Field Crops Research, 24 (1990) 163-171 163 Elsevier Science Publishers B.V., Amsterdam - - Printed in The Netherlands

Nitrogen fixation and yield of cowpea ( Vigna unguiculata) as influenced by cultivar and

Bradyrhizobium strain

K.O. Awonaike 1'4, K.S. Kumarasinghe 2 and S.K.A. Danso 2'3

~ Institute of Agricultural Research and Training, Obafemi A wolowo University, P.M.B. 5029, Ibadan, Nigeria

2Joint FAO/IAEA Division, P.O. Box 100, A-1400 Vienna, Austria

(Accepted 14 January 1990)

ABSTRACT

Awonaike, K.O., Kumarasinghe, K.S. and Danso, S.K.A., 1990. Nitrogen fixation and yield of cowpea ( Vigna unguiculata) as influenced by cultivar and Bradyrhizobium strain. Field Crops Res., 24: 163-171.

The nitrogen (N2) fixed and yield of three field-grown cowpea varieties, Ife Brown, Ife BPC and AFB 1757, uninoculated or inoculated with one of two cowpea Bradyrhizobium strains, were exam- ined at physiological maturity. All plants were well nodulated, and the amounts of N2 fixed were not appreciably influenced by inoculation; the average values were I 16, 87 and 74 kg N ha- ~ for Ife BPC, Ife Brown and AFB 1757, respectively. While there were no varietal differences in the dry-matter yields of pods (with enclosed seeds), differences occurred in the total dry-matter yields, due to the large variations in vegetative matter, with Ife BPC generally giving the highest yield. Except for Ife BPC for which inoculation resulted in increased vegetative-matter yield, inoculation had no effect on dry-matter yields. Because of the higher vegetative yield of Ife BPC, the net soil N balance after re- moval of pods and seeds averaged+ 30.3 kg N/ha, compared t o - 10.8 a n d - 8.5 kg N/ha for Ife Brown and AFB 1757, respectively. Thus, although harvest index of Ife BPC was no higher than that of the other cultivars, nor was harvest index enhanced by inoculation, the economic returns, assessed in terms of pod yield plus the soil-N-restoring ability, was highest for Ife BPC, while inoculated Ife BPC was of greater economic value than when uninoculated.

I N T R O D U C T I O N

Legumes are important in the farming systems of the sub-humid and hu- mid-tropical regions of the developing countries, because they provide a rel- atively inexpensive dietary source of protein and serve as organic nitrogen fertilizer for cereal crops grown either in a relay or as rotational crops, partic- ularly when the straw is not harvested.

3Author to whom correspondence should be addressed. 4present address: IAEA Agricultural Laboratory, A2444 Seibersdorf, Austria.

0378-4290/90/$03.50 © 1990 Elsevier Science Publishers B.V.

164 K.O. AWONAIKE ET AL.

In contrast to newly introduced legumes which often respond to inocula- tion in tropical soils (Kang, 1975; Ayanaba, 1977; Owiredu and Danso, 1988 ), several workers have reported that the nodulation and yield of cowpea ( Vigna unguiculata (L.) Walp.), an important crop in the sub-humid and humid tropics, were not enhanced by inoculation (Ezedinma, 1964; Doku, 1969; Kang et al., 1977; Rhodes and Nangju, 1979).

Besides inoculating legumes with highly efficient Rhizobium strains, sev- eral other approaches may be used to improve nitrogen fixation. These in- clude selection of cultivars with superior ability for Nz fixation, or of strains and cultivar combinations best matched to a particular environment (Nut- man, 1984 ). Although cultivar differences in N2 fixation have been examined for many legume species, there are few reports on this for cowpea.

The economic value of a food crop is traditionally based on the harvested seeds. This approach, however, ignores the contribution of the unharvested vegetative parts as biofertilizers. In the absence of chemical fertilizers, plant residues are valuable for maintaining soil fertility. It is therefore essential that total crop yield, not only that of seeds, be examined in breeding programmes.

This study examined the effect of inoculation on nitrogen fixation and yield of reproductive (pods plus seeds) and vegetative parts of three cowpea culti- vars, and the residual N effect in the soil after removal of pod plus seeds.

MATERIALS A N D M E T H O D S

The experiment was conducted on the experimental field of the Institute of Agricultural Research and Training, Moor Plantation, Ibadan, Nigeria (7°23'N, 3°56'E) in the late planting season of 1986. The soil is classified within the Apomu Series as an Oxic Paleustalf (Murdoch and Moorman, 1974); some of its physical and chemical properties are shown in Table 1. Due to the low P and K content of the soil, a basal application of 50 kg P and 50 kg K ha - t , as single superphosphate and muriate of potash respectively, was made at the time of planting.

The treatments consisted of inoculating Ife Brown, Ife BPC and AFB 1757

TABLEI

Some physical and chemical properties of Apomu soil

Physical (%) Chemical

Sand Silt Clay Organic Total pH Available Exchangeable carbon N (in P (ppm) cations (meq/kg)

water) Bray- 1 P Ca K Na

63 20 17 0.84 0.07 6.3 3.9 36.7 4.2 2.7

NITROGEN FIXATION IN COWPEA 165

cowpea varieties with Bradyrhizobium strain 1 Rc 252 (designated BR2) or 1Rc 430 A (BR4). For the uninoculated treatment (BRI), any nodules formed were attributed to the indigenous Bradyrhizobium strain (s). Maize ( Zea mays L. ) was used as the reference crop for estimating N2 fixed in the cowpea crop. A complete randomized block design with four replicates was used. Each sub- plot measured 1.6 × 1.2 m with a plant spacing of 0.4 m between rows and 0.2 m within rows. Immediately after planting, 20 kg N ha-1 in the form of a solution of ammonium sulphate containing 5% ~SN atom excess was sprayed uniformly over the plots.

At physiological maturity (57 days after planting), all above-ground por- tions of the plants within the central 0.8 X 0.8 m 2 of each plot were harvested, divided into the vegetative (leaves, stems and peduncles) and reproductive parts (pods with enclosed seeds), and dried at 70°C to constant weight. Seeds were not separated from the pods, since the usual farming practice in this area involves the removal and non-return of not only seeds, but also the empty pods to the field. Harvest index was therefore estimated as the relative yields of pods (with enclosed seeds) and total dry-matter.

Plant samples were milled to < 0.2-mm size and analysed for %N on a Kjel- dahl digest (Eastin, 1978) and the N isotopes ratio analyses were performed on a VG-isogas mass spectrometer using the Dumas Combustion method (Fielder and Proksch, 1975 ). Nitrogen fixation for the whole plant was quan- tified using the following equation of Fried and Middelboe ( 1977 ):

weighted %~5N atom excess in cowpea~ %N2fixed= 1 ~ 0 / ~ a - i - o o m - ~ s - [ n - m - - ~ z e ] X 1 0 0 (1)

I

total N2 fixed = %N2 fixed X total N in cowpea (2) 100

Nitrogen fixed in the individual organs (for calculating the N balance in soil ) used the same equations, substituting the %~5N atom excess in the organ in question for that of the weighted %~5N atom excess in the whole plant. Soil N was estimated as the difference between total N in the plant and N derived from fixation plus from fertilizer. The N balance in soil was estimated by assuming that any soil N harvested along with the pods plus seeds constitutes a loss to the soil system, while any N2 fixed in the vegetative parts is a gain. Root N contribution was not considered, however, and was assumed to be small. All data collected were analysed statistically using the one-way analysis computer programme of Mohan and Blane ( 1985 ).

RESULTS

Dry-matter yield The dry-matter yields are shown in Table 2. Except for the uninoculated

treatment in which the total dry-matter yields of all three varieties were sire-

166 K.O. AWONAIKE ET AL.

TABLE2

Dry-matter (t h a - t ) in different plant parts of three varieties of cowpea when uninoculated (BRI), or inoculated with Bradyrhizobium strain 1Rc 252 (BR2) or IRc 430A (BR4)

Inoculum/variety Vegetative Reproductive Total Harvest parts parts index

(%)

Uninoculated Ife Brown 3.9c ~ 3.2a 7.1 abc 45 Ife BPC 4. lbc 2. la 6.7abc 39 AFB 1757 2.6d 2.4a 5.0cd 48 Mean 3.5_+0.8 2.7_+0.4 6.3+ 1.1 44-+5

BR2 Ife Brown 3.1 cd 2.5a 5.6bcd 45 Ife BPC 5.4a 2.4a 7.8a 31 AFB 1757 2.6d 2.4a 5.0cd 48 Mean 3.7-+ 1.5 2.4+0.1 6.1 _+ 1.5 41 + 9

Ba, Ife Brown 2.6d 2.6a 5.2cd 50 Ife BPC 5.2a 2.2a 7.4ab 30 AFB 1757 2.1d 2.3a 4.4d 52 Mean 3.3-+ 1.7 2.4_+0.2 5.7+ 1.6 44+ 12

Cultivar means Ife Brown 3.2 _+ 0.7 2.8 + 0.4 6.0 + 1.0 47 -+ 3 Ife BPC 4.9+0.7 2.4-+0.2 7.3_+0.6 33_+5 AFB 1757 2.4+0.3 2.4_+0.1 4.8+0.3 4 9 + 2

~Values in a vertical column followed by the same letter(s) are not significantly different using the Duncan's Multiple Range Test ( P < 0.05 ). The mean values are shown with standard deviations.

ilar, Ife BPC gave higher yields (average 37%) than the other two varieties. There were no varietal differences in the yield of pods (with enclosed seeds), inoculated or not, and bradyrhizobial inoculation did not enhance pod yields.

There were, however, some significant varietal differences in the vegeta- tive-matter yields, as well as some inoculation effects. For example, Ife BPC consistently gave the highest vegetative-matter yield and AFB 1757 the lowest (P< 0.05 ). There was little difference in the vegetative-matter yield between Ife BPC and Ife Brown when not inoculated. With inoculation, however, the vegetative-matter yield of Ife BPC was enhanced but not that of Ife Brown. Consequently, with inoculation, the vegetative-matter yield of Ife BPC was significantly higher than for both Ife Brown and AFB 1757.

On average, AFB 1757, which had the lowest total dry-matter, had the high- est harvest index, while Ife BPC, with the highest total dry-matter, had the lowest harvest index.

Total N yield Data on N yields are shown in Table 3. For the uninoculated treatments,

the total N in Ife Brown was significantly ( P < 0.05 ) higher than in AFB 1757,

NITROGEN FIXATION IN COWPEA 167

TABLE 3

N yield (kg h a - t ) in different plant parts of three varieties of cowpea when uninoculated (BR~), or inoculated with Bradyrhizobium strain 1Rc 252 (BR2) or 1Rc 430A (BR4)

Inoculum/variety Vegetative Reproductive Total parts parts

Uninoculated Ife Brown 63.3bc ~ 144.3a 207.6a Ife BPC 74.8b 83.3ab 158.1abc AFB 1757 45.5c 80.5b 126.0c Mean 61.2+ 14.8 102.7_+6.1 163.9_+41.1

BR2 Ife Brown 56.0bc 83.3ab 139.3bc Ife BPC 115.3a 85.0ab 200.3a AFB 1757 48.3bc 82.0ab 130.3bc Mean 73.2-+36.7 83.4_+ 1.5 156.6-+38.1

BR4 Ife Brown 41.3c 77.8b 119. lc Ire BPC 112.8a 76.3b 189.1a AFB 1757 39.3c 82.8ab 122.1c Mean 64.5-+41.9 79.0_+3.4 143.4_+39.6

Cultivar means Ife Brown 53.5+ 11.2 101.8_+36.9 155.3_+46.4 Ife BPC 101.0 + 22.7 81.5 _+ 4.6 182.5 _+ 21.9 AFB 1757 44.4_+4.6 81.8_+ 1.2 126.1 +4.1

Walues in a vertical column followed by the same letter(s) are not significantly different using the Duncan's Multiple Range Test ( P < 0.05 ). The mean values are shown with standard deviations.

TABLE 4

Estimates of soil N balance (kg N ha - ~ ) following the removal of only the pods and seeds of cowpea from the field

Inoculum strain Mean

BRm BR2 BR4

Ife Brown - 2 6 - 2 . 8 - 3 . 6 - 10.8 Ire BPC 15 31.2 44.7 30.3 AFB 1757 - 14 - 14.5 3.0 - 8 . 5 Mean - 8 . 3 4.6 14.7

but not than in Ife BPC (Table 3 ). In the inoculated treatments, however, Ife BPC accumulated more N than Ife Brown. Also, in contrast to when unino- culated, the differences between total N yields of inoculated Ife BPC and AFB 1757 were significant, while the N yields of inoculated Ife Brown and AFB 1757 were similar. Nitrogen partitioning showed that, on average, seeds plus

168 K.O. AWONAIKE ET AL.

TABLE5

The percentage and total N2 fixed in three varieties of cowpea when uninoculated (BR~), or, inocu- lated with Bradyrhizobium strain 1Rc 252 (BR2) or 1Rc 430A (BR4)

Inoculum/variety N: fixed

(%) (kgha -~)

Uninoculated Ire Brown 57abc I 105.8ab Ire BPC 68.3ab 112.3ab AFB 1757 53.8bc 67.8c Mean 59.7 _+ 7.6 95.3 + 24.0

BR2 Ire Brown 57.Sabc 80.8abc Ire BPC 58.0abc 116.8ab AFB 1757 51.8c 66.0c Mean 55.9_ + 3.5 87.9 _+ 26.1

a ~ Ire Brown 62.3abc 74.3bc Ife BPC 64.0abc 120.3a AFB 1757 70.3a 87.3abc Mean 65.5 _+ 4.2 94.0_+ 23.7

Cultivar means Ife Brown 59.0_+ 2.9 87.0+ 16.3 Ife BPC 63.4 + 5.2 116.5 -+ 4.0 AFB 1757 58.6_+ 10.2 73.7_+ 11.8

JValues in a vertical column followed by the same letter(s) are not significantly different using the Duncan's Multiple Range Test (P<0 .05) . The mean values are shown with standard deviations.

pods contained about 56% of the N in the above-ground parts. This propor- tional distribution of N in plant parts sometimes differed greatly between cul- tivars, and was also occasionally influenced by inoculation treatments (Table 4 ). For example, the vegetative parts of inoculated Ife BPC, in contrast to the uninoculated treatment, contained more N than the seeds. Also, the N contri- bution from the atmosphere in the herbage of Ife BPC and AFB 1757 in- creased by about 60 and 48% respectively, while there was no effect in Ife Brown. On the other hand, inoculation did not influence the N contribution from the atmosphere to the reproductive parts of Ife BPC and AFB 1757, while it was associated with a reduction in Ife Brown of about 40%.

The soil N balance estimates w e r e - 2 6 kg N ha-~ for uninoculated Ife Brown, - 14 kg N ha - ~ for AFB 1757, and + 15 kg N ha - ~ for Ife BPC (Table 4 ). Inoculation further enhanced the N balance in soil sown with Ife BPC by more than 200%.

Nitrogen fixation Data o n N 2 fixed in the various cultivars and treatments are shown in Table

5. Irrespective of inoculation or cultivar differences, cowpea derived more

NITROGEN FIXATION IN COWPEA 169

than 50% of its N from N2 fixation, with a mean of 92 kg N ha- l for all treatments. When uninoculated, no significant varietal differences in %N2 fixed were observed even though Ife BPC was slightly superior to the others. Similarly, only very small differences in %N2 fixed occurred between the in- oculated varieties. The differences between the lower total N2 fixed in uni- noculated or BR2-inoculated AFB 1757 and the other varieties was significant (P< 0.05 ). Although total N2 fixed in uninoculated Ife Brown was, on aver- age, about 37% higher than in any of the corresponding inoculated treat- ments, because of the high variability associated with the dry-matter and N yields, these apparently large differences were not significant. Total N2 fixed in the other varieties was not affected by inoculation.

D I S C U S S I O N

Unless the inoculated strains were excluded from forming nodules, then our finding that both the inoculated and uninoculated treatments, on average, fixed N2 to the same extent suggests that adequate numbers of effective cow- pea bradyrhizobia existed in this soil. Further indication of this was provided by the non-quantitative observations made, showing that the uninoculated plants bore as many nodules, of similar intense pink interior coloration, to those of the inoculated treatments (data not presented). Similar reports on the abundance of effective cowpea bradyrhizobia in tropical soils have been provided by SeUschop (1962), Ezedinma (1964) and Doku ( 1969 ).

One factor that has received little attention is how cowpea inoculation re- sponses are influenced by plant genotype. Our results and those of Pulver et al. ( 1978 ) indicate some interesting strain × genotype interactions in these tropical soils, leading to responses in some cultivars but not in others. For example, cultivar AFB 1757 was the only one that gave significantly higher %N2 fixed when inoculated with strain BR4 (but not with the BR2 strain). Also, cultivar differences occurred in dry-matter and total N yield, often in- fluenced by the bradyrhizobial strain.

It is interesting that the response of Ife BPC to inoculation was reflected only in increased vegetative matter yield but not in the reproductive parts. Consequently, inoculation resulted in a lowered harvest index. A similar find- ing was reported for Phaseolus vulgaris (Awonaike et al., 1980) and cowpea (Summerfleld et al., 1976 ) under controlled environmental conditions.

Because economic return is usually measured in terms of grain-yield, a new cultivar may not appeal to the farmer if seed yield is not increased. The agron- omist, however, is interested in the subsequent effect of cultivation on soil fertility, in addition to grain-yield. This could, therefore, lead to a cultivar being preferred over others even if grain-yields are similar. This was the case in our study. Although the pod plus seed yields of Ife BPC and the other two cultivars were similar, harvest index differed. Ife BPC had the lowest harvest

170 K.O. AWONAIKE ET AL.

index, because of the higher vegetative-matter yield. Thus, we est imated that the net N balance in soil if the pods plus seeds (but not the vegetative mat ter ) were removed from the field would be a gain of some 30 kg N h a - ~ for Ife BPC, compared to the net negative balance for either Ife Brown ( - 10.8 kg N h a - ~ ) or AFB 1757 ( - 8.5 kg N h a - i ). Our results therefore show that, while the cultivation of some cowpea varieties or the adopt ion of some agronomic practices may result in little or no net gains in soil N, others may add appre- ciable amounts. For example, although the inoculation of Ife BPC did not increase pod plus seed yield, the higher net gain in soil N ( + 15, + 31, + 45 kg N h a - ~ when uninoculated, inoculated with BR2, or with BR4, respectively) can be made use of by a cereal crop grown subsequently or in relay, and may thus far outweigh the cost o f the inoculum input.

ACKNOWLEDGEMENT

The study was partly supported through an International Atomic Energy Agency Technical Assistance Project ( N I R / 5 / 0 1 3 ; R A F / 5 / 0 1 0 ) awarded to the Institute of Agricultural Research and Training, Ibadan, Nigeria. The Bradyrhizobium strains were supplied by Dr. K. Mulongoy of the Interna- tional Institute of Tropical Agriculture, Ibadan, Nigeria. Ms. H. Axmann and staff o f the IAEA Laboratory in Seibersdorf performed the isotopic analysis and Ms. R. Rossi typed the manuscript.

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