Journal of Scientific & Industrial Research Vol. 64, June 2005, pp. 393-402
Citrus in terms of soil and water salinity: A review
Ashutosh A Murkute1, Satyawati Sharma1,* and S K Singh2
1Centre for Rural Development and Technology, Indian Institute of Technology Delhi, New Delhi110 016
2Division of Fruits and Horticultural Technology, Indian Agricultural Research Institute, Pusa, New Delhi 110 012 Received 03 September 2004; revised received 15 March 2005; accepted 24 May 2005
Citrus, one of the important fruit crops of the tropics and subtropics, is grouped as a glycophyte. The deleterious effects of salt stress lead to reduction in fruit yield and quality. The response of different citrus species to different salt(s) further brings differential responses, when fruit quality is concerned. The possible mechanisms (physical, nutritional, biochemical), which plants adapt to sustain salt stress, might provide an indication to plant breeders and biotechnologists to proceed further in crop improvement. The present study is an attempt to review the literature and explore the possible mechanisms of salt tolerance on sustainability of salinity by citrus. The role of endomycorrhizal fungi in citrus under salt stress is also discussed.
Keywords: Citrus, Salinity, Abiotic stress, Arbuscular Mycorrhizal fungi, Tissue culture
Introduction
In India, of the geographical area of about 329 m ha, the cultivated area is only 156 m ha. The per capita availability of land has declined from 0.89 ha in 1951 to 0.37 ha in 1991, and is projected to slide down to 0.19 ha in 20351. The sizable fertile areas are going out of cultivation due to salinity and sodicity. In India, 10.1 m ha area is reported to be salt affected2, where cultivation of salt tolerant plant species could be the best-suited alternative.
Citrus, one the most important fruit crops of India, forms an essential commercial commodity for several industries (Fig. 1). Apart from susceptible to several biotic stresses (insect pests, pathogens like Phytophthora root rot, citrus tristeza virus and nematodes), citrus is highly sensitive to soil salinity and pH. About 13 percent decrease of citrus yield per each 1 dS m-1 increasein salinity above 1.4 dS m-1, the threshold value of electrical conductivity of saturated soil extract, has been reported3. Chloride toxicity appears to be the main reason for reduction in its fruit yield although the possibility of osmotic component cannot be excluded. Also, the possible nutrient imbalance due to salinity can lead to physiological and biochemical disturbances which eventually may lead to impair yield and/or fruit quality4. Commercially, citrus is propagated by shield budding
on the rootstocks of desired characters. Rootstocks influence tree vigor, water relations, cold hardiness, mineral nutrition, hormonal balance and fruit yield and quality. Therefore, the study on differences in potential of rootstocks to transport water and nutrients becomes important especially in budded trees5. The plant growth in saline environment is affected due to:
(i) Water deficit; (ii) Effects on plant metabolism; and (iii) Ion toxicity and nutritional imbalance. The tree growth and fruit yield are impaired at a soil salinity of about 2 dS m-1 without any concomitant expression of leaf symptoms6,7. Though, citrus is highly salt sensitive crop, differences in tolerance do exist among species8. The present attempt tries to gather the information on work done so far on various citrus genotypes and their performance under salinity conditions to understand the possible mechanisms for salt stress tolerance.
Seed Germination under Salinity
Seed germination is greatly affected by salinity (Table 1)9. Soil salinity may prove detrimental to seed germination due to reduced water uptake and excessive absorption of ions till their toxicity lasts. It affects the emergence either by decreasing the osmotic potential of the soil solution to a point, which will retard or prevent the intake of water or become toxic to embryo and seedling10. Salt tolerance is not a constant character in citrus rootstocks but varies with the stages of seedling development11. The existed reduction or
complete inhibition of germination
________________
*Author for correspondence Tel: 91-11-2659-1116
E-mail: satyawatis@hotmail.com
Fig. 1Industrial potential of citrus Table 1Effect of salinity on seed germination in vivo
Sr No Citrus species Salt(s)/ Magnitude of salinity Effect of salinity Ref
1 Trifoliate orange, Cleopatra and Bakraie mandarin
NaCl, Na2SO4, PEG (0, 250, 500, 750 J/kg water potential)
Germination rate reduced; however, final germination not affected.
14 2 Citrus aurantium, C. volkameriana, C.
sinensis, C. reshni, C. jambhiri, C.
sinensis x Poncirus trifoliata and C.
paradis x P. trifoliata.
NaCl (50, 100 mM) + CaSO4 (5mM)
Salinity affected the emergence of first seedling, emergence spread, time to 50%
emergence and final emergence.
10
3 Sour orange, Volkamer lemon, Rangpur lime and Cleopatra mandarin
NaCl (1000-9000 ppm) Reduced seed germination as follows: Sour orange, 13; Volkamer lemon, 8.7; Rangpur lime, 3; and Cleopatra mandarin, 19%. The number of days for seed germination increased. Maximum tolerable salinity was:
Volkamer lemon & Rangpur lime, 9000;
Sour orange & Cleopatra mandarin, 600 ppm.
12
4 Troyer citrange, Volkamer lemon, Sour orange, Cleopatra mandarin and Flying Dragon
NaCl (50, 100 mM) in vitro seed germination
No germination at 100 mM. At 50 mM, germination was: Flying Dragon, 97.7;
Troyer citrange, 98%.
31
5 C. grandis cv. Pingshanyou and C.
reticulata cv. Fuju
NaCl (20-40 mM/l) No effect on seed germination, sprout length and radical length. Germination time increased.
32
MURKUTE et al: CITRUS IN TERMS OF SOIL & WATER SALINITY 395 potential under salt stress could be attributed to the
salinity that progressively diminishes the endogenous phytohormones (gibberellins, auxins, cytokinins), the main factors for controlling germination. Also, salinity increases the level of natural inhibitors12. Plant Growth Parameters
In all, the effect of salt stress on the growth parameters may be seen as reduced plant height, stem diam, leaf area, root length, fresh and dry weight and senescence, if salinity increases beyond tolerance limit. However, the tolerance may vary according to species13,14, salt combination and concentration and seedling age13,15,16. Reduction in growth parameters at increasing salinity levels can, in some instances, be attributed to salinity-induced adverse change in leaf water relations reducing photosynthesis, dehydration of proteins and protoplasm to a lower extent17. The decreased growth might also be because of osmotic effect of salt on root and toxic effect of accumulated ions in the plant tissues18,19. The reduction in relative growth was reported to be more dependent on scion, whereas the defoliation was more rootstock dependent20. The addition of calcium (CaSO4) to saline solution significantly decreased the adverse effect of NaCl on shoot growth21 and the treatment of Paclobutrazol compensated the deleterious effects of salinity on root growth (root size, number of lateral roots, dry weight of roots)12.
Nutritional Aspects
In general, the mineral accumulations either increase or decrease (Table 2) as a result of salt stress21,22. However, the effect was found dependent on scion-rootstock combination20. The NaCl addition in growth media increased N, P and K and reduced Ca and Mg in most of the rootstocks, whereas the addition of CaSO4 in saline medium did not affect the N, P, K and Mg but obvious increase in Ca in some rootstocks revealed that the salt quality and quantity also affect the nutritional imbalance10. The mineral concentrations in the plant organs (leaf, stem, root) analyzed in two citrus genotypes (Sour orange, Macrophylla) proved that the mineral accumulation varies in the different organs of the plant23 apart from species and salt concentration. There was no linear trend found in accumulation of micronutrients;
however, higher salinity (≥60 mM) decreased only Cu and Mn accumulation at the whole plant level24. Na and Cl Accumulation and K Substitution
Some citrus species while growing on saline conditions absorb large quantities of chloride and
sodium in their leaves. The concentration of Cl, Na and K in leaves on salt treated plants of all rootstocks varied according to age or position of leaves on the plant25. The higher leaf Cl in salt treated plants than control of trifoliate orange appeared to be balanced largely by the accompanying higher concentrations of accumulated K, however, in sweet orange, K concentration did not increase significantly in salt treated plants. Walker & Douglas26 revealed the reduction in K with increase in salinity and thus, there was evidence of limited Na/K exchange and possible sequencing of Na. Also, increasing Ca (2-8 mM) in growth medium did not alter either shoot growth or levels of Cl-, Na+ and K+ in all rootstocks tested. The salt induced loss of K and increase in Na+ with increasing salinity could not be prevented by addition of Ca. pH (5-8) could not affect the shoot growth and ion uptake in Rangpur lime26.
Leaf chloride analysis indicated that Rangpur lime and Cleopatra mandarin rootstocks restricted the uptake and/or transport of Cl to shoots16. However, comparatively high concentrations of Na were accumulated in mature leaves of all rootstocks during salt treatment and the leaf K did not alter by salinity and remained same as that of control. Rootstocks (Cleopatra mandarin and Rangpur lime), which maintained ability for chloride exclusion during treatment with 150 mM NaCl, were unable to restrict Na, indicated that apparently separate mechanism existed which regulate the uptake and/or transport of Cl and Na in citrus. Thus, this apparent inability of rootstocks grown on salinity to satisfactorily exclude sodium represents the most likely limitation to the extent to which salt tolerance can be improved. The upper limit of Cl– concentration in roots of citrus grafted plants seems to differ from each scion- rootstock combination, because Cl– accumulation in leaves and Cl– transport to scion could affect Cl– concentration in roots20. However, distribution of Na+ between plant organs was found scion–
rootstock dependent with limited transport from root to shoot.
Leaf chloride concentrations decreased with seedling age and were negatively co-related with seedling dry weight within same cultivars, but increased basipetally in all cultivars27. Walker28 revealed that the Na exclusion and K-Na selectivity in salt treated rootstocks appeared to possess a greater ability to withdraw sodium from the xylem in the proximal root and basal stem and sequester it in both
the wood and the bark of these regions suggesting that the lower content of salt treated plants resulted from net loss of K from plant rather than reduced uptake.
Release of K into the xylem in exchange for Na is implied by the significant increase in K concentration.
However, reports also show that the scion-rootstock combination did not affect Na and K concentrations in leaves22.
The addition of CaSO4 (5 mol m-3) to the saline solution (50 mol m-3) reduced Na and/or Cl concentration in shoots of Sour orange, Troyer citrange, Swingle citrumelo, Ridge pineapple sweet orange and Rough lemon but did not affect Na and Cl contents in roots of any of the rootstocks10. Also, none of the several rootstocks tested could exclude Na or Cl from its shoots. However, Na and Cl exclusion capacities of some citrus rootstocks were lost at the saline solution having osmotic potential of –0.20 MPa (~50mol m-3) or higher29.
The relative rates of mineral accumulation by shoot and the relative rates of mineral transport from the root to shoot of Cl and/or Na, increased in seedlings
of Sour orange and Macrophylla grown in Cl and/or Na treatment23. Na stressed seedlings decreased accumulation of Ca in Sour orange and Ca and K in Macrophylla compared to control. Also, leaf injury symptoms associated with Na in both genotypes may be due to reduced uptake and transport of Ca. The K deficiency, caused due to Na stress, could be the reason of reduced CO2 assimilation rates23. The ‘Fine Root Turnover’ phenomenon in which continuous root formation in plants apparently used by the plants to remove the excess ions and delay onset of accumulation in this tissue has been well established in Poncirus trifoliata24. The high Cl accumulation was more toxic than Na in leaves whereas the latter is more toxic to fine roots and limited K substitution. It is proposed that the accumulation of Na or/and Cl in plant organs is also a species dependent phenomenon30.
Physiological Aspects: Water Relation / Stomatal Conductance and Gas Exchange
Physiological disturbances due to salt treatment in some cases might be attributed to salinity-induced
Table 2Effect of salinity on nutritional parameters
Sr No Citrus species Salt(s)/ Magnitude of salinity Effect of salinity Ref
1 Navel orange and Clementine on Troyer citrange and Cleopatra mandarin
NaCl (0-60 mM) Sharp decrease in N accumulation and dependent on scion rootstock combination.
Little difference in P and significant decrease in Ca, Mg, and K contents in leaves and roots of all combinations.
20
2 C. aurantium NaCl (0, 40 mM) + CaSO4, CaCl2
and KCl (1, 5, 7.5 mM)
Significantly reduced the leaf Ca, Mg and K and no significant differences were found in P, Fe, Mn, Zn and Cu contents.
21
3 Verna lemon trees on Macrophylla, Cleopatra mandarin and Sour orange
NaCl (6, 12, 20, 28 mol m-3 Cl) Linear increase in P. Leaves from Macrophylla generally had higher total N under than those from other two combinations. No change in Mg but Ca decreased.
22
4 Cleopatra mandarin and Volkamer lemon
6.13 dS m-1 salinity K, Ca and Mg in low range; Zn, Cu, Mn, and Fe in sufficient range and not affected by saline treatments. N increased with salinity.
18
5 C. aurantium, C. volkameriana, C.
sinensis, C. reshni, C. jambhiri, C.
sinensis x P. trifoliata and C.
paradisi x P. trifoliata.
NaCl (50, 100 mM) +CaSO4
(5mM)
N, P, K, Ca and Mg reduced in most rootstocks; addition of CaSO4 did not affect N, P, K and Mg but increase in Ca in some rootstocks.
10
6 Sour orange and Macrophylla Na without Cl, Cl without Na and NaCl (40 mM)
Ca and Mg in leaves, K and Ca in roots and K in stem decreased and Mg in the root and stem increased. Cu and Mn in leaves; Ca, Mg, Cu, Mn, Zn in the stem;
and Mg and Zn in the roots increased; total N, P and Fe in general reduced.
23
7 P. trifoliata NaCl (0, 30, 60, 90, 120 mM) K and P increased in roots and leaves. No differences for Ca and P in stem and Mg in structural roots.
24
MURKUTE et al: CITRUS IN TERMS OF SOIL & WATER SALINITY 397
changes in water relations (Table 3). Specifically, turgor dependent processes (photosynthesis) are likely to be affected if the osmotic adjustment is insufficient.
Stomata of mature citrus leaves can remain open at low leaf water potentials by maintaining higher turgor in guard cells than mesophyll cells38. However, the citrus genotypes appeared to have different ability for turgor maintenance during salt stress16,33.
Osmotic potentials decreased as leaves matured and responses to salinity were found rootstock dependent34. The net gas exchange characteristics of mature leaves were unaffected by salinity but mature leaves had lower stomatal conductance and internal CO2 concentration, resulting in higher water use efficiency than young leaves, regardless of salinity.
On the contrary, Nieves et al17 reported that the salt treated plants had significantly lower stomatal conductance than control and there was no difference related to rootstock or scion.
The leaf stomatal density was affected by the rootstocks under salt stress37. Leaf water potential, stomatal conductance and photosynthesis were reduced more in grafted plants and choice of rootstocks (Cleopatra mandarin, Troyer citrange) had little effect on salt induced parameters36. Thus, the
reduction in gas exchange parameters and growth at increasing salinity levels depended more on the scion type than on Na and Cl concentrations in leaves.
Otherwise, leaf injury and defoliation were closely correlated to leaf Cl concentration39.
Biochemical Aspects
Biochemical indicators (proline, plant pigments, sugars) showed varied response to salinity (Table 4).
Nolte et al41 studied 55 cultivated and wild species of Aurantioideae for proline and betaine analysis and found that some species accumulated proline alone and some accumulated proline and proline-betaine (20-100 µmol g-1 dry mass) under salt stress. Proline level increased during winter42,43, and was found to be species dependent17.
The plant pigment contents decrease in response to salt stress in several citrus rootstocks13,34. Zekri35 reported the loss of chlorophyll due to Cl accumulation. The reduced photosynthetic ability under salinity is due to stomata closure and suppression of specific enzymes that are responsible for the synthesis of photosynthetic pigments. The reduction of chlorophyll contents is mainly due to destruction of chlorophyll biosynthesis and reduction
Table 3 Effect of salinity on physiological parameters
Sr No Citrus species Salt(s)/ Magnitude of
salinity
Effect of salinity Ref
1 Valencia on Cleopatra mandarin, Rangpur lime, Sweet orange, Rough lemon, Trifoliate orange and Carrizo citrange
NaCl (0-150 mM) Marked reduction in leaf water potential and also reduction in leaf osmotic potential. Stomatal resistance increased and showed only partial recovery after cessation of salt treatment.
16
2 Etrog citron (C. medica) NaCl (0-150 mM) Showed complete stomatal recovery when stress relieved.
33 3 Valencia grafted on Trifoliate
orange and Sweet orange rootstocks
NaCl (10, 14, 20 mol m-3 Cl)
Osmotic potentials decreased. Responses to salinity found rootstock dependent. Net gas exchange remained unaffected but mature leaves had lower stomatal conductance and internal CO2
concentration.
34
4 Verna and Fino lemon on Sour orange and Macrophylla
NaCl (2, 40 and 80 mM) Stomatal conductance lowered than control and there was no difference related to rootstock or scion. Leaf water potential was higher than in control in all scion-rootstock combinations.
17
5 Sour orange and Cleopatra mandarin
NaCl (-0.10, -0.20, -0.35 MPa)
Root hydraulic conductivity, stomatal conductance and evapotranspiration significantly reduced.
35 6 Cleopatra mandarin and Volkamer
lemon
NaCl +CaCl2 3:1 (6.13 dS/m)
Whole plant transpiration rate and photosynthetic rate reduced.
18 7 Navel orange Clementine on
Cleopatra mandarin and Troyer citrange
NaCl (0, 20 40, 60 mM) Leaf water potential, stomatal conductance and photosynthesis reduced. Reduction in gas exchange parameters at increasing salinity levels depended more on scion type than on Na and Cl in leaves.
36
8 Satsuma mandarin on Trifoliate orange and Troyer citrange
NaCl (2.0, 3.5, 5.0, 6.5 dS/m)
Stomatal density, photosynthetic rate and water use efficiency reduced.
37
in magnesium, iron and manganese12. The beneficial effect of Paclobutrazol (PP333) under salinity on the accumulation of chlorophyll, carotenoids and phytohormones has also been documented12.
Considerable variations in accumulation of soluble sugars in response to salt stress are also evident at inter specific, intra-specific and even among all lines, which are salt tolerant44. Lower osmotic potential of plant cells would increase the capability of plants to absorb saline substrates. Hence, increased concentration of reducing and total sugars in response to salinity could be attributed as osmotic adjustment to lower down the osmotic potential of plant cells44. Salinity and Mycorrhizal Fungi
Mycorrhizal symbioses have been found to improve the ability of stress tolerance in certain plant species. Duke et al45 reported the accumulation of phosphorus, dry matter and betaine during NaCl stress on Carrizo citrange colonized with mycorrhizal fungi (Glomus intraradices). High root-infection (77-83%) was observed with control and values dropped (53%) at 100 mM NaCl. Shoot and root dry weight of AM plants was significantly high over non-treated plants at all salt treatments. Proline-betaine was found linearly related to high NaCl concentrations regardless of plant mycorrhizal status. Levy et al46 reported reduced VAM infection in deep layers due to increasing the salinity of the irrigation water (EC=1.1- 2.7 dS m-1). Harmond et al47 reported that the mycorrhizal colonization remained unaffected under salinity stress and reduced the hydraulic conductivity of roots, leaf water potential, stomatal conductance and net assimilation of CO2 of AM and non-AM
seedlings to a similar extent in pineapple sweet orange, Carrizo citrange and Sour orange. AM plants of Carrizo citrange and Sour orange accumulated more chloride in leaves than non-AM plants. The observed reduction in N and P contents due to salinity was at par in AM and non-AM plants.
Graham & Syvertsen48 confirmed that salinity (30, 60 mM NaCl) did not affect the mycorrhizal colonization but reduced the growth of non- Mycorrhizal plants, though not significantly, than AM inoculated plants of Sour orange and Sweet orange.
Mycorrhiza did not affect the growth of either species even under non-stress condition. Though the concentration of N, Ca and Mg decreased, Na and Cl increased due to salt stress and remained at par with VAM plants except Cl. The total chloride was greater in VAM plants at control and 30 mM NaCl concentration. The leaf Cu concentration was significantly higher in AM plants. Mycorrhiza increased Cl and Zn and decreased Mn in roots of both the species irrespective of salinity. The root Cu of either species was increased by mycorrhiza under non-stress condition but salinization reduced it in mycorrhizal roots. Mycorrhizal fungi appeared to be functioning in P uptake under salinity stress, as moderate levels of NaCl did not reduce P concentration in AM plants. Also, the reduced hydraulic conductivity of roots and transpiration of shoots was irrespective of mycorrhization and similar in either species.
Salinity treatments tended to reduce the activity of peroxidase, while it did not affect the polyphenol oxidase. On the contrary, mycorrhizal infection did not significantly alter the peroxidase activity but
Table 4Effect of salinity on biochemical parameters
Sr No Citrus species Salt(s)/ Magnitude of salinity Effect of salinity Ref
1 Valencia orange (C. sinensis) on P.
trifoliata and C. sinensis
NaCl (10, 14, 20 mol/m3) Total chlorophyll reduced. 34 2 Verna and Fino lemon (C. limon) on
Sour orange (C. aurantium) and Macrophylla (C. macrophylla)
NaCl (2, 40, 80 mol/m3) Chlorophyll a, b and a+b reduced.
Proline increased.
17
3 Sour orange (C. aurantium) and Cleopatra mandarin (C. reticulata)
NaCl (osmotic potential of soils -0.10, -0.20, – 0.35 MPa)
Total chlorophyll reduced by 56% in Sour orange and 11% in Cleopatra mandarin by first salinity level (-0.10 MPa).
35
4 Hamlin sweet orange (C. sinensis) NaCl, KCl and NaNO3 (0, 15, 45, 50, 60 mM)
NaCl and KCl increased proline; NaNO3 did not affect proline.
40 5 Sour orange (C. aurantium), Volkamer
lemon (C. volkameriana), Rangpur lime (C. limonia) and Cleopatra mandarin (C.
reticulata)
NaCl (4000, 5000 ppm), NaCl 5000 ppm + PP333 (50, 100 ppm)
Chlorophyll a, b, a+b and carotenoids reduced. Cytokinins, gibberellins, auxins decreased.
12
MURKUTE et al: CITRUS IN TERMS OF SOIL & WATER SALINITY 399
appreciably increased the leaf polyphenol oxidase in Sour orange44. A marked decrease in starch and total carbohydrates and significant increase in reducing, non-reducing and total soluble sugars, with increasing salinity (NaCl+CaCl2, 1:1) was also observed. The total chlorophyll and chlorophyll-a responded negatively to salinity but chlorophyll-b was not greatly affected. However, significantly increased total chlorophyll and chlorophyll-b in response to mycorrhizal inoculation under control could not be retained under salinity44.
Fruit Quality and Yield under Salinity
Salinity ultimately affects the fruit quality and yield (Table 5). The yield reduction due to salinity stress could be understood as reduction in the number of fruits per tree and the ion toxicity was substantiated for the deterioration of quality of fruits49. Bielorai et al50 revealed that high sodium absorption ratio (SAR) and high Cl reduced the yield (9%) of Marsh Seedless grapefruit and high ESP (exchangeable sodium percent) did not specifically affect yield. The salinity- affected changes in peel characters could be associated with the loss of water in the albedo through reduced osmotic potential51. Despite unaffected juice content, total soluble solids, density and titrable acidity increased in several citrus species as a response to increasing salinity68; however, the selection of rootstocks influenced the quality fruits differently.
Salinity and Tissue Culture
The use of plant tissue culture to incorporate disease resistance into crop plants and to select mutants that are tolerant to toxins, herbicides, salinity and environmental stresses has been proved54. The progress in selection of NaCl tolerance in C. sinensis of ovular callus enlightened the use of cell line selection for salinity tolerance in citrus, a highly salt sensitive fruit crop55. Gamma irradiated (8-16 kR) callus performed well as compared to non- irradiated when subjected to salinity (0.2 M NaCl), though the difference was non-significant and found stable after removal of selection pressure. Several callus lines of Shamouti sweet orange and one cell line of Sour orange have been reported to grow in the presence of NaCl by repetitive exposure to the medium containing salt56. The NaCl tolerant cells also found to perform well on the selection pressure of Na2SO4 and K2SO4, but performed poorly with KCl.
Ben-Hayyim & Kochba57 revealed growth characteristics and stability of tolerance of citrus callus cells subjected to NaCl stress. The non-tolerant selected callus cell lines did not show any gain in weight and growth when exposed to 0.2 M NaCl, unlike tolerant line. Cells, which performed well in NaCl stress, had capacity to withstand in the presence of various Na+ salts with other cations (Na2SO4, NaNO3, NaBr). On the contrary, replacing Na+ with other cations gave rise to various degrees of growth inhibition, indicating that the nature of cation in the resistance to salt stress is rather important factor.
Table 5Effect of salinity on fruit yield and quality
Sr No Citrus species Salt(s)/ Magnitude of salinity Effect of salinity Ref
1 Valencia orange Cl and SO4 Reduced fruit yield.
6 2 Valencia orange (C. sinensis) 1.7 dS/m (no salt), 3.8 dS/m (5mM each
of CaCl2 + Na2SO4 + MgSO4), 5.7 dS/m (10mM each of CaCl2 + Na2SO4 + MgSO4)
Reduction in fruit number, not size.
Rind thickness decreased and maturity delayed. Total soluble solids (TSS):Titrable acidity(TA) unaffected.
49
3 Marsh Seedless grapefruit (C.
paradisi)
Salt combination,
EC- (2.7dS/m), SAR-10.3 (mol/m3)1/2
Reduction in fruit yield (9%). 50
4 Shamouti orange Cl (100, 250 and 450 mg/l) Fruit yield decreased (13%) with
increased salinity (100-450 mg/l Cl).
52 5 Verna lemon(C. limon) budded on
C. aurantium, C. reticulata, C.
macrophylla
Saline irrigation water with EC- 1.2, 2.2, 3.8 and 5.2dS/m
Peel thickness increased. TSS, density and titrable acidity of juice increased, juice content remained unaffected.
68
6 Fino-49 (C. limon) on C.
macrophylla
NaCl 15 mM (EC- 2.5 dS/m, SAR- 4.5) and 30 mM (EC-4dS/m, SAR- 9) Control (1dS/m, SAR-1.6)
Fruits/tree, fruit juice and yield decreased. Peel and pulp increased. TA and TSS decreased but TSS:TA unchanged.
53
7 Star Ruby grape fruit (C. paradisi) on C. reticulata and C. sinensis x P. trifoliata
NaCl (3, 15 30 dS/m) Reduced fruit yield. 30
To introduce salt tolerance, selection for salt tolerant genetic variants in tissue culture (spontaneous or induced mutation), which have regenerative capacity, may provide useful starting material for conventional breeding. Ben-Hayyim & Kochba58 studied the aspects of salt tolerance in NaCl tolerant selected stable cell lines of C. sinensis. NaCl tolerant callus line when removed from selection pressure for at least four passages retained the same capacity of tolerance to 0.2 M NaCl. The recorded Na+ and Cl- uptake were also found low than the salt sensitive cell line. KCl again proved as growth suppressing when substituted to NaCl. The use of Na2SO4 and K2SO4
revealed that the toxic effects were that of accumulated chloride only. The presence of Ca2+ in growth medium was also essential for proper growth.
The electron micrographs proved that the salt tolerant cells had very big vacuoles as compared to salt sensitive cells. It was concluded that the tolerance of cells to NaCl stress was due to partial avoidance of toxic elements by variants.
The presence of NaCl during embryogenesis affects the growth regulators balance. It also enhances the requirement of gibberellic acid for the normal heart shaped embryo formation. However, the interactions of NaCl with cytokinins or gibberellins are not yet clear59. The ideal procedure for the development of salt tolerant saplings in vitro was to keep the selected stress throughout the regeneration process59. The NaCl tolerant cell line required same salt to produce green embryos that was not obtained without NaCl. The greening of white embryos could be done by the addition of cytokinins (BAP 0.2 mg-l) or abscisic acid (0.1 mg-l) in the solid glycerol medium. The addition of NaCl into growth medium resulted in callus proliferation and death of embryos.
With several transfers of normal embryos to kinetin medium (0.1–1 mg l-l) gave the plantlets. In the absence of kinetin, some embryos developed roots spontaneously and shoot development was arrested completely. The saplings developed in the presence of NaCl (0.2 M) yielded some normal looking leaves, but no internodes. NAA could induce only rooting but further shoot development did not occur. Beloualy &
Bouharmont60 could differentiate the plantlets from salt tolerant cell lines of Poncirus trifoliata.
Several cytological changes related with the salt tolerance occur in the embryogenic callus. Electron microscopy observations of salt tolerant (0.17 M NaCl) embryogenic calli of C. limon revealed the salt
tolerant calli had thick cell walls, ring shaped mitochondria, increased content of lipid bodies and parallel accumulation of rough endoplasmic reticulum61. The thick cell walls may be as a mechanism to preserve water potential of the cell walls against high saline external medium. The significant increase in the proline and sugar contents in C. aurantium callus salinity (137 mM NaCl) would be an indicator of the sensitivity to saline stress62.
The plant survival against cytotoxic effects depends upon the presence of reduced mineral molecules and antioxidant enzymes. Adoption of cells is associated with reduced cell expansion even though the turgor pressure of cells is maintained. Piqueras et al63 revealed that the growth of salt tolerant cells was reduced to five folds as compared to control. In the salt tolerant cells, Na+ and Cl- concentration were higher than control cells. However, Ca2+, Mg2+, K+ contents were decreased in salt tolerant cells. The concentration of total sugars, proline and betaine increased two folds in the salt tolerant cell mass over the control. While evaluating the saline stress and cell toxicity using cell suspension culture of cv. Carvalhal, Lima-Costa et al64 reported no great differences in protein contents during cell growth. This can be attributed as a strong indication that the protein metabolism has not been changed by different salt shock.
Future Perspectives
Apart from considering the mechanism for nutritional imbalance and their optimization, salinity should be tackled by using several other means. It is suggested that not only ion content of leaf tissues, but also ion content and mass production of all tissues should be considered when the salinity tolerance of citrus and related genera is to be characterized65. Plants treated with Pachlobutrazol (PP333)44 and abscisic acid66, showed high reduction in deleterious effects of soil salinity, may be apprehended as plant growth hormones yield some fruitful results in salinity tolerance66. The results suggest that abscisic acid plays a role in modifying citrus physiological behaviour in response to salinity and could be helpful in their acclimatization to saline conditions. The new concept of using interstock to reduce toxic ion accumulations in leaves of budded citrus trees67 has enormous potential to deal with soil salinity, but the mechanism is yet to be understood completely and needs more attention. The plant cell and tissue culture and transgenics form alternative means, which is yet
MURKUTE et al: CITRUS IN TERMS OF SOIL & WATER SALINITY 401 left under exploited as far as salinity tolerance of
perennial plant species are concerned and can give a long awaited success.
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