J. Mirzaei 1, and M. Moradi 2 ABSTRACT

Transcription

1 J. Agr. Sci. Tech. (2016) Vol. 18: Single nd Dul Arbusculr Mycorrhiz Fungi Inoculum Effects on Growth, Nutrient Absorption nd Antioxidnt Enzyme Activity in Ziziphus spin-christi Seedlings under Slinity Stress J. Mirzei 1, nd M. Mordi 2 ABSTRACT Ziziphus spin-christi re distributed in rid nd semi-rid regions of world. Most of these res re subjected to soil slinity. So, the im of this study ws to find out the effects of different NCl concentrtions on Z. spin-christi seedlings growth, in the presence of number of single nd dul AMF inoculums, to provide some informtion bout possible effects of AMF under slinity condition. For this purpose, the study ws conducted in nursery using 4 3 fctoril scheme (4 slinity levels; 0, 50, 100 nd 150 mm nd 3 mycorrhiz tretments; non-mycorrhizl plnt, G. fscicultum nd Funneliformis mossee+g. fscicultum inoculted plnts). Our results reveled tht slinity hs negtive effects on root length coloniztion, growth prmeters, chlorophyll content, nutrients bsorption nd results in N, proline, superoxide dismutse, peroxidse nd ctlse ctivity increment but mycorrhizl plnts mintined higher growth chrcteristics, chlorophyll content, nutrients, root coloniztion nd enzymes ctivity, proline nd N ion reduction nd these effects in dul inoculum were significntly different from single inoculum fungi. According to the results dul-inoculted plnts re more tolertive to slinity compred to single-inoculted nd not-inoculted seedlings. Indicting tht Z. spin-christi plnttion in sline soil would give us the best result if we use dul inoculted plnts. It could be sid tht in sline soils Z. spin-christi dulinoculted with mycorrhiz is tolertive nd more efficient. Keywords: Enzyme, Mycorrhiz, Proline, Slinity, Ziziphus spin-christi. INTRODUCTION One third of the world is rid nd semirid which is chrcterized by receiving less thn 400 mm of nnul precipittion (Wickens, 1998). In this vst re of the world soil slinity is the mjor problem ffecting forest plnttions estblishment. One of the common plnt species is Ziziphus spinchristi which is used for plnttion of these regions. Z. spin-christi is distributed in vst regions of the world including North Afric, 1 Deprtment of Forest Science, Fculty of Agriculture, University of Ilm, Ilm, Islmic Republic of Irn. Corresponding uthor; emil: j.mirzei@mil.ilm.c.ir 2 Deprtment of Forestry, Fculty of Nturl Resources, Behbhn Khtm Al-Anbi University of Technology, Islmic Republic of Irn. South Europe, the Mediterrnen, South nd Est of Asi nd the Middle Est (Yossef et l., 2011) nd most of these res re fcing slinity conditions. To reduce the negtive effects of slinity, Arbusculr Mycorrhizl Fungi (AMF) could serve s useful strtegy. Becuse, first they re known s microorgnisms tht cn increse plnt torrent under biotic nd biotic stresses (Hrynkiewicz nd Bum, 2012), second these fungus hve ssocition with more thn 80% terrestril plnt species (Smith nd Red 2008). Reserches showed 1845

2 Mirzei nd Mordi tht plnt tolernce is enhnced by the AM fungi in slt condition (Peng et l., 2010; Abdel Ltef nd Choxing, 2011, 2014) but the mechnism re not cler nd lso the bility of different AMF vried to llevite slt stress (Al-Krki et l., 2001). There re reports bout some mechnisms such s incresing dry weight, nutrition cquisition, wter uptke (Evelin et l., 2009) nd chlorophyll content (Giri nd Mukerji 2004) which cn cuse n increse in slinity tolernce of plnts. Although AMF cn llevite slinity stress but they could be negtively ffected by slinity stress in coloniztion, sporultion nd hyph growth (Porcel et l., 2012) which could depend on different AMF nd host plnt species (Crvlho et l., 2001). They re lso effective mechnisms for incresing ntioxidnt enzyme ctivity (Zhu et l., 2010; Abdel Ltef nd Choxing, 2011). In the dry nd slt condition plnts re subjected to low wter vilbility resulting in oxidtive stress induction but mycorrhizl fungi cn decline rective oxygen species nd increse drought stress tolernce (Foud et l., 2014) by inducting ntioxidnt enzymes tht cn protect plnts from oxidtive condition (Mittler, 2002). The mjor problem of plnttions of some prts of the world is soil slinity. So, the objective of this study ws to find out the effects of NCl concentrtions on Z. spinchristi seedlings in the presence of number of single nd dul AMF inoculums to provide some informtion bout possible effects of AMF on nutrients bsorption, growth prmeters, chlorophyll content, proline nd enzyme ctivity of Z. spinchristi seedlings. We hypothesized tht, using dul rbusculr mycorrhizl fungi (Glomus mossee+g. fscicultum) s Tble 1. Physico-chemicl chrcteristics of soil used in this experiment. inoculum could hve better results in llevitive slinity condition compred to single (G. fscicultum) mycorrhizl fungi inoculum nd non-mycorrhizl plnts. MATERIALS AND METHODS Growth Conditions nd Methodology In order to overcome seed dormncy, seed cots were scrified by nil clippers. Seeds were germinted in mix of cly, snd nd perlite (2:1:1 v/v) (Tble1). Seedling were plnted in nursery using 4 3 fctoril scheme experiment design (4 slinity levels nd 3 mycorrhizl tretments). Growth condition of seedlings ws djusted s nturl photoperiod condition (13 hours of light), between Februry nd August of 2013 (6 months). There were 20 plnts selected for ech tretment, but for enzyme nd nutrient elements nlysis only 6 plnt per tretment ws used. Ech pot ( cm) contined sterilized soil by utoclve (30 minutes t 121 C) nd 100 grms of single (G. fscicultum) nd dul (G. mossee+g. fscicultum) AM inoculum. The inoculum contined t lest 50 spore per grm soil, hyphe nd root frgments. Plnts were estblished for 5 weeks before being subjected to 4 NCl levels (0, 50, 100 nd 150 mm NCl) (Zmni et l., 2011) by ddition of slt solution to soil with the irrigtion wter. Coloniztion, Growth Prmeters, Nutrient Anlysis nd Enzymtic Assys Growth indictors (height nd bsl dimeter) nd shoots nd roots dry weight Soil chrcteristics Vlue Soil chrcteristics vlue ph 7.32 N (g kg -1 ) 0.12 EC (ds m -1 ) 0.52 P (ppm) 19.6 Orgnic crbon (%) 1.5 K (ppm) 601 Clcium (%) 5.4 Mg (ppm) 0.6 N (g kg -1 )

3 Arbusculr Mycorrhiz Fungi nd Z. spin-christi (Meloni et l., 2004) ws mesured six months fter plnttion time. Root length coloniztion ws performed ccording to the method of Phillips nd Hymn (1970). Dry Weight (DW) of shoot (leves nd stems) nd roots were obtined by oven-drying t 70ºC for 72 hours (Meloni et l., 2004). Ionic content including totl NNitrogen (N) ws estimted by semi-micro Kjeldhl method (Nelson nd Sommers, 1982) nd the other nutrients [totl potssium (K), totl Phosphorus (P) nd sodium (N)] were determined by tomic bsorption spectrophotometry (UV/VIS 9000) ccording to Xu et l. (2006). Lef snd roots were grounded in liquid nitrogen for enzyme ssys nd extrction ws performed with ice-cold 50 nm phosphte buffer (ph 7.0). Crude extrcts were obtined by the method of Mirzei nd Yousefzdeh (2013). The totl superoxide dismutse (SOD) (Ginnopolitis nd Ries, 1977), Peroxidse (POD) nd Ctlse ctivity (CAT) (Ckmk nd Horst, 1991) were determined. Proline content ws determined by the ninhydrin nd sulfoslisilic cid method (Btes et l., 1973). Chlorophyll content of semi-mture lefs were determined using spectrophotometer (UV/VIS 9000) t 645 nd 663 nm for chlorophyll nd b. Experimentl Design nd Sttisticl Anlysis SPSS softwre (version 19) ws used for dt nlyzing, ll prmeters were nlyzed through two-wy ANOVA for min effects (S, Slinity nd M, rbusculr mycorrhizl inocultion). Mens comprison ws done by Duncn s test t 0.05 confidence level. RESULTS Coloniztion Assessment The men root length coloniztion for dul inoculum tretments were 59, 57, 45 nd 44 percent t 0, 50, 100 nd 150 mm concentrtion of NCl, respectively. Our results indicted 25 percent reduction of root length coloniztion in dul inoculum by slinity stress. These results were 40, 36, 23 nd 18 percent in single inoculum respectively which cused 55 percent decrese in root length coloniztion. As shown in Figure 1, dul rbusculr mycorrhizl fungi hd better result in root length coloniztion compred to single mycorrhizl fungi. In both single nd dul mycorrhizl inoculums the highest nd lowest coloniztion rte were observed in control nd 150 mm respectively. In dul Root cloniztion rte (%) b Single-AM Dul-AM bc bc c c d d Slt stress (mm) Figure 1. Root coloniztion rte in different NCl concentrtions. (Mens followed by the sme letters re not significntly different t P< 0.05 (n= 20)]. 1847

4 Mirzei nd Mordi inoculum, there ws no significnt difference (P> 0.05) in root length coloniztion between control nd 50 mm. In ddition to tht there ws decresing trend from 50 mm to 150 mm in dul inoculum but this trend ws not significnt (P> 0.05) between 100 nd 150 mm. According to our results root coloniztion in single inoculted plnts t ll levels of slinity is generlly lesser thn dul inoculted seedlings. The lowest coloniztion rte for both single nd dul inoculums ws observed in 100 nd 150 mm for single inoculum (Figure 1). Growth Prmeters Growth prmeters in AMF treted seedlings were incresed. The increse in these prmeters in dul inoculted plnts ws significntly higher thn single inoculted seedlings (Tble 2). In control tretment (0 mm NCl) dul inocultion cused incresing in ll studied growth prmeters significntly except shoot dry weight. There is no significnt difference between single inocultion nd non-treted seedlings in mentioned prmeters (Tble 2). As slinity incresed, growth prmeters decesed, nd this loss in control ws higher thn single inoculted, nd in single inoculted it ws higher thn dul inoculted seedlings. According to the results, growth prmeters (shoot nd root dimeter, shoot nd root dry weight) in AMF treted plnts t ll levels of slinity hve been incresed. Although the increse in growth t high level of slinity cused by dul inocultion is higher thn single nd non- inoculted seedlings significntly (Tble 2), we found tht in low levels of slinity there were no significnt differences between single inoculted plnts nd non- mycorrhizl in growth prmeters, but in higher levels of slt these indictors in single inoculted plnts were higher thn non-mycorrhizl plnts. Chlorophyll Content The highest vlue of chlorophyll content both for nd b were observed in control. Increse in slinity levels from 0 to 150 mm cused reduction of both nd b chlorophyll content significntly. At control, dul inoculted seedlings hd the highest level of chlorophyll content in comprison to single nd non-inoculted plnts (Figures 2 nd 3). In double inoculted plnts, chlorophyll content, t 0 nd 50 mm, 1.7 times nd in 100 nd 150 mm, hs been 2 times higher Tble 2. Effect of slt stress nd rbusculr mycorrhiz on growth prmeters of Z. spin-christi. Slt stress AM Height Dimeter Shoot dry Root dry inocultion (cm) (mm) (g) (g) Single-AM 23.1±4.5 bc 3.23±0.31 b 7.12± ±0.45 b 0 Dul-AM 32.8± ± ±0.25 b 4.52±0.38 Non-AM 28.8±4.8b 1.33±0.25 c 5.02±1.5 b 1.52±0.38 d Single-AM 8.3±1.6de 2.47±0.26 b 3.2± ± Dul-AM 13±2.02de 3.17±0.41 b 4.1±0.38 bc 3.38±0.36 b Non-AM 15.4± ± ±0.38 ef 0.62±0.24e Single-AM 6.3 ±1.14ef 2.55 ±0.4 b 2.34 ±0.48 de 1.6±0.2 d e 100 Dul-AM 16.8± ±0.37 b 3.24± ±0.2 c Non-AM 7.8±2.9de 0.61± ±0.43 f 0.3±0.15 e Single-AM 3.1±0.46e 1.5±0.26 c 2.34±0.45 de 0.68±0.22 e 150 Dul-AM 12±0.44 de 2.6±0.22 b 3.44± ±0.23 Non-AM 7.4±1.98de 0.2±0.17 d 0.72±0.21 f 0.04±0.02 e Mens±SE, sme letters t ech tretment re not significntly different t P< 0.05 (n= 20). 1848

5 Arbusculr Mycorrhiz Fungi nd Z. spin-christi Clorophyll (Mg g-1 fresh weight) bc b e Single-AM Dul-AM Non-AM b b e e e e b Slt concentrtion (mm) () e Clorophyll b (Mg g-1 fresh weight) b Single-AM Dul-AM Non-AM bc b b b d Slt concentrtion (mm) Figure 2. Men chlorophyll nd b content t different slinity levels. [Brs represent stndrd error, mens followed by the sme letters re not significntly different t P< 0.05 (n= 6)]. thn tht in non-inoculted plnts. The sme trend ws observed for single inoculted plnts in comprison to control. Just like chlorophyll, the highest mount of chlorophyll b ws observed in dul inoculted plnts. Tble 3. Effect of slinity nd rbusculr mycorrhiz on some lef nutrients content. Slt stress Nutrients nd N Absorption By incresing the slinity level, the seedling shoot N content in control ws reduced but in mycorrhizl plnts dverse effects of slinity were llevited. Dul inoculted plnts hd the highest level of N content t ll levels of slinity, single inoculted nd non-inoculted plnts hd AM inocultion Shoot N (g kg -1 ) Root Shoot P (ppm) Root (b) lesser level of N content respectively. Although slinity results in N content reduction but dul inoculted plnts cn enhnce N content in comprison to nonmycorrhizl plnts. Our results reveled tht in 150 mm slinity tretment dul inoculted plnts hd 93 percent more N content in comprison to non-mycorrhizl plnts. This result showed how these fungi cn enhnce N bsorption in higher slinity levels. Like shoot N content, the highest vlue of N content for root ws observed in non-slinity level, in the presence of dul inoculum. Also for root N content t nonslinity level there ws no significnt difference between single, dul nd non mycorrhizl tretments (Tble 3) but dul Shoot K (ppm) Single-AM 2.92±0.3b 2.22±0.48bc 4.05±0.65 b 3.71±0.24b 3.09± ±0.83b 0 Dul-AM 4.62± ± ± ± ± ±0.76 Non-AM 2.72± ±0.36bc 3.05±0.47b 3.31±0.2bc 2.31± ±0.83b Single-AM 2.74± ±0.44bc 3.72±0.57bc 3.05±0.5bc 2.36± ±0.5b 50 Dul-AM 3.84±0.32b 2.38±0.28bc 3.82±0.3bc 3.55±0.3bc 3.46± ±0.5 Non-AM 2.74± ±0.1 c 2.72±0.3e 2.2± ± ±0.4bc Root Single-AM 2.13±0.29 de 1.62±0.24bc 1.72±0.38ef 1.53±0.27d 1.3± ± Dul-AM 3.52±0.37bc 2.88± ±0.42def 2.2± ± ±0.16bc Non-AM 2.32±0.2 de 1.62±0.24bc 1.32±0.38fgh 1.13±0.17d 1.74± ±0.32d Single-AM 2.2±0.37 de 1.58±0.39bc 0.58±0.17gh 1.51±0.67d 0.79± ± Dul-AM 3.1±0.37b 2.48±0.39b 1.28±0.17fgh 2.2± ± ±0.07bc Non-AM- 1.6±0.23 e 1.58±0.39bc 0.28±0.12 h 1.27±0.59d 0.45± ±0.09d Mens±SE, sme letters t ech tretment re not significntly different t P< 0.05 (n= 6). 1849

6 Mirzei nd Mordi inoculted plnts in 100 mm nd more slinity levels hd significntly higher mount of N in roots. As shown in Tble 3 the increse in slinity cused P content deficit in shoots. The lowest reduction ws observed in dul inoculted plnts but in non-mycorrhizl plnts the highest chnges in P content hppened. In 150 mm, P content of dul inoculted plnts ws bout 78 percent higher thn non-mycorrhizl plnts. This increse in 0 slinity level ws only 40 percent which indictes the importnce of dul inocultion in P bsorption t higher levels of slinity. Root P content showed the sme results, but there re no significnt differences of root P content between 100 nd 150 mm of slt. As shown in Tble 3, K ws lso evluted. The increse in slinity to 150 mm cused reduction in shoot K content but not significntly. In ll tretments under slinity stress, mycorrhiz inocultion in comprison to non-mycorrhizl plnts cused decrese in K bsorption but this chnge ws not significnt. According to the results shoot K content in dul mycorrhizl plnts t 150 mm slinity, is t lest 73 percent more thn non-mycorrhiz plnts. Similr results bout N nd P indicte the importnce of dul AMF inocultion t higher levels of slinity. Unlike shoot, the root K content hd significnt decrese by incresing the slinity level. Mycorrhizl tretments hd the sme results in roots s shoot in K bsorption under slinity stress. In ddition to tht dul inoculum hd more effect in root K bsorption compred to single mycorrhiz inoculum. As shown in Figures 4 nd 5, N content incresed by incresing the slinity but mycorrhizl inocultion cused significnt reduction in N content t ll levels of slinity in comprison to control. There ws significnt difference between dul inoculted plnts with single nd nonmycorrhizl seedlings in N reduction t higher levels of slinity. Antioxidnt Enzyme The ctivity of some ntioxidnt enzymes known s the most importnt enzymes expressed in response to slinity stress were evluted both for root nd shoot. Our results reveled tht shoot nd root CAT ctivity incresed by incresing the slinity levels. In ddition to tht, there is n incresing trend for CAT in both root nd shoot but this is not significnt in shoot. There ws no significnt difference between mycorrhizl nd non-mycorrhizl plnts in CAT ctivity both in root nd shoot (Tble 4). As our results showed POD ctivity in shoot incresed by incresing the slinity levels. The highest level of POD ctivity in shoot ws observed in 150 nd 100 mm of slt but there ws no significnt difference Single-AM Dul-AM Non-AM Lef N (gr/kg) d b b bc bc b bc d Slt concentrtion (mm) Root N (gr/kg) de Single-AM Dul-AM Non-AM b b b bc bc bc e Slt concentrtion (mm) () Figure 4. Men lef N () nd root N (b) content t different slinity levels. [Brs represent stndrd error, mens followed by the sme letters re not significntly different t P< 0.05 (n= 6)]. (b) 1850

7 Arbusculr Mycorrhiz Fungi nd Z. spin-christi between 0, 50 nd 100 mm slinity level. Similr to shoot, incresing in slinity level results in root POD increment but dul mycorrhiz inocultion cused POD reduction (Tble 4). The highest SOD ctivity ws observed t 150 mm slt in non-mycorrhizl plnts nd the lowest SOD ctivity for both shoot nd root ws observed in dul mycorrhizl seedlings t control (Tble 4). Proline Content As shown in Tble 5 in both shoot nd root tissue proline content ws incresed significntly by incresing the slinity level. The highest proline content ws observed both for shoot nd root t 150 mm of slt. At ll levels of slinity for both root nd shoot, the lowest proline content ws observed in non-mycorrhizl, single inoculted nd dul mycorrhizl plnts respectively. DISCUSSION Slinity stress hd dverse effects on Z. spin-christi seedlings growth, nutrients bsorption nd AMF symbiosis. As we know slinity is mjor problem in rid nd semirid prts of the world (Ruiz-Lozno et l., 2012). Dt nlysis for slinity stress nd mycorrhiz inocultion reveled tht rbusculr mycorrhizl fungi llevite negtive effects of slinity stress in Z. spinchristi nd such llevition effects were more in dul rbusculr mycorrhizl inoculted plnts. Therefore these kinds of fungi cn be used s bio meliortor under slinity condition (Chndrsekrn et l., 2014). According to the results this meliortion effect is higher in dul AMF inoculted plnts. Root Coloniztion In this study the ppliction slinity suppressed the root length coloniztion in both single nd dul mycorrhiz inoculted plnts especilly t higher levels of slinity but this suppression in dul inoculted plnts is lesser thn single ones. As Jhromi et l. (2008) mentioned our results showed significnt decrese in root length coloniztion in non-mycorrhizl plnts t 100 nd 150 mm slinity levels. Decrese in single inoculted ws more thn dul inoculted plnts which indictes tht multiple inocultion under slinity stress could result in better root coloniztion rte. 1851

8 Mirzei nd Mordi Tble 5. Effect of slinity nd rbusculr mycorrhiz on free proline content. Slt stress AM inocultion Proline Shoot Root Single-AM 3.33±0.69de 1.73±0.23f 0 Dul-AM 2.23±1.65ef 0.92±0.26f Non-AM 4.63± ±0.32de Single-AM 2.56±0.48ef 4.16± Dul-AM 1.66±1.37f 2.86±0.52ef Non-AM 4.06± ±1.41b Single-AM 5.23±0.73bc 6.43±0.94b 100 Dul-AM 4.13± ±0.99bc Non-AM 6.53±0.51b 7.73±2.21b Single-AM 6.75±1.36b 7.75±0.79b 150 Dul-AM 5.45±0.94bc 6.05±1.02b Non-AM 7.85± ±1.02 Mens±SE, sme letters t ech tretment re not significntly different t P< 0.05 (n= 6). It is possible tht different levels of coloniztion in dul nd single mycorrhiz inocultion re becuse of different rbusculr mycorrhiz behvior (Klironomos et l., 1993). Growth Prmeters In ccordnce to Johnson-Green et l. (2001) our results showed tht slinity hd negtive effects on plnts growth nd biomss production. But s it is shown mycorrhizl fungi inocultion results in llevition of dverse effects on plnt growth prmeters under slinity stress. In lmost ll studied prmeters in inoculted plnts, either single or dul, there ws no significnt difference between 0 nd 50 mm of slt. It probbly shows tht Z. spinchristi could tolerte 50 mm of slt. It could be sid tht there re no significnt differences between 50, 100 nd sometimes even 150 mm of slt in dul inoculted plnts. It simply shows tht using dul mycorrhizl inocultion in sline soil would result in better biomss production nd growth rte in Z. spin-christi. In higher slinity levels growth prmeters significntly decresed nd since in our results higher slinity levels led to n insignificnt decrese of nutrients, it might be the reson for growth prmeters reduction (Evelin et l., 2009). Nutrient nd N Absorption Generlly by incresing the slinity, the nutrient content decresed, but in this study N content in both shoot nd root ws incresed by incresing the slinity levels. As shown in results mycorrhizl tretment especilly dul inocultion results in higher nutrient content both in root nd shoot which could be due to stbility of membrne by incresing minerl bsorption (Beltrno et l., 2013) in slinity levels compred to nonmycorrhizl plnts. It could be sid tht decrese in N content in mycorrhizl plnts in comprison to non-mycorrhizl plnts is due to positive effects of mycorrhizl in K bsorption which cn keep high K/N rtio (Chndrsekrn et l., 2014). In this study non-mycorrhizl plnts hd lower NPK nd higher N content which is in greement with Beltrno et l. (2013). The increse in minerl uptke by mycorrhiz is the min mechnism for plnts fcing limiting conditions (Moucheshi 1852

9 Arbusculr Mycorrhiz Fungi nd Z. spin-christi et l., 2012) nd by this mechnism plnts tolerte unfvorble conditions like slinity (Estrd et l., 2013). Chlorophyll Content The increse in chlorophylls nd b compred to non-mycorrhiz plnts might be becuse of positive effects of mycorrhizl on N bsorption t higher levels of slinity compred to control (Evelin et l., 2009; Grg nd Chndel, 2011) nd in the present study incresing nutrients bsorption in mycorrhizl plnts were observed compred to non-mycorrhizl plnts. This finding is in greement with other studies (Sheng et l., 2008; Beltrno et l., 2013) which report positive effects of mycorrhizl fungi on chlorophyll content under slinity stress. Free Proline Proline is free mino cid tht plys n importnt role in keeping osmtic pressure (Ruiz-Lozno et l., 2012) nd the higher proline content shows the higher osmtic pressure. Arbusculr mycorrhizl fungi hve known to be good mechnism to decrese proline content in stress conditions (Sheng et l., 2008; Abdel Ltef nd Choxing, 2014). Similr to other studies (Beltrno et l., 2013) free proline content is incresed with incresing slinity levels which cuses better osmotic sttues of plnts (Mrtinez et l., 1995; Abdel Ltef et l., 2009). We found out tht proline content ws higher in root compred to shoot which indicte tht roots need more proline to overcome osmtic pressure nd bsorb wter from the soil (Ruiz-Lozno et l., 2012) nd this is lso in greement with findings of Zhu et l. (2010). Antioxidnt Enzymes Some ntioxidnt enzymes including CAT, POD nd SOD by incresing slinity levels nd in the presence of mycorrhizl fungi were evluted. Our study reveled tht slinity incresed ntioxidnt ctivity in both roots nd shoots of Z. spin-christi seedlings which is in greement with findings of Ghorbnli et l. (2004). Also similr to some other findings (Wu et l., 2006; Zhu et l., 2010), our study showed tht AMF reduces the ctivity of ntioxidnt in high slinity levels in comprison to control. As reson for this we could sy tht ntioxidnt enzymes upregultion is one of the plnts defense mechnisms ginst oxidtive dmge cused by slinity stress, lso proline is one of the comptible solutes contributing osmoregultion of cell wll (Ahnger et l., 2014). But s we mentioned in the results, AMF inocultion resulted in higher proline content compred to control which could possibly result in fewer plnt needs for ntioxidnt enzymes production in comprison to control becuse proline cn ct s hydroxyl rdicl scvenger (Ali et l., 1995). The increse in proline content under biotic stresses such s slinity results from protein degrdtion (Bgdi nd Shw, 2013). In this study proline nd ntioxidnt ctivity decresed in the presence of mycorrhiz especilly dul mycorrhizl fungi which indictes tht these fungi hve the bility to decrese protein degrdtion under slinity stress. CONCLUSIONS Z. spin-christi is ntive species of rid nd semi-rid prts of the world. This species cn tolerte soil slinity up to 50 mm but dul mycorrhizl fungi inocultion levels would be good strtegy to hve successful plnttion in higher slinity levels. These results suggest tht by dul mycorrhiz inocultion G. mossee+g. fscicultum could hve better results in llevition effects of mycorrhiz in Z. spinchristi seedlings under high slinity levels. Mycorrhizl plnts (single nd dul inoculted) mintined higher growth 1853

10 Mirzei nd Mordi prmeters, chlorophyll content, nutrients, growth prmeters nd root coloniztion. Our results indicte tht dul mycorrhiz inoculted plnts re less susceptible to slinity conditions compred to single inoculted plnts nd non-mycorrhizl plnts. Finlly AMF especilly dul mycorrhizl inocultion cn increse the tolernce of Z. spin-christi in high slinity levels nd could be n effective mechnism for hving successful plnttion in rid nd slt conditions. It could be concluded tht G. fscicultum s single mycorrhiz inoculum nd G. mossee+g. fscicultum s dul mycorrhiz inoculums hd significnt effects in llevition of dverse effects of slinity conditions for Z. spinchristi seedlings but these llevitive effects were higher in dul mycorrhiz inoculum. REFERENCES 1. Abdel Ltef, A. A. nd Choxing, H Effect of Arbusculr Mycorrhizl Fungi on Growth, Minerl Nutrition, Antioxidnt Enzymes Activity nd Fruit Yield of Tomto Grown under Slinity Stress. Sci. Hortic., 127: Abdel Ltef, A. A. nd Choxing, H Does the Inocultion with Glomus mossee Improve Slt Tolernce in Pepper Plnts? J. Plnt Growth Regult., 33: Abdel Ltef, A. A., Shddd, M. A., Ismil, A. M nd Abou Elhmd, F. M Benzyldenine Cn Allevite Sline Injury of Two Roselle Cultivrs vi Equilibrtion Cytosolutes Including Anthocynin. Int. J. Agri. Biol., 11: Ahnger, M. A., Tygi, S. R., Wni, M. R. nd Ahmd, P Droughttolernce: role of orgnic osmolytes, growth regultors,nd minerl nutrients. In: Physiologicl Mechnisms nd Adpttion Strtegies in Plnts under Chnging Environment, (Eds.): Ahmd, P. nd Wni, M. R. Springer, New York, NY, 1: Al-Krki, G. N., Hmmd, R., Rusn, M Response of Two Tomto Cultivrs Differing in Slt Tolernce to Inocultion with Mycorrhizl Fungi under Slt Stress. Mycorrhiz, 11: Ali, K. V., Prsd, S. K. nd Srdhi, P. P Effect of Zinc on Free Rdicls nd Proline in Brssic nd Cjnus. Phytochem., 39: Bgdi, D. L. nd Shw, B. P Anlysis of Proline Metbolic Enzymes in Oryz stiv under NCl Sstress. J. Environ. Biol., 34: Btes, L. S., Wldern, R. P. nd Teve, I. D Rpid Determintion of Free Proline for Wter Stress Studies. Plnt Soil, 39: Beltrno, J., Ruscitti, M., Arngo, M. C. nd Ronco, M Effects of Arbusculr Mycorrhiz Inocultion on Plnt Growth, Biologicl nd Physiologicl Prmeters nd Minerl Nutrition in Pepper Grown under Different Slinity nd P Levels. J. Soil. Sci. Plnt Nutr., 13(1): Ckmk, I. nd Horst, W Effect of Aluminium on Lipid Peroxidtion, Superoxide Dismutse, Ctlse nd Peroxidse Activities in Root Tip of Soyben (Glysinmx). Plnt Phisiol., 83: Crvlho, L. M., Correi, P. H., Mrtins- Louco, A Arbusculr Mycorrhizl Fungl Propgules in Slt Mrsh. Mycorrhiz, 14: Chndrsekrn, M., Boughtts, S., Hu, S., Oh, S. H. nd S, T. A Met-nlysis of Arbusculr Mycorrhizl Effects on Plnts Grown under Slt Stress. Mycorrhiz, 24(8): El-Swify, S.A Soil nd Wter Slinity. In: Plnt Nutrient Mngement in Hwii s Soils, Approches for Tropicl nd Subtropicl Agriculture, (Eds.): Silv, J. A. nd Uchid, R. College of Tropicl Agriculture nd Humn Resources, University of Hwii, Mno, PP Estrd, B., Aroc, R., Bre, J. M., Ruiz- Lozno, J. M Ntive Arbusculr Mycorrhizl Fungi Isolted from Sline Hbitt Improved Mize Antioxidnt Systems nd Plnt Tolernce to Slinity. Plnt Sci., 201: Evelin, H., Kpoor, R. nd Giri, B Arbusculr Mycorrhizl Fungi in Allevition of Slt Stress: A Review. Ann. Bot., 104: Foud, M.O., Esshibi, A., Benhib, L. nd Qddoury, A Effectiveness of Arbusculr Mycorrhizl Fungi in the Protection of Olive Plnts ginst Oxidtive 1854

11 Arbusculr Mycorrhiz Fungi nd Z. spin-christi Stress Induced by Drought. Spn. J. Agric. Res., 12(3): Grg, N. nd Chndel, S The Effects of Slinity on Nitrogen Fixtion nd Trehlose Metbolism in Mycorrhizl Cjnuscjn (L.) Mill sp. Plnts. J. Plnt. Growth. Regul., 30: Ghorbnli, M., Ebrhimzdeh, H. nd Shrifi, M Effects of NCl nd Mycorrhizl Fungi on Antioxidnt Enzymes in Soyben. Biol. Plnt, 48: Ginnopolitis, C.N. nd Ries, S.K Superoxidedismutse. I. Occurrence in Higher Plnts. Plnt. Physiol., 59: Giri, B., Mukerji K. G Mycorrhizl Inoculnt Allevites Slt Stress in Sesbni egyptic nd Sesbni grndiflor under Field Conditions: Evidence for Reduced Sodium nd Improved Mgnesium Uptke. Mycorrhiz, 14: Hrborne, J.B Nitrogen Compounds. In: Phytochemicl Methods: A Guide to Modern Techniques of Plnt Anlysis, (Ed.): Hrborne, J. B. Chpmn nd Hll, London, PP Hrynkiewicz, K. nd Bum, C The Potentil of Rhizosphere Microorgnisms to Promote the Plnt Growth in Disturbed Soils. In: Environmentl Protection Strtegies for Sustinble Development, (Eds.): Mlik, A. nd Grohmnn, E. Springer Science+Business Medi BV, PP Jhromi, F., Aroc, R., Porcel, R., Ruiz- Lozno, J. M Influence of Slinity on the In vitro Development of Glomus intrrdices nd on the In vivo Physiologicl nd Moleculr Responses of Mycorrhizl Lettuce Plnts. Microb. Ecol., 55: Johnson-Green, P., Kenkel, N. C. nd Booth, T Soil Slinity nd Arbusculr Mycorrhizl Coloniztion of Puccinelli nuttllin. Mycol. Res., 105: Klironomos, J. N., Moutoglis, P., Kendrick, B., Widden, P A Comprison of Sptil Heterogeneity of Vesiculr Arbusculr Mycorrhizl Fungi in Two Mple-forest Soils. Cn. J. Bot., 71: Ldeiro, B Sline Agriculture in the 21st Century: Using Slt Contminted Resources to Cope Food Requirements, J. Bot., Lu, Y., Wng, G., Meng, Q., Zhng, W. nd Dun, B Growth nd Physiologicl Responses to Arbusculr Mycorrhizl Fungi nd Slt Stress in Dioecious Plnt Populus tomentos. Cn. J. For. Res., 44(9): Mrtinez, C. A., Guerrero, C. nd Moreno, U Diurnl Fluctutions of Crbon Exchnge Rte, Proline Content, nd Osmotic Potentil in Two Wter-stressed Potto Hybrids. Rev. Brs. Fisiol. Veg., 7(1): Meloni, D. A., Gulott, M. R., Mrtínez, C. A. nd Oliv, M. A The Effects of Slt Stress on Growth, Nitrte Reduction nd Proline nd Glycinebetine Accumultion in Prosopislb. Brz. J. Plnt. Physiol., 16(1): Mirzei, J. nd Yousefzdeh, H Peroxidse, Superoxide Dismutse nd Ctlse Activities of the Pistcikhinjuk Seedlings under Drought Stress. Ecopersi, 1(4): Mittler, R Oxidtive Stress, Antioxidnts nd Stress Tolernce. Trend. Plnt. Sci., 7: Moucheshi, A., Heidri, B. nd Assd, M. T Allevition of Drought Stress Effects on Whet Using Arbusculr Mycorrhizl Symbiosis. Int. J. Agri. Sci., 2(1): Nelson, D. W. nd Sommers, L. E Totl Crbon, Orgnic Crbon nd Orgnic Mtter. Prt 2. In: Methods of Soil Anlysis, (Eds.): Pge, A. L., Miller, R. H. nd. Keeney, D. R. Americn Society of Agronomy nd Soil Sci., Mdison, Wisconsin, PP Porcel, R., Aroc, R., Ruíz-Lozno, J.M Slinity Stress Allevition Using Arbusculr Mycorrhizl Fungi. A Review. Agron. Sustin. Dev., 32: Peng, J., Li, Y., Shi, P., Chen, X., Lin, H. nd Zho, B The Differentil Behvior of Arbusculr Mycorrhizl Fungi in Interction with Astrglus sinicus L. under Slt Stress. Mycorrhiz, 21: Phillips, J. M. nd Hymn, D. S Improved Procedures for Clering Roots nd Stining Prsitic nd Vesiculr-rbusculr Mycorrhizl Fungi for Rpid Assessment of Infection. Trns. Br. Mycol. Soc., 55: Ruiz-Lozno, J. M., Porcel, R., Azcon, C. nd Aroc, R Regultion by Arbusculr Mycorrhize of the Integrted Physiologicl Response to Slinity in Plnts: New Chllenges in Physiologicl nd Moleculr Studies. J. Exp. Bot., 63(11):

12 Mirzei nd Mordi 38. Sheng, M., Tng, M., Chn, H., Yng, B., Zhng, F. nd Hung, Y Influence of Arbusculr Mycorrhize on Photosynthesis nd Wter Sttus of Mize Plnts under Slt Stress. Mycorrhiz, 18: Smith, S. E., Red, D. J Mycorrhizl Symbiosis, Ed 3. Aemic Press, New York. 40. Wickens, G. E Ecophysiology of Economic Plnts of Arid nd Semi-rid Lnds. In: Adpttions of Desert Orgnisms, (Ed.): Cloudsley-Thompson, J. L. Springer-Verlg, Berlin, Heidelberg. 41. Wu, Q. S., Xi, R. X. nd Zou, Y. N Rective Oxygen Metbolism in Mycorrhizl nd Non-mycorrhizl Citrus (Poncirus trifolit) Seedlings Subjected to Wter Stress. J. Plnt. Physiol., 163: Xu, J., Li, H. D., Chen, L. Q., Wng, Y., Liu, L. L., He, L., Wu, W. H A Protein Kinse, Intercting with Two Clcineurin B-like Proteins, Regultes K + Trnsporter AKT1 in Arbidopsis. Cell, 125: Yossef, H. E., Khedr, A. A. nd Mhrn, M. Z Heptoprotective Activity nd Antioxidnt Effects of El Nbk (Zizyphus spin-christi) Fruits on Rts Heptotoxicity Induced by Crbon Tetrchloride. Nt. Sci., 9(2): Zmni, S, Nezmi, M. T., Bybordi, A., Behdd, M. nd Khorshidi, M. B Effect of Different NCl Slinity on Antioxidnt Enzyme Activity nd Reltive Wter in Winter Cnol (Brssic. npus). J. Res. Agr. Sci., 7 (1): Zhu, X., Song, F. nd Xu, H Influence of Arbusculr Mycorrhiz on Lipid Peroxidtion nd Antioxidnt Enzyme Activity of Mize Plnts under Temperture Stress. Mycorrhiz, 20: : ) 4 3 G. :. G. mosse + G. fsicultum) fsicultum

13 Arbusculr Mycorrhiz Fungi nd Z. spin-christi