Effect of different phosphorus and sulfur levels on nitrogen fixation and uptake by chickpea (Cicer arietinum L.)
Efecto de diferentes niveles de fósforo y azufre en el nitrógeno consumido y fijado por el garbanzo (Cicer arietinum L.)
Muhammad Islam1*, Saleem Mohsan1, Safdar Ali2
1 National Fertilizer Development Centre, Street # 1 Sector H-8/1 Islamabad-44000 Pakistan.*Author for correspondence: ( islamuaf@gmail.com).
]]> 2 Department of Soil Science and Soil and Water Conservation, Arid Agriculture University, Rawalpindi- 46300 Pakistan.
Received: april, 2011.
Approved: december, 2011.
Abstract
Nitrogen fxing capacity of leguminous plants can be increased by the supply of adequate amounts of nutrients, especially phosphorus and sulfur. Some nutrients have direct involvement in the process of nitrogen fixation while others afect by improving growth of host plant. Field experiments were conducted at two different locations in northern rainfed Punjab, Pakistan, to assess the amount of nitrogen fixation and nitrogen uptake by chickpea (Cicer arietinum L.) during crop growing seasons 2006-2007 and 2007-2008. Treatments were: phosphorus levels (0, 40 and 80 kg P2O5 ha-1); sulfur sources (gypsum and ammonium sulfate) and levels (0, 15 and 30 kg S ha-1). The experimental design was randomized complete block with split-split plot arrangement: phosphorus levels in main plots, sulfur sources in sub-plots and sulfur levels in sub-sub-plots. Phosphorus (80 kg P2O5 ha-1) increased nitrogen fixed and uptake by 33 and 31 % over control. However, effect of phosphorus on nitrogen derived from atmosphere was not signifcant while that of sulfur was signifcant. Sulfur (30 kg S ha-1) increased nitrogen derived from atmosphere, amount of nitrogen fixed and nitrogen uptake by 6, 25 and 17 % over control. Ammonium sulfate treatments caused signifcantly higher amount of nitrogen fixed and higher nitrogen uptake as compared to gypsum treatments, although both sulfur sources were similar regarding percent nitrogen derived from atmosphere. A strong positive correlation (R= 0.98**) between amount of nitrogen fixed and nitrogen uptake at both locations was observed. There was a direct effect of sulfur on the process of nitrogen fixation, whereas phosphorus mainly afected growth of chickpea.
Key words: Cicer arietinum L., ammonium sulfate, gypsum, natural abundance technique, percent nitrogen derived from atmosphere.
Resumen
]]> La capacidad de fijación del nitrógeno de las leguminosas puede aumentar suministrando cantidades adecuadas de nutrientes, especialmente fósforo y azufre. Algunos nutrientes participan directamente en el proceso de fijación del nitrógeno, mientras que otros mejoran el crecimiento de la planta huésped. Los experimentos de campo se llevaron a cabo en dos lugares diferentes en la zona norte de temporal, en Punjab, Pakistán, para evaluar la cantidad de nitrógeno fijado y la absorción de nitrógeno por el garbanzo (Cicer arietinum L.) durante las temporadas de cultivo 2006-2007 y 2007-2008. Los tratamientos fueron: fósforo (0, 40 y 80 kg de P2O5 ha-1), fuentes (yeso y sulfato de amonio) y niveles de azufre (0, 15 y 30 kg S ha-1). El diseño experimental fue de bloques completos al azar con arreglo de parcelas sub-subdivididas: niveles de fósforo en las parcelas principales, fuentes de azufre en las sub-parcelas y niveles de azufre en las sub- sub-parcelas. El fósforo (80 kg P2O5 ha-1) aumentó la absorción y fijación de nitrógeno en 33 y 31 % sobre el testigo. Sin embargo, el efecto del fósforo en el nitrógeno derivado de la atmósfera no fue signifcativo, mientras que el del azufre sí lo fue. El azufre (30 kg S ha-1) aumentó el nitrógeno derivado de la atmósfera, la cantidad de nitrógeno fijado y su absorción en 6, 25 y 17 % sobre el testigo. Los tratamientos con sulfato de amonio aumentaron signifcativamente la cantidad de nitrógeno fijado y la absorción de nitrógeno comparados con los tratamientos de yeso, aunque ambas fuentes de azufre fueron similares con respecto al porcentaje de nitrógeno proveniente de la atmósfera. Hubo una fuerte correlación positiva (R= 0.98**) entre la cantidad de nitrógeno fijado y la absorción de nitrógeno en ambas localidades. Hubo un efecto directo del azufre en el proceso de fijación de nitrógeno, mientras que el fósforo afectó principalmente el crecimiento del garbanzo.Palabras clave: Cicer arietinum L., sulfato de amonio, yeso, técnica de abundancia natural, porcentaje de nitrógeno proveniente de la atmósfera.
INTRODUCCIÓN
Chickpea (Cicer arietinum L.) is an important pulse crop of rainfed areas in semiarid/arid climate. Average chickpea yield in Pakistan is 685 kg ha-1 (Government of Pakistan, 2010) which is very low compared to China (2.4 Mg ha-1), Canada (1.9 Mg ha-1) and USA (1.7 Mg ha-1) (FAO, 2009). This low yield is due to genetic, agronomic and environmental factors and inadequate fertilization is the key among them.
Sulfur (S) is becoming defcient in soils due to introduction of high yielding varieties, use of high grade S free fertilizers and reduced emission of S from industrial units (Khalid et al., 2009a; Scherer, 2009). Therefore, it is important to study the changes in concentration of this element under different conditions of soils, climate, crop species and cropping systems.
An adequate supply of mineral nutrients to legumes enhances nitrogen (N) fixation (Ganeshamurthy and Reddy, 2000). Thus, S availability increases tissue N concentration of many legumes (Claro-Cortes et al., 2002). Whether this is due to a direct effect on symbiotic N fixation or an effect on the host plant growth is not very clear. Furthermore, S defciency may afect N fixation because it is a constituent of ferredoxin and enzymes, such as nitrogenase (Fukuyama, 2004).
In Pakistan, studies about crop response to S application are limited to oilseeds and their oil contents. Research regarding interaction of phosphorus (P) and S and their role in legume's growth, N fixation and nutrient uptake is very rare (Islam et al., 2009). It may be speculated that supply of adequate amount of nutrients to legumes may result in increased amount of N fixation. Therefore, this study was conducted to assess the interactive effect of S and P application on N fixation and N uptake by chickpea crop under rainfed conditions of northern Punjab, Pakistan.
MATERIALS AND METHODS
]]> Field experiments were conducted using chickpea cultivar Balkassar 2000 at: 1) Barani Agricultural Research Institute (BARI), Chakwal (sandy loam, pH 7.6, AB-DTPA extractable P 3.0 mg kg-1, CaCl2 extractable SO4-S 6.4 mg kg-1); and 2) a farmer's field Talagang, district Chakwal (loamy sand, pH 7.7, AB-DTPA extractable P 1.4 mg kg-1, CaCl2 extractable SO4-S 7.5 mg kg-1); during crop growing seasons 2006-2007 and 2007-2008. Both experimental sites are located at 32.5 °N, 72.4 °E. The experimental design was a randomized complete block with a split-split-plot arrangement (plot size 1.5×3.5 m at BARI Chakwal, and 1.8×4 m at farmer's field Talagang); P rates (0, 40 and 80 kg P2O5 ha-1) were in main plots, S sources in sub-plots (gypsum and ammonium sulfate) and S rates (0, 15 and 30 kg S ha-1) in sub-sub-plots. As a result, there were 18 treatments which were replicated three times. Starter dose (26 kg ha-1) of N was applied as urea; in S treatments, urea dose was adjusted after taking into consideration the addition of N from ammonium sulfate (AS). Phosphorus was applied as triple super phosphate. All the fertilizers were applied as basal dose. Chickpea crop was sown maintaining 30 cm row to row distance. Crop was grown under rainfed conditions and no supplemental irrigation was applied. Samples of plant dry matter tissue of legume and non legume reference plant were taken for δ15N determination (Unkovich et al., 2008).Nitrogen derived from atmosphere (Ndfa) =100 × (δ15N (soil N) - δ15N legume N)/ [δ 15N (soil N)-B]
where d15N (soil N) is commonly obtained from a non N fixing reference plant grown in the same soil as the legume; B is the δ15N of the same N fixing plant when grown with N as the sole source of N and its value is -2.0 (Kyei-Boahen et al., 2002).
Legume N uptake (kg ha-1) = legume dry matter yield (kg ha-1) × N in plant tissue (%)
Amount of N fixed (kg ha-1) =legume N uptake (kg ha-1) × Ndfa × 1.5*
* 1.5 factor was used to include an estimate for contribution by underground N (Rochester et al., 1998).
At physiological maturity, crop from 1 m2 in the middle of each plot was harvested separately. The plant samples were dried and data were recorded for seed, straw and total dry matter yield. Representative samples of 100 g from both seed and straw were collected from bulk sample, oven dried and ground and analyzed for N (Ryan et al., 2001).
Data on all observations were subjected to analysis of variance (ANOVA) by using software MSTATC. Treatment means were compared by least signifcant diference (LSD; p≤0.05) test.
Simple linear correlation analysis was also performed to study the relationship between variables.
]]> RESULTS AND DISCUSSION
The precipitation during crop growing season (October to March) at experimental sites (Chakwal and Talagang) was 385 and 362 mm during the first crop growing season (2006-2007), and 90 and 30 mm during the second crop growing season (2007-2008). The long term average (1977-2009) annual precipitation is 630 mm for Chakwal and 450 mm for Talagang. Tw o third of total rainfall occurs in moonsoon (July to September).
Two locations and years differed significantly with respect to N fixation and uptake (Table 1). Percent N derived from atmosphere, amount of N fixed and N uptake were signifcantly higher in the frst growing season, as compared to the second one, due to the occurrence of a larger rainfall in both locations during 2006-2007, which is similar to results reported by Hayat et al. (2008). Habtemichial et al. (2007) also observed that during wetter crop growing season, there was better nodulation and nitrogen uptake by faba bean (Vicia faba L.) crop. Likewise, amount of N fixed and N uptake were higher at Talagang as compared to Chakwal (Table 1). Low level of total organic carbon (1.8 mg g-1) and nitrate N (5.6 ug g-1) in Talagang soil before starting the experiment might be a reason for the higher amount of N fixed.
The year × location interaction effect was significant. The treatment (P levels, S sources and S levels) × location and treatment × year interaction effects were significant for amount of N fixed and N uptake, but they did not affect Ndfa. Therefore, the results are presented for each year and location separately.
Nitrogen derived from atmosphere
Treatment with P did not change (p>0.05) Ndfa at Chakwal (Table 2), and a similar trend was observed at Talagang during 2007-2008. However, during 2006-2007 P application (80 kg P2O5 ha-1) resulted in a signifcant increase of Ndfa. Analysis of data pooled over location and year did not show any signifcant effect of P application on Ndfa.
The observations with chickpea regarding Ndfa appear to support the finding that P deficiency affects the process of N fixation through its secondary effects on plant growth, rather than a direct involvement in nitrogenase functioning. Similar results regarding effect of P application on Ndfa were reported in chickpea by Islam and Ali (2009) and white lupin (Lupinus albus) by Schulze et al. (2006). However, contrary to these findings, Hayat et al. (2008) report an increase up to 32 % in Ndfa due to application of P (80 kg P2O5 ha-1) using mung bean (Vigna radiate) and mash bean (Vigna mungo) as test crop. There is an increase in nodule number and nitrogenase activity with P application which results in an increased Ndfa (Fatima et al., 2007). Similar to our findings, Tang et al. (2001) did not observe any role of P in nodule functioning and nitrogenase activity, but the amount of N fixed increased with P application in bean (Phaseolus vulgaris). It seems that the role of P in N fixation varies with crop, growing conditions and time of measurement.
Results (Table 2) show no signifcant effect (p>0.05) of S sources on N derived from the atmosphere. Results regarding effect of different S sources on Ndfa, have not been reported previously, although there are comparisons about their effect on overall plant growth. Ryant and Skladanka (2009) point out that gypsum and ammonium sulfate show similar effects on dry matter yield of forage grass and soil S status after crop harvest. Application of S signifcantly increased Ndfa at both locations, except during 2007-2008 at Talagang (Table 2). Rainfall was very low (30 mm) during the second crop growing season at Talagnag, as compared to first one (365 mm). Drought stress resulted in retarded growth of plant and lower response to S application. There was no effect (p>0.05) of S level at Chakwal, but there were diferences (p≤0.05) at Talagang during 2006-2007. Higher dry matter yield at Talagnag during 2006-2007 (data not shown) due to favourable climatic conditions might have resulted in higher S requirement of plant.
]]> Increase in Ndfa due to S application is in agreement with the findings of Habtemichial et al. (2007) that the Ndfa increased from 55 to 70 % in pods and from 44 to 56 % in faba bean (Vicia faba) straw when applying 30 kg S ha-1 as potassium sulfate. Sulfur application enhances N requirement of plant which results in increased nodule number, nodule weight, Ndfa and amount of N fixed (Jamal et al., 2010a, 2010b). Scherer et al. (2006) report similar results for peas (Pisum sativum) and they conclude that N fixation is very sensitive to photosynthetic assimilates and reduced supply of carbohydrate results in low N fixation in S starved plants.Amount of nitrogen fixed
Phosphorus application signifcantly increased the amount of N fixed at both locations (Table 3), and response to P application was higher (36 % increase over control) at Talagang as compared to Chakwal (30 % increase over control). This higher response of Chickpea crop at Talagang might be due to the fact that Talagang soil had lower available P as indicated by fertility status of soil samples taken before starting the experiment.
Data pooled over locations and years indicated that amount of N fixed increased from 42 to 56 kg ha-1 as a response to P application (0 to 80 kg ha-1 (Table 3), which was mainly due to improvement in host plant growth. These results are in line with the findings of Somado et al. (2006) who observed that neither nodulation nor Ndfa was significantly affected by P application, but rather total N accumulation was enhanced due to an improved biomass yield.
Ammonium sulfate treated plots had higher amount of N fixed as compared to gypsum treatments at both locations (Table 3), since the effect of applying 30 kg S ha-1 as gypsum was signifcantly lower than 30 kg S ha-1 as ammonium sulfate (Table 4). This lower response of chickpea to gypsum application may be due to slow release of S from this source (Girma et al., 2005). After several experiments, Ghosh et al. (2000) conclude that for immediate S defciency relief, readily soluble sources like ammonium sulfate outclassed less soluble sources such as gypsum; they also observed that in calcareous soils, gypsum was less efective as compared to ammonium sulfate.
Sulfur treatments were in descending order of S2 (30 kg S ha-1)> S1 (15 kg S ha-1) > S0 (0 kg S ha-1) regarding amount of N fixed at both locations (Table 3). Increases in amount of N fixed due to S application were reported by Scherer et al. 2008. Sulfur deficiency results in low leghemoglobin content which might be one of the reasons for low nitrogenase activity (Scherer, 2008). Lower and higher levels of P and S difered significantly regarding amount of N fixed, since the P×S level interaction was significant at Chakwal but not at Talagang. Data pooled over locations and years indicated that the highest amount of N (61 kg a-1) was fixed with treatment P2S2 (80 kg P2O5 and 30 kg S ha-1) followed by P2S1 (80 kg P2O5 and 15 kg S ha-1) and the lowest in control (Table 6).
Nitrogen uptake
Different P and S levels had significant effect on N uptake at both locations (Table 5). Data pooled over locations and years indicated that N uptake increased from 55 to 72 kg ha-1 and from 59 to 69 kg ha-1 as P application rate was increased from 0 to 80 kg P2O5 ha-1 and S application rate from 0 to 30 kg S ha-1. Increase in N uptake due to P application confirms the findings of Shu-Jie et al. (2007) and Reed et al. (2007). Tis increment in N uptake might be attributed to a better plant growth.
Sulfur sources significantly affected (p≥0.05) N uptake which was higher for ammonium sulfate as compared to gypsum (Table 5). The treatment receiving 30 kg S ha-1 as gypsum was significantly lower than 30 kg S ha-1 as ammonium sulfate (Table 7). Contrary to these findings, Khalid et al. (2009b) report no significant difference between gypsum and ammonium sulfate regarding seed yield of Brassica napus, although ammonium sulfate caused a higher S concentration in plant tissue. The effect of different S sources also varies with the crop, soil and climatic conditions. Gypsum and single super phosphate are superior to AS in coarse textured soils due to benefcial role of calcium and due to less leaching losses as sulfate (Till, 2010). In our study ammonium sulfate was better than gypsum due to: 1) low rainfall caused less leaching of sulfate from ammonium sulfate especially during 2007-2008; and 2) soils in both locations were S deficient and therefore S was needed immediately.
The P by S level interaction was significant at Chakwal but not at Talagang when data were pooled over locations and years. Statistical analysis of the data combined over location and years indicated that the highest N uptake (76 kg ha-1) was recorded for P2S2 (80 kg P2O5 and 30 kg S ha-1) followed by P2S1 (80 kg P2O5 and 15 kg S ha-1) and the lowest in control (Table 6).
]]> Increased N uptake as a result of S application may also be due to an increment in protein synthesis and then in photosynthesis (Zhao et al., 2008). In the absence of S, amino acids cannot be transformed into proteins, which results in reduced N acquisition (Varin et al., 2009). In different crops there is a significant positive correlation of N and S content with protein content in seeds (Bahmanyar and Poshtmasari, 2010). In fact N fixation and N uptake show a direct relationship which is also confrmed by a strong positive correlation between these two variables (Table 8).
CONCLUSIONS
Application of P and sulfur significantly increased N fixed and N uptake. The effect of P application on N fixation was due to an effect on host plant growth, whereas sulfur was directly involved in N fixed since a deficiency of this element will reduce Ndfa. Ammonium sulfate was a more efficient source of S as compared to gypsum regarding the effect on N fixed. Sulfur should be included in fertilizer recommendation along with P in order to optimize sustainability of soil resources.
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