Journal of Applied Botany and Food Quality 89, 21 - 28 (2016), DOI:10.5073/JABFQ.2016.089.003 1Botany Department, Faculty of Agriculture, Fayoum University, Fayoum, Egypt 2Irrigation Department, Centro de Edafología y Biología Aplicada del Segura (CEBAS-CSIC), Murcia, Spain 3Botany Department, National Research Centre, Dokki, Giza, Egypt 4Biology Department, Faculty of Sciences, Albaha University, Albaha, Suadi Arabia Growth, heavy metal status and yield of salt-stressed wheat (Triticum aestivum L.) plants as affected by the integrated application of bio-, organic and inorganic nitrogen-fertilizers Mostafa M. Rady1*, Oussama H. Mounzer2, Juan José Alarcón2, Magdi T. Abdelhamid3, Saad M. Howladar4 (Received September 4, 2015) * Corresponding author Summary Efforts have been made to use the integrated application of bio-, organic and inorganic nitrogen (N)-fertilizers to decrease waste ac- cumulation, and to minimize nutrient losses and yield contamination with heavy metals for human nutrition and health. Therefore, a field experiment was conducted to assess the effect of integrated applica- tions of organic manures, bio-fertilizer and/or mineral-N fertilizers on growth, yield, some chemical constituents and shoot and yielded grain heavy metal contents of wheat (Triticum aestivum L. cv. Sakha 93) plants grown under salinity stress (ECe = 7.84 dS m-1). Results showed that, the treatment comprised of ⅓NH4NO3 (55 kg N ha-1) + Cerealine (bio-fertilizer; 4 Kg ha-1) + cattle manure (10 t ha-1) was found to be most effective, producing the best status of growth characteristics, osmoprotectants concentrations, essential nutrient contents, shoot heavy metal concentrations, and grain yield and its content of heavy metals compared to the all other treatments. The treatment comprised of Cerealine (4 Kg ha-1) + cattle manure (20 t ha-1) was occupied the second order. We can recommend to use the integrated treatment of ⅓NH4NO3 (55 kg N ha-1) + Cerealine (bio- fertilizer; 4 Kg ha-1) + cattle manure (10 t ha-1) effectively in saline soils to improve wheat growth and yield with minimum contents of heavy metals for human health and nutrition. Introduction Wheat (Triticum aestivum L.) is the most important cereal crops used for human nutrition in most countries worldwide. It is a moderately salt-tolerant crop (MAAS and HOFFMAN, 1977), and is often grown on newly-reclaimed saline soils in Egypt. On the other hand, the cultivation of wheat will be restricted or even prevented in such soils if their salinity reached about 10 dS m-1 due to the severe reduction in wheat growth and productivity. Salinity stress is one of the major problems of agriculture in arid and semiarid regions. Salt stress affects plant physiology due to the osmotic and ionic stresses at both the cellular level and whole plant. It generates a physiological drought by affecting the plant water re- lations (MUNNS, 2002). The accumulation of toxic salts in the leaf apoplasm leads to dehydration, loss of turgor and the death of cells and tissues. Photosynthesis is one of the most severely affected pro- cesses during salinity stress, mediating by decreased levels of chlo- rophyll (RADY and MOHAMED, 2015) and the inhibition of various enzymes, including Rubisco (SOUSSI et al., 1998). In addition, lipid peroxidation and the antioxidant system of the plant have been re- ported to be stimulated by salt stress (RADY et al., 2013; SEMIDA and RADY, 2014). All these, and other altered processes lead to poor plant growth and productivity. Saline soils have low fertility and a poor structure with a high bioavailability of heavy metals (ACOSTA et al., 2011) to be available for plant roots. Heavy metal bioavailability can be minimized via biological immobilization and stabilization methods using a range of organic fertilizers (OFs), such as cattle manure (CM) and poul- try manure (PM). The beneficial effects of OFs, which are sourced locally, are an increase soil quality, total C and N content of the soil and soil-waterholding capacity, alteration of soil pH, elevation of soil humus and soil aggregation, and water infiltration properties (DOMAGALA-SWIATKIEWICZA and GASTOL, 2013; YAGIOKA et al., 2014; ABDOU and MOHAMED, 2014; BOBUL’SKA et al., 2015), and a reduction in the bioavailability of heavy metals (ANGELOVA et al., 2013). Although some studies have reported that organic farming tends to reduce plant growth and yield (JANSEN et al., 2015), an im- portant one of the OFs, CM application was found to increase the growth and yield of many crops without any undesirable impacts on the environment (SUTHAR, 2009; RADY, 2011). Poultry manure (PM) has long been recognized as perhaps the most desirable of these OFs because of its high nitrogen content. The application of OFs and other organic amendments to farmland is an economical and environmentally sustainable mechanism for increasing crop pro- duction and reducing bioavailability of heavy metals (YOUNIS et al., 2015). Cerealine (Bio-fert) is a commercial product of biofertilizer contains Azospirillum sp. For decreasing high doses of chemical fer- tilizer, the integrative effect of Bio-fert, OFs and chemical fertili- zers is a promising tool to increase wheat productivity, get cleaner product with low undesirable high doses of heavy metals and other pollutants (SUSHILA et al., 2000). Different strategies are being employed for attaining optimum growth and productivity under saline conditions. One of them is the proper management of nitrogen (N) fertilizer (RADY, 2012). To alleviate deleterious salinity effects such as a reduction in growth and yield, and an increase of bioavailability and plant absorption of heavy metals, different N-forms are being used. In view of these reports, it was postulated that the integrated application of bio-, or- ganic- and mineral-N fertilizer could alleviate the adverse effects of salt stress on the growth and productivity of wheat. Thus, the major objective of this study was to examine the extent to which N-forms could ameliorate the effects of salt stress on growth, ion accumula- tion, heavy metal uptake, and yield of wheat crop. Materials and methods Plant material, treatments and growth conditions A field experiment was conducted at the Experimental Farm of the Faculty of Agriculture, Fayoum University, Egypt. The main charac- teristics of the experimental soil, organic manures and irrigation wa- ter (Southeast Fayoum; 29° 17´N; 30° 53´E) used in this study were determined according to JACKSON (1973) and WILDE et al. (1985) and are shown in Tab. 1. Healthy seeds of wheat (Triticum aestivum L. cv. Sakha 93) were ob- tained from The Crop Research Institute, The Agricultural Research Center, Giza, Egypt, and were sown on 15 November 2009 and final harvest was done on 20 May 2010. Seeds were washed with distilled 22 M.M. Rady, O.H. Mounzer, J.J. Alarcón, M.T. Abdelhamid, S.M. Howladar Tab. 1: Physical and chemical properties of experimental soil, organic manures and irrigation water. Soil analysis Organic manure analysis Property Values Property Cattle Poultry Sand% 76.8 1m3 wt. (kg) 738 498 Silt% 18.1 Moisture% 70.0 66.7 Clay% 5.1 pH 7.50 6.34 Texture Sandy loam EC (dSm-1) 5.76 5.21 *pH 7.68 *O.C.% 34.0 43.6 EC (dS m-1) 7.84 *O.M.% 47.6 69.8 Total N% 0.63 C/N ratio 23:1 22:1 Total P% 0.36 Total N% 1.47 1.99 Total K% 1.25 Total P% 0.50 0.78 CaCO3% 11.1 Total K% 1.32 1.52 Fe (ppm) 16.9 CaCO3% 2.86 3.26 Mn (ppm) 8.4 Fe (ppm) 946 1928 Zn (ppm) 7.2 Mn (ppm) 429 876 Cu (ppm) 6.4 Zn (ppm) 146 230 Pb (ppm) 12.8 Cu (ppm) 68.2 98.2 Cd (ppm) 13.6 Pb (ppm) 12.2 31.2 Ni (ppm) 11.9 Cd (ppm) 5.91 18.0 Cr (ppm) 9.04 Ni (ppm) 7.13 16.9 Se (ppm) 6.17 Cr (ppm) 5.81 17.3 Co (ppm) 4.96 Se (ppm) 4.95 13.0 Co (ppm) 5.01 11.9 *pH (1:1 suspension), O.C.% = Organic carbon %, O.M.% = Organic mat- ter%. water, sterilized in 1% (v/v) sodium hypochlorite for approx. 2 min, washed thoroughly again with distilled water, and left to dry at room temperature (25 ºC) for approx. 1 h before sowing. Fourteen treat- ments in a randomized complete block design were used with three replications, generating 42 plots. The area of each plot was 10.5 m2 (3 m × 3.5 m). The distance between plots was 1 m from all sides to avoid treatments overlapping. The treatments imposed in the present investigation were detailed in Tab. 2. The experiment was designed to examine the influence of N-form (organic, bio- and/or inorganic) fertilizer on growth, heavy metal uptake, nutrients concentration and yield of wheat plants grown at salinity with EC = 7.84 dS m-1. PM and CM were added to soil be- fore sowing, during soil preparation. They spread on the soil surface, and then mixed into the soil-surface layer. Ammonium nitrate and urea quantities according to treatments were added before sowing. A quantity of 360 kg ha-1 of single super phosphate (15.5 % P2O5) was added before sowing as recommended dose. Cerealine is a commer- cial product of biofertilizer contains Azospirillum sp. produced by General Organization of Agriculture, Egypt, and was added before sowing at rate of 4 Kg ha-1. All other agricultural practices for wheat production were conducted according to the recommendations of the Egyptian Ministry of Agriculture and Land Reclamation. Plant sampling, and growth and yield measurements At fourteen weeks after sowing, plants were sampled to study the growth characteristics, and to perform the chemical analyses in leaves or shoots. At harvest, yield parameters were estimated. The following growth and yield parameters were measured: Plant height (cm), number (No.) of leaves plant-1, leaf area plant-1 (cm2), No. of tillers plant-1, shoot dry weight (DW) plant-1 (g), 1000-grain weight (g) and grain yield ha-1 (ton). Chemical analyses At fourteen weeks after sowing, the fifth leaf or shoot of four se- lected plants were collected from each experimental unit for chemi- cal determinations. The concentrations of chlorophylls and carote- Tab. 2: Components of the 14 treatments used in the present experiment Treatment Nitrogen (N) source Amount of N source ha-1 Amount of N ha-1 T1 (Control)* Not found Not found Not found T2 (Recom.)** Ammonium nitrate (NH4NO3) 475 kg (33.5 % N) 160 kg T3 (Recom.)** Urea (NH2-CO-NH2) 350 kg (46 % N) 160 kg T4 Poultry manure (PM1) 10 t 199 kg T5 Cattle manure (CM1) 20 t 294 kg T6 Bio-N (Cerealine) 4 Kg ……… T7 ½NH4NO3 + PM2 237.5 kg (33.5 % N) + 5 t 80 kg + 99.5 kg*** T8 ½NH4NO3 + CM2 237.5 kg (33.5 % N) + 10 t 80 kg + 147 kg*** T9 ½NH4NO3 + Cerealine 237.5 kg (33.5 % N) + 4 kg 80 kg + ……… T10 ½NH2-CO-NH2 + PM2 175 kg (46 % N) + 5 t 80 kg + 99.5 kg T11 ½NH2-CO-NH2 + CM2 175 kg (46 % N) + 10 t 80 kg + 147 kg T12 ½NH2-CO-NH2 + Cerealine 175 kg (46 % N) + 4 kg 80 kg + …….. T13 Cerealine + CM1 4 kg + 20 t ……. + 294 kg T14 ⅓NH4NO3 + Cerealine + CM2 158.3 kg + 4 kg + 10 t 55 kg + …... + 147 kg * The control treatment was not received any N-fertilizer. ** The recommended dose of N for wheat from either ammonium nitrate or urea as the Egyptian Ministry of Agriculture and Land Reclamation. *** Calculated from the Tab. 1 (Analyses of cattle and poultry manures). Effect of N form on wheat productivity and its quality 23 noids of the fresh upper fifth leaf were extracted by acetone 80 %, and were then measured colorimetrically using the method of ARNON (1949). The concentration of free proline was determined colorime- trically by using acid ninhydrin reagent with the sulfosalicylic acid (3 %)-extract as outlined by BATES et al. (1973). The concentration of total free amino acids was determined according to MUTING and KAISER (1963). The concentration of total soluble sugars was deter- mined in ethanolic extract with freshly-prepared anthrone reagent [150 mg anthrone + 100 ml of 72 % (v/v) sulphuric acid] according to IRIGOYEN et al. (1992). The contents of nutrients were determined in digested samples according to JACKSON (1973). The N content was measured using the semi-micro Kjeldahl method (BREMNER, 1965). The P content was determined colorimetrically by the molyb- denum blue method (CHAPMAN and PRATT, 1961). The contents of K and Na were assessed using flame photometry (JACKSON, 1973). The Ca content, and the all tested heavy metals (Pb, Cd, Ni, Cr, Se, Co, Zn and Cu) contents were determined using a Perkin-Elmer Model 3300 Atomic Absorption Spectrophotometer (CHAPMAN and PRATT, 1961). Statistical analysis The data were subjected to the analysis of variance (ANOVA) ap- propriate to the randomized complete block design according to the procedure outlined by GOMEZ and GOMEZ (1984). The significant differences among treatments were compared using the Fisher’s least-significant difference test (LSD) at P ≤ 0.05. Results Influences of N-fertilizer form on plant height, No. of leaves plant-1, leaf area plant-1, No. of tillers plant-1, shoot DW plant-1 of wheat grown under saline soil conditions are shown in Tab. 3. The treat- ment comprised of ⅓NH4NO3 (55 kg N ha-1) + Cerealine (4 Kg ha-1) + cattle manure (CM2; 10 t ha-1) was found to be most effective, pro- ducing the highest values of the all mentioned growth characteristics compared to the all other treatments. The treatment comprised of Cerealine (4 Kg ha-1) + cattle manure (CM1; 20 t ha-1) was occupied the second order. The treatment of the recommended mineral-N dose of ammonium nitrate or urea was in a parallel line with each of the treatments of ½Urea (80 kg N ha-1) + PM2 (5 t ha-1), ½Urea (80 kg N ha-1) + CM2 (10 t ha-1) and ½Urea (80 kg N ha-1) + Cerealine (4 Kg ha-1) in the third order compared to the other treatments, including the control. Fig. 1 shows the influences of N-fertilizer forms on the concentra- tions of total chlorophyll, total carotenoids, free proline, total so- luble sugars and free amino acids in wheat leaves under saline soil conditions. The treatments comprised of ⅓NH4NO3 (55 kg N ha-1) + Cerealine (4 Kg ha-1) + cattle manure (CM2; 10 t ha-1) and Cerealine (4 Kg ha-1) + cattle manure (CM1; 20 t ha-1) were found to be most effective, producing the highest values of the all aforementioned parameters, occupying the first and the second orders, respectively compared to the all other treatments. Tab. 4 shows the effects of N-fertilizer forms on nutrient contents (nitrogen; N, phosphorus; P, potassium; K, calcium; Ca and sodium; Na), and K/Na and Ca/Na ratios in shoots of wheat plants grown under saline soil conditions. The treatment comprised of ⅓NH4NO3 (55 kg N ha-1) + Cerealine (4 Kg ha-1) + cattle manure (CM2; 10 t ha-1) was found to be most effective, producing the highest values of the contents of N, P, K and Ca and the ratios of K/Na and Ca/Na, and the lowest values of Na contents compared to the all other treatments. The treatments comprised of Cerealine (4 Kg ha-1) + cattle manure (CM1; 20 t ha-1) and ½NH4NO3 (80 kg N ha-1) + cattle manure (CM2; 10 t ha-1) were occupied the second and third orders, respectively compared to the other treatments, including the control. The effects of N-fertilizer forms on the concentrations of heavy metals (lead; Pb, cadmium; Cd, nickel; Ni, chromium; Cr, selenium; Se, cobalt; Co, zinc; Zn and copper; Cu) in shoots and yielded grains of wheat plants grown under saline soil conditions are shown in Tab. 5 and 7. The treatments comprised of ⅓NH4NO3 (55 kg N ha-1) + Cerealine (4 Kg ha-1) + cattle manure (CM2; 10 t ha-1) and Cere- aline (4 Kg ha-1) + cattle manure (CM1; 20 t ha-1) were found to be most effective, producing the lowest values of the concentrations of all heavy metals, except some fluctuations compared to the all other treatments. On the other hand, the treatment of ½Urea (80 kg N ha-1) + poultry manure (PM2; 5 t ha-1) was found to generate the highest concentration values of all heavy metals, followed by all treatments with urea either solely or combined with poultry manure, or cattle manure, and even with Cerealine. N-fertilizer form effects on the yield parameters (1000-grain weight and grain yield ha-1) of wheat plants grown under saline soil condi- tions are shown in Tab. 6. The treatments comprised of ⅓NH4NO3 (55 kg N ha-1) + Cerealine (4 Kg ha-1) + cattle manure (CM2; 10 t ha-1) and Cerealine (4 Kg ha-1) + cattle manure (CM1; 20 t ha-1) were found to be most effective, producing the highest values of the 1000-grain weight and grain yield ha-1, occupying the first and the second orders, respectively compared to the all other treatments. Discussion The salinity of soil (ECe = 7.84 dS m-1; Tab. 1) may affect the plant metal uptake through the toxic effects of the Na+ and Cl- ions. The Tab. 3: Effect of N-fertilizer form applications on plant height, number (No.) of leaves plant-1, leaf area plant-1, and shoot dry weight (DW) plant-1 of wheat grown under saline soil conditions Treatments Plant height No. of leaves Leaf area Shoot plant-1 DW plant-1 (cm) plant-1 (cm2) (g) T1 58.4e* 5.11e 20.1g 6.1e T2 78.4c 6.89c 37.8cde 11.5c T3 78.4c 6.79c 36.7cde 11.3c T4 76.9cd 6.68c 35.3de 11.1c T5 77.1cd 6.61c 34.9e 11.0c T6 70.7d 5.98d 30.3f 9.9d T7 80.1c 6.95c 37.9cd 11.6c T8 78.4c 6.78c 36.9cde 11.3c T9 81.6bc 7.09bc 38.8c 11.8bc T10 77.8c 6.66c 35.5de 11.1c T11 80.1c 6.73c 35.7de 11.2c T12 80.0c 6.96c 37.8cde 11.6c T13 87.8b 7.63b 41.8b 12.7b T14 96.3a 8.37a 46.6a 14.0a *Values are means (n = 9), and mean values in each column followed by a dif- ferent lower-case-letter are significantly different by Fisher’s least-significant difference test (LSD) at P ≤ 0.05. T1: Control (without any N-fertilizer); T2: Recommended N as ammonium nitrate (160 kg N = 476 kg NH4NO3 ha-1 with 33.5 % N); T3: Recommen- ded N as Urea (160 kg N = 348 kg ha-1 urea [(NH2)2CO] with 46 % N); T4: Poultry manure (PM1 = 10 t ha-1); T5: Cattle manure (CM1 = 20 t ha-1); T6: Bio-N (Cerealine at rate of 4 Kg ha-1); T7: ½NH4NO3+PM2 (PM2 = 5 t ha-1); T8: ½NH4NO3+CM2 (CM2 = 10 t ha-1); T9: ½NH4NO3+Cerealine; T10: ½Urea+PM2; T11: ½Urea+CM2; T12: ½Urea+Cerealine; T13: Cerealine+CM1; T14: ⅓NH4NO3+Cerealine+CM2. 24 M.M. Rady, O.H. Mounzer, J.J. Alarcón, M.T. Abdelhamid, S.M. Howladar Na+ ions may release heavy metals from the sediment to the soil solution, thereby increasing the heavy metal accumulation in the plant organs (Tab. 5 and 7). In arid regions including Egypt, soils and irrigation water are characterized by high salinity levels that may aggravate the problem of heavy metal pollutions. In addition, high levels of chloride ions-containing irrigation water might increase soil heavy metal mobilization through the formation of metal- chloride complexes (MCLEAN and BLEDSOE, 1992; GHALLAB and USMAN, 2007). They also reported that the consequences of soil sa- linity can increase heavy metal solubility and availability, depen- ding on the composition of soil solution and its dominant inorganic ligands. The soil applications of organic manures have been shown to improve soil fertility, and to reduce salt stress and the bioavail- ability of heavy metals to plant roots (WEGGLER et al., 2004; OK et al., 2011). It has been also reported that the application of organic residues to heavy metal-contaminated soils increased soil organic matter content, improved the chemical and biological properties, and decreased the heavy metal bioavailability to plant roots (OK et al., 2011). Moreover, the application of organic manures to heavy metal- contaminated soil reduced the metal toxicity and consequently in- creased soil microbial activities (USMAN et al., 2013). In the present study, unlike the poultry manure the addition of cattle manure (CM) as a full alternative or a partial alternative to mineral fertilizers [i.e., the treatment comprised of ⅓NH4NO3 (55 kg N ha-1) + Cerealine (4 Kg ha-1) + cattle manure (CM2; 10 t ha-1) and the treatment com- prised of Cerealine (4 Kg ha-1) + cattle manure (CM1; 20 t ha-1)] was significantly increased wheat growth characteristics (Tab. 3), nutritional status (Tab. 4), photosynthetic pigments and osmopro- tectants (Fig. 1) and grain yield (Tab. 6), and significantly decreased the concentrations of heavy metals in plant shoots and yielded grains (Tab. 5 and 7). The most interesting finding is that these treatments were found to produce higher grain yield than average grain yield productivity of Egypt (6.452 t ha-1) in 2009, compared to 7.85 and 6.66 t ha-1 of these two treatments, respectively. Our results showed that the addition of CM caused a significant increase in shoot mineral nutrients (Tab. 4). This finding may be attributed to the release of nutrients with the decomposition of this organic manure (WANG et al., 2014). Results showed also that soil salinity caused increases in the bioavailability of heavy metals, as indicated by a significant increase in the wheat shoot and grain heavy metal concentrations in the control and the treatments con- tained inorganic N fertilizers and/or poultry manure (Tab. 5 and 7). This was observed in the treatment comprised of urea (inorganic N) + poultry manure (T10) which resulted in the highest heavy metal concentrations in wheat plants. This increase in plant heavy metal concentration may be due to poultry manure application that has shown to increase the soil concentration of heavy metals (HANČ et al., 2008; ARROYO et al., 2014). In addition, the frequent use of chemical fertilizers such as urea and pesticides in common agricul- tural fields can also cause an increase in the concentration of heavy metals in soil and water (HADI et al., 2014) and, consequently, in plants grown on this soil. However, the addition of CM significant- ly decreased shoot and grain heavy metal concentrations of wheat plants. These results are in accordance with those of AHMAD et al. (2011), noting that farm manure amendment significantly decreased Cd and Pb content in wheat grains. The lower heavy metal avail- ability in the farmyard amended soils could be mainly related to the heavy metal immobilization through sorption, chelation, and seques- tration by the solid and soluble organic matter. Indeed, soil heavy metal availability can be decreased by adding the organic materi- als, particularly CM through heavy metal transformation from more readily available forms to less available forms such as fractions as- sociated with organic materials, carbonates, or heavy metal oxides, causing a reduction in heavy metal uptake by plants (OK et al., 2011; BOLAN et al., 2003). NARWAL and SINGH (1998) observed that the Cd, Cu, Ni and Zn in soils decreased with increasing the addition of farmyard manure and peat that correspond with our results by soil application with CM (Tab. 5 and 7). Soil organic matter may con- tribute to the adsorption and immobilization of heavy metals, re- stricting its solubility and bioavailability. Therefore, organic manu- res such as CM may have an important role in improving the quality and minimizing the metal toxicity of contaminated soils under heavy metals and salinity stress (USMAN, 2015). It has been speculated that the application of organic manure to metal-contaminated soil may create a good environmental condition for mitigating the toxicity in- duced by high heavy metal concentrations (USMAN et al., 2013). The application of CM in combination with a partial inorganic fer- tilization and/or bio-fertilization (Cerealine) found to significant- ly increase wheat growth characteristics (plant height, number of Fig. 1: Effect of N-fertilizer form applications on the leaf concentrations of total chlorophyll, total carotenoids, free proline, total soluble sugars, and amino acids in wheat plants grown under saline soil conditions. *Mean values are presented in form of vertical bars. Bars with a dif- ferent lower-case-letter are significantly different by Fisher’s least- significant difference test (LSD) at P ≤ 0.05. T1: Control (without any N-fertilizer); T2: Recommended N as ammonium nitrate (160 kg N = 476 kg NH4NO3 ha-1 with 33.5 % N); T3: Recommended N as Urea (160 kg N = 348 kg ha-1 urea [(NH2)2CO] with 46 % N); T4: Poultry manure (PM1 = 10 t ha-1); T5: Cattle manure (CM1 = 20 t ha-1); T6: Bio-N (Cerealine at rate of 4 Kg ha-1); T7: ½NH4NO3+PM2 (PM2 = 5 t ha-1); T8: ½NH4NO3+CM2 (CM2 = 10 t ha-1); T9: ½NH4NO3+Cerealine; T10: ½Urea+PM2; T11: ½Urea+CM2; T12: ½Urea+Cerealine; T13: Cerealine+CM1; T14: ⅓NH4NO3+Cerealine+CM2. Effect of N form on wheat productivity and its quality 25 Tab. 5: Effect of N-fertilizer form applications on the concentrations of heavy metals [lead (Pb), cadmium (Cd), nickel (Ni), chromium (Cr), selenium (Se), cobalt (Co), zinc (Zn), and copper (Cu)] in shoots of wheat plants grown under saline soil conditions Treatments Pb Cd Ni Cr Se Co Zn Cu (mg kg-1 DW) T1 2.85g* 4.16f 2.51g 3.27gh 1.26bcd 2.25f 138i 55.2ij T2 3.86e 5.61d 3.40e 4.41e 1.20de 3.06d 182f 71.6f T3 6.24c 8.96b 5.49c 7.12c 1.25bcde 4.94b 229bc 92.1bc T4 7.38b 9.80a 6.56b 8.39b 1.12e 5.95a 238b 94.8ab T5 2.64gh 3.82fg 2.33gh 3.01h 1.41a 2.11fg 132i 52.6jk T6 2.85g 4.13f 2.50g 3.45g 1.25bcde 2.24f 151h 60.3hi T7 5.75d 8.13c 5.00d 6.70d 1.20de 4.55c 212de 84.3de T8 3.48f 5.05e 3.05f 4.00ef 1.37abc 2.75e 164g 65.1gh T9 3.45f 4.99e 3.10f 3.94f 1.26bcd 2.73e 165g 66.2g T10 7.83a 9.98a 6.82a 9.12a 1.18de 6.03a 256a 98.7a T11 5.70d 8.24c 4.97d 6.58d 1.30abc 4.47c 202e 80.4e T12 5.88d 8.53c 5.16d 6.67d 1.24cde 4.66c 218cd 87.0cd T13 2.52h 3.68g 2.19h 2.86h 1.30abc 1.96g 117j 47.4k T14 2.02i 3.21h 1.81i 2.33i 1.38ab 1.59h 101k 40.8l *Values are means (n = 9), and mean values in each column followed by a different lower-case-letter are significantly different by Fisher’s least-significant dif- ference test (LSD) at P ≤ 0.05. T1: Control (without any N-fertilizer); T2: Recommended N as ammonium nitrate (160 kg N = 476 kg NH4NO3 ha-1 with 33.5 % N); T3: Recommended N as Urea (160 kg N = 348 kg ha-1 urea [(NH2)2CO] with 46 % N); T4: Poultry manure (PM1 = 10 t ha-1); T5: Cattle manure (CM1 = 20 t ha-1); T6: Bio-N (Cerealine at rate of 4 Kg ha-1); T7: ½NH4NO3+PM2 (PM2 = 5 t ha-1); T8: ½NH4NO3+CM2 (CM2 = 10 t ha-1); T9: ½NH4NO3+Cerealine; T10: ½Urea+PM2; T11: ½Urea+CM2; T12: ½Urea+Cerealine; T13: Cerealine+CM1; T14: ⅓NH4NO3+Cerealine+CM2. Tab. 4: Effect of N-fertilizer form applications on the contents of various elements [nitrogen (N), phosphorus (P), potassium (K), calcium (Ca), and sodium (Na)] and the ratios of Ca:Na and K:Na in shoots of wheat plants grown under saline soil conditions Treatments N P K Ca Na Ca/Na K/Na (%) (%) (%) (%) (%) ratio ratio T1 1.85h* 0.24i 1.53h 0.13h 0.30a 0.43h 5.1g T2 2.29defg 0.42gh 2.20efg 0.23e 0.22c 1.05de 10.0def T3 2.26efg 0.42gh 2.16fg 0.21f 0.24b 0.88fg 9.0ef T4 2.24fg 0.40h 2.12g 0.19g 0.24b 0.79g 8.8f T5 2.24fg 0.42gh 2.14g 0.21f 0.22c 0.95ef 9.7def T6 2.22g 0.40h 2.11g 0.19g 0.24b 0.79g 8.8f T7 2.42cd 0.51c 2.41bc 0.25cd 0.18e 1.39c 13.4c T8 2.46c 0.51c 2.40bc 0.26c 0.18e 1.44c 13.3c T9 2.35cdefg 0.46ef 2.29cdef 0.24de 0.22c 1.09d 10.4d T10 2.38cdef 0.48de 2.33cde 0.25cd 0.22c 1.14d 10.6d T11 2.39cde 0.50cd 2.36bcd 0.25cd 0.18e 1.39c 13.1c T12 2.30defg 0.44fg 2.22defg 0.24de 0.22c 1.09d 10.1de T13 2.68b 0.55b 2.49b 0.28b 0.14f 2.00b 17.8b T14 2.91a 0.60a 2.71a 0.31a 0.10g 3.10a 27.1a *Values are means (n = 9), and mean values in each column followed by a different lower-case-letter are significantly different by Fisher’s least-significant dif- ference test (LSD) at P ≤ 0.05. T1: Control (without any N-fertilizer); T2: Recommended N as ammonium nitrate (160 kg N = 476 kg NH4NO3 ha-1 with 33.5 % N); T3: Recommended N as Urea (160 kg N = 348 kg ha-1 urea [(NH2)2CO] with 46 % N); T4: Poultry manure (PM1 = 10 t ha-1); T5: Cattle manure (CM1 = 20 t ha-1); T6: Bio-N (Cerealine at rate of 4 Kg ha-1); T7: ½NH4NO3+PM2 (PM2 = 5 t ha-1); T8: ½NH4NO3+CM2 (CM2 = 10 t ha-1); T9: ½NH4NO3+Cerealine; T10: ½Urea+PM2; T11: ½Urea+CM2; T12: ½Urea+Cerealine; T13: Cerealine+CM1; T14: ⅓NH4NO3+Cerealine+CM2. 26 M.M. Rady, O.H. Mounzer, J.J. Alarcón, M.T. Abdelhamid, S.M. Howladar leaves plant-1, leaf area plant-1 and shoot dry weight plant-1; Tab. 3), positively reflecting in yield parameters. This finding agrees with earlier reports of BAIYERI and TENKOUANO (2007) and NDUKWE et al. (2011) that animal manure is a valuable source of crop nutrients and organic matter, which can improve the soil biophysical condi- tions making the soil more productive and sustainable for plant growth. Growth and yield improvements in plants grown in soil amended with a combination of bio-, organic and inorganic ferti- lizers might be due to nutrient availability, particularly N, P and P ions (MAMAN and MASON, 2013) that agreed with our results ob- tained under salt stress (Tab. 4). Organic manures not only provide nutrients by making them more available for plants but also add to the most important constituent of the soil humus, which provides excellent substrate for plant growth. This could be attributed to the fact that the nutrients in the organic fertilizers were released gra- dually through the process of mineralization (MEHEDI et al., 2012), maintaining optimal soil levels over prolonged periods of time. Some of the organic substances released during the mineralization may act as chelates, which help in the absorption of essential ions and other micro-nutrients (LOBO et al., 2012). They added that, organic mate- rials could have formed complex (or chelate), preventing the preci- pitation of P, reduced the P-sorption capacity of the soil, enhanced P availability, improved P-recovery or resulted in better utilization by plants. Organic materials add carbon into the soil, provides substrate for microbial growth, and subsequent microbial activity. The turn- over resulting from the decomposition of organic materials improves C and N mineralization rates, and enzyme activities, which affect nutrient cycling and availability to the plants (SMITH et al., 1993). In addition, VANLAUWE et al. (2002) reported that the combination of organic and inorganic fertilizers result into synergy and improved conservation and synchronization of nutrient release and crop de- mand, leading to increased fertilizer efficiency and higher growth and yields. The combined treatment of CM (10 t ha-1) + ⅓NH4NO3 Tab. 6: Effect of N-fertilizer form applications on the 1000-grain weight and grain yield of wheat plants grown under saline soil conditions Treatments 1000-grain weight (g) Grain yield ha-1 (ton) T1 29.5d* 3.18g T2 43.9bc 5.40cdef T3 43.6bc 5.32cdef T4 42.7bc 5.26def T5 42.4bc 5.23ef T6 41.0c 5.11f T7 44.5bc 5.68cde T8 44.3bc 5.55cdef T9 44.9bc 5.80c T10 44.2bc 5.63cde T11 44.1bc 5.53cdef T12 44.4bc 5.74cd T13 45.3b 6.66b T14 49.9a 7.85a *Values are means (n = 9), and mean values in each column followed by a dif- ferent lower-case-letter are significantly different by Fisher’s least-significant difference test (LSD) at P ≤ 0.05. T1: Control (without any N-fertilizer); T2: Recommended N as ammonium nitrate (160 kg N = 476 kg NH4NO3 ha-1 with 33.5% N); T3: Recommend- ed N as Urea (160 kg N = 348 kg ha-1 urea [(NH2)2CO] with 46% N); T4: Poultry manure (PM1 = 10 t ha-1); T5: Cattle manure (CM1 = 20 t ha-1); T6: Bio-N (Cerealine at rate of 4 Kg ha-1); T7: ½NH4NO3+PM2 (PM2 = 5 t ha- 1); T8: ½NH4NO3+CM2 (CM2 = 10 t ha-1); T9: ½NH4NO3+Cerealine; T10: ½Urea+PM2; T11: ½Urea+CM2; T12: ½Urea+Cerealine; T13: Cerealine+CM1; T14: ⅓NH4NO3+Cerealine+CM2. Tab. 7: Effect of N-fertilizer form applications on the concentrations of heavy metals [lead (Pb), cadmium (Cd), nickel (Ni), chromium (Cr), selenium (Se), cobalt (Co), zinc (Zn), and copper (Cu)] in harvested grains of wheat plants grown under saline soil conditions Treatments Pb Cd Ni Cr Se Co Zn Cu (mg kg-1 DW) T1 1.31de* 1.40de 1.89fgh 1.18e 0.93c 0.38g 141efg 56.3efg T2 1.36de 1.48d 1.99def 1.24de 0.87cd 0.42f 150ef 59.8e T3 4.26c 2.34c 4.63c 2.58c 0.71e 0.58c 194c 77.5c T4 6.12b 2.99b 6.02b 3.16b 0.64f 0.79b 212b 84.6b T5 1.20e 1.26e 1.75hi 1.04f 1.00b 0.33h 128h 51.4g T6 1.30de 1.38de 1.85fghi 1.18e 0.92c 0.38g 140fg 56.1efg T7 1.42d 1.56d 2.09d 1.30d 0.81d 0.47d 168d 67.2d T8 1.30de 1.39de 1.84ghi 1.19de 0.92c 0.38g 141efg 56.0efg T9 1.33de 1.43de 1.92efg 1.20de 0.90c 0.40fg 144ef 57.6e T10 7.10a 3.52a 7.31a 3.48a 0.58f 0.91a 244a 94.9a T11 1.36de 1.49d 1.98defg 1.24de 0.87cd 0.43ef 152e 60.7e T12 1.40d 1.55d 2.07de 1.29de 0.82d 0.46de 170d 68.1d T13 1.20e 1.24e 1.72i 1.04f 1.02b 0.32h 131gh 52.1fg T14 1.01f 0.93f 1.53j 0.91g 1.11a 0.27i 114i 46.2h *Values are means (n = 9), and mean values in each column followed by a different lower-case-letter are significantly different by Fisher’s least-significant dif- ference test (LSD) at P ≤ 0.05. T1: Control (without any N-fertilizer); T2: Recommended N as ammonium nitrate (160 kg N = 476 kg NH4NO3 ha-1 with 33.5% N); T3: Recommended N as Urea (160 kg N = 348 kg ha-1 urea [(NH2)2CO] with 46% N); T4: Poultry manure (PM1 = 10 t ha-1); T5: Cattle manure (CM1 = 20 t ha-1); T6: Bio-N (Cerealine at rate of 4 Kg ha-1); T7: ½NH4NO3+PM2 (PM2 = 5 t ha-1); T8: ½NH4NO3+CM2 (CM2 = 10 t ha-1); T9: ½NH4NO3+Cerealine; T10: ½Urea+PM2; T11: ½Urea+CM2; T12: ½Urea+Cerealine; T13: Cerealine+CM1; T14: ⅓NH4NO3+Cerealine+CM2. Effect of N form on wheat productivity and its quality 27 + Cerealine was found to significantly increase wheat growth and productivity and significantly decrease plant and grains heavy metals concentrations. These results show that it is important to choose the suitable N source and its suitable amount to sustain the agricultural productivity, by minimizing the concentrations of heavy metals in the edible parts for human nutrition and health. 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