Rhizobium inoculant for soybean - Springer Link

Plant and Soil 79, 2 3 5 - 2 4 0 (1984). Ms. 5509 9 1984 Martinus Nijhoff/Dr Ir Junk Publishers, The Hague. Printed in the Netherlands.

R h i z o b i u m i n o c u l a n t for s o y b e a n Mekong Delta

[Giycine m a x

(L.) Merrill] in

I. Response of soybean to Rhizobiurninoculation TRAN PHUOC DUONG 1, CAO NGOC DIEP 2, NGUYEN TRI KHIEM 3, NGUYEN HUU HIEP 2, NGUYEN VAN TOI 4, NGUYEN VAN LICH* and LE THI KIEU NHAN ~

Tropical Biological Nitrogen Research Center (TBNRCJ University of Can Tho, Can Tho, Hau Giang, Viet Nam Received 27 May 1983. Revised October 1983

Key words Acid soils Inoculant Rhizobium Soybeans

Summary Rhizobial inoculation trials were conducted in an acid heavy clay soil in Mekong Delta, Viet Nam, using peat based inoculants produced locally and the commercial granular product of Nitragin Co., Wisconsin, USA. The pH of these soils ranged from 4.5 to 5.1. Two soybean cultivars, MTD 6 and MTDIo , were tested as host plants. There were no significant differences between locally made inoculant treated plants and the uninoculated controls in both cultivars. But, the Nitragin inoculation improved all plant characteristics examined in both cultivars. Grain yields of Nitragin inoculated plants of cultivar MTD~ and cultivar MTDao were 6.5 and 5.5 times as much as those of the controls; protein content of grain increased 11 and 16 percent, respectively. Well nodulated plants had shorter life cycles, flowering durations, and days to flowering. The Rhizobium symbiosis resulted in an additional 153 kg grain-N/ha. These studies show that a surface coated commercial multistrain inoculant can be used to successfully grow soybeans in the acid, heavy clay soils of the Mekong Delta.

Introduction Nitrogen fertilizer is a costly import to countries that do not have surplus natural gas. Thus, improved use of natural biological nitrogen fixation could be a less costly alternative for their agricultural production. The soybean-Rhizobium sybiosis has been shown to fix substantial amounts of dinitrogen 2 to sustain its own growth. However, this important potential of the soybean plant will not be realized unless the plant is nodulated by compatible Rhizobium. Field observations (Duong, unpublished data) showed that soybeans established on paddy soils in many parts of the Mekong Delta in Viet Nam had few or were devoid of nodules. The inoculation trial reported here was conducted on moderately acid, heavy paddy soil not previously cropped with legumes. t Principal Investigator and Director TBNRC; 2 Research Associates TBNRC; 3 Statistician, Faculty of Agricultural Economics, University of Can Tho; 4 Undergraduate students.

235

DUONGETAL.

236 Materials and methods

A commercial granular multistrain inoculant of Nitragin Co. (Wisconsin, USA) kindly supplied by the InTSOY program (University of Illinois), and a peat based inoculant, ViDana, produced locally were used. Two soybean cultivars, MTD~ and MTDI0 , supplied by the Soybean Center, University of Can Tho were used as test plants. Cultivar MTD6 was selected from local varieties for its short life cycle and high-yielding characteristics. Cultivar MTD1o is a progeny of a cross between cultivar Wisconsin CBF.11 and cultivar Santa Maria originating from the USA and Brasil, respectively. The inoculation trial was conducted on a heavy-textured clay soil. The soil was classified as belonging to an undifferentiated alluvial great group 4. The surface soil had a moderate acidity and high clay content (Table 1). The soil had been intensively cropped with high-yielding flooded rice at least twice a year. The main water source was rain and the only fertilizer used for the rice crop was urea at a rate of 80-100 kg N/ha/crop. There were three treatments with four replicates for each soybean cultivar as follows: (1) Uninoculated control, (2) ViDana inoculated, and (3) Nitragin inoculated. Soil preparation was done by a small walk-behind tractor. Phosphorus and potassium fertilizers were band dressed to seed rows one day prior to planting at the rates of 40 kg P2Os and 30kg K20 per hectare as thermophosphate (15% P2Os) and KCI (60% K20), respectively. Nitrogen fertilizer was not used. Soil was sprayed with liquid fungicide (Kitazin 50 EC) ten days before planting. Soybean seed, ammonium molybdate (at a rate of 0.5 kg Mo/ha) and inoculants (at a rate of 5 kg/ha of Nitragin or ViDana, where applicable) were mixed with a small amount of water to facilitate the coating of inoculant and Mo on the seeds just prior to planting. The plot size was 3.2 X 5 m with 5 m rows spaced at 40 cm and interrows spaced at 10 cm, two plants/pot. The final population of soybean plants was 500,000 plants/ha. There was a total of 24 plots arranged in a split-plot design with inoculants and cultivars in the main plots and subplots, respectively. During the first 10 days after planting (DAP), soybean seedlings were watered twice a day by hand-carried can; this watering frequency was reduced to once a day from the 10th day until 10 days before harvesting. Plants were protected against insects (Melanogromya phaseoH, Etiella ]inckinella, Spodoptera litura) and fungi by spraying with Azodrin 50 EC, Methyl Parathion 50 EC, Basudin 10 G and Kitazin 50 EC. Seed was sown on December 15, 1979 and grain was harvested on March 22, 1980. All plant samples were taken by randomized digging of five plants in the areas within the guarded rows and 0.5 m from each end of the plots. The first examination for nodulation was done at 15 DAP, the second when 80% of plants had flowers, and the third when 80% of plants reached their ripening stage. Final grain yields were determined by harvesting all plants within the guard rows and 0.5 m from two ends of each plot. Plants were dried, processes by hand and cleaned grains were weighed to determine final grain yields. Total N of grain was determined

Table 1. Chemical and mechanical analysis of the upper 0 - 2 0 cm soil layer

pH Total C Total N Available P as P20 s Total as K20 Exchangable K as K~O Mg Ca CEC

Chemical analysis

Mechanical analysisa

4.5-5.1 a 1.21%a 0.163% a 10 m~/100 g of soilb 2.4%u 0.65 meq/100 g of soilb 12.3 meq/100g of soilb 9.27 meq/100g of soilb 22.7 meq/100g of soilb

Clay 65.5% Silt 33.4% Sand 1.1%

asoil samples taken from experimental site were analyzed by Department of Soil Science, University of Can Tho. bUnpublished data of Mekong Delta project, 1974.

SOYBEAN INOCULATION IN ACID SOILS

237

by the microkjeldahl method and the protein content of the soybean grain was calculated using the 6.25 index.

Results and discussion During the first 15 DAP, plants of all treatments grew normally and had a healthy appearance. However, all plants, inoculated plants as well as uninoculated controls, showed signs of nitrogen deficiency after 15 DAP; their leaves turned pale yellowish green. Nodule development was observed on all plants treated with Nitragin taken at 15 DAP; most of nodules clustered in the upper 5 cm portion of the tap roots indicating prompt infection. There was no sign of nodule initiation either on uninoculated controls or ViDana treated plants although ViDana inoculant has been shown to effectively nodulate another cultivar, DH-4, in another experiment conducted at the same site z. A host genotypic effect on nodulation may be the cause 6, but an inoculant handling problem can not be ruled out. From 23 DAP until maturity, distinct differences developed among plants of different treatments. Leaves of plants that received Nitragin inoculant developed dark green color within the week of 23 DAP to approximately 30 DAP indicating that their root nodules had been actively fixed dinitrogen 3 and supplying the host plant with sufficient Table 2. Effects of at flowering stage

Rhizobium/aponicum on

some agronomic characteristics of soybean plants

Soybean cultivars

Inoculant

Days to floweringa

Flowering period (days)b

Plant height at flowering (cm)

Nodules/ plant

Maximum nodule diameter (mm)

MTD 6

None ViDana Nitragin

35.00 35.00 33.00

21.00 20.25 16.25

16.85 16.85 22.35

0.50 0.30 29.10

0.20 0.50 8.95

MTD1o


39.25 40.00 37.25

23.50 21.75 15.50

17.60 17.70 22.55

0.40 0.90 34.70

0.15 1.30 8.75

LSD e

5% 1% 5% 1%

0.90 1.36 0.59 0.85

1.35 2.04 1.63 2.34

1.97 2.99 0.70 1.01

6.96 10.54 5.65 8.12

0.95 1.45 0.86 1.23

LSD d a b e d

Days from sowing to when 80% of plants had flowers. Days from when 80% of plants had flowers to when 80% of plants ceased to flower. LSD for comparison between Rhizobium inoculant treatments. LSD for comparison between soybean cultivars.

DUONG ET AL.

238

Table 3. Effect of Rhizobium inoculants on some agronomic characteristics of soybeans at ripening stage Soybean cultivars

lnoculant

Life cycle (days) a

Plant height at maturity (cm)

Effective branches/ plant b

Effective nods/ ptant e

100-seed weight (g)d

Protein content in grain (%)

MTD~


86.25 88.00 83.50

21.25 21.10 39.60

1.20 1.77 3.60

3.77 4.62 8.76

12.75 13.75 15.75

38.19 37.44 42.44

MTDIo


93.50 93.50 88.00

25.42 25.45 39.57

1.90 1.95 3.17

5.50 6.00 8.00

10.88 11.12 13.00

34.75 38.19 40.50

LSD e

5% 1% 5% 1%

1.71 2.61 1.52 2.12

1.77 2.69 1.48 2.13

0.69 1.03 0.71 1.01

1.27 1.92 0.65 0.94

2.13 3.23 1.27 1.83

LSD f

a Days from sowing to harvesting. b Branches which had pods. e Nods which had pods at two ends on the main stem. dGrains from different plots of the same treatment were mixed and analyzed by microKjeldahl method for nitrogen content. e LSD for comparison between Rhizobium inoculant treatments. f LSD for comparison between cultivars.

nitrogen. ViDana treated plants and the uninoculated controls remained pale yellowish green throughout their growth cycle. Further differentiation between Nitragin treated plants and plants of other treatments was observed on samples of the second sampling of roots taken when 80% of the plants had flowers (Table 2). Nitragin treated plots had an average of 29.1 (cultivar MTD6)and 34.7 (cultivar MTDIo) nodules per plant. More than 80% of these nodules clustered on the 6 cm upper most portion of the tap roots and the nodules had an intense pink color indicating that they were actively fixing dinitrogen s. Plants treated with ViDana and uninoculated controls had none or few small scattered nodules. This nodulation pattern was also observed by Tin 7. Differences between treatments were observed on other plant characteristics examines in both cultivars such as days to flowering (DF), flowering duration (FD), plant height, effective branches per plant, etc. (Tables 2, 3, and 4). Well nodulated plants had a much shorter life cycle than those in other treatments. They also produced higher grain yields, higher 100-seed weight and had higher grain protein content (Tables 3 and 4). Nitragin treated plants of cultivar MTD6 and MTDI0 produced grain yields 6.5 and 5.5 times, respectively, greater than those of the uninoculated controls. The Rhizobium symbiosis had

239

SOYBEAN INOCULATION IN ACID SOILS Table 4. Effect of Rhizobium inoculants on soybean grain yield (kg/ha) Cultivar

Inoculant

Replication I

II

III

IV

MTD6

None ViDana Nitra~n

520 a 491 2674

548 a 694 2910

317 582 2666

MTDIO


442 499 2979

719 703 2745

393 576 2410

CVa 9.24% b 13.33%

Total

Mean X

253 421 2540

1638 2188 10,790

479 547 2697

411 469 2760

1965 2247 10,894

491 562 2723

LSDb 5% 140kg/ha 1% 212 kg/ha A N O VA

Variables

DF

SS

MS

F calculated

Replication Inoculant Error a Cultivars Inoculant X cultivar Error b Total

3 2 6 1 2 5 19

231486.1666 26047.0833 78533.5834 10004.1666 5149.0834 136214.7500 26509003.8333

77162.0555 13023.0417 13089.9306 10004.1666 2574.5417 27242.9500

5.89 a 995.02 b

Table F 5%

< 1 < 1

4.76 5.14 161 200

1% 9.7 10.9 405.2 499.9

a Contaminated plots were treated as missing data. Contamination with Rhizobium from other plots was noticed from 50 DAP by the development of dark green color of leaves and by appearance of nodules. b LSD for comparison between Rhizobium inoculant treatments.

apparently supplemented the host plant with 153 kg/ha of dinitrogen in both cultivars to produce the extra amounts of grain (subtracting grain N of the controls from those of Nitragin treated plants). There were significant differences between uninoculated controls of cultivars MTD6 and MTDlo for DF, FD, plant height, effective branches and effective nods per plants, plant life cycles, 100-seed weight, etc. (Tables 2 and 3). But, most of these differences disappeared in the Nitragin treated plants of both cultivars. The final grain yield in each set of treatments in both cultivars were not significantly different from each other (Table 4). Also, no significant difference was observed between ViDana treated plants and uninoculated controls in both cultivars. Conclusion The Mekong paddy soil, after years being cropped with rice, appears to be devoid of Rhizobium japonicum. Any native Rhizobia present

240

SOYBEAN INOCULATION IN ACID SOILS

could not produce significant legume grain yield. However, the multistrain inoculant from Nitragin Co. is successfull in these acid, heavy clay soils to provide effective inoculation such that the soybean host provided a satisfactory grain yield. References 1

2 3 4 5

6 7 8

Freire J R 1977 Inoculation of soybeans. 3 3 5 - 3 7 9 . I n Exploiting the Legume-Rhizobium Symbiosis in tropical Agriculture. Eds. J M Vincent, A S Whitney and J Bose. Proceedings of a workshop held at Kahului, Maui, Hawaii, August 2 3 - 2 8 , 1976. Dung Tran nhan 1980 Effects of plant population density and plant spacing on grain yields of soybean cultivar DH 4 . Agro. Eng. Thesis, Univ. of Can Tho. Hardy R W F, Burns R C, Herbert R R, Holsten R D and Jackson E R 1971 Biological nitrogen fixation: A key to world protein. Plant and Soil, special vol. 1 9 7 1 , 5 6 1 - 5 9 0 . Moormann F R 1961 The soils of the Republic of Viet Nam. Directorate of Studies and Research in Agronomy, Forestry and Animal Husbandry, Ministry of Agriculture, Saigon. Munns D N 1977 Soil acidity and related factors. 211-236. In Exploiting the LegumeRhizobium symbiosis in tropical Agriculture. Eds. J M Vincent, A S Whitney and J Bose. Proceedings of a workshop held at Kahului, Maui, Hawaii, August 2 3 - 2 8 , 1976. Rai R and Singh S N 1979 Interaction between chickpeas (Cicaarietinurn Linn.)genotypes and strains of Rhizobium sp. J. Agric. Sei. Camb. 9 2 , 4 3 7 - 4 4 1 . Tin Chu Huu 1980 Rhizobium /aponicum inoculation on Glycine max in Viet Nam. Soybean Genetics Newsletter. 7, 9 7 - 1 0 0 . Weber D F, Caldwell B E, Sloger C and Vest H G 1971 Some USDA studies on the soybean-Rhizobium symbiosis. Plant and Soil, Spec. Vol. 1971, 2 9 3 - 3 0 4 .