potassium application in majority of the

parameters, while the maximum reduction in various growth parameters of two

genotypes was observed under natural saline field condition. The tolerant and

sensitive genotypes also varied significantly. Salinity is one of the major abiotic

stresses that adversely affect crop productivity and quality. Various approaches like

engineering techniques and the use of amendments as well as mineral nutrients are

advocated to improve plant survival under salt stress (Marschner, 1995).

Nevertheless, plant species and their genotypes differ genetically in their ability to

adapt to salt stress environment (Rozeff,; Wahid et al., 1997). Characteristics like dry

matter production, Na+ accumulation and K+: Na+ ratio have been considered a useful

guide to assess salt tolerance and selection of genotypes on this basis is an important

strategy to minimize growth reductions in saline soils (Santa-Maria and Epstein,

2001). The decrease in dry matter production of two maize genotypes in the presence

of saline growth medium (ECe 10 dS m-1) was due the ion toxicity as Na+ displaced

K+ and resulted in metabolic imbalances which reduced growth and yields (Zhu,

2002). Chinnusamy et al. (2005) also reported that under salt stress, the predominant

cause of reduced plant growth appeared to be ion toxicity rather than osmotic stress.

Ion cytotoxicity was caused by the displacement of K+ by Na+ in biochemical

reactions and conformational changes and the loss of functions of proteins as Na+

ions penetrated the hydration shells and interfered with non-covalent interactions

between their amino-acids. The magnitude of decline in dry matter production among

maize genotypes varied possibly because of their differential selectivity for K+ over

Na+ (Ashraf, 2002; Curtain and Naidu, 1998).

Growth reduction due to salinity is mainly attributed to water deficit which

results in lowering water potential in the root zone, nutritional imbalance and specific

ion toxicity arising from higher internal concentration of Na+ (Greenway and Munns,

1980). In the present study, salinity caused a significant reduction in growth

CXIII

parameters of all the maize genotypes, while a significant increase in growth was

observed with the addition of K (200 kg ha-1 soil) to saline growing media.

The increase in growth is mainly associated with the improvement of K

nutrition under saline condition (ECe 10 dS m-1). It has been reported by Zhao et al.

(1991) that K nutrition significantly promoted adventitious roots. The effect of

salinity alone was significantly higher than that of salinity + K addition (200 kg ha-1

soil) in most of parameters studied. The tolerant (S-2002) and sensitive (Akbar)

genotypes also varied significantly regarding different growth parameters under the

saline treatment. In the previous experiment conducted in pot-culture soil with same

conditions of saline growth medium (ECe 10 dS m-1) and K application, similar

results were reported for growth parameters (plant height, shoot fresh and dry weight,

grain yield), leaf ionic concentrations (Na+, K+, Cl- and K+: Na+ ratio) and

biochemical parameters.

Maize genotypes S-2002 and Akbar were used as tolerant and sensitive

genotypes, respectively. The plant height of S-2002 was 1.45 and 1.70 times higher

than Akbar under saline and saline + K (200 kg ha-1) treatments, respectively.

Genotype S-2002 performed better than Akbar for grain yield production. The yield

of S-2002 was about 1.75 times higher under saline field condition and 1.69 times

higher than Akbar under (saline + K at 200 kg ha-1).

In the saline field, Akbar had 23.7 mol m-3 and 29.7 mol m-3 higher Na+

concentration in leaf sap than S-2002 under saline field and saline + K (200 kg ha-1)

K treatment conditions respectively. On the other hand S-2002 had 1.71 and 2.01

times higher K+ concentration and 2.24 and 2.83 times higher K+: Na+ ratio under

saline field and saline field + potassium supply at 200 kg ha-1. In other words, the

higher concentration of Na+ and lower concentration of K+ and K+: Na+ ratio was

observed in the leaf sap of Akbar. A number of studies have shown that the growth

performance of plants growing under saline condition depends on their ability to

minimize the accumulation of toxic Na+ and to have a high concentration of K+ and

K+: Na+ ratio in their actively growing leaves (Schatchman and Munns, 1992; Rashid

et al., 1999; Saqib et al., 1999; Ashraf, 2004; Bastias et al., 2004; Sharma et al.,

CXIV

2005). Higher K+ and K+: Na+ ratio and lower Na+ concentration helped S-2002 in

maintaining relatively higher transpiration rates and stomatal conductance and

consequently a relatively higher growth. The better performance of S-2002 under

saline field conditions and with addition of K (200 kg ha-1) to the saline field, might

be due to restricted uptake and transport of Na+ and better regulation of K+ uptake.

Carden et al. (2003) found that the salt tolerant variety maintained a 10-fold lower

cytosolic Na+ and in the root cortical cells than the more sensitive variety. These

results are also in accordance with the findings of Wenxue et al. (2003) who reported

that a salt tolerant barley genotype maintained significantly lower Na+ concentrations

but higher K+: Na+ ratio in young leaf blades and young sheath tissues than the salt

sensitive one when exposed to salt stress.

Osmotic adjustment or accumulation of solutes by cells is a process by

which water potential of a cell can be decreased without an accompanying decrease

in cell turgor. It is a net increase in solute contents per cell that is independent of the

volume change that results from loss of water (Taiz and Zeiger, 2002). Osmotic

adjustment in plants subjected to salt stress can occur by the accumulation of high

concentrations of their inorganic ions or organic solutes (or both). Data for organic

solutes and proline (Table 4.29) showed that S-2002 had higher concentrations of

proline and soluble sugars under K (200 kg ha-1) treated saline (ECe 10 dS m-1) field

and saline field alone as compared to the salt sensitive genotype Akbar. Proline

accumulates in larger amounts than other amino acids in salt stressed plants (Wyn

Jones, 1981; Ashraf, 1994b; Ali et al., 1999; Abraham et al., 2003). Although both

organic and inorganic solutes play a crucial role in osmo-regulation of higher plants

subjected to saline conditions, their relative contribution varies among species,

among cultivars and even between different compartments within the same plants

(Greenway and Munns, 1980; Ashraf, 1994a; Ashraf and Bashir, 2003).

Data for protein contents in maize seeds/grains showed that S-2002 had

significantly more protein contents under potassium-treated saline field and saline

field alone as compared to Akbar. The increase in protein contents of S-2002 under

K-supplied saline field is due to the improved K nutrition of plants, because K plays

CXV

an important role in the transfer of nitrate from roots to shoots and leaves. Nitrate in

plants is reduced first to amines and then incorporated into amino acids to ultimately

form the protein. Both factors nitrate and protein contents are important contributors

to crop quality, yield and oil contents of maize. These results are in accordance with

the findings of Kaya et al. (2001b) who reported that additional supply of potassium

to a saline medium increased the yield of cucumber and pepper, while Cakmak

(2002) concluded that additional potassium supply under conditions of salt stress

substantially alleviated the stress and resulted in an increase in crop yield. While

assessing the role of KNO3 in ameliorating the adverse effects of salt on melon, Kaya

et al. (2007) found that addition of supplementary KNO3 to the rooting medium

helped the melon plants to avoid Na+ toxicity, improved cell membrane stability and

Ca2+, K+ and N uptake. From these reports, it is clear that addition of nutrients in soil

may improve the plant mineral nutrient status that results in growth enhancement

under saline conditions. However, plants have to utilize a sufficient amount of

metabolic energy for uptake of nutrients.

Addition of K at different levels of salinity inhibited the uptake of Na+ and

improved juice quality of sugarcane. Wahid (2004) demonstrated that potassium had

a positive correlation with extractable juice and brix percentage. While working with

sunflower, Akram et al. (2007) reported that foliar application of different K

inorganic salts considerably improved the ion homeostatic conditions. They also

found that improved K nutrition due to foliar application of K protected the cell

membrane as estimated by extent of ion leakage. Improved tissue K status of

sunflower plants regulated the plant photosynthetic activity through stomatal

movement. They concluded that all K application improved the growth of sunflower

plants, however; the effectiveness of K salts in improving growth depends upon a

number of factors including concentration of salt, plant developmental stage at which

applied, and accompanying anion in specific salt. Thus, application of inorganic salts

(K fertilizers) can be beneficial to improve crop salt tolerance.

Sodium concentration in shoot of genotypes S-2002 negatively correlated

with potassium concentration and potassium: sodium ratio, total soluble sugars, and

proteins. In case of Akbar, sodium concentration negatively correlated with K+ and

K+: Na+, oil content and total soluble sugar. On the other hand, Na+ concentration had

CXVI

a positive correlation with Cl- concentration and a negative correlation with protein

content. The oil content of genotype Akbar positively correlated with crude fiber, and

total soluble sugar. In case of S-2002 crude fiber, protein, and total soluble sugars

were positively correlated with oil content (Tables 4.30 and 4.31).

Table 4.30 Pearson correlation in maize genotypes Akbar and S-2002 Na+ K+ K+:

Na+ Cl- Oil

content Protein Total

soluble sugar

Na+ -0.874 0.023

-0.946 0.004

0.892 0.017

-0.836 0.038

-0.752 0.085

-0.906 0.013

K+ -0.856 0.030

0.974 0.001

-0.957 0.003

0.780 0.067

0.846 0.034

0.799 0.057

K+: Na+ -0.919 0.010

0.986 0.000

-0.962 0.002

0.862 0.027

0.852 0.031

0.852 0.031

Cl- 0.809 0.051

-0.967 0.002

-0.934 0.006

-0.875 0.023

-0.941 0.005

-0.893 0.017

Oil content

-0.886 0.019

0.975 0.001

0.968 0.002

-0.942 0.005

0.923 0.009

0.737 0.095

Protein -0.884 0.019

0.984 0.000

0.982 0.000

-0.944 0.005

0.994 0.000

0.761 0.079

Total soluble sugar

-0.829 0.041

0.883 0.020

0.864 0.027

-0.961 0.002

0.877 0.022

0.867 0.025

The values above the diagonal line are for Akbar and those below for S-2002 genotype.

Below the Pearson correlation, is the P-value. Significance: P-value = 0.05

Table 4.31 Pearson correlation in maize genotypes Akbar and S-2002

Oil content Crude fiber

Protein Total soluble sugar

Proline

Oil content 0.795 0.059

0.923 0.009

0.737 0.095

-0.663 0.151

Crude fiber

0.936 0.006

0.762 0.078

0.766 0.075

-0.875 0.022

Protein

0.994 0.000

0.923 0.009

0.761 0.079

-0.823 0.044

Total soluble sugar

0.877 0.022

0.781 0.066

0.867 0.025

-0.733 0.097

Proline

-0.825 0.043

-0.633 0.178

-0.840 0.036

-0.932 0 .007

The values above the diagonal line are for Akbar and those below for S-2002 genotype. Below the Pearson correlation, is the P-value. Significance: P-value = 0.05

CXVII

Chapter- 5

SUMMARY

Maize being the highest yielding cereal crop in the world is of significant

importance for countries like Pakistan, where rapidly increasing population has

already out stripped the available food supplies. In Pakistan maize is third important

cereal after wheat and rice. Maize accounts for 4.8% of the total cropped area and

3.5% of the value of agricultural output. The bulk (97%) of the total production come

from two major provinces, NWFP, accounting for 57% of the total area and 68% of

total production. Punjab contribute 38% acreage with 30% of total maize grain

production. Very little maize 2-3% is produced in the province of Sindh and

Balochistan. Though not included in Pakistan official statistics maize is an important

crop of Azad Jammu Kashmir (AJK) with about 0.122 million hectare of maize being

planted during kharif season. Similarly a very growing and high yielding sector of

maize, the spring maize area and production in Punjab is not accounted for , which

covers around 0.070 million ha with about 0.50 million tons of maize grain being

produced.

Maize (Zea mays L.) occupies a key position as one of the most important

cereals both for human and animal consumption and is grown under irrigated

conditions of arid and semi-arid regions of the world. Maize grain has high food

value and its oil is used for cooking purposes while green fodder is quite rich in

protein. In Pakistan, maize is the third most important cereal after wheat and rice.

Being an important “Kharif” crop, maize is grown on about one million hectares with

a total yield of about 3.13 million tons and an average yield of 3264 kg ha-1

(Anonymous, 2008).

The present study was planned to assess the salt tolerance of maize

genotypes and role of potassium in improving the salt tolerance with the following

objectives:

Assessment of salt tolerance of various maize genotypes in solution

culture.

CXVIII

Effect of potassium supply in alleviating salt stress in maize genotypes in

solution as well as soil culture.

Role of potassium in improving growth, quality, yield and biochemical

parameters of maize genotypes grown under salt stress.

Fifteen synthetic and hybrid maize genotypes were screened in Hoagland’s

nutrient solution at control, 70 and 100 mol m-3 NaCl. A wide variation was observed

between synthetic and hybrid genotypes. The synthetic genotypes produced more

shoot fresh and dry weights, accumulated lower concentrations of Na+ and Cl- in leaf

sap and maintained higher K+: Na+ ratio under saline conditions. Hybrid genotypes

produced less shoot fresh and dry weights and higher Na+ and Cl- concentrations in

leaf sap as well as maintained lower K+/Na+ ratio. Therefore, on the basis of better

performance, synthetic genotypes were used for further studies.

The seeds of selected genotypes (SL-2002, EV 1098, EV 6098, S-2002,

Agati 2000, Akbar, EV 5098) from the first study were grown in ½ strength

Hoagland’s nutrients solution. Three K levels (1.0, 5.5 and 8.0 mM K) were used at

control, 70 and 100 mol m-3 NaCl. Shoot fresh and dry weights of maize genotypes in

all saline treatments decreased consistently with increasing NaCl concentration in the

growth medium. Addition of K significantly alleviated the toxic effects of Na+ and

improved plant growth in maize genotypes under saline conditions. Salt sensitive

genotypes were less responsive to added K than salt tolerant genotypes. The

beneficial effect of K was more pronounced under saline conditions than under no-

salinity conditions.

To investigate and verify the role of K in mitigating the deleterious effects

of salinity, two maize genotypes, S-2002 and Akbar previously (in solution culture)

tested and identified as salt tolerant and salt sensitive, respectively were grown in a

pot culture to study yields and biochemical parameters. For this study, normal

textured soil was salinized using NaCl to develop an ECe of 10 dS m-1, while the

original soil was used as control. The original concentration of K (100 mg kg-1) in

soil was kept as a control, while the 2nd level of K was 200 mg kg-1 soil.

CXIX

There was a significant difference under both K levels in the salt tolerant

genotype in non-saline (control) and saline conditions. Addition of salts (ECe 10 dS

m-1) significantly reduced plant height and shoot fresh and dry weights of both

genotypes, but this reduction was more pronounced in the salt sensitive than in the

salt tolerant genotype on absolute or relative basis. Although salinity reduced shoot

fresh and dry weights, the application of potassium induced a higher biomass

production in the salt tolerant genotype than the salt sensitive one. The decrease in

grain yield of the salt tolerant genotype (S-2002) was less as compared to the salt

sensitive genotype (Akbar). The addition of K (200 mg kg-1) to the growth medium

produced more grain yield in maize genotype S-2002 than in Akbar. There was a

significant difference in growth and grain yield under both K levels (100 and 200 mg

kg-1 soil) in salt tolerant genotype under non-saline (control) and saline (ECe 10 dS m-

1) conditions. Salinity increased Na+ concentration in the leaf sap but with the

application of potassium, the salt tolerant genotype had lower concentrations of Na+

compared to the salt sensitive one. Addition of K improved dry matter, K+

concentration, yield, biochemical and photosynthetic parameters of both genotypes

under non-saline and saline conditions. The addition of 200 mg K kg-1 soil resulted in

significantly higher K+ concentrations in leaves of genotype S-2002 than in Akbar.

The addition of K 200 mg K kg-1 soil resulted in significantly higher K+

concentrations in leaves of genotype S-2002 than in Akbar.Addition of K to soil

improved stomatal conductance, transpiration rate, water potential, seed oil contents,

protein contents, total soluble sugars and crude fiber in salt tolerant genotype (S-

2002) as compared to the salt sensitive genotype (Akbar). S-2002 maintained its leaf

water potential even at saline (ECe 10 dS m-1) conditions. The highest concentration

of proline was found in the salt tolerant maize genotype (S-2002) under saline

environment, while the addition of K had no significant effects on the proline

contents in leaves of the salt sensitive genotype (Akbar).

The behavior of the tolerant (S-2002) and sensitive (Akbar) genotypes was

further tested in natural saline field having ECe 9.95-10.60 dS m-1. Potassium

concentration in the soil (i.e. 70-85.50 mg kg-1) was kept as control and 200 kg ha-1

was applied as K treatment. Salinity had a significant effect on plant height total

biomass and grain yield of both genotypes when grown under natural field saline

CXX

conditions. With the application of potassium (200 kg ha-1) growth response in terms

of plant height, total biomass, grain yield as well as biochemical parameters like total

soluble sugars, protein contents, crude fiber and oil contents of both genotypes varied

significantly compared to the control. Salt tolerant (S-2002) genotypes had better

improvement in all growth parameters compared to salt sensitive (Akbar) genotype.

Addition of K at the rate of 200 kg ha-1 to the saline field had no significant effects on

proline concentration. Under natural salt affected field conditions, salinity alone and

in combination with potassium application (200 kg ha-1) caused a significant effect on

the concentration of Na+, K+, K+: Na+ and Cl- concentration in leaf sap of maize

genotypes.

It was concluded that addition of K increased the crop growth, yield,

biochemical parameters and crop quality under saline conditions. Salt affected land

could be made productive through a better nutrient management and by growing salt

tolerant genotypes. In pot experiment, salinity (ECe 10.0 dS m-1) reduced plant height

up to 9% in S-2002 and 32% in Akbar. Reduction in grain yield (g plant-1) was up to

16% and 29% in genotypes S-2002 and Akbar, respectively. Salinity reduced oil

contents up to 15% in S-2002 while, reduction in Akbar was 21% however, with the

addition of K (200 mg kg-1 soil) oil contents in S-2002 were increased up to 14%

while, in Akbar the increase was 18% under non-saline condition while under saline

condition (ECe 10.0 dS m-1) the increase in oil contents of S-2002 was 10% compared

to Akbar having 7% increase.

Prospective

Genotypes S-2002 was a promising genotype under saline conditions, and can be used for the development of more salinity-tolerant genotypes.

CXXI

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Abraham, E. G. Rigo, G. Szekely, R. Nagy, C. Koncz and L. Szabados. 2003. Light-dependent induction of proline biosynthesis by abscisic acid and salt stress is inhibited by brassinosteroid in Arabidopsis. Plant. Biol. 51: 363-372.

Abrol, I. P., J. S. P. Yadov and F. I. Massiud. 1988. Salt affected soils and their management. Soil Resours. Manage. Conser. Ser., FAO Land and water Dev. Div. Bul. 39.

Akhtar, J., A. Shahzad, R. H. Qureshi and M. Aslam. 1998. Performance of selected wheat genotypes grown under saline and hypoxic environment. Pak. J. Soil Sci. 15: 146-153.

Akhtar, J., A. Naseem, K. Mahmood, S.Nawaz, R.H.Qureshi and M.Aslam. 2001. Response of some selected wheat genotypes to salinity. Growth and Ionic relations. Pak. J. Soil Sci. 19: 1-7.

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APPENDICES STUDY-1 Appendix 1 ANOVA TABLES

SHOOT LENGTH SOV df SS MS F P

S 2 17856.53 8928.27 2277.62 0.0000 SL 1 17756.60 17756.60 4529.74 0.0000 SQ 1 289.00 289.00 73.72 0.0000 G 14 16077.96 1148.43 292.97 0.0000 Syn Vs Hyb 1 7765.40 7765.40 1980.97 0.0000 Amng Syn 6 6338.40 1056.40 269.49 0.0000 Amng Hyb 7 1974.20 282.03 71.95 0.0000

S×G 28 910.58 32.52 8.30 0.0000 G×SL 14 802.20 57.30 14.62 0.0000 G×SQ 14 108.37 7.74 1.97 0.0286 Syn Vs Hyb (12.5) 1 1725.10 1725.10 440.08 0.0000 Syn Vs Hyb (70) 1 2641.40 2641.40 673.83 0.0000 Syn Vs Hyb (100) 1 3564.30 3564.30 909.26 0.0000 Among Syn (12.5) 6 1735.62 289.27 73.79 0.0000 Among Syn (70) 6 2081.90 346.98 88.52 0.0000 Among Syn (100) 6 2737.62 456.27 116.40 0.0000 Among Hyb (12.5) 7 1363.29 194.76 49.68 0.0000 Among Hyb (70) 7 670.00 95.71 24.42 0.0000 Among Hyb( 100) 7 469.29 67.04 17.10 0.0000

Error 90 352.67 3.92 Total 134

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Appendix 2 ROOT LENGTH SOV df SS MS F P

S 2 4168.84 2084.42 374.22 0.000SL 1 4119.70 4119.70 739.62 0.000SQ 1 49.10 49.10 8.82 0.003G 14 39517.16 2822.65 506.76 0.000 Syn Vs Hyb 1 35659.00 35659.00 6401.97 0.000 Amng Syn 6 2678.19 446.37 80.14 0.000 Amng Hyb 7 1179.76 168.54 30.26 0.000

S×G 28 177.60 6.34 1.14 0.315 G×SL 14 99.85 7.13 1.28 0.235 G×SQ 14 77.75 5.55 1.00 0.463 Syn Vs Hyb (12.5) 1 11969.00 11969.00 2148.83 0.000 Syn Vs Hyb (70) 1 11908.00 11908.00 2137.88 0.000 Syn Vs Hyb (100) 1 11782.00 11782.00 2115.26 0.000 Among Syn (12.5) 6 1157.24 192.87 34.63 0.000 Among Syn (70) 6 693.90 115.65 20.76 0.000 Among Syn (100) 6 948.57 158.10 28.38 0.000 Among Hyb (12.5) 7 502.00 71.71 12.88 0.000 Among Hyb (70) 7 335.16 47.88 8.60 0.000 Among Hyb( 100) 7 398.29 56.90 10.22 0.000

Error 90 501.33 5.57 Total 134

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Appendix 3 Shoot Fresh Weight SOV df SS MS F P S 2 550.65 275.33 248.04 0.000 SL 1 545.87 545.87 491.77 0.000 SQ 1 4.78 4.78 4.31 0.040 G 14 4205.30 300.38 270.61 0.000 Syn Vs Hyb 1 1676.40 1676.40 1510.27 0.000 Amng Syn 6 1314.61 219.10 197.39 0.000 Amng Hyb 7 1214.34 173.48 156.29 0.000 S×G 28 125.83 4.49 4.05 0.000 G×SL 14 122.10 8.72 7.86 0.0000 G×SQ 14 3.74 0.27 0.24 0.997 Syn Vs Hyb (12.5) 1 958.23 958.23 863.27 0.000 Syn Vs Hyb (70) 1 502.85 502.85 453.02 0.000 Syn Vs Hyb (100) 1 307.53 307.53 277.05 0.000 Among Syn (12.5) 6 532.19 88.70 79.91 0.000 Among Syn (70) 6 400.43 66.74 60.12 0.000 Among Syn (100) 6 396.43 66.07 59.52 0.000 Among Hyb (12.5) 7 542.22 77.46 69.78 0.000 Among Hyb (70) 7 398.81 56.97 51.33 0.000 Among Hyb( 100) 7 292.47 41.78 37.64 0.000 Error 90 99.55 1.11 Total 134

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Appendix 4 SHOOT DRY WEIGHT SOV df SS MS F P S 2 10.66 5.33 266.50 0.000 SL 1 10.00 10.00 500.00 0.000 SQ 1 0.66 0.66 33.00 0.000 G 14 162.33 11.60 579.75 0.000 Syn Vs Hyb 1 70.33 70.33 3516.50 0.000 Amng Syn 6 56.04 9.34 467.00 0.000 Amng Hyb 7 35.95 5.14 256.79 0.000 S×G 28 1.88 0.07 3.35 0.000 G×SL 14 1.13 0.08 4.04 0.000 G×SQ 14 0.74 0.05 2.64 0.002 Syn Vs Hyb (12.5) 1 29.29 29.29 1464.50 0.000 Syn Vs Hyb (70) 1 19.29 19.29 964.50 0.000 Syn Vs Hyb (100) 1 22.30 22.30 1115.00 0.000 Among Syn (12.5) 6 18.37 3.06 153.08 0.000 Among Syn (70) 6 18.11 3.02 150.92 0.000 Among Syn (100) 6 19.67 3.28 163.92 0.000 Among Hyb (12.5) 7 13.04 1.86 93.14 0.000 Among Hyb (70) 7 10.93 1.56 78.07 0.000 Among Hyb( 100) 7 13.22 1.89 94.43 0.000 Error 90 1.37 0.02 Total 134

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Appendix 5 Root Fresh Weight SOV df SS MS F P S 2 44.68 22.34 413.70 0.000 SL 1 43.07 43.07 797.65 0.000 SQ 1 1.61 1.61 29.78 0.000 G 14 721.52 51.54 954.39 0.000 Syn Vs Hyb 1 403.58 403.58 7473.70 0.000 Amng Syn 6 195.60 32.60 603.69 0.000 Amng Hyb 7 122.35 17.48 323.67 0.000 S×G 28 4.00 0.14 2.65 0.000 G×SL 14 3.41 0.24 4.51 0.000 G×SQ 14 0.60 0.04 0.79 0.680 Syn Vs Hyb (12.5) 1 146.39 146.39 2710.93 0.000 Syn Vs Hyb (70) 1 143.83 143.83 2663.52 0.000 Syn Vs Hyb (100) 1 114.58 114.58 2121.85 0.000 Among Syn (12.5) 6 59.50 9.92 183.64 0.000 Among Syn (70) 6 65.96 10.99 203.58 0.000 Among Syn (100) 6 70.69 11.78 218.18 0.000 Among Hyb (12.5) 7 55.05 7.86 145.62 0.000 Among Hyb (70) 7 40.50 5.79 107.14 0.000 Among Hyb( 100) 7 29.05 4.15 76.84 0.000 Error 90 4.88 0.05 Total 134

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Appendix 6 ROOT DRY WEIGHT SOV df SS MS F P

S 2 2.07 1.04 34.50 0.000 SL 1 2.07 2.07 68.93 0.000 SQ 1 0.005 0.00 0.15 0.696 G 14 15.35 1.10 36.55 0.000 Syn Vs Hyb 1 8.05 8.05 268.43 0.000 Amng Syn 6 4.49 0.75 24.94 0.000 Amng Hyb 7 2.81 0.40 13.38 0.000

S×G 28 0.29 0.01 0.35 0.998 G×SL 14 0.21 0.02 0.50 0.927 G×SQ 14 0.08 0.01 0.19 0.999 Syn Vs Hyb (12.5) 1 3.47 3.47 115.67 0.000 Syn Vs Hyb (70) 1 2.16 2.16 71.90 0.000 Syn Vs Hyb (100) 1 2.50 2.50 83.47 0.000 Among Syn (12.5) 6 1.45 0.24 8.06 0.000 Among Syn (70) 6 1.52 0.25 8.44 0.000 Among Syn (100) 6 1.60 0.27 8.86 0.000 Among Hyb (12.5) 7 1.38 0.20 6.57 0.000 Among Hyb (70) 7 0.84 0.12 4.00 0.000 Among Hyb( 100) 7 0.72 0.10 3.43 0.002

Error 90 0.28 0.003 Total 134

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Appendix 7 Na+ CONC. SOV df SS MS F P

S 2 42237.50 21118.75 35197.9 0.000 SL 1 41057.20 41057.20 68428.6 0.000 SQ 1 1180.40 1180.40 1967.33 0.000 G 14 6548.40 467.74 779.57 0.000 Syn Vs Hyb 1 4598.60 4598.60 7664.33 0.000 Amng Syn 6 1633.50 272.25 453.75 0.000 Amng Hyb 7 316.40 45.20 75.33 0.000 S×G 28 2893.00 103.32 172.20 0.000 G×SL 14 2673.30 190.95 318.25 0.000 G×SQ 14 219.60 15.69 26.14 0.000 Syn Vs Hyb (12.5) 1 13.73 13.73 22.88 0.000 Syn Vs Hyb (70) 1 2900.80 2900.80 4834.67 0.000 Syn Vs Hyb (100) 1 3586.90 3586.90 5978.17 0.000 Among Syn (12.5) 6 2.73 0.46 0.76 0.604 Among Syn (70) 6 954.55 159.09 265.15 0.000 Among Syn (100) 6 1439.68 239.95 399.91 0.000 Among Hyb (12.5) 7 2.24 0.32 0.53 0.807 Among Hyb (70) 7 252.46 36.07 60.11 0.000 Among Hyb( 100) 7 288.29 41.18 68.64 0.000 Error 90 54.20 0.60 Total 134

CL

Appendix 8 K+ CONC. SOV Df SS MS F P S 2 225.47 112.74 955.38 0.000 SL 1 222.10 222.10 1882.20 0.000 SQ 1 3.37 3.37 28.56 0.000 G 14 2086.81 149.06 1263.20 0.000 Syn Vs Hyb 1 1597.40 1597.40 13537.29 0.000 Amng Syn 6 476.37 79.40 672.84 0.000 Amng Hyb 7 13.06 1.87 15.81 0.000 S×G 28 11.10 0.40 3.36 0.000 G×SL 14 7.27 0.52 4.40 0.000 G×SQ 14 3.84 0.27 2.32 0.008 Syn Vs Hyb (12.5) 1 571.39 571.39 4842.29 0.000 Syn Vs Hyb (70) 1 546.39 546.39 4630.42 0.000 Syn Vs Hyb (100) 1 481.64 481.64 4081.69 0.000 Among Syn (12.5) 6 165.29 27.55 233.46 0.000 Among Syn (70) 6 170.55 28.43 240.89 0.000 Among Syn (100) 6 143.23 23.87 202.30 0.000 Among Hyb (12.5) 7 11.76 1.68 14.24 0.000 Among Hyb (70) 7 6.52 0.93 7.89 0.000 Among Hyb( 100) 7 1.13 0.16 1.37 0.228 Error 90 10.66 0.118 Total 134

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Appendix 9 K+/Na+

SOV df SS MS F P S 2 111.17 55.59 5053.18 0.000 SL 1 101.40 101.40 9218.18 0.000 SQ 1 9.77 9.77 888.18 0.000 G 14 39.93 2.85 259.29 0.000 Syn Vs Hyb 1 25.83 25.83 2348.18 0.000 Amng Syn 6 13.99 2.33 211.97 0.000 Amng Hyb 7 0.11 0.02 1.43 0.203 S×G 28 33.05 1.18 107.31 0.000 G×SL 14 29.72 2.12 192.99 0.000 G×SQ 14 3.33 0.24 21.62 0.000 Syn Vs Hyb (12.5) 1 47.61 47.61 4328.18 0.000 Syn Vs Hyb (70) 1 1.09 1.09 99.09 0.000 Syn Vs Hyb (100) 1 0.74 0.74 67.27 0.000 Among Syn (12.5) 6 21.08 3.51 319.39 0.000 Among Syn (70) 6 1.07 0.18 16.21 0.000 Among Syn (100) 6 0.76 0.13 11.52 0.000 Among Hyb (12.5) 7 0.48 0.07 6.23 0.000 Among Hyb (70) 7 0.15 0.02 1.95 0.071 Among Hyb( 100) 7 0.001 0.00 0.01 1.000 Error 90 0.97 0.011 Total 134

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Appendix 10 Cl- Conc. SOV df SS MS F P S 2 27647.89 13823.95 2770.33 0.000 SL 1 27463.58 27463.58 5503.72 0.000 SQ 1 184.00 184.00 36.87 0.000 G 14 6202.32 443.02 88.78 0.000 Syn Vs Hyb 1 5403.40 5403.40 1082.85 0.000 Amng Syn 6 476.70 79.45 15.92 0.000 Amng Hyb 7 322.20 46.03 9.22 0.000 S×G 28 2682.39 95.80 19.20 0.000 G×SL 14 2251.94 160.85 32.24 0.000 G×SQ 14 430.46 30.75 6.16 0.000 Syn Vs Hyb (12.5) 1 14.82 14.82 2.97 0.088 Syn Vs Hyb (70) 1 3848.30 3848.30 771.20 0.000 Syn Vs Hyb (100) 1 3774.20 3774.20 756.35 0.000 Among Syn (12.5) 6 4.87 0.81 0.16 0.985 Among Syn (70) 6 210.01 35.00 7.01 0.000 Among Syn (100) 6 459.81 76.64 15.36 0.000 Among Hyb (12.5) 7 2.45 0.35 0.07 0.999 Among Hyb (70) 7 380.94 54.42 10.91 0.000 Among Hyb( 100) 7 189.26 27.04 5.42 0.000 Error 90 448.86 4.99 Total 134

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ANOVA TABLES STUDY-2

Appendix 11 Analysis of Variance for SL/Plant height Source DF Seq SS Adj SS Adj MS F P salinity 2 61153 61112 30556 534.15 0.000 potash 2 127635 127273 63636 1112.43 0.000 Var 6 29534 29534 4922 86.05 0.000 Error 177 10125 10125 57 Total 187 228447 Appendix 12 Analysis of Variance for Root length. Source DF Seq SS Adj SS Adj MS F P salinity 2 6931.9 6932.8 3466.4 449.70 0.000 potash 2 7439.0 7417.9 3708.9 481.17 0.000 Var 6 5024.5 5024.5 837.4 108.64 0.000 Error 177 1364.4 1364.4 7.7 Total 187 20759.8 Appendix 13 Analysis of Variance for SFW Source DF Seq SS Adj SS Adj MS F P salinity 2 9477.5 9501.1 4750.6 57.25 0.000 potash 2 33859.5 33692.6 16846.3 203.03 0.000 Var 6 37677.5 37677.5 6279.6 75.68 0.000 Error 177 14686.6 14686.6 83.0 Total 187 95701.1 Appendix 14 Analysis of Variance for SDW Source DF Seq SS Adj SS Adj MS F P salinity 2 196.59 195.26 97.63 125.76 0.000 potash 2 598.98 597.43 298.71 384.79 0.000 Var 6 470.09 470.09 78.35 100.92 0.000 Error 177 137.41 137.41 0.78 Total 187 1403.06 Appendix 15 Analysis of Variance for RFW Source DF Seq SS Adj SS Adj MS F P salinity 2 1054.8 1049.6 524.8 63.39 0.000 potash 2 7356.7 7333.6 3666.8 442.93 0.000 Var 6 4258.7 4258.7 709.8 85.74 0.000 Error 177 1465.3 1465.3 8.3 Total 187 14135.4

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Appendix 16 Analysis of Variance for RDW Source DF Seq SS Adj SS Adj MS F P salinity 2 28.523 28.458 14.229 91.15 0.000 potash 2 150.652 149.997 74.999 480.45 0.000 Var 6 83.060 83.060 13.843 88.68 0.000 Error 177 27.630 27.630 0.156 Total 187 289.865 Appendix 17 Analysis of Variance for Na Source DF Seq SS Adj SS Adj MS F P salinity 2 134937 135169 67584 517.79 0.000 potash 2 12901 12834 6417 49.16 0.000 Var 6 27888 27888 4648 35.61 0.000 Error 177 23103 23103 131 Total 187 198829 Appendix 18 Analysis of Variance for K Source DF Seq SS Adj SS Adj MS F P salinity 2 39367.4 39680.3 19840.2 361.80 0.000 potash 2 2720.9 2733.4 1366.7 24.92 0.000 Var 6 27316.6 27316.6 4552.8 83.02 0.000 Error 177 9706.2 9706.2 54.8 Total 187 79111.1 Appendix 19 Analysis of Variance for K+ : Na+ Source DF Seq SS Adj SS Adj MS F P salinity 2 156.712 157.117 78.559 865.07 0.000 potash 2 9.935 9.929 4.964 54.67 0.000 Var 6 31.596 31.596 5.266 57.99 0.000 Error 177 16.074 16.074 0.091 Total 187 214.317 Appendix 20 Analysis of Variance for Cl Source DF Seq SS Adj SS Adj MS F P salinity 2 38905.8 38928.7 19464.4 121.95 0.000 potash 2 19847.4 19796.9 9898.4 62.01 0.000 Var 6 34553.7 34553.7 5759.0 36.08 0.000 Error 177 28251.8 28251.8 159.6 Total 187 121558.8

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Appendix 21 ANOVA TABLES STUDY-3 Shoot length SOV df SS MS F P G 1 4676.04 4676.04 426.65 0.000 S 1 1305.38 1305.38 119.10 0.000 K 1 345.04 345.04 31.48 0.000 GS 1 345.04 345.04 31.48 0.000 GK 1 77.04 77.04 7.03 0.017 SK 1 12.04 12.04 1.10 0.310 GSK 1 0.04 0.04 0.00 0.952 Error 16 175.33 10.96 Total 23 6935.96

Appendix 22 Shoot Fresh Weight & dry weight

Shoot Fresh Weight Shoot dry weight SOV df SS MS F P SS MS F P

G 1 43958.00 43958.00 563.56 0.000 2220.10 2220.10 670.73 0.000S 1 6712.00 6712.00 86.05 0.000 212.59 212.59 64.23 0.000K 1 27160.00 27160.00 348.21 0.000 962.54 962.54 290.80 0.000

GS 1 905.00 905.00 11.60 0.004 66.97 66.97 20.23 0.000GK 1 5960.00 5960.00 76.41 0.000 202.36 202.36 61.14 0.000SK 1 141.00 141.00 1.81 0.197 0.10 0.10 0.03 0.864

GSK 1 47.00 47.00 0.60 0.448 14.96 14.96 4.52 0.049Error 16 1248.00 78.00 52.93 3.31 Total 23 86101.00 3732.56

Appendix 23 1000 seed weight

SOV df SS MS F G 1 81.881 81.881 6993.41 S 1 1.220 1.220 104.16 K 1 2.620 2.620 223.79

GS 1 0.490 0.490 41.87 GK 1 1.426 1.426 121.79 SK 1 0.021 0.021 1.79

GSK 1 0.000 0.000 0.01 Error 16 0.187 0.012 Total 23 87.845

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Appendix 24 Na+ concentration

SOV df SS MS F P G 1 770.67 770.67 39.77 0.000 S 1 1837.50 1837.50 94.81 0.000 K 1 280.17 280.17 14.46 0.002

GS 1 308.17 308.17 15.90 0.001 GK 1 8.17 8.17 0.42 0.525 SK 1 32.67 32.67 1.69 0.213

GSK 1 2.67 2.67 0.14 0.716 Error 16 310.00 19.38 Total 23 3550.00

Appendix 25 K+ CONC.

SOV df SS MS F P G 1 1261.50 1261.50 81.39 0.000 S 1 522.67 522.67 33.72 0.000 K 1 2773.50 2773.50 178.94 0.000

GS 1 0.67 0.67 0.04 0.838 GK 1 121.50 121.50 7.84 0.013 SK 1 54.00 54.00 3.48 0.080

GSK 1 16.67 16.67 1.08 0.315 Error 16 248.00 15.50 Total 23 4998.50

Appendix 26 K+: Na+

SOV df SS MS F P

G 1 9.16 9.16 88.93 0.000 S 1 11.72 11.72 113.79 0.000 K 1 11.47 11.47 111.36 0.000

GS 1 0.19 0.19 1.84 0.190 GK 1 1.69 1.69 16.41 0.001 SK 1 0.55 0.55 5.34 0.035

GSK 1 0.05 0.05 0.49 0.514 Error 16 1.65 0.10 Total 23 36.48

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Appendix 27 Cl- CONC.

SOV df SS MS F G 1 1335.04 1335.04 196.62 S 1 1584.37 1584.37 233.34 K 1 57.04 57.04 8.40

GS 1 247.04 247.04 36.38 GK 1 30.38 30.38 4.47 SK 1 22.04 22.04 3.25

GSK 1 35.04 35.04 5.16 Error 16 108.67 6.79 Total 23 3419.63

Appendix 28 Stomatal conductance

SOV df SS MS F P G 1 11235.50 11235.50 1256.77 0.000 S 1 791.40 791.40 88.52 0.000 K 1 991.50 991.50 110.91 0.000

GS 1 3.10 3.10 0.35 0.567 GK 1 258.20 258.20 28.88 0.000 SK 1 25.80 25.80 2.89 0.108

GSK 1 35.30 35.30 3.95 0.064 Error 16 143.00 8.94 Total 23 13483.90

Appendix 29 Transpiration rate

SOV df SS MS F P G 1 10.481 10.481 2096.16 0.000 S 1 0.073 0.073 14.52 0.002 K 1 0.454 0.454 90.76 0.000

GS 1 0.008 0.008 1.62 0.223 GK 1 0.005 0.005 0.96 0.342 SK 1 0.002 0.002 0.48 0.500

GSK 1 0.045 0.045 9.02 0.009 Error 16 0.081 0.005 Total 23 11.148

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Apendix-30 Water potential

SOV df SS MS F P G 1 0.304 0.3040 380.00 0.000 S 1 0.154 0.1540 192.50 0.000 K 1 0.029 0.0290 36.25 0.000

GS 1 0.084 0.0840 105.00 0.000 GK 1 0.003 0.0028 3.52 0.079 SK 1 0.002 0.0024 3.00 0.102

GSK 1 0.004 0.0038 4.69 0.046 Error 16 0.013 0.0008 Total 23 0.593

Appendix 31 Seed Oil Contents

SOV df SS MS F P G 1 9.88 9.88 796.91 0.000 S 1 1.09 1.09 88.09 0.000 K 1 4.51 4.51 363.44 0.000

GS 1 0.01 0.01 0.44 0.519 GK 1 1.93 1.93 155.38 0.000 SK 1 0.04 0.04 2.85 0.112

GSK 1 0.02 0.02 1.94 0.183 Error 16 0.20 0.01 Total 23 17.67

Appendix 32 Crude fiber

SOV df SS MS F P G 1 0.440 0.440 220.05 0.000 S 1 0.010 0.010 5.00 0.040 K 1 0.065 0.065 32.55 0.000

GS 1 0.032 0.032 15.77 0.001 GK 1 0.008 0.008 4.22 0.057 SK 1 0.012 0.012 5.85 0.028

GSK 1 0.022 0.022 11.10 0.004 Error 16 0.032 0.002 Total 23 0.621

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Appendix 33 protein contents

SOV df SS MS F P G 1 26.670 26.670 703.70 0.000 S 1 2.018 2.018 53.26 0.000 K 1 0.992 0.992 26.18 0.000

GS 1 0.180 0.180 4.76 0.044 GK 1 0.045 0.045 1.19 0.292 SK 1 0.012 0.012 0.32 0.579

GSK 1 0.054 0.054 1.43 0.249 Error 16 0.607 0.038 Total 23 30.579

Appendix 34 Total soluble sugars

SOV df SS MS F P G 1 1.088 1.0880 340.00 0.000 S 1 0.152 0.1520 47.50 0.000 K 1 0.165 0.1650 51.56 0.000

GS 1 0.017 0.0165 5.16 0.037 GK 1 0.103 0.1027 32.09 0.000 SK 1 0.000 0.0002 0.06 0.804

GSK 1 0.002 0.0022 0.69 0.418 Error 16 0.051 0.0032 Total 23 1.578

Appendix 35 Proline contents

SOV df SS MS F P G 1 4949.03 4949.03 404.00 0.000 S 1 283.59 283.59 23.15 0.000 K 1 87.78 87.78 7.17 0.017

GS 1 47.38 47.38 3.87 0.067 GK 1 15.11 15.11 1.23 0.283 SK 1 2.76 2.76 0.23 0.641

GSK 1 0.00 0.00 0.00 0.998 Error 16 195.95 12.25 Total 23 5581.65

ANOVA TABLES STUDY-4 Shoot length/Plant height

CLX

Appendix 35 SOV df SS MS F P Block 2 43.11 21.56 1.88 0.232

G 1 1950.75 1950.75 170.07 0.000 K 1 184.08 184.08 16.05 0.007

G×K 1 114.08 114.08 9.95 0.020 Error 6 68.83 11.47 Total 11 2360.92

Appendix 36 Yield

SOV df SS MS F P Block 2 0.105 0.053 2.76 0.144

G 1 32.329 32.329 1701.53 0.000 K 1 0.755 0.755 39.74 0.001

G×K 1 0.1027 0.103 5.41 0.061 Error 6 0.1159 0.019 Total 11 36.4076

Appendix 37 Na+ CONC.

SOV df SS MS F P Block 2 73.5 36.75 4.55 0.063

G 1 1850.08 1850.08 228.97 0.000 K 1 310.08 310.08 38.38 0.001

G×K 1 2.08 2.08 0.26 0.630 Error 6 48.5 8.08 Total 11 2284.25

Appendix 38 K+

SOV df df SS MS F P Block 2 2 30.17 15.09 1.62 0.274

G 1 1 2945.33 2945.33 316.36 0.000 K 1 1 1200.00 1200.00 128.89 0.000

G×K 1 1 481.33 481.33 51.70 0.000 Error 6 6 55.83 9.31 Total 11 11 4712.67

Appendix 39 K+: Na+

CLXI

SOV df SS MS F P Block 2 0.1638 0.082 2.64 0.149

G 1 9.2928 9.293 299.77 0.000 K 1 3.6963 3.696 119.24 0.000

G×K 1 2.2533 2.253 72.69 0.000 Error 6 0.1851 0.031 Total 11 15.5913

Appendix 40 Cl- concentration

SOV df SS MS F P Block 2 26.00 13.00 14.61 0.005

G 1 1925.33 1925.33 2163.29 0.000 K 1 300.00 300.00 337.08 0.000

G×K 1 0.33 0.33 0.37 0.563 Error 6 5.33 0.89 Total 11 2257.00

Appendix 40 Total soluble sugars SOV df SS MS F P Block 2 0.017817 0.008909 3.54 0.097 G 1 0.3072 0.3072 121.90 0.000 K 1 0.1083 0.1083 42.98 0.001 G×K 1 0.012033 0.012033 4.78 0.072 Error 6 0.015117 0.00252 Total 11 0.460467 Appendix 41 Protein contents

SOV df SS MS F P Block 2 0.0014 0.0007 0.03 0.973

G 1 10.9061 10.9061 422.72 0.000 K 1 5.1221 5.1221 198.53 0.000

G×K 1 1.092 1.092 42.33 0.001 Error 6 0.155 0.0258 Total 11 17.2767

Appendix 42 oil contents

CLXII

SOV df SS MS F P Block 2 0.0192 0.01 0.72 0.526

G 1 11.0017 11.00 821.02 0.000 K 1 6.8252 6.83 509.34 0.000

G×K 1 3.2344 3.23 241.37 0.000 Error 6 0.0806 0.01 Total 11 21.1611

Appendix 43 Crude fiber

SOV df SS MS F P Block 2 0.002217 0.001 0.49 0.633

G 1 0.267008 0.267 119.09 0.000 K 1 0.069008 0.069 30.78 0.001

G×K 1 0.007008 0.007 3.13 0.127 Error 6 0.01345 0.002 Total 11 0.358692

Appendix 44 Proline contents

SOV df SS MS F P Block 2 33.57 16.785 2.84 0.135

G 1 1632.87 1632.87 276.76 0.000 K 1 150.52 150.52 25.51 0.002

G×K 1 21.87 21.87 3.71 0.102 Error 6 35.38 5.90 Total 11 1874.2