the effects of sodium sulphate and sodium chloride on growth, morphology

8/9/2019 The effects of sodium sulphate and sodium chloride on growth, morphology

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The effects of sodium sulphate and sodium chloride on growth morphology

photosynthesis and water use efficiency of

henopodium rubrum

Plant Physiology Research Group Department of Biological Sciences University of Calgary Calgary Alta.

Canada N IN4

Received June 14, 1989

W A R N E , ., G U Y ,R . D . ,

ROLLINS,

. , and REI D, D. M. 1990. The effects of sodium sulphate and sodium chloride on

growth, morphology, photosynthesis, and water use efficiency of

Chenopodium rubrum.

Can. J Bot. 68: 999-1006.

The effects of sodium sulphate and sodium chloride salinity on the anatomy, water relations, and photosynthesis of Chen-

opodium rubrum L. were compared. Low concentrations of either salt stimulated growth, but higher concentrations resulted

in large decreases in dry weight and leaf area. Leaf succulence and the number of layers of palisade cells were increased,

but these effects were more pronounced with NaCl than with N S0 4. Stomatal density was reduced at low to moderate

salinities, but then increased again at high salinity. Stomatal size was reduced at all salinities. Increasing salinity had no

great effect on photosynthetic rates except with older plants grown at the highest level of Na'SO,. Stomatal cond uctan ce

decreased at all salinities. This reduced transpiration and led to increased intrinsic water use efficiency. Total tissue stable

carbon isotope ratios also indicated that water use efficiency was improved.

Chenopodium rubrum

adjusted osmotically by

accumulating electrolytes from the nutrient solution and by synthesizing glycinebetaine. Plants in NaCl limited osmotic

adjustment more than those growing in Na2 S0,. Despite this, N S0 4 was more dama ging than NaCl and caused earlier leaf

senescence at high concentrations.

W A R N E , ., G U Y ,R . D . ,

ROLLINS,

. , et REID,D. M . 1990. The effects of sodium sulphate and sodium chloride on growth,

morphology, photosynthesis, and water use efficiency of Chetiopodiurn rubrum. a n . . B ot . 6 8 9 99 -1 00 6.

Les effets de la salinitt du sulfate de sodium et du chlorure de sodium sur I'anatomie, les relations hydriques et la

photosynthkse d u

Chenopodium rubrum

sont compares. Les basses concentrations de I'un ou I'autre sel stirnulent la crois-

sance , mais les concentrations plus Clevtes causent de s diminutions importantes de la masse sec he et de la surface des feuilles.

La succulence des feuil les et le nom bre de couches d e cellules palissadiques augm entent, mais ces effets sont plus pr onon cts

avec NaCl q u'avec Na 2S0 4. La den sit t stomatique est r tdu ite aux salinites basses B modtrtes, mais, par la suite, augmente

B forte salinitt. La dimension des stomates est rtdu ite toutes les salinitts. L'augm entation de la salinitt n'a pa s d'effet

important sur les taux de photosynthkse sauf chez les plantes plus igtes au niveau le plus ClevC de NaZSO,. La conductance

stomatique dim inue B toutes les salinitCs. Ceci rtd uit la transpiration et conduit une augmentation d e I'efficacitC de l 'uti-

lisation d e I'eau. Les ratios en isotope de carbone stable des tissus indiquent aussi que I'effic acitt d e I'utilisation de I'eau

est amCliorte.

Cherlopodium rubrum

s'ajuste osmotiquement par I'accumulation des Clectrolytes de la solution nutritive et

par la synthkse de glycinebetaine. Les plantes dans la solution de NaCl limitent I'ajustement osmotique plus que celles en

prtsen ce de Na2 S04. En dtp it de ce fai t , Na,S04 est plus domm ageable que NaCl et cause une stnescenc e foliaire plus

hitive aux concentrations Clevtes.

[Traduit par la revue]

ntroduction

Within the Chen opodiaceae are found many species that are

highly tolerant to salinity. Typically, these species are able to

survive at soil osmotic potentials below . 5 MPa (Sharma

1 9 8 2 ) .

The exposure of halophytes to salinity often induces

alterations in morphology and (or) anatomy (Poljakoff-Mayber

1 9 7 5 ) . Less apparent effects also occur at the physiological

level. Several chenopods accumulate large quantities of inor-

ganic ions in their leaf tissues as soil salt concentrations

increase. This is usually accompanied by the accumulation of

glycinebetaine. Though low or moderate salinity may stimu-

late growth, high salinity results in decreased growth. How-

ever, the photosynthetic rate of many chenopods is only

slightly affected by chronic salinity stress. Many chenopods

also have low er transpiration rates compared w ith other plants

and may therefore use water more efficiently.

Chenopodiurn rubrum is one of several halophytic cheno-

pods found on saline soils in western Canada. Many of these

soils contain a high proportion of sodium sulphate (Dodd et

al. 1 9 6 4 ; Lilley 1 9 8 2 ) . Very little information is available

'Present address: Department of Forest Sciences, The University

of British Columbia, Vancouver, B .C ., Canada V6T 1 W5.

'Author to whom correspondence should be addressed.

regarding the effects of Na2S0, salinity on various aspects o

plant growth, with most studies utilizing only sodium chloride

This paper compares the effects of both Na2S0, and NaCl o

grow th, succulence, stomata1 size and nu mb er, accumulatio

of glycinebetaine, water relations, intrinsic water use effi

ciency, and photosynthetic capacity of C . rubrum.

Methods and materials

Seeds of

C. rubrum

L. (obtained from several plants that in tur

originated from seed collected near Nanton, Alberta) were scarifie

and sown on to 4-inch (40.8-cm ) pots contain ing granite grit No.

(Imasco). Pots were suspended by their rims from Lucite covers i

deep plastic trays (six pots per tray) containing 5 L of nutrient solu

tion. Th e lower cm of each pot was submerged in the solution

which was maintained at constant level by the automatic addition o

deionized water. Solutions were continuously aerated to keep them

well mixed and to prevent anaerobic conditions. Nutrient solution

consisted of half-strength Hoagland's solution (Hoagland and Amo

1950) with salt (either NaCl or anhy drou s Na,S04 ACS certified

added to obtain the solute potentials desired in any particular exper

iment. Media solute potentials (MSP) were confirmed by psychro

metry. Th e solute potential of half-strength Hoa gland 's solution

about -0. 03 MPa. For our purposes, this small contribution is n

included in further references to MSP. Plants were allowed to adju

to increased salinities by incrementally dropping MSP by no mor

Printcd in anada Imprim e au anada


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1000 CAN. J. BOT.

than -0 .2 MPa every 2 days. Solutions were changed weekly after

maximum salt concentrations were reached. Plants were maintained

at the final salinities for at least 2 weeks prior to harvesting.

When an Econaire (Winnipeg) growth chamber was used,

daymight

temperatures were 22:12 C. The light source was Sylvania gro-lux

lamps, which provided an irradiance (PAR) of 313 IJ.Em-' s- at

plant height. Irradiance was increased to 483 IJ.Em-' s- ' for the

photosynthesis experiments. Photoperiod was 16 h, and humidity was

46% during the day and 75% at night. Treatments were represented

by only a single tray of pots each, but experiments were repeated

where warranted and tray positions within the growth chamber were

randomized each time.

In experiments performed in the greenho use, irradiance ranged from

about 220 IJ.Em -Z s- on overcast days to about 1250 IJ.E n-, s-

on sunny days. Relative humidity was variable, ranging from 30 to

40% during the day and up to 70% at night. Temperature varied from

approximately 23C on overcast days to 33OC on sunny days and

dropped to a minimum of 14C at night. All treatments were repre-

sented by on e tray in each of two randomized block s (i.e., 12 plants

per treatment).

Unless otherwise noted, measurements of foliar characteristics refer

to the leaf from the 7th node up from the base of the plant. Succu lence

(water content per unit leaf area) was measured by cutting leaf disks

of equal area with a cork borer and determining the difference between

fresh and dry weights. One disk was cut from near the middle of the

7th leaf from each of six plants per treatment. Major veins were

avoided. Changes in the size and number of layers of leaf cells were

determined by exam ining leaf cross-sections under a microscope. Leaf

samples were taken near the centre of the leaf and fixed in 3% glu-

taraldehyde buffered at pH 6.8 with 0.05 M phosphate buffer. After

dehydration, tissues were embedded in LKB Historesin and sections

cut with a glass knife on an LKB 2218 microtome. Mounted sections

were stained with periodic acid -Schiff's reagent (Feder and O'Brien

1968) and counterstained with toluidine blue

0

Leaf thickness was

estimated from freehand and cross-sections.

Numbers and sizes of stomata were determined from nail polish

leaf impressions (modified from Sampson, 1961) examined under a

Reichert projecting microscope (magnification 800 X . Stomata1 sizes

were measured lengthwise and classified into two categories: large

(>3 2 IJ.M) r small (


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WARNE ET AL. 100

TABLE

. Effect of NaCl and NqSO, salinity on leaf area (one surface), stornatal size, stornatal density, and number of stomata per leaf f

the 7th leaf up from the base on the main axis of C. ru rlitn

Stomata1 density

No. of stomata per leaf

Mean area Stornatal size

Solute potential

(no./mm2)

per leaf ( >32 km

X

10-7

Salt

MPa (cm')

long) Abaxial

Adaxial

Abaxial Adaxial

Control

0 14.97k 1.99 64.7 22 .3 152.9k 3.3

105.8k4.0

226.9k29.6 165.8221.6

NaCl .4 15.38k2.09 43.3?4.8 92. 0k 1.9 6 5. 2k 2. 0 147.0224.0 105.8k 18.0

1.0 10. 112 1.22 14.9 2.7 105.5?2.1 63. 9k 1.4 107.4k 14.2 66.5 k9 .6

-2.1 6.75 0.92 0.7 k0 .7

124.4k2.9 79.72 2.0

86. 8k 12.4 56.1 k8 .5

N S04 .4 19.04k2.41 41.2&7.7 104.7k 2.5 75. 92 2.6 200.3 22 7. 2 155.2225.4

1.0 9.1350.67 8.4 k2. 5 110.62 2.6 75.2 k 1.9 99.7k 6.9 68.2k5.1

-2.1 4.27 0.55 0 209.6 4. 131. 823. 2 86 .5 k 10.7 55 .4 27 .2

NOTE:

Each datum represents the mean + SE of 12 plants or, for effects on stomatal size (abaxial leaf surface only), six plants per treatment. For density determinations, stoma

were counted in five different areas on each leaf.

NaCl

MEDI A SOLUT E POT ENT I AL MPa)

FIG.2. Effects of NaCl or Na,SO, on leaf succulence. Data points

represent means k SE of six plants per treatment.

part to the development of larger cells in the palisade layer an

an increase in the number of cell layers. Leaves of contro

plants averaged 2.3

+

0.1 (SE) layers of palisade cell

whereas at MS P of -0 .8 MP a, this increased to 2.96 .0

layers with NaCl and to 2.89 + 0.1 with N S04. The spong

meso phyll lay er a136 increased in thicknes s in NaCl trea

ments, but very little increase was observed in Na,S0

treatments.

Stomatal numbers atid sizes

In view of the fact that the growth of plants declined as sa

concentrations increased, it seemed reasonable to investigat

the effects of salt on stomatal number and size, since an

changes would presumably influence photosynthetic carbo

gain. Stomatal densities on both abaxial and adaxial leaf sur

faces decreased at moderate salinities, but increased again a

higher salinities (Table 1). This trend was particularly eviden

in the Na,S0 4 treatment where density was actually higher

-2. 1 MPa than in the control treatment. However, there wa

a tendency towards larger leaves at low salinity before lea

area decreased again at higher salinities. T hes e trends appeare

to be at least partially independent, as evidenced by the fa

that their combined action resulted in a reduction in the tota

number of stomata per leaf across all salt treatments (Tab

1). A change in the size stomata was also observed (Table 1

In the absence of salt, 65 of stomata were greater than 3

p,m in length . Aperture length was reduced as M SP decreas ed

and O. l9

with

Na S04 (both

at

M P a )

The ratio

such that there were very

few

stomata of this size seen at .

decreased again

at MSP

.6 MPa'

The

low

MPa, Trends were virtually identical for both salts. Interes

irradiance within the growth cham ber (compared with the nat-

ural environment) might have prevented or precluded a more

ingly, at - 0. 4 MP a, stomatal size was reduced relative

substantial response. Hence, an experiment was conducted in

controls, even though stomatal density was lowe r and leaf are

the greenhouse to investigate whether o r not irradiance in com-

was greater.

bination with salinity would influence the expression of pros-

trate growth. No such effect was found.

smotica accumulation

Like many halophytic chenopods,

C.

rubrum readily accu

Succulence

Increasing salt concentrations had a significant effect on leaf

succulence (Fig. 2). Both salts caused substantial increases

from 0 to - 0. 8 MP a, but NaCl had a much greater effect

beyond this point. Leaf thickness also increased as MSP

decreased and again, NaCl had the greatest effect (data not

shown ). For examp le, leaf thickness at MS P of 1.0 MPa

was increased over controls by 50 in NaC1-treated plants and

by 21 in Na,S04-treated plants. This increase was due in

mulates large quantities of salt within its tissues. Ash conten

increased almost linearly with decreasing MSP, ranging fro

14 of leaf dry weight in controls to 40 at the highest sa

concentrations. Results were the same for both salts (data n

shown). A ccumulation of glycinebetaine was also similar wi

both Na,S04 and NaCl (Fig. 3). Data in Fig. 3 are presente

in terms of organic matter (i.e ., dry weight less the ash weigh

to correct for the bias caused by the high salt content of th

leaves. The glycinebetaine content of plants grown at MSP o


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1002 CAN.

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VOL. 68, 1990

M E DI A S O LU TE P O TE N TI AL M P ~ )

FIG.3. Leaf glycinebetaine content of C. rubrum grown in NaCl

and N% S04. Data points represent means SE of four plants per

treatment. Error bars not visible where smaller than the symbol size.

.1 MPa was about 420 pmol per gram of organic matter

(equivalent to 4.9% of the leaf biomass).

Water relations

To determine the extent and pattern of osmotic adjustment

in C. rubrum water and solute potentials of leaf tissue were

measured. Results presented in Fig. 4 show that as salt con-

centration increased, both

*,

and Ts ecreased and were

maintained below the MSP. Estimated leaf

Tp

emained rel-

atively constant in NaCl treatments, but increased with

increasing Na,SO, concentrations. As MSP below .4 MP a,

leaf 9 nd Ts ere lower in N% S0 4 treatments than in NaCl

treatments. For example, at 1.6 MPa Na2S0 4, the leaf Tw

was .84 MP a, whereas plants at .0 MPa NaCl had a

Tw f only .5 MP a. Superficially, then, it would appear

that plants grown in

N% S04 should have had an advantage

over those grown in NaC1. As already described, this was not

the case.

Photosynthetic CO, assimilation and related variables

Results already presented showed changes in stomatal num-

bers and length that, in addition to possible changes in stomatal

opening, could influence gas exchange. Under steady-state

conditions, A was about 22 pm ol CO, m-'s-' for plants

grown without salt (Fig. 5 . This rate decreased slightly at

-0 .4 MPa (for both salts) but increased again at 1 0 and

.0 MP a (NaCl) to a level similar to that of the controls. At

1.6 MPa N%SO,, however, the photosynthetic rate dropped

to approximately 10 pmol CO, m-'s ,perhaps reflecting the

poor condition of plants at this treatment level.

At all salt concentrations there was a significant decrease in

stomatal conductance (Fig. 6A) to a level approximately one-

third that of the con trols. Decreased conductance can limit the

rate of CO, assimilation. In C. rubrum however, was only

modestly reduced at most salt concentrations (the notable

MEDIA SOLUTE POTENTIAL

M P ~

FIG. 4

Wate r relations of C.

rubrum

leaves from 50-day-o ld plan

grown in N %S 04 and NaC1. Each data point represents the mean

S E of three to five samples per treatment. D ashed diagon al lines re

resent the isomotic limit, where leaf water or solute potent

equals media solute potential.

0

Y ;

?

9, ;Ys.

rror bars n

visible where smaller than the symbol size.

exception being the 1.6 MP a Na 2S0 4 (treatment). Cons

quently, CJC, ratios were reduced at all salinities other tha

the .6 MPa Na 2S 04 treatment (Fig. 6B). The fact that

was m uch lower (and yet the C i of plants from this treatme

did not differ from unsalinated controls) implies that photo

synthesis was somehow impaired at the biochemical leve

Trends in WU E (Fig. 6C) closely mirrored CJC,, as would

expected.

The photosynthesis experiments were repeated under ide

tical conditions with the on e modification that the plants we


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T

AL.

100

MED IA SOLU TE POTEN TIAL MPa)

FIG. 5 . Steady-state net p hotosyn thetic rates of leaves of 50-d ay-

old C. rubrum plants grown in NaCl and Ns SO ,. Data points rep-

resent means SE of six plants per treatment.

1 week youn ger at the time of measurem ent. Results were very

similar to those shown in Figs. 5 and 6 except that the effects

of the .6 MP a Na,SO, treatmen t on A conductance,

C,/C,, and water use efficiency we re not significantly differen t

from the

0

MPa NaCl treatment. The effects on CJC, are

presented in Table 2 Measurem ents were repeated 7 days later,

by which time the leaves of .6 MPa N S0 4 plants were

beginning to show visual damage. These plants were again

behaving as shown in Figs. 5 and 6. It thus appears that N S0 4

at high con centrations induces earlier leaf senescence that does

NaCl.

Carbon isotope analysis

Isotopic composition of C. rubrum leaf tissue is presented

in Fig. 7 . Genera lly, less negative 613C values were obta ined

with increasing salt concentrations. Plants grown with NaCl

appeared to have slightly greater changes in 613C than those

grown with N S0 4, but there was no significant difference

(by t-test) between the slopes of regression lines for both sets

of data.

iscussion

At low to moderate salinities, similar effects on growth of

C. rubrum were obtained with both NaCl and N S0 4. For

example, small amou nts of the salt promoted growth and higher

concentrations inhibited growth. Th is pattern is typical of many

halophytes. Morphological changes induced by the two salts

were also similar though different in degree. Hence, stomata1

density responded in a complex fashion, but trends were still

more or less the same for both NaCl and N S0 4. Leaf suc-

culence was increased with either salt, but m ore so with NaCl.

Similar observations were made by Poljakoff-Mayber (1975).

Increased succulence is frequently observed in halophytes

exposed to increasing salinity and various suggestions have

M ED IA S O LU T E P O TE N TI AL M P

FIG.6. Effects of increasing salt concentrations on various g

exchange parameters under steady-state conditions (same 50-day-o

plants as in Fig. 5).

(A)

Stomata1 conductance to diffusion of CO

(B) intercellular to ambient CO, ratio (CjC ,); (C) intrinsic water-u

efficiency (WU E). 0 aCL;

a

Na,SO,.


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1990

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25

M ED IA S O LU TE P O TE NT IA L M P ~ )

FIG.

7

Effect of increasing concentrations of NaCl or N SO, on the isotopic composition (613C value) of leaves of 50-day-old C .

rubrum

plants. Analyses were performed on pooled samples from six plants per treatment.

T A B L E

.

The effect of NaCl and Na,SO, salinity on inter-

cellular to ambient CO, ratio (C,/C,) for leaves from two

ages of C.

rubrurn

Solute potential C,/C, ratio

Salt MPa 43 days 50 days

NaCl 0

.4

1.0

.6

NOTE ns, not significantly different (by I-test) from the NaCl 0 MPa

treatment of the 43-day-old plants; , indicates significance at

P

the effects of sodium sulphate and sodium chloride on growth, morphology

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