**3. Description of soybean morphology and anatomy**

in ground, surface or atmospheric water, known as drought are highly susceptible to pests and diseases. Mattson and Haack [1] provided evidence on the occurrence of fungi and insect induced stalk rots, wilts and foliar diseases in plants caused by drought stress. The prevalence in disease outbreak occurred in water-stressed plants compared to the normal water stressfree plants. Estimations of yield losses in soybeans compiled by Wrather and Koenning in the United States from 1996 until 2007 indicated that, the role of pathogens such as soybean cyst nematode, phytophthora root and stem rot, as well as charcoal rot that affected seedling development was exacerbated by the physical environmental stress conditions [2]. Drought is, and continues to be an insidious hazard to plants, animals and human lives. Drought conditions in many regions worldwide are worsening due to various factors, some of which are

380–400 ppm, and alterations in hydrological cycles make drought a recurring natural hazard world-wide [3, 4]. In this regard, plants undergo permanent or temporary damage to their morphological architecture, and their anatomical and physiological processes when exposed to dry and hot conditions. According to Shao et al. [5], water stress effects can be extended in plants to alter gene expression, change cellular metabolism, cause reduction in mitotic cell division activities in mesophyll tissues and other organs, as well as to cause the decrease in stomatal conductance [6]. Scientific research showed that; drought stress causes imbalances in the natural status of the environment and drastically disrupts crop cultivation thus, threatening food security [7, 8]. Many regions have experienced the detrimental and severe effects of drought, particularly, populations in the developing countries. In the Southern African Development Community (SADC) region; poor rainfall conditions were recorded for the 2016/2017 agricultural season as a result of El Niño induced drought [9]. FAO's global information and early warning system in 2015 reported significant drought dating back to 1984 [10]. The area data covered regions such as the United States, Semi-Arido of Brazil, Eastern Europe and African countries where, severe drought causing food crisis across Ethiopia, Kenya and Somalia resulted into the deaths of over 1 million people. Therefore, the continuing drop into below-normal annual rainfalls and increasing temperatures create the relevance to study and understand the morphological/anatomical changes that plants undergo to cope with environmental stresses. In cultivated crops such as soybean (*Glycine max* L.), this would minimise limitations that adversely affect plant growth, and the improvement of this crop for yield purposes [11], as well as counteracting against factors that negatively influence the nutritional

level, currently estimated at about

caused by climate change. The increase in atmospheric CO2

10 Plant, Abiotic Stress and Responses to Climate Change

content and essential secondary metabolites synthesised in this plant.

**2. Analyses of soybean responses to water deficit stress**

Plants experience water deficit stress when the amount of water in the cells and surrounding becomes limiting to growth and development. To investigate these effects, a study was conducted to primarily assess the influence of water stress on the growth of soybean; morphologically and anatomically, under greenhouse conditions. According to Lisar et al. [8] water deficit is caused by prolonged water shortage. In order to examine this stress, reduction in the frequency of irrigation was performed by limiting watering to once a week (WT 1) and once Plants are responsible for a number of essential ecological services. Plants are the main primary source of foods for humans and animals, supply oxygen, timber, medicine and also have ornamental value. The multiple and complex processes involving genetic, morphological, anatomical, physiological and biochemical mechanisms are responsible for the goods and services that plants provide. These functions are made possible by the architecture of the plant's internal and external structures. Soybeans like other legumes and non-leguminous plants display different types of internal and external growth forms that functions together to provide these services. The external form include indeterminate, determinate and semideterminate morphological growth habits, which typically take place in both the early and late maturity groups of varieties grown for commercial and subsistence farming [15]. Soybean plants with determinate growth terminate their vegetative growth stage during the onset of the reproductive stage. In contrast, indeterminate varieties continue growing even during flower setting and anthesis. Anthesis is the period in which flowers developed during the reproductive stage of the plant's life cycle begin to open. According to the NDSU [15] the semi-determinate growth habit lies between the polarity and growth of the other two growth habits (determinate and indeterminate form). The vegetative parts of soybean include the stem, leaves and the soil submerged roots. A few types of leaves can be found in soybean. The plant has trifoliate leaves, which are photosynthetic foliage with three leaflets. They have protective scale leaves which covers and protect young immature flowers before anthesis.

outermost single layer of cells derived from the protoderm, and in soybean it covers the plant for its entire life cycle. The three main types of epidermal cells found in soybean include trichomes and microscopic guard cells as well as the subsidiary cells of the stomata (**Figure 2b**–**d**). This layer of elongated and compactly arranged cells functions to protect soybean against water loss and harsh external environmental factors, including pathogens. Trichomes are unicellular or multicellular hairs occurring on shoot system of plants. On the roots, hairs are called root hairs. In leaves, this layer of cells is followed by the palisade parenchyma and spongy mesophylls.

Water Stress: Morphological and Anatomical Changes in Soybean (*Glycine max* L.) Plants

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13

**Figure 2.** Examples of microscopic cross-section in roots and stems of soybean plants. (a) Formation of pith canal as a result of water stress in WT 1 plants. (b) Broadening of canal and changes on stem cortex tissue in WT 2. (c) Control plants showing unaffected pith and cortex. (d) Cross-section of WT 2 root showing rupturing of the stele, protoxylem (PX) and metaxylem (MX). (e) Cross-section of WT 1 root showing marks of lateral roots (left right arrow). (f) Root section taken from the control showing expanded thickened xylem tissue and reduced cortex. (g) Close view of xylem tissue from the control plant. (h) Close view of parenchymatous pith as indicated on (c), (arrows indicate intercellular spaces of the parenchyma. (i) Soybean cortical tissue of the stem showing phloem fibres (left right arrow), collenchyma

(solid arrow) and a single layer of epidermis (dashed arrow).

These scales are small bracts which appear subtending the yellow or purple flowers of soybeans [16]. The special leaf types constituting the floral parts or inflorescence (raceme) can also be found. The vegetative stage is furthermore characterised by erect elongated stems, axillary buds, some viewed immediately above the cotyledons at the axil, unifoliate buds and the terminal buds (**Figure 1a–c**). Both young and old stems of soybeans are heavily covered by the epidermal hairs (trichomes) (**Figure 1d**). Even though soybean plants produce primary roots, originating from the seedling's embryo; the roots have many branching secondary roots that slightly resemble fibrous root system in monocots. Most of the lateral roots are concentrated at the upper part of the root zone. As in most of the dicotyledonous plants, soybean's body is made up of the three main tissue systems: dermal, ground and vascular (**Figure 1e**, **f**). The epidermis as the dermal tissue is the

**Figure 1.** Overview of soybean plant morphology and anatomy. (a) Vegetative first trifoliate (V1) stage. (b) Example of cotyledons and axillary buds at the axil. (c) Trifoliate leaves showing adaxial-abaxial leaf surfaces. (d) Example of soybean stem with epidermal hairs. (e) A micrograph of soybean stem cross-section. (f) A micrograph of soybean root cross-section.

outermost single layer of cells derived from the protoderm, and in soybean it covers the plant for its entire life cycle. The three main types of epidermal cells found in soybean include trichomes and microscopic guard cells as well as the subsidiary cells of the stomata (**Figure 2b**–**d**). This layer of elongated and compactly arranged cells functions to protect soybean against water loss and harsh external environmental factors, including pathogens. Trichomes are unicellular or multicellular hairs occurring on shoot system of plants. On the roots, hairs are called root hairs. In leaves, this layer of cells is followed by the palisade parenchyma and spongy mesophylls.

The plant has trifoliate leaves, which are photosynthetic foliage with three leaflets. They have protective scale leaves which covers and protect young immature flowers before anthesis.

12 Plant, Abiotic Stress and Responses to Climate Change

These scales are small bracts which appear subtending the yellow or purple flowers of soybeans [16]. The special leaf types constituting the floral parts or inflorescence (raceme) can also be found. The vegetative stage is furthermore characterised by erect elongated stems, axillary buds, some viewed immediately above the cotyledons at the axil, unifoliate buds and the terminal buds (**Figure 1a–c**). Both young and old stems of soybeans are heavily covered by the epidermal hairs (trichomes) (**Figure 1d**). Even though soybean plants produce primary roots, originating from the seedling's embryo; the roots have many branching secondary roots that slightly resemble fibrous root system in monocots. Most of the lateral roots are concentrated at the upper part of the root zone. As in most of the dicotyledonous plants, soybean's body is made up of the three main tissue systems: dermal, ground and vascular (**Figure 1e**, **f**). The epidermis as the dermal tissue is the

**Figure 1.** Overview of soybean plant morphology and anatomy. (a) Vegetative first trifoliate (V1) stage. (b) Example of cotyledons and axillary buds at the axil. (c) Trifoliate leaves showing adaxial-abaxial leaf surfaces. (d) Example of soybean stem with epidermal hairs. (e) A micrograph of soybean stem cross-section. (f) A micrograph of soybean root

cross-section.

**Figure 2.** Examples of microscopic cross-section in roots and stems of soybean plants. (a) Formation of pith canal as a result of water stress in WT 1 plants. (b) Broadening of canal and changes on stem cortex tissue in WT 2. (c) Control plants showing unaffected pith and cortex. (d) Cross-section of WT 2 root showing rupturing of the stele, protoxylem (PX) and metaxylem (MX). (e) Cross-section of WT 1 root showing marks of lateral roots (left right arrow). (f) Root section taken from the control showing expanded thickened xylem tissue and reduced cortex. (g) Close view of xylem tissue from the control plant. (h) Close view of parenchymatous pith as indicated on (c), (arrows indicate intercellular spaces of the parenchyma. (i) Soybean cortical tissue of the stem showing phloem fibres (left right arrow), collenchyma (solid arrow) and a single layer of epidermis (dashed arrow).

The palisade and spongy cells are specialised tissues used by all eudicot plants with C3 pathway for photosynthesis and gaseous exchange in leaves, respectively [17]. Soybean is one of the C3 plants which undergo photosynthetic carbon reduction and do not have a CO2 concentrating mechanism. It differs with grain crops such as maize, rice, sorghum and wheat C<sup>4</sup> plants that concentrate CO2 by not salvaging carbon lost during photorespiratory carbon oxidation (PCO) cycle [17]. But, the palisade and spongy tissues of soybean form the mesophyll, a ground tissue system of a leaf, which plays a critical role in carboxylation, reduction and regeneration processes during photosynthesis. In roots and stems, the fundamental (ground) tissue consists of non-protective and non-conductive simple cells of parenchyma, collenchyma and sclerenchyma (**Figure 2e** and **f**). Evert and Eichhorn [18] referred to this tissue system as the one most dominated by parenchyma cells, which are by far considered the most common ground tissue of the pith and cortex in roots and stems of soybean and other eudicots, as well as in the monocots. The vascular system is made up of conducting strands of phloem and xylem. These are principal water and food conducting tissue in all vascular seedless and seed plants.

#### **3.1. Morphological changes due to water deficit stress**

The morphological evidence gathered in this study has shown that soybean growth is highly sensitive to water deficit stress. All plants exposed to water deficit presented significant changes in their shoot and root morphology. Complete reduction in the number of new branches per plant, initiation of leaves and expansion of the lamina (measured by estimated leaf area) and the number of trifoliate leaves per plant was observed. Decreases in the assessed morphological characteristics were more predominant in plants subjected to stress for longer periods (WT 2) than those watered once a week (WT 1). Soybean cultivar Dundee, TGx 1740-2F, TGx 1835-10E and Peking produced significantly similar mean number of trifoliate leaves (about 4.0–5.0) in WT 2, when compared to about 5.0–6.0 trifoliate leaves obtained in WT 1 (**Table 1**). Leaf rolling and flipping were observed in some of the older leaves as a result of induced water stress. The negative effects of water stress on new leaf and branch formation was also reported by Mabulwana [16]. Jaleel et al. [19] similarly added that, water stress decreases the elongation and expansion of stems and leaves. In contrast to observations made in all water stressed plants, the control exhibited normal shoot growth and the highest number of trifoliate leaves (**Table 1**).

According to Nosalewicz and Lipiec [20] suppression on the growth and distribution of the roots by water stress could also lead to the reduction in shoot growth. As the vegetative shoot growths appeared diminished by induced stress, roots in water-stressed plants became more elongated and branched than in the control. Root phenotype in the control appeared shallow and less branched than in WT 1 and WT 2 plants. However, plants which had irrigation reduced to once in 15 days (WT 2) had deep root phenotype compared to plants irrigated once a week (WT 1). Insufficient water supply for WT 2 plants with deep root development, and moderately stressed plants (WT 1), both demonstrated clear morphological changes. All cultivars in WT 2 also exhibited severe nutrient deficiency symptoms (the entire leaf with chlorosis and marginal necrosis) and stem wilting. These symptoms were accompanied by adverse growth effects and survival frequency of 0% when the experiment was terminated (**Table 2**). Water deficit stress ultimately led to the severe damage to shoots of WT 2 plants, with no possible indication of recovery. In WT 1 plants, moderate to severe deficiency symptoms

**Soybean** 

**Treatment plants 1**

**Treatment plants 2**

**Control plants**

**genotypes**

**Mean no. of** 

**Average** 

**Stomatal** 

**Mean no. of** 

**Average leaf** 

**Stomatal** 

**Mean no. of** 

**Average** 

**Stomatal density (no.** 

**leaf area** 

**of stomata/ cm2**

**)**

**fully developed** 

**trifoliate leaves**

**(cm2**

**)**

**area (cm2**

**)**

**density (no. of** 

**stomata/cm2**

**)**

**fully developed** 

**leaf area** 

**density (no. of** 

**fully developed** 

**trifoliate leaves**

Dundee

LS 677 LS 678 Peking

TGx

6.0c

37.5e

154e

5.0a

40.1d

163e

12.5e

16.1e

163e

1740-2F

TGx

6.0c

60.1f

167f The leaf area of central individual leaflets in soybean cultivars were estimated using the general Eq. LA

by linear regression forcing the regression intercepting line to be zero using Table Curve software (Richter et

Values within columns followed by different alphabets are statistically significant at p

days), Water Treatment 2 (WT 2); reduced to once in 15

reduced to once a week (After 7

**Table 1.** experiment.

4.0e

30.7e

155f

 = k ×

 ≤

Developmental patterns in the leaves of water stressed and unstressed soybean plants measured immediately after the termination of the water deficit stress

11.0f

39.8f (L.W) where LA, leaf area; k, is the 'adjustment factor' estimated

al. [14]), L, length of the leaflet and W, leaflet width.

0.05 confidence level. For Water Treatment 1 (WT 1), irrigation frequency was

days and the Control, watering depended upon moisture availability in the soil.

Water Stress: Morphological and Anatomical Changes in Soybean (*Glycine max* L.) Plants

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15

171f

1835-10E

8.0d

43.9d

181d

4.5d

36.5c

143d

15.5d

33.6d

256d

6.0c

40.1c

203c

6.0c

32.1b

167c

13.5c

57.7c

212c

7.0b

38.6b

191b

6.5b

32.1b

106b

14.5b

41.1b

213b

5.0a

55.1a

213a

5.0a

55.1a

112a

13.0a

50.0a

247a

**(cm2**

**)**

**stomata/ cm2**

**)**

**trifoliate leaves**


The palisade and spongy cells are specialised tissues used by all eudicot plants with C3

plants which undergo photosynthetic carbon reduction and do not have a CO2

principal water and food conducting tissue in all vascular seedless and seed plants.

**3.1. Morphological changes due to water deficit stress**

ing mechanism. It differs with grain crops such as maize, rice, sorghum and wheat C<sup>4</sup>

C3

that concentrate CO2

14 Plant, Abiotic Stress and Responses to Climate Change

way for photosynthesis and gaseous exchange in leaves, respectively [17]. Soybean is one of the

(PCO) cycle [17]. But, the palisade and spongy tissues of soybean form the mesophyll, a ground tissue system of a leaf, which plays a critical role in carboxylation, reduction and regeneration processes during photosynthesis. In roots and stems, the fundamental (ground) tissue consists of non-protective and non-conductive simple cells of parenchyma, collenchyma and sclerenchyma (**Figure 2e** and **f**). Evert and Eichhorn [18] referred to this tissue system as the one most dominated by parenchyma cells, which are by far considered the most common ground tissue of the pith and cortex in roots and stems of soybean and other eudicots, as well as in the monocots. The vascular system is made up of conducting strands of phloem and xylem. These are

The morphological evidence gathered in this study has shown that soybean growth is highly sensitive to water deficit stress. All plants exposed to water deficit presented significant changes in their shoot and root morphology. Complete reduction in the number of new branches per plant, initiation of leaves and expansion of the lamina (measured by estimated leaf area) and the number of trifoliate leaves per plant was observed. Decreases in the assessed morphological characteristics were more predominant in plants subjected to stress for longer periods (WT 2) than those watered once a week (WT 1). Soybean cultivar Dundee, TGx 1740-2F, TGx 1835-10E and Peking produced significantly similar mean number of trifoliate leaves (about 4.0–5.0) in WT 2, when compared to about 5.0–6.0 trifoliate leaves obtained in WT 1 (**Table 1**). Leaf rolling and flipping were observed in some of the older leaves as a result of induced water stress. The negative effects of water stress on new leaf and branch formation was also reported by Mabulwana [16]. Jaleel et al. [19] similarly added that, water stress decreases the elongation and expansion of stems and leaves. In contrast to observations made in all water stressed plants, the control exhibited normal shoot growth and the highest number of trifoliate leaves (**Table 1**).

According to Nosalewicz and Lipiec [20] suppression on the growth and distribution of the roots by water stress could also lead to the reduction in shoot growth. As the vegetative shoot growths appeared diminished by induced stress, roots in water-stressed plants became more elongated and branched than in the control. Root phenotype in the control appeared shallow and less branched than in WT 1 and WT 2 plants. However, plants which had irrigation reduced to once in 15 days (WT 2) had deep root phenotype compared to plants irrigated once a week (WT 1). Insufficient water supply for WT 2 plants with deep root development, and moderately stressed plants (WT 1), both demonstrated clear morphological changes. All cultivars in WT 2 also exhibited severe nutrient deficiency symptoms (the entire leaf with chlorosis and marginal necrosis) and stem wilting. These symptoms were accompanied by adverse growth effects and survival frequency of 0% when the experiment was terminated (**Table 2**). Water deficit stress ultimately led to the severe damage to shoots of WT 2 plants, with no possible indication of recovery. In WT 1 plants, moderate to severe deficiency symptoms

by not salvaging carbon lost during photorespiratory carbon oxidation

path-

plants

concentrat-

**Table 1.** Developmental patterns in the leaves of water stressed and unstressed soybean plants measured immediately after the termination of the water deficit stress experiment.


canals, which are usually formed in woody shrubs and trees. They are formed when the earliest vascular tissues, protoxylem, is destroyed by the formation of new metaxylem as the root or stem grows in diameter. In gymnosperms, these canals are instead used by the pine trees to store resin and they are more associated with the cortical tissue of the stems than the pith [25]. In stems of plants such as seedless vascular plants (horsetails), these canals are naturally formed to reduce the weight of the stem thus, increasing stem strength and resistance to buckling [18]. However, the formation of canals (breaking down of the soybean pith tissue) observed in roots and stems may have resulted from water stress. Furthermore, this may have possibly impacted negatively on the growth of plants, particularly when induced as a result

Water Stress: Morphological and Anatomical Changes in Soybean (*Glycine max* L.) Plants

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17

Even though the pith is poor in nutrients [26], the parenchyma cells can still function in storage of nutrients and water for the plant. Pallardy [27] suggested that, rapturing could also destroy the interconnectivity between the storage parenchyma of the pith with the cortex, disrupting short distance transport that occurs through the rays via secondary xylem. The variations in canal diameters between WT 1 and WT 2 (**Figure 2a**, **b**, including canal in the root- **d**) may be in response to the different water stress regimes or the genotype variability of the soybeans used. Canal diameters in soybean WT 2 plants were larger than the diameter observed in WT 1 plants (**Figure 2a**, **b**). Soybean cultivar LS 677 and LS 678 showed little resistance to the rapturing of the pith, compared to cultivar Peking and Dundee. This was the case, even though cultivar LS 677 and Peking were the only varieties more resistant to water stress treatment (WT 1). This could be both a genetically-linked response and the reaction or effects of water stress conditions to the tissue development. In cultivar Peking, TGx 1740-2F and TGx 1835-10E, pith canals appeared to be continuously cut from the central pith further to the cortical cells. This induced complete disruption of water transportation through some part of the xylems, xylem rays and nutrient transport by the phloem tissues. The cutting of water supply may have resulted in the poor survival rates observed in most of the cultivars (**Table 2**). But, the absence of pith canals in stems of the control plants furthermore suggests a relationship between water deficit and the change in anatomy of the soybean plants. When the imposed environmental stress reduced the

rate of tissue development, the length of xylem rays in roots was also reduced.

(ABA) on shoot and root growths during salinity and drought stress.

The reduction occurred when growth is affected by death of tissues and slowing down of metabolism as a result of the stress. Alteration in plant metabolism affect cell division, thus cell elongation and expansion is negatively affected as evidenced in **Figure 2d**, **e**). The xylem cell portion in the roots of water stressed plants was reduced compared to xylem tissue diameter in the control. Yamaguchi and Sharp [28] indicated that, water stress induce changes in root growth and cell length distribution which may be directly related to growth inhibition in roots, especially at root elongation zones. Another example is by Schuppler et al. [6] who also indicated the reduction on mitotic activity of mesophyll tissues in wheat (*Triticum aestivum*) seedlings subjected to mild water deficit. These reports indicate that, the lack of adequate water supply decreases the rate of cell division and tissue expansion in all plant organs, although root morphology may appear less affected in contrast to root anatomy. Munns and Sharp [29] made similar remarks following their investigation on the effect of abscisic acid

of severe water stress, like in WT 2.

**Percentage survival frequency** was calculated from the number of plants/ genotype that survive until the termination of the water stress deficit experiment.

Statistical significance among the values is designated by different superscript letters. Values within columns showing different letters are statistically varied (at 0.05) by ANOVA.

**Table 2.** Vegetative growth and flowering response of soybean plants subjected to water deficit stress conditions.

were observed. Soybean cultivar LS 677 and Peking showed some resistance with 15 and 30% survival rate (**Table 2**). A few plants in these two genotypes exhibited moderate stress effects among all the cultivars assessed. There were no differences in the lengths of root system and shoots observed in water stress resistant cultivars (LS 677 and Peking) in comparison with those severely affected (Dundee, LS 678, TGx 1740-2F and TGx 1835-10E) in both WT 1 and WT 2 plants.

Klamkowski and Treder [21] reported almost similar results in water stressed strawberry plants. In addition, there were no major differences, especially in root lengths that were observed between water stressed plants and the control. The report cited inhibition of growth by water stressed plants, involving decrease in root expansion as suggested by Boyer [22]. This claim probably led to the observed root phenotype in water stressed strawberry plants. This is in contrast with finding in this study and most of the other suggestions made on root phenotypes during water stress. In general, root formation has been found to increase in length during water stress, with roots growing deep into the soil in search for moisture [17, 19, 23, 24]. This further development in the root system is an attempt by plants to increase the uptake of water in order to sustain growth as observed in this study.

#### **3.2. Anatomical changes in response to water stress**

The WT 1 and WT 2 plants demonstrated a different internal anatomy compared to the control plants. Stem cortex of water stressed plant were generally smaller compared to the cortex in stems of plants in the control (**Figure 2**). However, vascular tissue thickening and expansion was observed in both the roots and stems of water stressed and control plants. The development of the secondary tissues in water stressed plants, especially the deposition of secondary xylem cells (as viewed in **Figure 2a**, **b**), was interrupted by the gradual rapturing of the pith which resulted in the formation of pith canals. Pith canals are hollow centres, called central canals, which are usually formed in woody shrubs and trees. They are formed when the earliest vascular tissues, protoxylem, is destroyed by the formation of new metaxylem as the root or stem grows in diameter. In gymnosperms, these canals are instead used by the pine trees to store resin and they are more associated with the cortical tissue of the stems than the pith [25]. In stems of plants such as seedless vascular plants (horsetails), these canals are naturally formed to reduce the weight of the stem thus, increasing stem strength and resistance to buckling [18]. However, the formation of canals (breaking down of the soybean pith tissue) observed in roots and stems may have resulted from water stress. Furthermore, this may have possibly impacted negatively on the growth of plants, particularly when induced as a result of severe water stress, like in WT 2.

Even though the pith is poor in nutrients [26], the parenchyma cells can still function in storage of nutrients and water for the plant. Pallardy [27] suggested that, rapturing could also destroy the interconnectivity between the storage parenchyma of the pith with the cortex, disrupting short distance transport that occurs through the rays via secondary xylem. The variations in canal diameters between WT 1 and WT 2 (**Figure 2a**, **b**, including canal in the root- **d**) may be in response to the different water stress regimes or the genotype variability of the soybeans used. Canal diameters in soybean WT 2 plants were larger than the diameter observed in WT 1 plants (**Figure 2a**, **b**). Soybean cultivar LS 677 and LS 678 showed little resistance to the rapturing of the pith, compared to cultivar Peking and Dundee. This was the case, even though cultivar LS 677 and Peking were the only varieties more resistant to water stress treatment (WT 1). This could be both a genetically-linked response and the reaction or effects of water stress conditions to the tissue development. In cultivar Peking, TGx 1740-2F and TGx 1835-10E, pith canals appeared to be continuously cut from the central pith further to the cortical cells. This induced complete disruption of water transportation through some part of the xylems, xylem rays and nutrient transport by the phloem tissues. The cutting of water supply may have resulted in the poor survival rates observed in most of the cultivars (**Table 2**). But, the absence of pith canals in stems of the control plants furthermore suggests a relationship between water deficit and the change in anatomy of the soybean plants. When the imposed environmental stress reduced the rate of tissue development, the length of xylem rays in roots was also reduced.

were observed. Soybean cultivar LS 677 and Peking showed some resistance with 15 and 30% survival rate (**Table 2**). A few plants in these two genotypes exhibited moderate stress effects among all the cultivars assessed. There were no differences in the lengths of root system and shoots observed in water stress resistant cultivars (LS 677 and Peking) in comparison with those severely affected (Dundee, LS 678, TGx 1740-2F and TGx 1835-10E) in both WT 1 and WT 2 plants.

Statistical significance among the values is designated by different superscript letters. Values within columns showing

**Table 2.** Vegetative growth and flowering response of soybean plants subjected to water deficit stress conditions.

Klamkowski and Treder [21] reported almost similar results in water stressed strawberry plants. In addition, there were no major differences, especially in root lengths that were observed between water stressed plants and the control. The report cited inhibition of growth by water stressed plants, involving decrease in root expansion as suggested by Boyer [22]. This claim probably led to the observed root phenotype in water stressed strawberry plants. This is in contrast with finding in this study and most of the other suggestions made on root phenotypes during water stress. In general, root formation has been found to increase in length during water stress, with roots growing deep into the soil in search for moisture [17, 19, 23, 24]. This further development in the root system is an attempt by plants to increase

The WT 1 and WT 2 plants demonstrated a different internal anatomy compared to the control plants. Stem cortex of water stressed plant were generally smaller compared to the cortex in stems of plants in the control (**Figure 2**). However, vascular tissue thickening and expansion was observed in both the roots and stems of water stressed and control plants. The development of the secondary tissues in water stressed plants, especially the deposition of secondary xylem cells (as viewed in **Figure 2a**, **b**), was interrupted by the gradual rapturing of the pith which resulted in the formation of pith canals. Pith canals are hollow centres, called central

the uptake of water in order to sustain growth as observed in this study.

**3.2. Anatomical changes in response to water stress**

**Soybean genotypes** **Mean plant height** 

16 Plant, Abiotic Stress and Responses to Climate Change

**Mean no. of branches**

**Flowering plants (%)**

**TP 1 TP 2 TP 1 TP 2 TP 1 TP 2 TP 1 TP 2 TP 1 TP 2**

Dundee 25.2a 24.1a 3.0a 3.0a — — — — — — LS 678 40.0b 26.4b 3.0a 3.0a — — — — — — LS 677 33.3c 26.5b 4.0b 3.0a 1.00a — 7.0a — 15.0a — Peking 24.2d 23.5a 4.0b 2.0b 15.0b — 3.0b — 30.0b — TGx 1740-2F 27.3e 21.0c 2.0c 3.0a — — — — — — TGx 1835-10E 26.1f 20.7d 3.0a 3.0a — — — — — — **Percentage survival frequency** was calculated from the number of plants/ genotype that survive until the termination

**Mean no. of pods produced**

**Survival frequency** 

**(%)**

**(cm)**

of the water stress deficit experiment.

different letters are statistically varied (at 0.05) by ANOVA.

The reduction occurred when growth is affected by death of tissues and slowing down of metabolism as a result of the stress. Alteration in plant metabolism affect cell division, thus cell elongation and expansion is negatively affected as evidenced in **Figure 2d**, **e**). The xylem cell portion in the roots of water stressed plants was reduced compared to xylem tissue diameter in the control. Yamaguchi and Sharp [28] indicated that, water stress induce changes in root growth and cell length distribution which may be directly related to growth inhibition in roots, especially at root elongation zones. Another example is by Schuppler et al. [6] who also indicated the reduction on mitotic activity of mesophyll tissues in wheat (*Triticum aestivum*) seedlings subjected to mild water deficit. These reports indicate that, the lack of adequate water supply decreases the rate of cell division and tissue expansion in all plant organs, although root morphology may appear less affected in contrast to root anatomy. Munns and Sharp [29] made similar remarks following their investigation on the effect of abscisic acid (ABA) on shoot and root growths during salinity and drought stress.
