Fitociencia

 

Effect of integrated application of sulphur and phosphorus on nitrogen fixation and nutrient uptake by Chickpea (Cicer arietinum L.)

 

Efecto de la aplicación de azufre–fósforo sobre la fijación de nitrógeno y la captación de nutrientes en garbanzo (Cicer arietinum L.)

 

Muhammad Islam^*, Safdar Ali

 

Department of Soil Science & SWC, PMAS– Arid Agriculture University, Rawalpindi–46300 Pakistan. *Autor responsable: (islamuaf@gmail.com)

 

Recibido: Julio, 2008.
Aprobado: Julio, 2009.

 

ABSTRACT

Nitrogen fixing capability of legumes can be enhanced by the supply of adequate amounts of nutrients, especially phosphorus (P) and sulphur (S). However, combined application of P and S may have synefgistic oí antognistic effects on crop yield, depending upon initial fertility status of soil, level of nutrients applied and test crop used. Field experiments were conducted at two different locations (Barani Agriculture Research Institute, Chakwal, and farmer's field, Talagang) in northern fainfed Punjab, Pakistan, to assess nodulation, nitrogen fixation and nutrient uptake by chickpea (Cicer arietinum L.) in response to application of P (0, 40 and 80 kg ha^–1) and of S (0, 15 and 30 kg ha^–1) in different combinations. Application of P and S significantly increased seed yield (27 to 41 % and 7 to 11 %) over control. Sulphur application increased N derived from ah– (% N_dfa) by 5 to 7 % over control; however, P application had no significant effect on N_dfa. Both P and S application increased N fixation (16 to 40 % and 15 to 22 %) over control. Nutrient uptake (N, P and S) increased significantly with the application of both P and S. There was a wreak correlation between P uptake and % N_dfa (R=0.28*) but a positive correlation between S uptake and % N_dfa (R=0.76**). Phosphorus (R=0.56") and sulphur (R=0.97**) uptake correlated positively with N fixation.

Key words: Legume, mineral nutrition, natural abundance technique, nitrogen derived from air.

 

RESUMEN

La capacidad de las leguminosas para asimilar el nitrógeno puede incrementarse si se suministran cantidades adecuadas de nutrientes, específicamente fósforo (P) y azufre (S). Sin embargo, la aplicación combinada de P y S puede tener efectos sinérgicos o antagónicos en el rendimiento del cultivo, dependiendo del estatus inicial de fertilidad del suelo, del nivel de nutrientes suministrados, así como del cultivo de prueba utilizado. Los experimentos de campo se realizaron en dos áreas (Barani Research Institute, Chakwal, y el campo agrícola de Talagang) de secano en el norte de Punjab, Pakistán, para evaluar nodulación, fijación de nitrógeno (N) y captación de nutrientes en el garbanzo (Cicer arietinum L.) en respuesta a la aplicación de P (0, 40 y 80 kg ha^–1) y de S (0, 15 y 30 kg ha^–1) en distintas combinaciones. La aplicación de P y S incrementó significativamente el rendimiento de semilla (27 % a 41 % y 7 % a 11 °/o) respecto al testigo. La aplicación de S aumentó el N derivado del aire (% N_dfa) en 5 % respecto al testigo; sin embargo la aplicación de P no tuvo un efecto significativo en N_dfa. La aplicación tanto de P como de S incrementó la fijación de N (16 % a 40 % y 15 % a 22 %) respecto al testigo. La captación de nutrientes (N, P y S) aumentó significativamente con la aplicación tanto de P como de S. Hubo una ligera correlación entre la captación de P y % N_dfa (R = 0.28*), y una correlación positiva entre la captación de S y % N_dfa (R=0.76**). La captación tanto de P (R=0.56**) como de S (R=0.97**) se correlacionó positivamente con la fijación de N.

Palabras clave: Leguminosas, nutrición mineral, método de abundancia natural, nitrógeno derivado del aire.

 

INTRODUCTION

Chickpea is the second most important pulse crop in the world, grown in at least 33 countries. Chickpea was grown on 1046 000 ha in Pakistan during 2007–08, with a production of 823 000 Mg (Economic Survey, 2008). Average yield was 787 kg ha^–1 which is very low as compared to the USA (1391 kg ha^–1), Canada (1427 kg ha^–1) and China (4135 kg ha^–1) (FAO, 2006). This low yield of the crop in Pakistan is primarily due to genetic, agronomic and economic factors. From an agronomic point of view, nutritional deficiency is likely to depress its nitrogen–fixing role and may in turn limit crop yield.

An adequate supply of mineral nutrients to legumes enhances nitrogen fixation (Ganeshamurthy and Reddy, 2000). For example, starter N stimulated early seedling growth and nodulation (Daramola et al, 1982); P and sulphur improve nodulation activity (Olivera et al, 2004; Scherer et al, 2008). Sulphur deficiency decreases the concentration of N in the shoots 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 plants is not very clear. In pot experiments with different legumes, Scherer and Lange (1996) found a lower N accumulation and a yield reduction when S was limiting. Therefore, S affects leguminous plant species growth through its effect upon N fixation (Varin et al, 2009). With S deficiency, amino acids and other N forms accumulate due to a lack of proteins synthesis which may have a feed–back repression on N fixation (Ahmad and Abedin, 2000). Furthermore, S deficiency may affect N fixation because of the relatively high S content of the nitrogenase (Mortenson and Thornley, 1979) and of ferredoxin (Fukuyama, 2004). It may be speculated that the synthesis of photosynthates is limited under S deficiency conditions, resulting in a reduced N₂ fixation (Scherer et al, 2006).

Under semarid conditions of Ethopia, an increase by 53 % and 95 % relative to control has been reported for nodule number and nodule fresh weight with application of 30 kg S ha^–1 using faba bean (Vicia faba L.) as test crop in a S deficient soil, whereas S application enhanced N_dfa from 55 % to 72 % and N fixation from 49 to 100 kg N ha^–1(Habtemichial et al, 2007). In England, three–fold increases in nodule weight and two–fold increases in N₂ fixation have been recorded in pot experiments on peas (Pisum sativum L.) (Scherer et al., 2006).

In many soil types especially calcareous soils, P is the most limiting nutrient for the production of crops (Jiang et al, 2006). As nitrogen fixing plants, legumes require P for adequate growth and nodulation (Tang et al, 2001a). The influence of phosphorous on symbiotic nitrogen fixation in legumes has received considerable attention, but its role in the process remains unclear (Tsvetkova and Georgiev, 2003). Somado et al. (2006) applied 90 kg P₂O₅ ha^–1 and recorded four–fold increases in N uptake of Crotalaria micans; they concluded that P application increased total N accumulated rather than % N_dfa.

Nutrient requirement of the crops depends upon many factors and balance between different nutrients is of great importance, since there may be synergistic or antagonistic effects of one nutrient on the other. Application of S increased N uptake in chickpea by 44.26 %, in black gram by 41.99 %, in linseed by 10.50 % and in wheat by 37–55 % (Dwivedi et al, 1996). Increase in P uptake has been observed with application of S containing fertilizers (Goos and Johnson, 2001; Togay etal, 2008).

In Pakistan, research on crop response to S application has been limited to oilseeds and their oil contents only. Research work regarding integrated use of P and S and their role in legume N fixation and nutrient uptake is very scarce. Therefore, the present study was conducted to assess the interactive effect of S and P application on nitrogen fixation and nutrient uptake by chickpea crop under rainfed conditions of Pothwar Plateau, Pakistan.

 

MATERIALS AND METHODS

Field experiments were conducted with chickpea at Barani Agriculture 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) 32.5°N, 72.4° E; and farmers field, Talagang (loamy sand, pH 7.7, AB–DTPA extractable P 1.4 mg kg^–1, CaCl₂ extractable SO₄–S 7.5 mg kg^–1) 32.5° N, 72.2° E, during the crop cycle 2006–07. The trial was carried out according to split plot arrangement in randomized complete block design (1.5×3.5 m plot at BARI, Chakwal; and 1.8×4 m farmer's field, Talagang) with P rates in main plots and S rates in subplots. There were nine treatments with combinations of P (0, 40, 80 kg ha^–1) and S rates (0, 15, 30 kg ha^–1). Starter dose of N (26 kg ha^–1) was applied as urea. Phosphorus was applied as triple super phosphate (TSP) and S as gypsum. All the treatments were replicated three times. Chickpea crop was sown in mid October, maintaining 30 cm row to row distance. All the fertilizers were applied at the time of sowing. Crop was grown under rainfed conditions and no irrigation water was applied. Rainfall data were recorded for both locations (Figure 1). Harvesting was done in the first week of May. Crop from a 1 m² area in the middle of each plot was harvested separately and data were recorded for seed yield after threshing. Representative samples of 100 g for both straw and grain separately were collected from bulk sample, oven dried, ground and analyzed for N and P (Ryan et al, 2001) and S (Verma et al, 1977). Titrate obtained from N determination was concentrated and used for (δ¹⁵N measurement by mass spectrometer (Peoples et al., 1989):

% N_dfa =100 × (δ¹⁵N (soil N) – δ¹⁵N legume N)/ (δ¹⁵N (soil N)–B)

where δ¹⁵N (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 –2.5 (Shah et al, 1997).

Legume kg N ha^–1 = legume total biomass kg ha^–1 × % N in plant

kg N fixed ha^–1=legume kg N ha^–1 × % N_dfa × l.5*

*1.5 factor was used to include an estimate for contribution by underground N (Peoples et al., 1989).

Data on all observations were subjected to analysis of variance (ANOVA) using the software MSTATC. Treatment means were compared by orthogonal contrasts.

 

RESULTS AND DISCUSSIONS

Seed yield

There was a significant interaction between P and S on seed yield at Chakwal but not at Talagang (Table 1). Different levels of P and S had significant effect on seed yield at both locations. Contrast of P₀ vs. P₁ and P₂ and P₁ vs. P₂ showed significant differences regarding seed yield being the highest (1380 kg ha^–1) in P₂S₂, less (1368 kg ha^–1) in P₂S₁ and least (942 kg ha^–1) in P₀S₀ (Tables 2 and 3).

Increase in seed yield due to P application is similar to those found by Aslam et al. (2000) and Hayat (2005). Adequate P nutrition enhances many aspects of plant physiology including photosynthesis, flowering, fruiting and maturation (Brady and Weil, 2005). Increase in yield due to S application may be due to the fact that S is related to photosynthesis of plants. Sulphur application increased rate of photosynthesis due to an increment in protein synthesis and maintenance of high chlorophyll content (Ahmad and Abedin, 2000). Similar results of increased seed yield due to application of S were found by Srinivasarao et al. (2004) and Raina and Tanawade (2005).

Nitrogen derived from air (% N_dfa)

Regarding % N_dfa , similar results were recorded at both locations (Table 1). The main effect of P as well as P by S interaction was not significant on % N_dfa . Sulphur application (15 and 30 kg ha^–1) resulted in significant increase of % N_dfa by 5 and 7 % at both locations (Table 4). Comparison of S₀ with S₁ and S₂ exhibited significant difference at both locations (Table 2). Difference between S₁ and S₂ was not significant at Chakwal, but it was significant at Talagang. These results are also confirmed by the correlation analysis as there was positive correlation between % N_dfa and S uptake (R = 0.76"), but R value was low (0.28*) between P uptake and % N_dfa (Table 9).

Similar results have been reported by Scherer and Lange (1996), Zhao et al. (1999) and Scherer et al. (2006). With S deficiency amino acids accumulate and protein cannot be synthesized, which may inhibit N fixation (Varin et al, 2009). Sulphur increased nitrogenase activity because of higher ferredoxin and ATP concentration in bacterioids of root nodules of legumes (Scherer et al., 2008). Increase in N fixation due to P application is comparable to those reported by Sanginga et al. (1996), Ankomah et al. (1996) and Somado et al. (2006). Phosphorous application affected total N accumulated rather than % N_dfa, which was due to an increase in growth of host plant due to P application rather than direct involvement of P in N fixation.

Nitrogen fixation

Individual effect of P and S was significant but combined application had no significant effect on N fixation at both locations (Table 1). Application of P (40 and 80 kg ha^–1) increased N fixation by 26 % and 40 % and 16 % and 40 % at Chakwal and Talagang (Table 4). Orthogonal contrast showed that higher (80 kg P₂O₅ ha¹) and lower level (40 kg P₂O₅ ha^–1) of P application differed significantly from each other as well as from P₀ (Table 2). Similarly, an increase of 15 % and 19 % and 16% and 22 % was recorded with S application (15 and 30 kg ha^–1) at Chakwal, and Talagang (Table 4). Lower (30 kg S ha^–1) and higher level (30 kg S ha^–1) of S were similar to each other but differed significantly from S₀ (Table 2). Furthermore, there was a positive correlation of both P (R=0.56**) and S uptake (R=0.97**) with N fixation (Table 9).

Phosphorus fertilization of legume species leads to increased nitrogenase activity, nodule number, nodule mass and plant N accumulation (Shu–Jie et al., 2007). Deteriorated phosphate metabolism leads to impairment of photosynthetic and nitrogen fixing metabolism (Liao and Yan, 1999). Nitrogen fixation is directly affected by S as it is an essential component of the ferredoxins, an iron–sulphur protein occurring in the chloroplasts. Ferredoxin has a significant role in NO₂^– and SO₄^–2 reduction, the assimilation of N₂ by root nodule bacteria and free living N—fixing soil bacteria (Havlin et al, 2007). Besides, root and nodule development on the roots of legumes is promoted by sulphur fertilization (Scherer, 2008; Scherer etal, 2008).

Nutrient uptake

Effect of P and S application was significant on N, P and S uptake at both locations (Table 5). Orthogonal comparison of P₀ vs. P₁ and P₂ indicated significant difference for nutrient uptake, but P₁ vs. P₂ showed no differences regarding P uptake (Table 6). Phosphorus application increased N uptake by 25 % and 38 % and 15 % and 36 % at Chakwal and Talagang (Table 7). There was a positive correlation of N uptake with both P (R = 0.65**) and S uptake (R = 0.97**) (Table 9). Shu–Jie etal. (2007) and Reed etal. (2007) reported similar results concerning increase in N uptake due P application. Increase in N uptake might be due to a better plant growth, especially roots. Root growth, and particularly development of lateral roots and fibrous rootlets, is encouraged by addition of P (Lopez–Bucio etal, 2002). Adequate P nutrition enhances many aspects of plant physiology including photosynthesis (Brady and Weil, 2005). This leads to increased N assimilation, and as a result, increased N uptake by the plant.

Increase in N uptake due to S application has been reported by Scherer et al. (2006) and Tiwari and Gupta (2006). Sulphur can readily alter primary and lateral root growth, modifying the overall root architecture (Kutz et al, 2002). In fact, nutrient ions act as signaling molecules for the regulation of root development (Lopez– Bucio et al, 2003). Increased N uptake may also be due to increased protein synthesis and subsequent photosynthesis (Sexton et al., 1998). Hence, it may be argued that increased sulphur availability resulted in better root and shoot growth and ultimately increased N uptake due to increased dry matter production.

There was a positive correlation between P uptake with S uptake (R=0.57**) (Table 9). Increase in P uptake due to P application was due to its increased availability in soil, a result similar to those reported by Amrani et al. (2001) and Kanwar and Paliyal (2002). Phosphorus application not only increased biomass production but also increased P concentration in plant tissue (Shu–Jie et al, 2007).

Increase in P uptake due to S application has been already reported by Tiwari and Gupta (2006). Besides, Bagayoko et al. (2000) and Scherer et al. (2008) have shown a positive effect of P and S application on root growth and morphology. Due to the acidifying effect of S oxidation, the availability of other nutrients like N and P is also influenced (Togay et al, 2008) Application of P and S increased uptake of these nutrients by the plant, which may be due to their increased availability in soil. It may be argued that better root growth resulted in increased uptake of these nutrients from soil (Zhao et al, 2008).

The highest increases among nutrients were recorded regarding S uptake up to 54 % and 22 % due to P and S application (Table 7). Lower (15 kg S ha^–1) and higher (30 kg S ha^–1) level of S application differed from S₀ regarding nutrient uptake but were similar to each other (Table 6). The P×S interaction was not significant regarding N and P uptake but showed significant effect on S uptake at Talagang (Table 5). The maximum S uptake (10.76 kg ha^–1) was recorded in P₂S₁ which was at par with P₂S₂ (10.19 kg ha^–1) and minimum (6.12 kg ha^–1) in P₀S₀ (Table 8). Bagayoko et al. (2000) and Tang et al. (2001b) have shown the positive effect of P and S application on root growth and morphology. Application of P and S increased uptake of these nutrients by the plant which may be due to their increased availability in soil. It may be argued that deeper root penetration resulted in increased uptake of these nutrients from soil.

 

CONCLUSION

Application of P and S significantly increased seed yield, N fixation and N uptake at both locations. The effect of P on % N_dfa was not significant but the effect of S was significant. It was observed that P application increased total N uptake, which contributed towards an increase in N fixation. Thus, the effect of P application on N fixation was indirect. Besides, the effect of S application on N fixation was direct since it increased N uptake as well as %N_dfa. Application of both P and S increased nutrient uptake, which may improve the quality of produce. Increased N₂ fixation due to application of P and S may lead to sustainability of soil resources.

 

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