Evaluation of Biological Nitrogen Fixation and Below Ground N Contribution of Grain Legumes Using 15N Techniques

1Kaizzi C.K., 2Rebecca Hood-Nowotny, and 3Charles S. Wortmann

1Kawanda Agricultural Research Institute, P.O. Box 7065 Kampala, Uganda; Email: kckaizzi@hotmail.com. 2Soil Science Unit, FAO/IAEA Laboratories, A-2444 Seibersdorf, Austria; 3University of Nebraska-Lincoln.

IntroductionNitrogen (N) deficiency often constrains crop productivity in Sub-Saharan Africa.Economic considerations make biological N fixation (BNF) an attractive N source for resource-poor farmers (Giller and Wilson, 1981; Van Cleemput, 1995). Most studies that evaluate BNF of legumes consider the above ground biomass only. Few have measured the fixed N in the macro-roots and in the soil, thereby underestimating the total N derived from the atmosphere (Ndfa). We evaluated Ndfa in four grain legumes and determined the below ground N contribution using the stem injection 15N technique (Russell and Fillery, 1996a,b).

100 x fixing-non excess N

15% atom

fixing excess N15

% atom-1 Ndfa%

soilbulk in N x total

roots of excess N15

% atom

soilbulk of excess N15

% atom RdfN(BS)

x100fertiliser excess N

15% atom

plant excess N15

% atom-1 Ndff%

soil eRhizospherin N x total

roots of excess N15

% atom

soil erhizospher of excess N15

% atom RdfN(Rh)

Materials and methodsSoybean [nodulating and non-nodulating], field peas, pigeon peas and cowpeas were evaluated in pots with 1 kg of a 1:1 soil-sand mixture in the greenhouse at FAO/IAEA Agricultural and Biotechnology Laboratories, Seibersdorf, Austria. Two trials were conducted.Trial 1. To measure Ndfa, 10 mg N kg-1 soil labelled (NH4)2SO4 at 5% atom excess 15N was applied to each pot. The non-

nodulating soybean was the reference crop. Trial 2. To measure below ground N contribution, 10 mg N kg-1 soil of ordinary (NH4)2SO4 fertilizer was applied to pots.

Each plant received 1 ml of 0.035M urea solution of 95% atom excess 15N using the stem injection technique. Stems were injected with 15N at 5 weeks after planting and biomass was harvested 23 days after injection. The bulk and rhizosphere soil were collected. The total N and 15N of the samples were determined by mass spectrometry. The equations below were used to calculate: N derived from the air (Ndfa) and fertiliser (Ndff), and N from exudates in the bulk soil [RdfN(BS)] and rhizosphere soil [RdfN(Rh)].In both trials, the shoots were cut at ground level and the recoverable roots were manually separated from the soil.

ResultsN derived from the atmosphere for four legumes• Soybean and field pea derived over 89% of their total N from the atmosphere. Ndfa is a valuable source in the low external input smallholder agriculture.• Cowpea and pigeon pea did not fix nitrogen. Distribution of legume N in plant and soil as determined by the stem injection technique•34 to 69% of legume N was in the above ground biomass (Fig.1). Harvesting for food or fodder of above ground biomass removes much of this N from field.•Below ground N varied by species from 31-66% of the total legume N (Table 1).•On average 43% of below ground N for soybean and field peas was Ndfa.• 13 to 42% of the total legume N was in the bulk and rhizosphere soil from legume exudates and more than the N in the recoverable root fraction.

Conclusions:•Up to 89% of legume N was Ndfa; Ndfa is potentially a valuable source of N for the low-input agriculture.•Below ground legume N was 31 to 66% of total legume N and therefore a significant N source for subsequent crops since much of the above ground legume biomass is either harvested for food or fodder. •Therefore, quantification of below ground legume N is important when evaluating cropping systems to maximise the benefits of legume N sources.

References:Giller, K.E., Wilson, K.J., 1991. Nitrogen Fixation in Tropical Cropping Systems. CAB International, Wallingford, 313p.Russel, C.A., Fillery, I.R.P., 1996a. In situ 15N labelling of lupin below-ground biomass. Aust. J. Agric. Res. 47, 1035-1046Russel, C.A., Fillery, I.R.P., 1996b, Estimates of lupin below-ground biomass nitrogen, dry matter and nitrogen turnover to wheat. Aust. J. Agric. Res. 47, 1047-1059Van Cleemput, O., 1995. Fertiliser, Sustainable Agriculture and Preservation of the Environment. In: IAEA (Ed.) Nuclear Methods in Soil-Plant Aspects of Sustainable Agriculture. Proceedings of an FAO/IAEA Regional Seminar for Asia and Pacific held in Colombo, Sri Lanka, 5-9 April 1993. IAEA-TECDOC-785. Vienna, Austria, pp. 7-16.

Acknowledgement:Financial support from the International Atomic Energy Agency (IAEA) through TC project UGA/05/025

Fig. 1. Relative distribution of nitrogen in shoots, roots, bulk soil (BS) and rhizosphere (Rh)

0%

20%

40%

60%

80%

100%

Soybean(fixing)

Soybean (non-fixing)

Cow peas Pigeonpeas Fieldpeas

Rel

ativ

e d

istr

ibu

tio

n o

f N

(%

)

Shoots

Roots

NdfR(BS)

NdfR(Rh)

Pigeon pea

Soybean (fixing)

Soybean (non fixing)

Field peas

Cowpea

Table 1. Distribution of N in plant parts, root exudates in soil and below ground N Legume Shoots

mg NRoots RdfN (BS)

mg N/kg soilRdfN (Rh) Total N

mgBelow ground Nas % of total N

Soybean fixing 60.4 (15.5)* 15.6 (4.4) 7.6 (0.38) 3.7(0.2) 87.4 31Soybean-non fixing 10.6(0.5) 7.6 (0.04) 10.4(4.2) 2.6 (0.4) 31.1 66Cowpeas 6.7 (1.1) 4.12(1.6) 4.7 (2.3) 1.21(0.2) 16.7 60Pigeon peas 2.5 (1.1) 0.68(0.35) 1.16(0.51) 0.10(0.03) 4.43 44

Table 1. Distribution of N in plant parts, root exudates in soil and below ground N Legume Shoots

mg NRoots RdfN (BS)

mg N/kg soilRdfN (Rh) Total N

mgBelow ground Nas % of total N

Soybean fixing 60.4 (15.5)* 15.6 (4.4) 7.6 (0.38) 3.7(0.2) 87.4 31Soybean-non fixing 10.6(0.5) 7.6 (0.04) 10.4(4.2) 2.6 (0.4) 31.1 66Cowpeas 6.7 (1.1) 4.12(1.6) 4.7 (2.3) 1.21(0.2) 16.7 60Pigeon peas 2.5 (1.1) 0.68(0.35) 1.16(0.51) 0.10(0.03) 4.43 44Field peas 31.8(14.5) 5.9(2.0) 13.1(8.2) 5.01(2.5) 55.9 43

*values in parenthesis are standard deviation

A cotton wick dipping in a vial containing 1ml of 0.035M urea solution at 95% atom excess 15N and passing through a hole pierced through the stem to evaluate below ground legume N.

The legumes used in the evaluation of Ndfa and below ground legume N. Soybean and field pea derived N from the atmosphere as indicated by the dark green leaves.