Figure
1. Weekly mean temperatures and daylengths during the growth of five soybean
varieties at three elevations on the
GROWTH AND YIELD RESPONSES OF Glycine max and Phaseolus
vulgaris TO MODE OF NITROGEN NUTRITION AND TEMPERATURE
CHANGES WITH ELEVATION
A DISSERTATION SUBMITTED TO THE GRADUATE DIVISION OF
THE
OF THE REQUIREMENTS FOR THE DEGREE OF
DOCTOR OF PHILOSOPHY
IN AGRONOMY AND SOIL SCIENCE
DECEMBER 1988
BY
THOMAS GEORGE
Dissertation Committee:
Duane P. Bartholomew, Chairman
B. Ben Bohlool
Paul W. Singleton
James A. Silva
Francoise M. Robert
ACKNOWLEDGEMENTS
My sincere gratitude goes to Dr. Duane P. Bartholomew, Dr. B. Ben Bohlool and Dr. Paul W. Singleton for their constant guidance, constructive criticisms and spirited support during this research. I am especially appreciative of the help given by Dr. Singleton and Dr. Bartholomew in the preparation of the manuscript. I am grateful to Dr. James A. Silva and Dr. Francoise M. Robert for their helpful suggestions and advice.
I would like to acknowledge Kevin Keane, Rick Koglin, Goeff Haines, Brian Duffy and Thomas Walker for their assistance in the field, the NifTAL Project staff for their untiring help, Robert Abaidoo for his participation in a part of this research.
Lastly, my heartfelt thanks goes to Maria Luz Caces, my best friend.
This research was supported in part by the U.S. Agency for International Development grants DAN-1406-G-SS-4081-00 (Indo/US Initiative on Science and Technology) and DAN-0613-C-00-2064-00 (NifTAL Project), National Science Foundation/Ecology Program, and the International Atomic Energy Agency.
ABSTRACT
Efficient exploitation of the legume-Rhizobium symbiosis in varied environments requires an understanding of the responses of both the host and the bacteria to management and environmental variables. The objective of this research was to study the growth, development and yield responses of soybean (Glycine max L. Merrill) and common bean (Phaseolus vulgaris L.) to mode of N nutrition and temperature at sites in an elevational transect differing in mean temperatures. In the first experiment five soybean varieties and their non-nodulating isolines from four maturity groups were grown after inoculating with three Bradyrhizobium japonicum strains. The proportion of nodules formed by each B. japonicum strain was not affected significantly by temperature, soil type, or soybean genotype. Soybean grain yield and total plant N decreased with increasing elevation. At a given site, the proportion of total plant N derived from symbiotic N2 fixation was approximately constant. Between sites, the proportion of fixed N was inversely related to soil N availability. Vegetative and total crop durations were prolonged by cool temperature. Maturity of soybean was hastened by N insufficiency. In a second experiment, inoculated and uninoculated soybean and common bean were grown with either 9, 120, or 900 kg N ha-1. As a result of extended crop durations at the higher elevation, total biomass production was similar between elevations in both legumes. Grain yield of common bean was similar at the cool and warm sites; in contrast the grain yield of soybean was greatly reduced at the cool site. Yield and total plant N of both legumes increased at mineral N levels of 120 and 900 kg ha-1 compared to 9 kg N ha-1. Significant symbiotic N2 fixation limited yield and total N assimilation in soybean. Common bean utilized proportionately more mineral N than soybean throughout growth and had a higher fertilizer N use efficiency than soybean during the vegetative phase. Common bean also had a lower N requirement and N assimilation was more uniform throughout growth when compared to soybean. Maximizing N derived from fixation in common bean may require altering its N assimilation characteristics.
TABLE OF CONTENTS
ACKNOWLEDGEMENTS . . . . . . . . . . . . . . . . . . iii
ABSTRACT . . . . . . . . . . . . . . . . . . . . . . iv
LIST OF TABLES . . . . . . . . . . . . . . . . . . . vii
LIST OF FIGURES . . . . . . . . . . . . . . . . . . . xii
CHAPTER I. INTRODUCTION . . . . . . . . . . . . . . 1
CHAPTER II. NODULATION AND INTERSTRAIN COMPETITION BY
Bradyrhizobium japonicum IN SOILS ALONG AN
ELEVATIONAL TRANSECT . . . . . . . . . . 6
CHAPTER III. EFFECT OF TEMPERATURE AND MATURITY GROUP
ON PHENOLOGY OF FIELD GROWN SOYBEAN . . 28
CHAPTER IV. YIELD, SOIL N UPTAKE AND N2 FIXATION BY
SOYBEANS FROM FOUR MATURITY GROUPS GROWN
AT THREE ELEVATIONS . . . . . . . . . . 48
CHAPTER V. GROWTH RESPONSES OF FIELD GROWN SOYBEAN
AND COMMON BEAN TO TEMPERATURE AT VARYING LEVELS OF N NUTRITION . . . . . . . . . 67
CHAPTER VI. NITROGEN ASSIMILATION AND N2 FIXATION IN
SOYBEAN AND COMMON BEAN AS AFFECTED BY
ELEVATION AND MINERAL N AVAILABILITY . . 89
SUMMARY AND CONCLUSIONS . . . . . . . . . . . . . . . 109
APPENDIX . . . . . . . . . . . . . . . . . . . . . . 114
LITERATURE CITED . . . . . . . . . . . . . . . . . . 123
LIST OF TABLES
Table Page
II-1 Soil and site characteristics of three
locations in an elevational transect on
the island of
II-2 Proportion of nodules formed by B. japonicum
strains from five soybean varieties grown in
three soils in the field and greenhouse . . . . . 22
II-3 Proportion of nodules formed by B. japonicum
strains from five soybean varieties grown in
the field and greenhouse . . . . . . . . . . . . 23
II-4 Number and dry weight of nodules from five
soybean varieties at 45 days after planting
at three different elevations . . . . . . . . . . 24
II-5 Number and dry weight of nodules from five
soybean varieties at 45 days after planting
in three different soils in the greenhouse . . . 25
II-6 Shoot dry weight of five soybean varieties
at 45 days after planting at three different
elevations . . . . . . . . . . . . . . . . . . . 26
II-7 Shoot nitrogen content of five soybean varieties
grown at three different elevations at 45 days
after planting . . . . . . . . . . . . . . . . . 27
III-1 Mean maximum and minimum temperatures during
growth of five soybean varieties at three
elevations on the island of
III-2 Days to first flower and physiological maturity
of nodulating isolines of five soybean varieties
grown at three elevations . . . . . . . . . . . . 42
III-3 Growing degree days accumulated during
vegetative growth and up to physiological
maturity for nodulating isolines of five
soybean varieties grown at three elevations . . . 43
III-4 Duration of seed-filling of nodulating isolines
of five soybean varieties at three elevations . . 44
III-5 Rate of seed dry matter accumulation of
nodulating isolines of five soybean varieties
grown at three elevations . . . . . . . . . . . . 45
III-6 Mean days to first flower and physiological
maturity for nodulating and non-nodulating
isolines of five soybean varieties grown at
two elevations . . . . . . . . . . . . . . . . . 46
III-7 Mean grain and shoot nitrogen concentrations of
nodulating and non-nodulating isolines and grain
yield of non-nodulating isolines of five soybean
varieties grown at three elevations . . . . . . . 47
IV-1 Characteristics of three sites in an elevational
transect on the island of
IV-2 Grain and stover yields of five nodulating
soybean varieties grown at three different
elevations . . . . . . . . . . . . . . . . . . . 60
IV-3 Total nitrogen assimilation by five nodulating
soybean varieties grown at three different
elevations . . . . . . . . . . . . . . . . . . . 61
IV-4 Symbiotically fixed nitrogen by five soybean
varieties grown at three different elevations . . 62
IV-5 Average rate of nitrogen assimilation by five
soybean varieties grown at three different
elevations . . . . . . . . . . . . . . . . . . . 63
IV-6 Percent of total nitrogen derived from symbiotic
fixation by five soybean varieties grown at
three elevations . . . . . . . . . . . . . . . . 64
IV-7 Comparison of estimated amounts of mineralized N
and actual soil nitrogen uptake by five soybean
varieties at three elevations . . . . . . . . . . 65
IV-8
Nitrogen uptake by
grown in three soils in the greenhouse at a mean
soil temperature of 27 C . . . . . . . . . . . . 66
V-1 Environmental characteristics at two sites in an
elevational transect on the
V-2 Days to first flower and physiological maturity
of inoculated soybean and common bean grown at
two elevations with three levels of N . . . . . . 82
V-3 Grain yield of soybean and common bean as
affected by elevation and N application . . . . . 83
V-4 Effect of elevation by legume species interaction
on harvest index and whole plant tissue N
concentration at physiological maturity . . . . . 84
V-5 Main effects of elevation, legume species and N
level on dry weights at different growth stages . 85
V-6 Main effects of elevation, legume species and
N levels on mean crop growth rate during
different growth phases . . . . . . . . . . . . . 86
V-7 Effect of elevation by legume species
interaction on leaf characteristics at different
growth stages . . . . . . . . . . . . . . . . . . 87
V-8 Effect of elevation and legume species on unit
leaf rate during different growth phases . . . . 88
VI-1 Elevation by legume by N level interaction on
N assimilation at physiological maturity . . . . 103
VI-2 Effect of elevation, N level and growth stage
on N2 fixation by soybean at different growth
stages . . . . . . . . . . . . . . . . . . . . . 104
VI-3 Effect of elevation, N application and legume
species on the rate of N assimilation during
crop cycle . . . . . . . . . . . . . . . . . . . 105
VI-4 Relative N assimilation during different growth
phases of soybean and common bean grown at two
elevations . . . . . . . . . . . . . . . . . . . 106
VI-5 Effect of legume and N levels on fertilizer N
use efficiency during the crop cycle . . . . . . 107
VI-6 Effect of legume and N levels on KCl-extractable
soil N during the crop cycle . . . . . . . . . . 108
A-1 Effect of elevation and N level on nodule dry
weight at end bloom . . . . . . . . . . . . . . 114
A-2 Days to flowering and physiological maturity
of the non-nodulating isoline of
grown at two elevations with three levels
of mineral N . . . . . . . . . . . . . . . . . . 115
A-3 Effect of N level on harvest index of legumes
grown at two elevations. . . . . . . . . . . . . 116
A-4 Grain yield of the non-nodulating isoline of
levels of mineral N . . . . . . . . . . . . . . 117
A-5 Total N assimilation by the non-nodulating
isoline of
with three levels of mineral N . . . . . . . . . 118
A-6 Effect of N level on the leaf area and leaf weight
of legumes at different growth stages . . . . . 119
A-7 Effect of N level on the unit leaf rate of
legumes at different growth stages . . . . . . . 120
A-8 Fertilizer use efficiency of legumes grown at two
elevations with three N levels . . . . . . . . . 121
A-9 KCl-extractable soil N after harvest of legumes
grown at two elevations with three N levels . . 122
LIST OF FIGURES
Figures Page
1 Weekly mean temperatures and daylengths during the
growth of five soybean varieties at three elevations
on the island of
Chapter 1
INTRODUCTION
Legumes respond to the environment in the same way other plants do, but what makes them physiologically distinct from other plants is their ability to harbor nitrogen-fixing rhizobia in a symbiotic relationship. The establishment of the legume-Rhizobium symbiosis starts with the infection of the legume host by compatible rhizobia, resulting in the formation of nodules.
Both symbiotic N2 fixation and mineral N assimilation contributes to the N requirement of legumes. The symbiosis is functional when the legume is stressed for nitrogen and, to some degree, compensates for decreased mineral N availability. However, mineral nitrogen suppresses symbiotic N2 fixation (Allos and Bartholomew, 1959; Chen and Phillips, 1977; Dart and Wildon, 1970; Oghoghorie and Pate, 1971). While N2 fixation is readily replaced by mineral N, evidence presented by Gibson and Pagan (1977), Manhart and Wong (1980), and Streeter (1985) do not show any direct inhibitory effect of mineral N on N2 fixation.
The general consensus regarding the N
nutrition of legumes is that legumes require N from both symbiotic and mineral
N sources for maximum yield (Bhangoo and Albritton, 1976; Harper, 1974). However, there are indications that legumes
relying on mineral N produce yields similar to or greater than symbiotic plants (Kato et al.
1984; Ryle et al. 1978; Silsbury, 1977).
Schweitzer and Harper (1985) found that increasing light interception by
a soybean canopy using reflectors increased nitrate reductase activity in
nitrate-supplied plants more than it increased nodule activity in inoculated
plants, and the nitrate-supplied plants consequently produced greater dry
weight. Also, Tanaka (1986) showed that
if photosynthetic activity decreased, N2 fixation decreases more
than mineral N absorption. Accordingly,
If there is sufficient N in the soil to meet the plant's requirement, and if assimilation from the soil is not limited by other factor(s), then symbiotically fixed N will not be a major source of N for the plant. Any factor which limits the availability, uptake or assimilation of soil N should favor N2 fixation. Low temperature is one factor which reduces the rate of soil N mineralization (Stanford et al., 1973; Cassmann and Munns, 1980), and uptake of available soil nitrogen (McDuff and Hopper, 1986; Tolley and Raper, 1985). Low temperature, therefore, may favor N2‑fixation over mineral N assimilation unless it decreases nitrogenase activity to the same degree that soil N assimilation is decreased.
Nitrogen nutrition presumably can range from complete dependence on mineral N to complete dependence on symbiotic N2 fixation. Plants dependent on N2 fixation or mineral N may respond differently to the environment because of the differences in their root morphology, N assimilation process, and N nutritional level. In the case of the symbiotic plant, the response may be further complicated by specific interactions between the legume host and the rhizobial microsymbiont. The interaction between the legume host and the rhizobial microsymbiont is specific, and the compatibility of the host with any one of the competing rhizobial strains may determine whether or not a productive symbiosis will be established. Thus, any attempt to correlate legume response to environment requires careful consideration of the mode of N nutrition and the N requirement of the plant.
Developing a holistic approach to legume productivity research requires the evaluation of the legume‑rhizobium system from soil, plant, and microbial perspectives in the field. Such an approach would render the results more relevant to productivity research and improve the degree of predictability of outcomes in varied environments.
Considering the complexity of environmental factors in agricultural ecosystems, the delineation of effects of single factors is often difficult. Although management variables may be made optimum and thus, eliminated as sources of variation, the effects of environmental variables such as temperature, photoperiod and irradiance are often confounded.
Temperature has direct and indirect
effects on biological or biochemical processes and in legumes, these responses
are readily modified by photoperiod (
Two experiments were conducted to study the effects of mode of N nutrition, temperature and other elevation-associated variables on the ecophysiological response of soybean and common bean. The approach taken was to study host-Rhizobium-environment interactions and plant genotype-temperature-mode of nitrogen nutrition interactions in soybeans adapted to a wide range of latitudes, and use the data thus obtained as the basis for a second experiment to evaluate the growth and yield responses of soybean and common bean to temperature under a range of mineral N availability conditions.
The Objectives of the experiments were:
1. Determine the effects of elevation-associated site characteristics (temperature, soil type) and soybean genotype on the competition pattern of Bradyrhizobium japonicum strains.
2. Determine the effects of temperature and mode of N nutrition on the phenology of a range of soybean maturity groups under similar photoperiodic regimes.
3. Assess the effects of site characteristics and soybean genotype on the contribution of N2 fixation to plant N.
4. Evaluate the growth responses of soybean and common bean to cool and warm environments at varying levels of soil N availability.
5. Evaluate the responses of soybean and common bean to mineral N application and contrast the N assimilation patterns of the two legumes.
Chapter 2
NODULATION AND INTERSTRAIN COMPETITION BY Bradyrhizobium
japonicum IN SOILS ALONG AN ELEVATIONAL TRANSECT
Abstract
A better understanding of the ecology of Bradyrhizobium
japonicum in relation to its host and its environment may aid in making
appropriate management decisions for soybean production in a wide range of
conditions. Success of one or more of
the competing rhizobial strains in forming nodules and establishing an
effective symbiosis is influenced by host genotype, soil type and
temperature. The effects of temperature
and soil type on interstrain competition, nodulation and nitrogen assimilation
in five soybean varieties belonging to maturity groups 00, IV, VI, and VIII
were investigated at three sites devoid of soybean rhizobia along an
elevational transect in
Introduction
The establishment of an effective and efficient symbiosis between rhizobia and the host legume is basic to viable legume production without mineral N fertilization. The symbiosis is modified by various environmental factors. An understanding of the effects of environmental factors on the symbiosis is therefore, essential for selecting adapted cultivars and management practices that will enhance symbiotic nitrogen fixation and yield.
Bradyrhizobium japonicum strains differ considerably in competitive ability, nodulation and nitrogen fixation (Abel and Erdman, 1964; Caldwell, 1969; Johnson and Means, 1964). The competition and nodulation characteristics of the strains have been shown to be influenced by the host genotype (Caldwell and Vest, 1968; Materon and Vincent, 1980).
Soil type may also influence the competition between strains for nodule sites. Dominance of different B. japonicum strains in soybean nodules from different soils have been reported (Damirgi et al., 1967; Ham et al., 1971; Johnson and Means, 1963; Keyser et al., 1984). Soil type greatly influenced the distribution of strains, with distinctly different populations of rhizobia in soybean nodules from six different soils (Damirgi et al., 1967). According to Ham et al. (1971), the presence of individual strains in soybean nodules was related to two or more of the soil properties, the most important of which was soil pH. Kosslak and Bohlool (1985) have reported that in soils devoid of native B. japonicum, the pattern of competition between two introduced strains remained the same regardless of soil type or soil amendments.
The recovery of specific rhizobial strains in the nodules, and nodulation and N2 fixation are also related to temperature. Caldwell and Weber (1970) found that the dominance of different serogroups in soybean nodules was altered by planting dates, and Weber and Miller (1972) concluded that the effect was due to differences in soil temperature. Low soil temperatures have also been shown to adversely affect both nodulation and nitrogen fixation (Lindeman and Ham, 1979; Waughman, 1977).
To understand the ecology of rhizobia in
relation to the host and its environment, experiments were conducted along an
elevational transect located on the island of
Materials and Methods
Experimental design: The treatments were five soybean varieties and three sites (elevations) of planting. All varieties were inoculated with a triple-strain inoculum consisted of three Bradyrhizobium japonicum strains. The experimental design was a randomized complete block with three replications at each site.
Selection
of sites: Three sites along an elevational transect on the slope of
Soil amendments: The soils were limed to a pH between 5.5 to 6.0 by incorporation of Ca(OH)2. Other amendments in kg ha-1 were 600 P as treble superphosphate, 300 K as K2SO4, 75 Mg as MgSO4, 15 Zn as ZnSO4, 2 Mo as Na2MO4 and 3 B as H3BO3.
Soybean varieties: The soybean varieties used were from four maturity groups (MG) and included Clay 00 (J.H. Orf, Agronomy and Plant Genetics, Univ. Minnesota), Clark IV (P. Cregan, USDA Nitrogen Fixation Lab., Beltsville), D68‑0099 VI and N77‑4262 VI, which are determinate and indeterminate selections from cv. Lee (T.E. Carter, Dept. of Crop Science, North Carolina State Univ.), and Hardee VIII (K. Hinson, Dept. of Agronomy, Univ. Florida).
Rhizobial
inoculum strains: Yeast extract-mannitol (Vincent, 1970) agar slants of the
serologically distinct rhizobial strains USDA 110, USDA 136b (CB 1809) and USDA
138 were obtained from the NifTAL Project collection,
Seed inoculation procedure: Gum arabic (40 g per 100 ml water) was applied to seeds at the rate of 3 ml 100 g-1 seed. The seeds were then coated with approximately 14 mg of the prepared peat inoculum per seed. The seeds were air dried and kept at 4 C overnight. Viable counts of rhizobia at planting averaged 1.5 x 107 per seed.
Plant
culture: All the sites were planted on
Weather
data collection.: Temperature data were recorded by Campbell Scientific
CR-21 microloggers (Campbell Scientific, Inc.,
Greenhouse experiment: An experiment was conducted in a greenhouse to separate the effect of soil type from that of temperature differences associated with site on rhizobial interstrain competition and nodulation in soybean. The experimental design was a randomized complete block with the five soybean varieties used in the field study and soils from the three field sites as treatments. There were three replications. Soils were amended as before. Nutrients applied in mg kg-1 of soil were 300 P, 150 K, 37.5 Mg, 7.5 Zn, 1 Mo, and 1.5 B. Seven seeds for each variety coated with inoculum prepared as above were planted in 3 liter pots filled with 2 kg of soil (oven dry basis). The pots were watered daily to a tension of 0.03 MPa with the aid of tensiometers. The plants were thinned to two per pot after 10 days. The average soil temperature in the pots was 27 C.
Sampling procedures: In the field experiment, twelve plants per replication were harvested at random from the two outer rows of each plot 45 days after planting. The plant tops were cut at the soil surface, oven dried at 70 C, weighed, ground, and the nitrogen content was determined by H2SO4 digestion followed by indophenol blue method (Keeny and Nelson, 1982). The roots were excavated and the nodules were collected after washing the roots free of soil, counted, dried at 70 C in a forced air oven, and weighed. Nodules from the plants grown in the greenhouse were also collected and dried 45 days after planting.
Strain identification: Occupancy of nodules by the three Bradyrhizobium sp. strains was determined by immunofluorescence using oven dried nodules (Somasegaran et al., 1983). At least 5%, but no fewer than 24 nodules, of the composite sample from each replication for the field experiment and 12 nodules per pot for the greenhouse experiment were selected at random. The nodules were placed in the wells of microtiter plates and rehydrated in the refrigerator overnight with a few drops of sterile water. The nodules were then crushed, smears were prepared and heat fixed on glass slides. The smears were treated with gelatin‑rhodamine isothiocyanate to control background fluorescence and non‑specific staining (Bohlool and Schmidt, 1968) and after drying, they were stained with strain‑ specific fluorescent antibodies (Schmidt et al., 1968). The slides were observed under epifluorescence microscopy for positive reactions. Positive reaction for more than one strain in the same nodule was counted as mixed infection.
Data
analysis: Data from the three sites were subjected to a combined analysis
(McIntosh, 1983). Nodule and plant
growth data as well as arcsine transformations of percent nodule occupancy from
both experiments were subjected to analysis of variance using SAS (SAS
Institute. 1982. SAS user's guide: Statistics. 1982 ed. SAS Institute Inc.,
Results
There were significant differences in the nodule occupancy by the strains used (Table 2). USDA 110 was the most competitive strain in both experiments; it formed 81% and 64% of nodules in the field and greenhouse, respectively. Strain USDA 138, which formed few nodules in the field, occupied the same number of nodules as USDA 136b in the greenhouse experiment. Both USDA 138 and USDA 136b were present in higher proportions in the greenhouse study than the field. However, there was no significant effect of site or soil on nodule occupancy in the two experiments.
Soybean varieties significantly differed in strain occupancy under both field and greenhouse conditions (Table 3). USDA 110 formed between 61% and 98% of the nodules in the field and 55% to 86% in the greenhouse study depending on cultivar. There was a positive correlation between maturity group and the proportion of nodules formed by USDA 110. Where nodulation by USDA 110 was reduced, increased nodulation by USDA 138 in the pot and by USDA 136b in the field was observed.
Nodule
number and dry weight were significantly greater at the 320 m site and both
parameters decreased with increasing elevation for all of the varieties (Table
4). Nodule weight declined more with increasing elevation than did nodule
number. Mean nodule number at
Shoot dry weight (Table 6) and total shoot nitrogen assimilation (Table 7) decreased with increasing elevation. There were no differences between varieties in shoot dry weight within each site. Total nitrogen content did not vary with varieties except at Kuiaha where late maturing cultivars (VI and VIII) assimilated less N at 45 days after planting than MG 00 and IV.
Discussion
In this study, the effect of soil type and temperature on nodulation and early growth of a wide range of soybean maturity groups was investigated. By introducing equal numbers of three B. japonicum strains into soils devoid of soybean rhizobia, the effects of soil type, soil temperature and host genotype on interstrain competition for nodule sites were evaluated.
Differences in nodule occupancy by the
three strains indicate that they have inherent differences in competitive
ability. Strain USDA 110, the
predominant strain in this study , has previously been reported to be a
superior competitor (Johnson and Means, 1964; Caldwell, 1969). Kosslak and Bohlool (1985) found USDA 110 to
be highly competitive against USDA 123 on soybeans grown in vermiculite and in
Hawaiian soils devoid of B. japonicum. However, in soils from the north central
region of the
Soil type did not affect the percent recovery of strains from either field or greenhouse nodules. Since there was no difference between soils in strain recovery in the greenhouse where all the soils were under the same temperature regime, we conclude that competitive attributes of strains are independent of soil type and, within a reasonable range, of soil temperature. This contradicts the view that interstrain competition is very much dependent on soil type (Damirgi et al., 1967; Johnson and Means, 1963) and soil temperature (Kvien and Ham, 1985; Weber and Miller, 1972). However, it may be noted that, in contrast to the above reports, no indigenous rhizobia were present in the soils of the present experiment. The results of this experiment agree with those of Kamicker and Brill (1986) who reported that competition patterns by indigenous strains were not related to soil type.
The varietal differences in strain recovery were related to soybean MG. Physiological development of the plant is not a contributing factor since plants within a MG were at the same growth stage in the three soils in the greenhouse but were at different stages of development in the field where decreasing temperature delayed development. Hardee was an exception to other varieties in that very few nodules where occupied by USDA 136b and USDA 138. This result may be partly explained by the fact that USDA 136b is incompatible with Hardee which carries the gene Rj2 (Materon and Vincent, 1980).
The reduction in nodule number and mass may be the direct consequence of decreasing soil temperatures as well as low temperature effect on plant growth. Decreased nodulation of soybeans at soil temperatures below 24 C has been reported as early as in 1921 by Jones and Tisdale. Gibson (1967) observed considerable delay in nodulation and a general decline in the rate of nodule formation by Trifolium subterraneum when root temperatures were below 22 C. Enhanced plant development and increased nodulation in soybeans with increasing temperatures within the range of 15 to 25 C have been reported previously (Kvien and Ham, 1985; Matthews and Hayes, 1982).
Despite a large decrease in nodule number between sites, the three strains occupied the same percentage of nodules in all sites. It has been suggested that infection of roots by rhizobia is less sensitive to low temperatures than nodule formation (Gibson, 1967; Kumarasinghe and Nutman, 1979). The results of this study indicate that competition among rhizobial strains for nodule sites is not related to the final number of nodules formed.
Large differences in shoot dry weight and nitrogen assimilation between sites may be attributed to the effects of soil and air temperatures on nodulation and plant growth. Plant growth was severely restricted by a 5 C lower average temperature at the highest site. There was also considerable delay in plant development with decreasing temperatures. At the lowest site, all varieties were beyond the flowering stage at 45 days with the early varieties Clay and Clark already forming seeds. At the highest site, plants with the exception of Clay and Clark had not flowered. Plant growth was directly reduced by the low average temperature at the high cool site, but cool temperature could have had an indirect effect by reducing nitrogen fixation and soil nitrogen absorption. Soil temperatures of 24 to 27 C are reported to be optimum for nitrogen fixation and nitrate absorption (Kuo and Boersma, 1971; Rufty et al., 1981) in soybeans. The temperature at the lowest site was in the vicinity of the reported optimum. The high nitrogen assimilation and growth at that site was consistent with the findings of others.
The present experiments identified USDA 110 as a better competitor than USDA 136b and USDA 138 for nodule occupancy regardless of soil type, soil temperature or MG of soybean. Nodulation and plant growth at 45 days after planting were drastically reduced by a 5 C decline in both soil and aerial temperatures. None of the nodulation and plant growth parameters were related to occupancy of strains in the nodules. Results indicate that strains superior in competitiveness can be identified for a broad range of host genotypes, soil types, and temperature.
Table
1. Soil and site characteristics of
three locations in an elevational transect on the
________________________________________________________________
Total Average temp‑
Site Soil Elevation rainfall erature
(Soil classification during during crop
series) crop growth growth
Soil Air
_______________________________________________________________ m mm C
Kuiaha Clayey, ferritic, (Haiku isohyperthermic 320 139 25.3 22.8
series) Humoxic Tropohumult
Haleakala Clayey, oxidic,
(Makawao isothermic 660 112 22.9 20.9
series) Humoxic Tropohumult
(
series) Entic Dystrandept
‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑
Table 2. Proportion of nodules formed by B. japonicum strains
from five soybean varieties grown in the field and greenhouse.1
________________________________________________________________
Nodules occupied by:
Site USDA 110 USDA 136b USDA 138
Field Pot Field Pot Field Pot
________________________________________________________________
- - - - - - - - - % - - - - - - - - -
Kuiaha 80.3 62.9 25.8 28.7 3.9 37.4
Haleakala 80.3 61.8 26.9 35.2 4.2 33.6
Mean 80.8A2 64.2a 27.6B 33.8b 4.1C 34.2b
________________________________________________________________
1 Values are means of 5 cultivars; Nodules with more than one
strain are counted for each strain testing positive.
2 Data followed by the same type letter for either field or pot
are not significantly different (P=0.05) by
range test.
Table 3. Proportion of nodules formed by B. japonicum strains from
five soybean varieties grown in the field and the greenhouse1.
________________________________________________________________
Nodules formed by
Variety USDA 110 USDA 136b USDA 138
Field Pot Field Pot Field Pot
_________________________________________________________________
Clay 60.7a2 55.1b 40.7a 38.0b 7.4a 47.9a
D68‑0099 87.5b 61.4b 26.9b 38.9b 0.5b 37.1b
N77‑4262 87.5b 60.5b 30.1b 52.4a 2.8b 33.4b
Hardee 97.7a 85.8a 8.3c 10.8c 0.9b 12.7c
________________________________________________________________
1 Values are means of three soils. Nodules with more than one
strain are counted for each strain testing positive.
2 Data followed by the same letter within a column are not
significantly different (P=0.05) by
test.
Table 4. Number and dry weight of nodules from five soybean
varieties at 45 days after planting at three different elevations.
________________________________________________________________
Variety Mean
Site Clay Clark D69‑0099 N77‑4262 Hardee
________________________________________________________________
- - - - Number plant-1 - - - -
- - - - (mg plant-1) - - - - -
Kuiaha 43 40 60 65 57 53A1
(289) (278) (294) (236) (249) (269A)
Haleakala 20 32 31 35 28 29B
(180) (216) (137) (134) (146) (162B)
(75) (76) (73) (73) (66) (73C)
Mean 27d1 32cd 38ab 43a 34bc
(181a) (190a) (168ab) (147b) (154b)
________________________________________________________________
1 Data followed by the same letter within a row or a column are
not significantly different (P=0.05) by
range test.
Table 5. Number and dry weight of nodules from five soybean
varieties at 45 days after planting in three different soils in
the greenhouse.
___________________________________________________________________
Soil series Variety Mean
(Site) Clay
___________________________________________________________________
- - - - Number plant-1 - - - -
- - - - (mg plant-1) - - - -
Haiku 27 30 34 45 42 35
(Kuiaha) (74) (102) (100) (114) (112) (100)
Makawao 31 45 35 29 35 35
(Haleakala) (91) (93) (91) (86) (96) (92)
(
Mean 26b1 34a 39a 37a 38a
(84b) (96ab) (94ab) (101a) (107a)
___________________________________________________________________
1 Data followed by the same letter within a row are not
significantly different (P=0.05) by
test.
Table 6. Shoot dry weight of five soybean varieties at 45 days
after planting at three different elevations. ________________________________________________________________
Site Variety
Clay
________________________________________________________________
- - - - - - g plant-1 - - - - - - -
Kuiaha 7.7 7.7 7.8 6.3 7.0 7.3A1
Haleakala 4.8 5.6 5.6 5.6 5.9 5.5B
Mean 4.5 4.8 4.8 4.3 4.6
________________________________________________________________
1 Data followed by the same letter within the column are not
significantly different (P=0.05) by
test.
Table 7. Shoot nitrogen content of five soybean varieties grown
at three different elevations at 45 days after planting.
________________________________________________________________
Site Variety
Clay Clark D68‑0099 N77‑4262 Hardee Mean
________________________________________________________________
- - - - mg N plant-1 - - - - -
Kuiaha 294a1 290a 232b 195b 218b 246A1
Haleakala 182a 198a 181a 178a 198a 187B
Mean 167a 171a 146ab 133b 147ab
________________________________________________________________
1 Data followed by the same lower case letter within a row or
same upper case letter within the column are not significantly
different (P=0.05) by
Chapter 3
EFFECT OF TEMPERATURE AND MATURITY GROUP ON
PHENOLOGY OF FIELD GROWN SOYBEAN
Abstract
The environmental adaptation of soybean (Glycine
max (L.) Merrill) genotypes depends in part on the effects of
temperature and photoperiod on the durations of phenological phases. The effects of temperature and nitrogen
nutrition on soybean phenology was studied at field sites with elevations of
320, 660, and 1050 m on the
Introduction
The adaptation of soybean genotypes to environment depends in part on how temperature and photoperiod affect ontogeny. While rate of growth is strongly influenced by temperature, rate of development and the duration of phenological phases in soybean are determined by both temperature and photoperiod.
Most reports concerning the effects of temperature on phenological development of soybean either used planting date as a variable (Lawn and Byth, 1973; Major et al., 1975a) or employed controlled environments (Hadley et al. 1984; van Schaik and Probst, 1958). Since the induction of flowering in soybean is photoperiod sensitive and most soybean genotypes are quantitative short‑day plants (Hadley et al. 1984; Lawn and Byth, 1973), the effects of temperature on soybean development in the field are often confounded with the effects of photoperiod which change with planting date and latitude. Even though controlled environment experiments may be useful in avoiding complex interactions between photoperiod and temperature, development of plants under such conditions may not be typical of what occurs in the field.
Hodges and French (1985) compiled data for soybean which showed that temperature was important during early growth and development. Once flowering occurred, both temperature and photoperiod influenced development. According to Lawn and Byth (1973) low temperature effects on phasic development of soybean were apparent only when photoperiodic effects were minimum. Major et al., (1975a) reported that although later maturity groups flowered after the early ones, there were no differences among maturity groups I through V in sensitivity to temperature at a constant photoperiod of 14h.
An elevational transect located on the
island of
Materials and Methods
Experimental design: Nodulating (nod) and non-nodulating (non-nod) isolines of five varieties of soybean representing four maturity groups (00, IV, VI, VIII) were grown at elevations of 320, 660, and 1050 m. The experimental design was a randomized complete block with three replications at each site.
Field and plant culture: The sources from which seeds of soybean varieties (nod isolines) were obtained were given in Chapter 2. The non-nod isoline of each variety used in the present study was also received from the same source as that of the nod isoline. Procedures followed for soil amendment, rhizobial inoculation, plant culture and temperature data collection were described in Chapter 2.
Sampling procedures: Phenological observations were made on a 1.0 m section of row in each plot. Plants were considered to have reached a particular phenological stage when fifty percent of the plants in the 1.0 m row reached that stage. Growth stage characterization was based on the system developed by Fehr et al., (1971).
Three meters of row from the center of the two central rows were harvested at physiological maturity. Non‑nod isolines were harvested at the same time as their fixing counterparts. Plants were cut at the soil surface and fresh weights for each plot were measured. A fresh weight subsample (15 to 25 plants) was then taken for dry weight determination. Samples were dried at 70 C and separated into grain and stover, which were ground separately and digested in H2SO4 after pre‑treatment with H2O2 (Parkinson and Allen,1975). Ammonium was determined in the digests by the indophenol blue method (Keeny and Nelson, 1982).
Durations of vegetative and seed‑filling phases were calculated, respectively, as days from planting to first flower (R1) and days to physiological maturity (R7) minus days to beginning seed (R5). Rate of seed dry matter accumulation was determined by dividing grain yield ha‑1 by seed‑fill duration. Growing degree days (Major et al., 1975b) were calculated using a base temperature of 7.8 C (Hadley et al., 1984).
Data
analysis: The data from the three sites were combined (McIntosh, 1983) and
subjected to analysis of variance using SAS (SAS Institute. 1982. SAS user's
guide: Statistics. 1982 ed. SAS Institute, Inc.,
Results
Mean maximum and minimum temperatures between the sites at 320 and 1050 m differed by about 4 C and 6 C, respectively, during the course of the experiment (Table 1). The decline in temperature from the start to the end of the experiment at a site was gradual and small (Figure 1). The photoperiod decreased from 13 h at planting to 11 h at the end of the experiment (Figure 1).
There were significant differences between sites and varieties in days to first flower (R1) and physiological maturity (R7) (Table 2). Both phenological events were delayed considerably at the higher elevations compared to the lowest elevation. The effect of increasing elevation was progressively greater on higher maturity groups. Flowering of Clay was only 12 days later at 1050 m than at 320 m while Hardee flowered 31 days later at the highest site compared to the lowest site. Similarly, Hardee flowered 15 days later than Clay at the 320 m site, but was 34 days later than Clay at 1050 m. Phenological development was consistent with maturity group number. Lower maturity groups flowered and matured more rapidly than higher maturity groups. Thus, within a site, each event was recorded first and last, respectively, for Clay and Hardee. The indeterminate maturity group VI variety N77 required a significantly greater number of days than the determinate VI variety D68 to reach both R1 and R7.
Growing degree days (GDD) required for vegetative growth for all varieties increased slightly with increasing elevation (Table 3). Because the seed filling period was approximately constant across the sites, GDD for the total crop duration actually decreased with increasing elevation. Late maturing varieties required a greater number of GDD to complete vegetative growth and reach maturity at all sites.
The differences in duration of the seed‑filling phase were much smaller than differences in durations of vegetative and total crop growth. Varieties differed significantly in duration of seed‑filling (stage R5 to R7) (Table 4). Differences between sites in seed‑fill duration, although significant, were small. Clay, the earliest maturing variety, had the shortest seed‑filling duration. The other maturity groups were inconsistent in this regard. When averaged across sites, Clark and Hardee had the longest seed‑fill period (Table 4) while N77 had the highest rate of seed dry matter accumulation (Table 5). The rate of seed dry matter accumulation declined significantly with increasing elevation and was lowest for the two early varieties at the 1050 m site.
Non‑nod isolines flowered later and matured earlier than their nodulating counterparts. When averaged over sites, the isolines of Clay and Clark were similar in days to R1 (Table 6), but non‑nod isolines of other varieties were late in flowering. Nod isolines had significantly higher concentrations of N in both the shoot and the grain (Table 7) than did the non‑nods. The yield of non‑nods were substantially lower than the nods.
Discussion
The delay in the occurrence of first flower (R1) and physiological maturity (R7) (Table 2) due to increasing elevation was assumed to be due to a 5 C decline in mean daily temperature from the lowest to the highest site. The increased number of days to flowering at the higher elevation, however, did not result in a similar increase in days to physiological maturity. A delay in flowering of soybean due to low temperature has been reported by many investigators (Hadley et al., 1984; Hesketh et al., 1973; Steinberg and Garner, 1936; van Schaik and Probst, 1958). According to Hadley et al., (1984) the reciprocals of the days taken to flower (rate of progress towards flowering) by soybean were linear functions of mean diurnal temperature in photoperiods shorter than the critical daylength. Assuming that the effect of the prevailing photoperiod on flowering was the same at the three sites, differences between sites in days to flowering could be due to differences in temperature. The consistent phenological responses of varieties to increasing elevation (Table 2) indicates that temperature was the dominant effect. However, critical day lengths for soybean are shorter in cooler temperatures and when photoperiods are longer than the critical value, the rate of progress towards flowering is a linear function of both temperature and photoperiod with no interaction between the two (Hadley et al., 1984). Thus, under a constant temperature regime and inductive photoperiods, soybean genotypes would flower about the same time regardless of their maturity group classification. In our experiment, flowering in order of progressively increasing maturity group number indicated that photoperiods may have been non‑inductive for the higher maturity groups at the beginning of the experiment. The plants at the highest elevation may have been subjected relatively more to non‑inductive photoperiods than those at the lowest elevation, further delaying flowering at that site. The overall effect was to increase the relative crop duration for successive maturity group numbers at higher elevations. Since Clay, maturity group 00, might be considered as relatively insensitive to photoperiod (Major et al., 1975a; Criswell and Hume, 1972), 12 and 43% delays in flowering at the intermediate and highest sites respectively may have been due entirely to lower temperatures.
The large increases in GDD required for vegetative development with increasing elevation for the higher maturity groups indicates the unsuitability of GDD as a predictor of phenological events in soybean under non‑inductive photoperiods. The present data also agree with the conclusion of Major et al., (1975b) that thermal unit methods, including GDD, only accurately predicted development of early maturing soybean varieties, i.e., varieties which were less sensitive to photoperiod.
The data indicate that the effects of temperature on the seed‑filling duration was minimal under similar N nutritional levels. The finding of Hesketh et al., (1973) that reproductive duration was relatively unaffected by temperatures in the range of 22 to 30 C may have been due to similar seed fill durations. Varietal differences in durations of seed‑filling were associated with differences in grain yield within a site; however, no consistency among sites was observed in this respect. On the other hand, a closer association between grain yield (can be calculated from Tables 4 & 5) and rate of seed dry matter accumulation (Table 5) was apparent. The above observation is in contrast to reports by others (Dunphy et al., 1979; Egli, 1975; Hanway and Weber, 1971) that grain yield is strongly related to seed‑fill duration with the rate of seed dry matter accumulation being a constant.
There was significant reduction in grain yield (can be calculated from Tables 4 & 5) of all varieties due to decreasing temperature. In general, yield increased with increasing duration of vegetative growth at each site, indicating a positive relationship between vegetative dry matter production and grain yield. Therefore, delay in phenological development up to the beginning seed stage (R5) due to decreasing temperature may be an adaptation by the plant to compensate for decreased rates of growth.
The differences in the response to temperature of nod and non‑nod isolines indicates how nitrogen nutrition can alter soybean phenological development. The net result of delayed flowering and hastened maturity by the non‑nods was shortened reproductive and seed‑fill durations. The significantly lower shoot and grain N concentrations of non‑nods than nods (Table 7) indicated N insufficiency in non‑nod plants. The early maturity of non‑nods was apparently due to nitrogen insufficiency. The cause of the delayed flowering of the non‑nod isolines is unknown although Huxley et al., (1976) reported that while the onset of flowering was substantially delayed by lower night temperature, low nitrogen levels also delayed flowering in soybean cv. TK5. Therefore, nitrogen nutrition may be another important consideration in comparing soybean phenology across temperature regimes.
The effects of temperature on phenological development of soybean under field conditions were demonstrated in this experiment. The results indicate that low temperature delayed flowering in soybean regardless of sensitivity to photoperiod. The extended vegetative growth due to decreasing temperature may be an adaptation by the soybean plant to compensate for decreased rates of growth. Duration of seed‑fill was relatively unaffected by temperature under similar N nutritional levels, but seed dry matter accumulation per day per unit land area was substantially decreased by cool temperature. Non‑nodulating isolines, in general, were late in flowering but early in maturity and their development was related to the degree of N insufficiency at the respective sites.
Table 1. Mean maximum and minimum temperatures during growth of
five
soybean varieties at three elevations on the
________________________________________________________________
Variety Maturity Temperature (C)
Group Max Min Max Min Max Min
320 m 660 m 1050 m
________________________________________________________________
Clay 00 26.81 19.8 25.7 17.1 23.1 14.2
D68 VI 26.5 19.8 25.3 17.0 22.9 14.0
N77 VI 26.4 19.8 25.3 17.0 22.5 13.7
Hardee VIII 26.2 19.6 25.3 16.8 22.5 13.7 ________________________________________________________________
1 Mean temperatures from planting to maturity.
Figure
1. Weekly mean temperatures and daylengths during the growth of five soybean
varieties at three elevations on the
Table 2. Days to first flower (R1) and physiological maturity
(R7) of nodulating isolines of five soybean varieties grown at
three elevations.
________________________________________________________________ First flower Physiological maturity
Variety 320 m 660 m 1050 m 320 m 660 m 1050 m
_______________________________________________________________ ----------------------days----------------------
Clay (00)1 27.7E 31.0E 39.7E 73.0E 77.7E 93.0E2
Clark (IV) 30.0D 35.0D 46.0D 87.0D 92.0D 107.0D
D68 (VI) 33.7C 41.3C 55.7C 90.0C 101.0C 112.0C
N77 (VI) 38.0B 49.0B 66.0B 94.0B 105.0B 128.0B
Hardee(VIII)42.3A 54.3A 73.7A 106.0A 117.0A 133.3A
Mean 34.3c2 42.1b 56.2a 90.0c 98.5b 114.7a ________________________________________________________________
1 Maturity grouping.
2 Means followed by the same upper case letter within a column
or lower case letter within a row for each category are not
significantly different (P=0.05) by
test.
Table 3. Growing degree days accumulated during vegetative growth
and up to physiological maturity for nodulating isolines of five
soybean varieties grown at three elevations.
________________________________________________________________
Vegetative phase Total crop duration
Varieties 320 m 660 m 1050 m 320 m 660 m 1050 m
________________________________________________________________
‑‑‑‑‑‑‑‑‑‑‑Growing degree days1‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑
Clay 432 426 444 1131 1060 1009
D68 523 564 616 1382 1348 1193
N77 588 667 723 1438 1402 1318
Hardee 651 740 811 1600 1550 1370
________________________________________________________________
1 Calculated using a base temperature of 7.8 C.
Table 4. Duration of seed‑filling1 of nodulating isolines of
five soybean varieties grown at three elevations.
______________________________________________________________
Elevation (m) Mean
Varieties 320 660 1050
______________________________________________________________
‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑days‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑
Clay 36.0 35.0 38.0 36.3D2
D68 42.3 45.0 40.3 42.6B
N77 39.0 39.3 44.3 40.9C
Hardee 47.3 46.0 43.3 45.6A
Mean 42.3a2 41.7b 42.4a
______________________________________________________________
1 Days from beginning seed (R5) to physiological maturity (R7).
2 Means followed by the same upper case letter within a
column and lower case letter within a row are not
significantly different (P=0.05) by
test.
Table 5. Rate of seed dry matter accumulation1 of nodulating
isolines of five soybean varieties grown at three elevations.
______________________________________________________________
Elevation (m) Mean
Varieties 320 660 1050
______________________________________________________________
‑‑‑‑‑‑‑‑‑‑‑‑kg ha‑1 day‑1‑‑‑‑‑‑‑‑‑‑‑‑‑
Clay 89.5 80.2 13.6 61.1D2
D68 101.0 82.7 47.1 76.9B
N77 90.5 89.6 58.2 79.4A
Hardee 102.1 78.3 43.3 74.6B
Mean 95.6a2 80.3b 38.4c
______________________________________________________________
1 Determined by dividing grain yield ha‑1 by duration of seed
fill.
2 Means followed by the same upper case letter within a
column and lower case letter within a row are not
significantly different (P=0.05) by
test.
Table 6. Mean days to first flower and physiological maturity
for nodulating (nod) and non‑nodulating (non‑nod) isolines of five
soybean varieties grown at two elevations.1 ________________________________________________________________
First flower (R1) Physiological maturity (R7)
Varieties Nod Non‑nod Nod Non‑nod
________________________________________________________________ ‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑days‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑
Clay 32.8a2 32.6a 75.3a 70.5b
D68 43.6b 46.6a 95.5a 90.5b
N77 51.0b 52.4a 99.5a 96.0b
Hardee 56.8b 62.1a 111.5a 106.3b
________________________________________________________________
1 Data averaged for the Kuiaha (32 m) and Haleakala (660 m)
sites.
2 Means followed by the same letter within a row for each
category are not significantly different
(P=0.05) by
multiple range test.
Table 7. Mean grain and shoot nitrogen concentrations of
nodulating (nod) and non‑nodulating (non‑nod) isolines and
grain yield of non‑nods of five soybean varieties grown at
three elevations.
____________________________________________________________
Nitrogen concentration
Shoot Grain Non‑nod
Varieties Nod Non‑nod Nod Non‑nod Grain yield ____________________________________________________________
‑‑‑‑‑‑‑‑‑‑‑‑‑‑%‑‑‑‑‑‑‑‑‑‑‑‑‑‑‑ kg ha‑1 %1
Clay 1.14a2 0.49b 6.46a 4.21b 826 37.9
D68 0.99a 0.44b 6.17a 3.52b 1262 38.3
N77 1.08a 0.36b 6.22a 4.05b 1253 39.0
Hardee 1.20a 0.59b 6.21a 4.80b 1046 30.4
____________________________________________________________
1 As a percentage of the grain yield of the nodulating isoline.
2 Means followed by the same letter within a row for either
shoot or grain are not significantly different(P=0.05) by
Chapter 4
YIELD, SOIL N UPTAKE AND N2 FIXATION BY SOYBEANS FROM
FOUR MATURITY GROUPS GROWN AT THREE ELEVATIONS
Abstract
The exploitation of the soybean-Bradyrhizobium
japonicum symbiosis in varied environments requires an understanding of
factors that may affect fixed and soil N assimilation. Temperature affects both soybean maturity and
N requirement, and soil N availability.
Five soybean varieties belonging to four maturity groups (00, IV, VI,
and VIII) and their respective non‑nodulating isolines were planted on
the same day at three sites along an elevational transect in
Introduction
Soybean plants can acquire a substantial portion of their N requirement through N2 fixation from symbiotic association with Bradyrhizobium japonicum. Both management and environmental variables affect the symbiosis. Any factor that affects the availability and plant uptake of soil N could directly influence N2 fixation and soybean yield. An understanding those factors that may differentially affect fixed and soil N assimilation could help enhance exploitation of the symbiosis.
Optimum temperature promotes growth, and symbiotic and soil N assimilation by soybean and other legumes. Increasing root temperature within the range of 15 to 27 C has been shown to increase growth and dry matter production in soybeans (Earley and Cartter, 1945; Matthews and Hayes, 1982; Trang and Giddens, 1980). Lowering both root and shoot temperature from 25 to 10 C decreased photosynthesis in soybean more than reducing shoot temperature alone (Taylor and Rowley, 1971). Nodule activity was reduced to the same extent if shoots or roots and shoots of soybean were exposed to 18 C temperature (Schweitzer and Harper, 1980). Harding and Sheehy (1980) showed that leaf growth of lucerne (Medicago sativa) was correlated with air temperature, while N2 fixation was correlated with root temperature. Temperature also affects soil N mineralization (Stanford et al., 1973; Cassman and Munns, 1980) and root‑uptake of mineralized N (Tolley and Raper, 1985). Therefore, temperature may directly and indirectly affect growth of soybean through effects on photosynthesis, N uptake and N2 fixation. Raper, et al., (1977) suggested that plants respond to temperature in such a way that N assimilation and photosynthate supply are in balance.
Due to the strong photoperiod sensitivity
of soybeans (Borthwick and Parker, 1939), assessment of temperature effects on
productivity under field conditions is often difficult. In the soybean producing regions of
The present investigation was undertaken to evaluate the effect of temperature on growth, phenology, (Chapter 3), yield, soil N uptake and N2 fixation by soybean varieties from four maturity groups. Results reported here include yield, soil N uptake and N2 fixation. By planting several varieties from a range of maturity groups at field sites along an elevational transect within the Maui Soil, Climate and Land Use Network: Mauinet, it was possible to minimize the effects of photoperiod and irradiance on soybean growth and development.
Materials and Methods
The experimental design, field and plant culture procedures, weather data collection and sampling procedures for yield and total N were described in Chapter 3. Only those procedures specific to this study are described here. Weather data for this study included irradiance measurements which were recorded by Campbell Scientific CR-21 micrologger (Campbell Scientific, Inc., Logan, UT) equipped with sensing probes for irradiance (Li-Cor LI 200ZC silicon pyranometer; Li-Cor, Inc., Lincoln, NE).
Total N and fixed N calculations: Total plant N was calculated by multiplying the percent tissue N concentration by total plant dry weight. The amount of N fixed was determined by subtracting the total plant N in the non‑nod isoline from its nod counterpart.
In vitro soil N mineralization: Soils from the three sites were sampled at planting to a depth of 20 cm to determine N mineralization potential of the soil under constant conditions. Samples of approximately 20 g air-dry soil were equilibrated on a pressure plate at 0.03 MPa for initial analysis and incubation (Cassman and Munns, 1980). Triplicate samples were extracted with 200 ml of 2 M KCl before and after incubation at 25 C in 250 ml flasks for two weeks. Flasks were covered with aluminum foil that was pierced with two pinholes. Water loss was negligible during incubation. Nitrate and NH4+-N were determined from the filtered extracts by cadmium reduction and indophenol blue methods, respectively (Keeny and Nelson, 1982).
Greenhouse experiment: Clark non‑nod soybean was planted in pots (two plants pot-1, three replications) containing 4 kg soil (oven-dry basis) from each of the three sites to measure plant uptake of N from the soils under a constant temperature regime. Soils were maintained at 0.03 MPa tension. The mean soil temperature during the experiment was 27 C. At 45 days from planting, plants were cut at the soil surface and analyzed for total N as described previously.
Data analysis: The data from the three sites were combined (McIntosh, 1983) and subjected to analysis of variance using SAS procedures (SAS Institute Inc., Box 8000, Cary, North Carolina 27511‑8000).
Results
Irradiance at the 660 m and 1050 m sites was 80 and 75% of that at the 320 m site (Table 1). Mean soil and air temperatures declined by about 5 C from the 1050 m to 320 m.
There were significant differences in grain and stover yields among sites and varieties (Table 2). Average grain yield and stover yields at the highest site were only 41 and 51%, respectively, of those at the lowest site. Yields were reduced at higher elevations to a greater extent in the early maturing varieties, Clay and Clark, than in later maturing varieties. In general, grain yields increased with maturity group number at a site. Variety N77‑4262, an indeterminate maturity group VI entree was less affected by low temperatures than other varieties. Hardee produced the highest average yield.
Total N assimilation (Table 3) and amounts of N fixed (Table 4) decreased with increasing elevation (Table 3 and 4). The effect of elevation was relatively greater on the early maturing variety, Clay. Total N and fixed N increased with increasing maturity group number. Varietal differences in the rate of N assimilation within a site were relatively small compared to the effect of temperature (Table 5).
The relative contributions from symbiotic
N2 fixation and soil N uptake to total N were significantly
different between sites (Table 6), but relatively constant for all varieties
within a site except for N77‑4262 at the 320 m site. The greatest soil N uptake was observed at
the Haleakala site (660 m) and the result was consistent with in vitro
mineralization measurements (Table 7).
Soil N uptake at
Discussion
By planting soybean at different elevations in the same latitude, any interactive effects between photoperiod and temperature on plant growth and development were minimized. Thus, the site effects on grain yield, symbiotic N2 fixation, and soil N uptake of soybean genotypes were assumed to be due to differences in temperature. It is unlikely that the 25% difference in irradiance between sites accounted for the reduction in yield and total plant N observed in this experiment. Despite similar irradiances at the 660 and 1050 m elevations, there were large differences in yield and N assimilation between these two sites. Since nutrients other than N and water were assumed to be non-limiting, the reduction in yield was attributed primarily to decreasing temperature with increasing elevation. Low temperatures may have reduced both photosynthesis and nutrient assimilation.
Seed yield was positively correlated with total N assimilation and the amount of nitrogen fixed. Crop durations increased with maturity group number as well as with decreasing temperature (Chapter 3). Increased crop duration was mainly due to an extended vegetative phase and was positively correlated with seed yield within a site. Thus, the increase in yield and total N with increasing maturity group number was due to increasing dry matter resulting from prolonged growth. Though total N and yield increased with maturity group, the rate of N assimilation (kg N ha-1 d-1) was not related to maturity differences between cultivars within a site. The large reductions in N assimilation per day due to decreasing temperature demonstrates the effect of temperature on crop growth and N requirement, and in turn on N assimilation, as suggested by Raper et al., (1977). Yield and N assimilation of short term varieties were reduced more by low temperature than longer maturing varieties, probably due to the short vegetative durations.
The fact that 97% of the plant N
requirement of nod isolines at
The proportions of N derived from fixation (Table 6) within a site were similar despite large differences in total N assimilation between maturity groups. Since the different maturity groups had similar average N assimilation rates per day within a site and soil N mineralization rate per day at a site was assumed to be constant, the proportion of total N derived from N2 fixation necessarily was constant. This is reasonable since soil N availability and uptake are functions of soil and temperature, and, within a site, temperature was relatively constant for the different maturity groups.
Soil N uptake at
The nod isolines at
Considering the complexity of the interplay of factors that may affect soil N availability and plant uptake, there are two reasonable temperature-related explanations for the low soil N uptake observed. During field preparation soil N might have been immobilized due to incorporation of residues from the previous grass vegetation, and immobilization may have been prolonged due to low soil temperatures. However, the primary tillage operations were carried out 4 weeks before planting at all sites, yet more soil N was available at the Haleakala site even though the soil temperature there was lower than that at the Kuiaha site. Another possibility might have been that soil N assimilation and N2 fixation by soybean are differentially tolerant to low temperature, such that root growth and uptake of soil N was reduced more than N2 fixation. Rufty et al., (1981) reported that root growth and nitrate absorption were less at 18 C than at 24 C. In contrast, soybean nodules have been reported to have maximum activity over the range of 15 and 30 C (Pankhurst and Sprent, 1976). Thus, in this experiment, N2 fixation may not have been reduced by low temperature to the same extent as was mineral N assimilation.
The different behavior of N77‑4262 is of interest. Although yield and N assimilation of N77‑4262 were relatively low at the lowest site, they were less affected by temperature than other varieties. This variety fixed similar amounts of N at the lowest and highest sites. The results indicate that this indeterminate variety may be suited for a wide range of temperature regimes at this latitude.
The results demonstrate the dependence of soybean growth and N assimilation on temperature. Decreasing temperatures caused greater restrictions on plant growth and dry matter production than on the functioning of symbiotic N2 fixation. The relative contribution of symbiotic N2 fixation was a function of soil N availability at a site regardless of crop duration and total N assimilation by different varieties. N assimilation per unit time was relatively constant for maturity groups within a site despite large differences in total N assimilation.
Table 1. Characteristics of three sites in an elevational transect
on
the island of
__________________________________________________________________
Site Soil Rainfall1 Temperature1 Irradiance1 (Elev., m) classification Soil Air __________________________________________________________________
mm C W m-2
Kuiaha Clayey, ferritic,
(320) isohyperthermic 415 24.4 22.6 262 Humoxic Tropohumult
Haleakala Clayey, oxidic,
(660) isothermic 407 22.6 20.6 211 Humoxic Tropohumult
(1050) skeletal, isomesic 678 19.6 17.8 194 ________________________________________________________________
1 All data averaged across the crop duration of all five
varieties at a site
Table 2. Grain and stover yields of five nodulating soybean
varieties grown at three different elevations under the same photoperiodic regime.
________________________________________________________________
Varieties
Site Clay Clark D68‑0099 N77‑4262 Hardee Mean (elevation, m) 001 IV VI VI VIII
_________________________________________________________________
------------------kg ha-1-----------------
Grain yield
Kuiaha 3223 4424 4273 3530 4833 4057A2
(320)
Haleakala 2803 3066 3721 3526 3601 3344B
(660)
(1050)
Mean 2181d2 2949c 3299ab 3212b 3437a
Stover Yield
Kuiaha 3105 3859 3661 3265 4117 3601A
Haleakala 2891 3381 3763 3829 4060 3585A
Mean 2194d 2877c 3171b 3197b 3570a
________________________________________________________________
1 Indicates maturity group designation of the variety.
2 Data followed by the same letter within the column or row are
not significantly different (P=0.05) by
range test.
Table 3. Total nitrogen assimilation by five nodulating soybean varieties grown at three different elevations.
________________________________________________________________
Varieties
Site
Clay
(elevation, m) 001 IV VI VI VIII
________________________________________________________________
--------------------kg N ha-1----------------
Kuiaha (320) 236 317 316 260 349 295A2
Haleakala (660) 199 246 271 258 276 250B
Mean 160d2 221c 238b 234bc 256a
________________________________________________________________
1 Indicates maturity group designation of the variety.
2 Data followed by the same letter within the column or row are
not significantly different (P=0.05) by
range test.
Table 4. Symbiotically fixed nitrogen by five soybean varieties
grown at three different elevations.
________________________________________________________________
Varieties
Site
Clay
(elevation, m) 001 IV VI VI VIII
________________________________________________________________
----------------------kg N ha-1---------------
Kuiaha (320) 199 258 268 180 281 237A2
Haleakala (660) 122 175 176 169 188 166B
Mean 121c2 177b 190ab 177b 203a
________________________________________________________________
1 Indicates maturity group designation of the variety.
2 Data followed by the same letter within the column or row are
not significantly different (P=0.05) by
range test.
Table 5. Average rate of nitrogen assimilation by five soybean varieties grown at three different elevations.
________________________________________________________________
Varieties
Site
Clay
(elevation, m) 001 IV VI VI VIII
________________________________________________________________
----------------kg N ha-1 day-1---------------
Kuiaha (320) 3.23 3.64 3.51 2.76 3.29 3.29A2 Haleakala (660) 2.56 2.67 2.68 2.45 2.36 2.55B
Mean 2.09b2 2.41a 2.44a 2.22b 2.24b
________________________________________________________________
1 Indicates maturity group designation of the variety.
2 Data followed by the same letter within the column or row are
not significantly different (P=0.05) by
range test.
Table 6. Percent of total nitrogen derived from symbiotic
fixation by five soybean varieties grown at three elevations.
________________________________________________________________
Varieties
Site Clay Clark D68‑0099 N77‑4262 Hardee Mean
(elevation, m) 001 IV VI VI VIII
________________________________________________________________
---------------------%----------------------
Kuiaha (320) 85 82 85 70 80 80B2 Haleakala (660) 62 71 65 66 68 66C
Mean 81ab2 84a 83a 78b 82ab
________________________________________________________________
1 Indicates maturity group designation of the variety.
2 Data followed by the same letter within the column or row are
not significantly different (P=0.05) by
range test.
Table 7. Comparison of estimated amounts of mineralized N and actual soil nitrogen uptake by five soybean varieties at three elevations.1
________________________________________________________________ Kuiaha, 320 m Haleakala, 660 m
Varieties Minera‑ Soil Minera‑ Soil Minera‑ Soil
lizable nitrogen lizable nitrogen lizable nitrogen
nitrogen uptake nitrogen uptake nitrogen uptake
________________________________________________________________
--------------------kg N ha-1--------------------
Clay 46.8 37.0
145.1 76.7 37.1
2.2
1 Mineralizable nitrogen estimated from the soil incubation study
(25 C, 0.03 MPa) and adjusted for soil temperature and the duration
of crop growth. [For details of calculation, see Stanford et al.
(1973, 1974)]. Actual soil N uptake is the N content of non‑nodulating plants.
Table
8. Nitrogen uptake by
_____________________________________________________________
Estimated rate Rate of nitrogen
Soils of nitrogen uptake by shoot2
mineralization1
_____________________________________________________________
-------ug N g soil-1 day-1--------
Kuiaha 0.39 1.02
Haleakala 1.29 1.31
_____________________________________________________________
1 Estimated from soil incubation study (25 C, 0.03 MPa).
2 Calculated from the N uptake from 4 kg of soil (oven
dry basis) by two plants after 45 days of growth.
Chapter 5
GROWTH RESPONSES OF FIELD GROWN SOYBEAN AND COMMON
BEAN TO TEMPERATURE AT VARYING LEVELS OF N NUTRITION
Abstract
An understanding of the adaptation of legumes to varied environments is a prerequisite for selecting appropriate species or cultivar for maximum yield in a new environment. To assess this adaptation in soybean (Glycine max (L.) Merrill) and common bean (Phaseolus vulgaris L.) to temperature at varying levels of N nutrition, these two legumes were evaluated. The two species were grown at elevations of 320 m and 1050 m with applied N levels 9, 120 and 900 kg N ha-1. The mean soil/air temperatures (C) during the experiment were 25/23 and 21/19 at the 320 and 1050 m elevations, respectively. Measurements of growth were made at end bloom (R2/R3), pod fill (R5/R6) and physiological maturity (R7). Higher elevation (low temperature) extended the vegetative growth period of both species an average of 44% and total crop duration 38%; low temperature had a relatively greater effect on soybean than on common bean. Application of 900 kg N ha-1 delayed the average days to R7 of soybean from 98 to 102 and common bean from 85 to 87. Average crop growth rate decreased by 26% at the 1050 m site compared to the 320 m site, but each legume attained similar dry weights at the two sites because the duration of growth was prolonged at the 1050 m site. Grain yield of soybean was higher than common bean at the 320 m site; the reverse was the case at the 1050 m site. Grain yield of soybean decreased by 30% at the 1050 m site due to a reduction in harvest index. In contrast to soybean, the grain yield of common bean was similar at the two elevations. Application of 900 kg N ha-1 increased the grain yield an average of 22% primarily by increasing the crop growth rate and total dry matter production. During vegetative growth, common bean accumulated 46% more dry weight than soybean due to a 34 % higher crop growth rate and a longer vegetative period than soybean. The crop growth rate during the early reproductive phase of soybean (13.9 g m-2 d-1) was 31% faster than common bean, yet the average growth rates of the two legumes over the crop duration were not significantly different. Total dry matter of soybean was 9% greater than common bean at R7, primarily due to a longer crop duration. Soybean had increased leaf production (leaf area and leaf weight ratio) and substantially reduced unit leaf rate compared to common bean at the 1050 m site. Nitrogen application increased the leaf area and crop growth rate of both legumes. The results indicate that extended growth duration and increased leaf production were the primary ways by which soybean and common bean compensated for reduced rate of growth at the higher elevation. The growth and yield of common bean was less affected by cool temperature than soybean.
Introduction
Understanding the effects of varied environments on growth and development of soybean (Glycine max (L.) Merrill) and common bean (Phaseolus vulgaris L.) may be helpful in selecting the appropriate species or cultivar for maximum grain yield in a new environment. Grain yield is determined by the total dry matter production and the proportion of total dry matter partitioned to reproductive tissue. Total dry matter is a function of the rate and duration of growth. Both the rate and the duration of growth as well as partitioning of dry matter within the plant are influenced by environmental variables.
Temperature affects growth, development,
and yield of legumes. Low temperature
decreases their growth rate by decreasing the rate of photosynthesis through
effects on structural and functional characteristics of leaves (
Among the nutrient variables, nitrogen is especially important for legumes because of their high N requirement and tissue N concentration. Photosynthesis and dry matter accumulation by soybean and common bean are positively correlated with tissue N concentration (Lugg and Sinclair, 1981; Ojima, et al., 1965; Ryle and Hesketh, 1969). A reduction in N supply decreases the growth rate of new tissue more than the rate of photosynthesis in older leaves (Raper and Peedin, 1978). Nitrogen stress also affects partitioning of N to seeds. Nitrogen limits the partitioning of dry matter and N to seeds relatively more than photosynthesis in soybean (Crafts-Brandner et al. 1984; Streeter, 1978).
The nitrogen requirement of legumes is met from both symbiotic and mineral N sources. The plant N status as influenced by symbiosis and mineral N availability have been ignored in studies of growth and development of legumes. Experiments conducted in 1985 (Chapter 3) showed that non-nodulated soybean plants grown under moderate levels of soil N matured earlier and yielded less than their nodulated counterparts due to N stress, regardless of the temperature regime. The high requirement of seeds for N, at a time when current assimilation of N from mineral and atmospheric sources is declining (Wych and Rains, 1979; Thibodeau and Jaworski, 1975), requires considerable remobilization of N from vegetative tissues to maintain seed filling in legumes (Sinclair and de Wit, 1976). Temperature and mineral N supply regulate both current assimilation and remobilization from vegetative tissues of N to seed in legumes.
It is clear that the effects of temperature and mineral N supply on growth, development and N nutrition are aspects of adaptability of legumes to environment. The purpose of the present study was to evaluate the response of soybean and common bean to temperature differences associated with elevation under varying levels of N nutrition.
Materials and Methods
Experimental design: The treatments were elevations of 320 and 1050 m, inoculated and uninoculated soybean, Glycine max (L) Merrill, and common bean, Phaseolus vulgaris (L), and nitrogen levels of 9, 120, and 900 kg N ha-1. The experimental design was a randomized complete block with three replications at each elevation. At each site, the treatments were arranged in a split-split plot with soybean and common bean assigned to main plots, inoculation to sub plots and the three levels of nitrogen to sub-sub plots.
Field
and plant culture: The characteristics of the two sites on the
Seeds of common bean, P. vulgaris cv. Brazil 2 (Centro Internacional de Agricultura Tropical) and soybean, G. max cv. Clark were planted at the 1050 m site on July 8, 1987 and at the 320 m site on July 15, 1987. The seeds for the inoculated treatment were coated with peat inoculants containing Rhizobium leguminosarum biovar phaseoli strains TAL 182, CIAT 632 and CIAT 899 for common bean and Bradyrhizobium japonicum strains USDA 110, USDA 136b, and USDA 138 for soybean. Seeds were sown in rows 60 cm apart in plots 9.5 by 3.0 m to attain a final population of 400,000 plants ha-1. Plots were watered to field capacity on the day of planting and reirrigated when tensiometer readings were approximately 0.02 MPa. Nitrogen as ammonium sulfate was applied in equal amounts after planting and at growth stages end bloom (R2/R3) and pod fill (R5/R6).
Plant sampling procedures: Measurements of growth and yield were made by harvesting plants approximately at growth stages end bloom, pod fill and physiological maturity (R7). Plant fresh weight was estimated by harvesting all plants at the soil surface from a 2.4 by 1.8 m area. Dry matter content of whole plants was established from a 10 to 15 plant sub sample after drying at 60 C. The sub samples at R7 were threshed for seed yield. Ground samples were digested by the salicylic acid modification of the Kjeldahl procedure (Keeney and Nelson, 1982) and total nitrogen in the digests was measured by the indophenol blue method (Keeney and Nelson, 1982).
Leaf area was measured with a Licor LI-3100 leaf area meter on leaves removed from 6 plants selected at random from each harvest. Leaves and stems were then dried (60 C) and weighed separately.
Growth analysis: Mean crop growth rate (CGR) was calculated by dividing dry matter accumulated by days between harvests. Leaf area index (LAI) was expressed as the leaf area per unit of land area. Leaf weight ratio (LWR) was calculated by dividing leaf dry weight by the total shoot dry weight. Approximate values of unit leaf rate (ULR) for each growth phase were calculated by the equation: ULR = Y / LAD, where Y is the dry matter increment during a given growth period and LAD is the leaf area duration for the same growth period (Hunt, 1982). LAD was calculated by the formula: LAD = (L1 + L2) (T2-T1) / 2, where L1 and L2 are the LAI at the beginning and end of the growth period and T1 and T2 are the number of days from the beginning and end of the growth period (Hunt, 1982).
Soil sampling: Extractable soil NH3-N and NO3-N were determined from soil samples collected to a depth of 20 cm 5 days after planting. Soil suspensions (1 part air dry soil: 7 parts 2M KCl) were agitated for one hour on a mechanical shaker. NH3-N and NO3-N were determined from filtered KCl extracts by the indophenol blue and the cadmium reduction methods, respectively (Keeny and Nelson, 1982).
Data analysis: The data from the inoculated segment of the experiment were subjected to analysis of variance procedures. Data from the uninoculated segment of the experiment were discarded because of nodulation observed in the uninoculated controls of common bean (see Appendix Table A-1).
Results and Discussion
The effects of temperature and N nutrition on growth and grain yield of soybean and common bean were studied at elevations of 320 m and 1050 m which provided approximately a 4 C differential in both soil and air temperatures (Table 1). The irradiance at the 1050 m site was 23% lower than the 320 m site, but the total solar radiation received during the total crop duration was highest at the higher site (Table 1). The difference in the irradiance between the sites was assumed to be insignificant for reasons discussed elsewhere (Chapter 4).
Only main effect means are presented and discussed unless significant interactions were encountered in the analysis of variance.
The vegetative and total durations were extended at the high cool site, with a relatively greater effect on soybean than common bean (Table 2). Although common bean flowered later than soybean, it matured earlier resulting in shorter reproductive and total durations compared to soybean. Flowering and maturity of soybean were also delayed by cool temperatures in the 1985 experiment (Chapter 3) and similar results have been obtained by others (Hadley et al., 1984; Hesketh et al., 1973). Days to flowering increased relatively more at the 1050 m site than reproductive or total duration for both legumes, a result similar to that obtained in 1985 (Chapter 3).
Nitrogen application did not affect days to flowering but slightly delayed maturity of both legumes. Maturity of soybean was delayed by 5% by application of 900 kg N ha-1 compared to a 2% delay in common bean. This result is in agreement with the conclusion reached in Chapter 3 that N-deficient non-nodulating soybean plants matured earlier than their nodulating counterparts (Chapter 3).
The effect of elevation and N nutrition on growth and development will ultimately affect the grain yield of legumes. Grain yield is dependent on vegetative growth (source) and reproductive growth (sink) (Summerfield and Wien, 1980) as well as on the proportion of total dry matter partitioned to seed. Grain yields of soybean and common bean in the present experiment were differentially affected by the site environment (Table 3). While soybean had higher grain yield than common bean at the 320 m site, the reverse was true at the 1050 m site. The maximum grain yield of soybean at the highest site was 39% lower than that at the 320 m site. Unlike soybean, grain yield of common bean was similar between elevations. Grain yield of both legumes generally increased with N application, but the effect of N on soybean yield at the 1050 m site was marginal.
Harvest indices of the two species were comparable at the 320 m site (Table 4). While dry matter partitioning to grain yield in common bean was relatively unaffected by low temperature, soybean proved to be quite sensitive. Despite similar total dry weights between elevations for both legumes (Table 5), grain yield of soybean decreased due to drastic reduction in harvest index at the higher site. Low temperature during the reproductive phase is reported to adversely affect pod and seed growth in soybean (Holmberg, 1973; Hume and Jackson, 1980; Seddigh and Jolliff, 1984). According to Thorne (1982), low temperature reduces the translocation of photosynthate into soybean seed. The substantial decrease in the whole plant tissue N concentration of soybean at the 1050 m site (Table 4) might have exacerbated the effect of low temperature on harvest index (Crafts-Brandner et al., 1984; Streeter, 1978).
For a given harvest index, increases in grain yield may be realized by increased total dry matter yield. Dry matter yield is a function of the duration of crop growth and crop growth rate. Total dry weights at the different growth stages (Table 5) between elevations were not different except at the pod fill stage. Crop growth rate during all three growth phases was significantly lower at the 1050 m site than at the 320 m site (Table 6). Similar final dry weights for the two legumes at both sites indicate that the prolonged duration of growth (Table 2) compensated for the reduced rates of growth at the higher elevation. Compensation for low temperature inhibition of growth has been demonstrated for soybean (Chapter 3; Shibles, 1980; Summerfield and Wien, 1980). The results of this experiment indicates a similar mechanism for common bean.
The pattern of growth of the two legumes was unaffected by temperature. Dry weight accumulation by common bean during vegetative phase was 46% greater than soybean (Table 6) due to a 34% increase in crop growth rate and an 18% longer vegetative phase. The crop growth rate of soybean during the early reproductive phase was faster than that of common bean. The final dry weight of soybean was 9% greater than common bean. Although total dry matter of soybean was greater than for common bean at maturity (Table 5), the difference between the two legumes was not proportional to the 17% greater crop duration of soybean, mainly due to the fact that the crop growth rate of soybean declined substantially during the late reproductive phase. Despite the differences in crop growth rates between soybean and common bean during different phases of growth, average crop growth rates of the two legumes for the entire crop were similar. Therefore, total crop period accounted for most of the differences in final dry weight between soybean and common bean.
Crop growth rate increased with increase in N applied (Table 6) and the differences effectively accounted for the observed differences in dry matter accumulation (Table 5). Crop duration was also extended by N application (Table 2) accounting in part for the increased dry weight.
A reduced photosynthetic rate at low temperature may be compensated for by increased dry matter allocation to leaves. Increased leaf production for soybean was observed at the 1050 m site (Table 7) where leaf area index and leaf weight ratio were substantially higher than at the 320 m site, especially at pod fill. In contrast, leaf area index and leaf weight ratio of common bean were similar at the 320 and 1050 m sites, an indication of its tolerance to cool temperature. Nitrogen application increased leaf area (see Appendix Table A-6) with no differential effect between the two legumes. The increased partitioning of dry weight into leaves observed at the cool site agrees with the observation of Wann and Raper (1984) that plants respond to changes in temperature by altering the partitioning of dry matter among organs. A higher leaf area retained for a longer period of time at the higher site may increase the total canopy photosynthesis and therefore, net dry matter accumulation.
Data in Table 8 indicate that the unit leaf rate, a measure of net photosynthesis per unit leaf area, was generally lower at the 1050 m site; unit leaf rate of soybean was relatively more affected than common bean. Both legumes had similar unit leaf rates during the vegetative phase, but soybean had significantly less leaf area and so had a lower crop growth rate and less dry weight accumulation than common bean. The greatest difference in unit leaf rate of soybean at the two elevations was during the early reproductive phase when the leaf area was also the greatest. The unit leaf rate during the early reproductive phase was substantially greater at the 320 m site for soybean than common bean, but both legumes had similar rates at the 1050 m site. In contrast to soybean, the unit leaf rate of common bean was relatively unchanged between growth phases. Unit leaf rate was unaffected by N application (see Appendix Table A-7) and therefore, differences in crop growth rate were primarily due to changes in leaf area with N application.
Conclusions: The responses of soybean and common bean to temperature and N application were evaluated in terms of growth and phenological development. Reduced rates of growth by both legumes at the higher elevation was offset primarily by prolonged crop duration. Greater partitioning of dry weight into leaves was an additional mechanism by which soybean responded to low temperature. In spite of equivalent dry weight production among cool and warm sites, grain yield of soybean was reduced at the cool site due to lowered harvest index. Growth and yield of common bean was more tolerant to low temperature than soybean
Table
1. Environmental characteristics at two
sites in an elevational transect on the
________________________________________________________________
Site Elevation Soil Extract- Temperature2 Solar radiation
classifi- -able Soil Air Irradi- Total3
-cation Soil N1 -ance2
________________________________________________________________
m g N kg-1 -- C -- W m-2 MJ
Kuiaha 320 Humoxic 0.05 25 23 243 1640
Tropohumult
Dystrandept
________________________________________________________________
1 KCl extractable NH3+ and NO3- at five days after planting in
plots receiving 3 kg N ha-1 as 15N tracer.
2 Average for the experimental period.
3 Total solar radiation for the experimental period.
Table 2. Days to first flower (R1) and physiological maturity
(R7) of inoculated soybean and common bean grown at two
elevations with three levels of N.
_________________________________________________________
Nitrogen R1 R7
applied1 Soybean Bean Soybean Bean
_________________________________________________________
kg N ha-1 - - - - - - days - - - - - -
320 m elevation
9 28 35 79 73
120 28 35 84 74
900 28 35 84 75
1050 m elevation
9 43 48 116 97
120 43 48 119 98
900 43 48 120 98
____________________________________________________________
1 Total applied split into equal applications at planting,
end bloom and pod fill.
Table 3. Grain yield of soybean and common bean as affected
by elevation and N application.
_____________________________________________________________
Elevation (m)
N applied1 320 m 1050 m
Soybean Common bean Soybean Common bean
_____________________________________________________________
kg N ha-1 - - - kg ha-1 - - -
9 2801 2865 2369 2814
120 3317 2928 2601 2981
900 4260 3315 2307 3293
____________________________________________________________
Analysis of variance
Source df Probability of significance by F-test
Elevation (E) 1 0.003
Legume (L) 1 0.310
Nitrogen (N) 2 0.002
E x L 1 0.003
E x N 2 0.031
L x N 2 0.596
E x L x N 2 0.032
1 Total applied split into equal applications at planting,
end bloom and pod fill.
Table 4. Effect of elevation by legume species interaction
on harvest index and whole plant tissue N concentration at
physiological maturity.
___________________________________________________
Legume Elevation (m)
species 320 1050
___________________________________________________
Harvest index (HI)
Soybean 0.48 0.36
Common bean 0.49 0.45
Tissue N concentration (TNC)
- - g N kg-1 - -
Soybean 38 32
Common bean 26 25
Analysis of variance
Source df Probability of significance by F-test
HI TNC
Elevation (E) 1 0.002 <0.001
Legume (L) 1 0.004 <0.001
E x L 2 0.009 <0.001
Table 5. Main effects of elevation, legume species, and
N level on total dry weights at different growth stages.
Treatment Growth stage
variable End bloom1 Pod fill2 Maturity3
- - - - - - - kg ha-1 - - - - - - -
Elevation
m
320 2018 5087 6641
1050 1947 4463 6777
Legume
Soybean 1614 4925 6983
Common bean 2351 4626 6435
Nitrogen level4
kg N ha-1
9 1806 4533 6140
120 1997 4814 6674
900 2143 4980 7313
Analysis of variance
Source df Probability of significance by F-test
Elev. (E) 1 0.599 0.001 0.673
Leg. (L) 1 0.001 0.223 0.041
Nitr. (N) 2 0.015 0.016 0.002
1 End bloom (R2/R3) harvest was 35 and 51 days after
planting (DAP) for soybean and 41 and 55 DAP for
common bean at 320 and 1050 m sites, respectively.
2 Pod fill (R5/R6) harvest was at 58 and 76 DAP for
soybean and 62 and 77 DAP for common bean at 320 and
1050 m sites, respectively.
3 Harvest was at physiological maturity (R7).
4 Total applied split into equal applications at planting,
end bloom and pod fill.
Table 6. Main effects of elevation, legume species, and N levels on
mean crop growth rate during different phases.
Treatment Growth phase
variable Vegetative1 Early repr.2 Late repr.3 Crop cycle4
- - - - - g m-2 d-1 - - - - -
Elevation
m
320 5.2 13.9 8.8 8.5
1050 3.7 10.6 8.4 6.3
Legume
Soybean 3.8 13.9 6.6 7.2
Common bean 5.1 10.6 10.7 7.6
Nitrogen level5
kg N ha-1
9 4.1 12.0 7.3 6.9
120 4.5 12.3 8.5 7.3
900 4.8 12.4 10.1 8.0
Analysis of variance
Source df Probability of significance by F-test
Elev. (E) 1 0.006 0.001 0.758 0.001
Leg. (L) 1 0.004 0.014 0.003 0.152
Nitr. (N) 2 0.008 0.750 0.252 0.012
1 Vegetative phase is the period from planting to end bloom (R2/R3).
2 Early reproductive phase is the period from end bloom (R2/R3) to pod
fill (R5/R6).
3 Late reproductive phase is the period from pod fill (R5/R6) to
physiological maturity (R7).
4 Crop cycle is the period from planting to physiological maturity.
5 Total applied split into equal applications at planting, end bloom and pod fill.
Table 7. Effect of elevation by legume species interaction on leaf
characteristics at different growth stages.
___________________________________________________________________
Legume End bloom1 Pod fill2 Maturity3
320 m 1050 m 320 m 1050 m 320 m 1050 m
___________________________________________________________________
Leaf Area Index (LAI)
Soybean 2.3 3.6 3.9 5.3 1.8 2.5
Common bean 3.7 3.8 3.6 3.6 1.4 1.7
Leaf Weight Ratio (LWR)
Soybean 0.48 0.52 0.25 0.37 0.09 0.16
Common bean 0.42 0.49 0.21 0.24 0.08 0.08
Analysis of variance
Source df Probability of significance by F-test
End bloom Pod fill Maturity
LAI LWR LAI LWR LAI LWR
Elev. (E) 1 0.015 <0.001 <0.001 0.004 0.100 0.002
Legume (L) 1 0.037 <0.001 0.012 <0.001 0.045 0.002
E x L 1 0.070 <0.025 0.036 0.002 0.389 0.007
1 End bloom (R2/R3) harvest was 35 and 51 days after planting
(DAP) for soybean and 41 and 55 DAP for common bean at 320 and
1050 m sites, respectively.
2 Pod fill (R5/R6) harvest was at 58 and 76 DAP for soybean and 62
and 77 DAP for common bean at 320 and 1050 m sites, respectively.
3 Maturity harvest was at physiological maturity (R7).
Table 8. Effect of elevation and legume species on unit leaf rate
during different growth phases.
____________________________________________________________________
Legume Vegetative1 Early repr.2 Late repr.3
320 m 1050 m 320 m 1050 m 320 m 1050 m
____________________________________________________________________
- - - - - - - g m-2 d-1 - - - - - - -
Soybean 3.65 1.92 5.15 2.65 2.82 1.34
Common bean 3.33 2.10 3.25 2.60 3.85 4.37
Analysis of variance
Source df Probability of significance by F-test
End bloom Pod fill Maturity
Elev. (E) 1 <0.001 <0.001 0.229
Legume (L) 1 0.543 0.002 0.005
E x L 1 0.094 0.003 0.050
1 End bloom (R2/R3) harvest was 35 and 51 days after planting
(DAP) for soybean and 41 and 55 DAP for common bean at 320 and
1050 m sites, respectively.
2 Pod fill (R5/R6) harvest was at 58 and 76 DAP for soybean and
62 and 77 DAP for common bean at 320 and 1050 m sites,
respectively.
3 Maturity harvest was at physiological maturity (R7).
Chapter 6
NITROGEN ASSIMILATION AND N2 FIXATION IN SOYBEAN AND COMMON
BEAN AS AFFECTED BY ELEVATION AND MINERAL N AVAILABILITY
Abstract
The relationships between N assimilation, mineral N availability, and N2 fixation in soybean (Glycine max L.) and common bean (Phaseolus vulgaris L.) grown under two temperature regimes were investigated. Soybean and common bean were grown at elevations of 320 and 1050 m where mean soil/air temperatures during the experiment were 25/23 and 21/19 C, respectively. Ammonium sulfate with an excess of 15N-isotope was used to provide mineral N levels of 9, 120, and 900 kg N ha-1 . Nitrogen assimilation increased as applied mineral N increased for both species at both sites. With 900 kg N ha-1, N assimilation by both species increased by 25% compared to treatments receiving 9 kg N ha-1. With 900 kg N ha-1, N assimilation by soybean at the 320 m site was 320 kg N ha-1 which was 67% higher than the maximum N assimilation by common bean. Total N of soybean at the 1050 m site was 31% less than that at the 320 m site; total N of common bean was similar at the two sites. Total N assimilation at the 1050 m site by both species was reduced far less than the reduction in the rate of N assimilation, because crop duration was greatly extended. The relative increases in N assimilation rates with increased mineral N in both species were generally greater during vegetative and late reproductive growth phases compared to the early reproductive phase. Nitrogen assimilation was most rapid during the early reproductive phase of soybean while for common bean it was relatively uniform throughout growth. The early reproductive phase of soybean accounted for 56% of total plant N assimilation but only 24% of total crop duration. N2 fixation by soybean followed a pattern similar to that of N assimilation; the period of maximum N assimilation coincided with the period of peak N2 fixation. Common bean, relied relatively more on mineral N assimilation than soybean during the vegetative phase partly due to greater fertilizer N use efficiency. Common bean can grow better in low N soil than soybean, given the typical lag time in the development of the symbiotic N2 fixing system. Symbiotic N2 fixation may limit plant N assimilation in both legumes.
Introduction
The N requirement of legumes can be met by mineral N assimilation or symbiotic N2 fixation, or both. However, symbiotic N2 fixation is not a major source of N during the early and late phases of growth. Symbiotic N2 fixation begins only after nodule formation, which is preceded by the colonization of the rhizosphere and the infection of legume roots by rhizobia (Hardy et al., 1971; Vincent, 1974). Dinitrogen fixation generally reaches a peak at early pod fill and then declines during the late reproductive phase (Lawn and Brun, 1974a; Latimore et al., 1977; Thibodeau and Jaworski, 1975). Therefore, mineral N is a major source of N for legumes during both the early and late growth phases.
The interaction between genetic yield potential and environment determines the N requirement of a legume. The contributions from symbiosis and mineral N to total plant N are determined by the legume N requirement and the soil N availability in a given environment. When soil N uptake is less than the legume N requirement, N2 fixation is promoted. The N2 fixation potential is approximately equal to the aggregate of deficits in mineral N uptake per day over the legume life cycle. The daily N requirement of the legume is dependent on its growth rate and developmental stage. Therefore, N2 fixation also depends on the ontogeny of the legume, its efficiency of mineral N uptake, soil N availability and yield potential.
Temperature is a major environmental variable that regulates yield potential, plant N requirement and soil N availability. Temperature regulates crop growth rate and crop development and, therefore, the rate of N assimilation. Experiments conducted in 1985 (Chapter 3) indicated that the reduction in total plant N in soybean due to decreased rates of growth and N assimilation under low temperature was partially compensated for by an extended crop duration. Temperature also regulates N uptake directly (Rufty et al., 1981; Tolley and Raper, 1985). Since temperature indirectly regulates plant N requirement by regulating growth and directly regulates mineral N uptake, it also regulates the N2 fixation potential.
Mineral N application in sufficient amounts replaces N2 fixation (Allos and Bartholomew, 1959). However, N2 fixation compensates to a degree for decreased mineral N availability, and therefore, there is a paradox in interpreting the response of legumes to mineral N application. Significant yield and total plant N increases are not always obtained in legumes in response to N fertilization (Bhangoo and Albritton, 1976; Cassman et al., 1981; Diebert et al., 1979; Lawn and Brun, 1974b; Piha and Munns, 1987; Summerfield et al., 1977). It has been suggested that both mineral and fixed N are required for maximum yield and N assimilation in legumes (Bhangoo and Albritton, 1976; Franco et al., 1979; Harper, 1974). However, considering the high energy requirement for the development, maintenance and functioning of the symbiotic system in comparison to mineral N assimilation (Finke et al., 1982; Pate et al., 1979; Ryle et al., 1978 & 1979), the yield potential of legumes relying totally on N2 fixation may be limited. Soybean grown in solution culture supplied with mineral N assimilates substantially greater amounts of N than do nodulated plants (Jones et al., 1981; Lathwell and Evans, 1951; Wych and Rains, 1979). Similar results have not been reported from field experiments. It appears that legumes have a greater yield potential than can be supported by the symbiosis. Attaining the genetic yield potential may require maintaining an adequate fertilizer N regimen throughout growth.
Legumes differ substantially in their N requirements. While comparisons between soybean and common bean under identical growing conditions are rare (Piha and Munns, 1987), the data for soybean (Chapter 4; Cassman et al., 1981; Herridge et al., 1984;) and common bean (Rennie and Kemp, 1983; Westerman et al., 1981) show that soybean has a higher N requirement than common bean. The higher N requirement of soybean is mainly due to a greater seed N concentration than common bean rather than greater dry matter accumulation or seed yield. Because of the contrasting N requirements of these two legumes, soil N availability that is sufficient to meet the N requirement of common bean may be inadequate to meet the soybean N requirement without significant N2 fixation. Accordingly, soybean meets a major portion of its N requirement through symbiosis in soils of low to moderate N availability (Chapter 4) while many reports indicate very little or no N2 fixation in common bean (El-Nadi et al., 1972; Graham, 1981) except under very low soil N conditions (Piha and Munns, 1987; Rennie and Kemp, 1983; Westerman et al., 1981).
It is clear that in order to maximize the N derived from fixation in legumes, it is important to understand the patterns of N assimilation for the two crops and how those patterns may be influenced by the interactions between environment, legume N requirement, and soil N availability. The contrasting N requirements and N2 fixation potentials of soybean and common bean provide an opportunity to identify N assimilation traits that are useful in plant breeding programs aimed at increasing N2 fixation. In the present experiment, the effects of elevation (temperature) and N regimes on the relationships between yield potential, N requirement, soil N uptake, and N2 fixation by soybean and common bean were studied.
Materials and Methods
Common bean (Phaseolus vulgaris L.) and soybean (Glycine max (L.) Merrill) were grown with and without inoculation at elevations of 320 and 1050 m on the island of Maui with applied nitrogen levels of 9, 120 and 900 kg N ha-1 as ammonium sulfate. The experimental design, site characteristics, field and plant culture, and sampling procedures for determining plant dry matter yield were described in detail previously in Chapter 5. Only those procedures which were specific to this investigation are described below.
N and 15N application: The total N applied was split into thirds with one-third applied just after planting, one-third at end bloom (R2/R3), and one-third at pod fill (R5/R6) stages. At each stage, aqueous solutions of 15N enriched ammonium sulfate were sprayed on 1.35 x 3.0 m microplots within each 9.5 x 3.0 m whole plot. The aqueous ammonium sulfate was enriched to provide 30 mg 15N m-2 in each microplot at each application. Aqueous solutions of natural 15N abundance ammonium sulfate was sprayed on the remaining area of the whole plots.
Plant sampling: At growth stages end bloom, pod fill and physiological maturity (R7), 10 to 15 plants were harvested from the center three rows of the five-row microplots by cutting them at the soil surface. Whole-plant fresh and dry weights were determined on the microplot subsamples. The plants were dried to a constant weight at 60 C. The dried microplot subsamples were ground and digested in H2SO4 with 1% (w:v) salicylic acid to convert tissue N, including nitrate, into ammonium. Ammonium-N in digest subsamples was measured by the indophenol blue method (Keeney and Nelson, 1982). Kjeldahl digests were steam distilled with 13 N NaOH and the distillates were collected in 0.02 N H2SO4. The distillates were then acidified to a pH of 4.5 to 5.0 and concentrated by evaporating on a hot plate under medium heat for mass spectroscopic analysis of 15N (Bergersen, 1980).
Sampling and extraction of soil N: Soil samples from the 15N microplots were collected to a depth of 20 cm (4 samples per microplot) at 5 days after planting, end bloom, pod fill and physiological maturity. Extractable soil NH3-N and NO3-N were determined as described in Chapter 5.
15N calculations: The amount of plant N derived from the atmosphere and fertilizer and the fertilizer use efficiency were calculated by equations developed for 15N dilution techniques (International Atomic Energy Agency, 1983). The atom% 15N excess was calculated with reference to the natural 15N abundance of air (0.3663 atom% 15N). The non-nodulating isoline served as the non-N2-fixing reference plant for soybean. N2 fixation could not be determined for common bean because some nodulation was observed in the uninoculated controls (see Appendix Table A-1). The non-nodulating isoline of soybean was judged to be an unsatisfactory reference crop for common bean because of differences between the two legumes in total mineral N uptake and the pattern of N assimilation.
Statistical analysis: The data from the inoculated segment of the experiment were subjected to analysis of variance.
Results and Discussion
The mean air temperatures during the experiment were 24 and 19 C at the 320 and 1050 m elevations, respectively (Chapter 5). The extractable N (NH4+ and NO3-) at planting was 0.05 and 0.10 g kg-1 soil at the 320 and 1050 m sites, respectively (Chapter 5).
In the present experiment, total N assimilation for soybean decreased significantly with increased elevation while N assimilation of common bean was relatively unaffected (Table 1). Nitrogen assimilation by both legumes increased significantly with increasing applied mineral N. The results indicate that up to 900 kg N ha-1, N assimilation by both soybean and common bean was limited by mineral N availability. Nitrogen assimilation for both legumes increased by 25% for plants fertilized with 900 kg N ha-1 compared to plants receiving only 9 kg N ha-1. Although the elevation by N interaction was not significant, the increase in total N of soybean in response to mineral N application was less pronounced at the 1050 m site. The data demonstrate that maximum N assimilation by soybean and common bean can only be realized when sufficient mineral N is applied. Reports on total plant N increases in the field due to mineral N applications have been inconsistent and the total N of non-N2 fixing plants has seldom exceeded that of an N2-fixing counterpart (Bhangoo and Albritton, 1976; Cassman et al., 1981; Diebert et al., 1979; Harper, 1974; Lawn and Brun, 1974b; Piha and Munns, 1987; Weber, 1966; Westerman et al., 1981). The total amount of mineral N applied or the application regimen used in other experiments may have been inadequate to maintain sufficient N in the soil solution to meet plant N assimilation potential. High levels of N in the root zone may increase crop yield by permitting a plant to partition relatively more dry matter to shoots than to roots (Cassman et al., 1980; Tolley-Henry and Raper 1986; Vessey and Layzell, 1987) compared to plants that rely on symbiotic N2 fixation. In this study, N2 fixation by soybean declined with N application, but was still appreciable at 900 kg N ha-1 at the warm site (Table 2). It appears that, unless fertilizer use efficiency can be improved, more than 900 kg ha-1 mineral N needs to be applied to completely suppress N2 fixation and obtain maximum N assimilation by soybean under warm growing conditions.
Mineral N application increased the total N assimilated by increasing the average rate of N assimilation by both species (Table 3). The average rate of N assimilation over the duration of the crop was 25% higher at 900 kg N ha-1 than it was at 9 kg N ha-1; however, the same response was not observed at all growth phases. The increase in the rate of N assimilation with increasing N applied was generally greatest during the vegetative and late reproductive phases. The rate of N assimilation did not increase with applied N during the early reproductive phase except for common bean at the 320 m site. Since the N2 fixation by soybean was limited during the vegetative and the late reproductive phases (Table 2), mineral N availability during the early and late growth periods is critical to attain maximum N assimilation.
Low temperature reduces rate of growth and N assimilation, but this effect can be compensated for by extended crop duration. The results indicate that the rate of N assimilation at the 1050 m site was reduced by as much as 53% in soybean and 24% in common bean compared to the 320 m site (Table 3). Yet, no reduction in total N assimilation was observed at the 1050 m site in common bean and the reduction in soybean was only 31% compared to the 320 m site (Table 1). Therefore, total N assimilated over the crop cycle was less affected by low temperature than the rate at which N was assimilated.
The relative N assimilation in relation to the relative duration of different growth phases of soybean and common bean is displayed in Table 4. Soybean and common bean were found to have distinct N assimilation patterns over time. On average, 50% of the total N assimilated by common bean occurred during vegetative growth, the period of which accounted for 56% of its total crop duration, indicating that growth and N assimilation were proportional. The above relationship between N assimilation and growth phase duration was continued up to physiological maturity of common bean. In contrast, only 28% of the total N was assimilated by soybean during its vegetative phase, a period that accounted for 44% of its total crop duration. The slow rate of N assimilation during the vegetative phase of soybean was followed by a period of rapid N assimilation in the early reproductive phase when 56% of the total N was assimilated in 24% of its total crop duration. The differences in the pattern of N assimilation between soybean and common bean in the present experiment generally prevailed across sites, however, the patterns were less pronounced at the 1050 m site. The pattern of N assimilation observed in the present experiment may be characteristic to these legumes.
The relationship between growth phase and N assimilation for these legumes seems to influence their N2 fixation potentials. The relative contributions of symbiotic N2 fixation and mineral N assimilation to total plant N are determined by the N requirement of the legume, the soil N availability and the efficiency of mineral N uptake. Under a given level of soil N availability, the relative importance of the two modes of N nutrition is determined by the N assimilation characteristics of the legume. The lower per day N requirement of common bean may limit N2 fixation compared to soybean under conditions of equal mineral N availability. The relationship between the time course of N assimilation by the legume and the ontogeny of the symbiotic N2 fixing system may also have an influence on the amount of N2 fixed. The very rapid N assimilation during the relatively short early reproductive phase of soybean was also the time when N2 fixation reached a maximum (Table 2). Additionally, isotope analysis indicated that the early N requirement of common bean was mostly met by soil N assimilation. The fertilizer N use efficiency of common bean was about two times higher than soybean at end bloom, but was similar to soybean both at pod fill and physiological maturity (Table 5). Consistent with the higher fertilizer N use efficiency of common bean during vegetative growth, the extractable soil N was significantly lower in plots of common bean than soybean at end bloom (Table 6). The extractable soil N in plots of both legumes were similar at pod fill, but was lower under soybean than common bean at physiological maturity. Therefore, common bean had a higher reliance on mineral N uptake than soybean during vegetative growth and utilized proportionately more mineral N throughout growth. The higher mineral N uptake efficiency of common bean during vegetative growth when the symbiosis is still establishing gives the plant an advantage over soybean under low soil N.
Conclusions: Attaining maximum N assimilation by common bean and soybean in the field requires greater mineral N levels applied at more frequent intervals than has been a common practice in other experiments. The data presented here show that significant N2 fixation limits total N assimilation by soybean. The pattern of N assimilation in soybean was characterized by low N assimilation during vegetative growth, followed by rapid assimilation during the early reproductive phase. Nitrogen assimilation by common bean was relatively uniform throughout the growth cycle. Nitrogen assimilation by soybean followed a pattern similar to the ontogeny of the symbiotic N2-fixing system; the period of maximum N assimilation coincided with the period of peak N2 fixation. Common bean, instead relies more on mineral N assimilation due to higher mineral N use efficiency than soybean, especially during the vegetative phase. Maximizing N derived from fixation in common bean may require altering its N assimilation characteristics.
Table 1. Elevation by legume by N level interaction on N
assimilation at physiological maturity.
______________________________________________________________
N applied Elevation (m)
320 1050
Soybean Common bean Soybean Common bean
______________________________________________________________
(kg ha-1)1 - - - - - kg N ha-1 - - - - -
9 241 142 209 149
120 252 147 229 170
900 320 191 220 192
______________________________________________________________
Analysis of variance
Source df Probability of significance by F-test
______________________________________________________________
Elevation (E) 1 0.067
Legume (L) 1 <0.001
Nitrogen (N) 2 <0.001
E x L 1 0.010
E x N 2 0.011
L x N 2 0.983
E x L x N 2 <0.083
1 Total applied split into equal applications at planting, end
bloom and pod fill.
Table 2. Effect of elevation and N level and growth stage on
N2 fixation1 by soybean at different growth stages.
_____________________________________________________________
N applied Growth stage
End bloom Pod fill Maturity
_____________________________________________________________
- - -kg N ha-1- - -
(kg ha-1)2 320 m elevation
9 37 161 168
120 25 117 109
900 20 29 41
1050 m elevation
9 13 70 67
120 9 55 49
900 0 0 0
Analysis of variance
Source df Probability of significance by F-test
Elevation (E) 1 0.003 0.001 0.007
Nitrogen (N) 2 0.124 0.001 0.003
E x N 2 0.685 0.307 0.474
_____________________________________________________________
1 Determined by 15N-dilution using non-nodulating isoline of
soybean as reference.
2 Total applied split into equal applications at planting, end
bloom and pod fill.
Table 3. Effect of elevation, N application and legume species on the rate of N assimilation during crop cycle.
__________________________________________________________
N applied Elevation (m)
320 1050
Soybean Common bean Soybean Common bean
(kg ha-1)1 - - - - g m-2 d-1 - - - -
Vegetative phase (Veg.)2
9 0.17 0.19 0.11 0.12
120 0.19 0.20 0.14 0.12
900 0.22 0.28 0.15 0.16
Early reproductive phase (ER)3
9 0.67 0.22 0.50 0.28
120 0.64 0.20 0.50 0.38
900 0.68 0.28 0.49 0.29
Late reproductive phase (LR)4
9 0.15 0.16 0.07 0.11
120 0.16 0.20 0.08 0.10
900 0.34 0.15 0.04 0.20
Crop Cycle (CC)5
9 0.31 0.20 0.18 0.15
120 0.30 0.20 0.19 0.17
900 0.38 0.25 0.18 0.19
__________________________________________________________
Analysis of variance
Source df Probability of significance by F-test
Veg. ER LR CC
Elevation (E) 1 0.003 0.288 <0.001 <0.001
Legume (L) 1 0.098 <0.001 0.568 0.001
Nitrogen (N) 2 <0.001 0.879 0.040 <0.001
E x L 1 0.085 0.008 0.027 0.003
E x N 2 0.219 0.206 0.409 0.010
L x N 2 0.268 0.674 0.531 0.781
E x L x N 2 0.645 0.632 0.002 0.211
1 Total applied split into equal applications at planting,
end bloom and pod fill.
2 Vegetative phase is the period from planting to end
bloom.
3 Early reproductive phase is the period from end bloom to
pod fill.
4 Late reproductive phase is the period from pod fill to
physiological maturity.
5 Crop cycle is the period from planting to physiological
maturity.
Table 4. Relative N assimilation1 during different growth
phases of soybean and common bean grown at two elevations.
_____________________________________________________________
Growth Elevation (m)
phase 320 1050
Soybean Common bean Soybean Common bean
_____________________________________________________________
- - - - - - - - - % - - - - - - - - -
Vegetative 25 58 31 42
(43)2 (56) (44) (56)
Early repr. 56 30 57 41
(28) (28) (21) (22)
Late repr. 19 12 12 17
(29) (16) (35) (22)
_____________________________________________________________
Analysis of variance
Source df Probability of significance by F-test
Vegetative Early repr. Late repr.
Elevation (E) 1 0.157 0.120 0.297
(0.055) (<0.001) (<0.001)
Legume (L) 1 <0.001 <0.001 0.654
(<0.001) (<0.001) (<0.001)
E * L 1 0.003 0.039 0.052
(0.249) (0.003) (0.375)
_____________________________________________________________
1 Amount of N assimilated during the growth phase expressed
as a percentage of total N at physiological maturity.
2 Value in parenthesis is growth phase duration expressed as a
percentage of total crop duration.
Table 5. Effect of legume and N levels on fertilizer N use efficiency1 during the crop cycle.
_____________________________________________________________
N applied Growth stage
End bloom Pod fill Maturity
Soybean Bean Soybean Bean Soybean Bean
_____________________________________________________________
(kg ha-1)2 - - - - - - - % - - - - - - -
9 7.1 14.8 31.0 34.5 35.4 37.1
120 10.4 16.0 34.3 33.0 42.7 40.1
900 10.1 18.2 19.4 17.4 18.4 13.8
Analysis of variance
Source df Probability of significance by F-test
Legume (L) 1 0.001 0.971 0.437
Nitrogen (N) 2 0.033 <0.001 <0.001
L x N 2 0.510 0.128 0.355
_________________________________________________________________
1 Fertilizer N use efficiency is determined by using 15N isotope
as tracer and is the percent of applied fertilizer nitrogen
assimilated by the plant.
2 Total applied split into equal applications at planting, end
bloom and pod fill.
Table 6. Effect of legume and N levels on KCl-extractable
soil N during the crop cycle.
_____________________________________________________________
N applied Growth stage
End bloom Pod fill Maturity
Soybean Bean Soybean Bean Soybean Bean
_____________________________________________________________
(kg ha-1)1 g N kg-1 soil
9 0.029 0.017 0.019 0.014 0.016 0.016
120 0.026 0.025 0.023 0.022 0.015 0.019
900 0.101 0.075 0.155 0.140 0.173 0.215
Analysis of variance
Source df Probability of significance by F-test
Legume (L) 1 0.001 0.971 0.437
Nitrogen (N) 2 <0.001 <0.001 <0.001
L x N 2 0.034 0.290 <0.001
_____________________________________________________________
1 Total applied split into equal applications at planting,
end bloom and pod fill.
SUMMARY AND CONCLUSIONS
Experiments were conducted to study the growth and yield responses of soybean (Glycine max L. Merrill) and common bean (Phaseolus vulgaris L.) to mode of nitrogen nutrition and temperature. Three sites in an elevational transect (320, 660, 1050 m) differing in mean annual temperatures were used to conduct two experiments.
In the first experiment, five varieties of G. max belonging to four maturity groups (00, IV, VI and VIII) were planted at the three sites to determine i) the effect of temperature, soil type and host genotype on nodulation by a mixed inoculum composed of three serologically distinct Bradyrhizobium japonicum strains; ii) the phenology of soybean as affected by temperature, maturity group and nitrogen nutritional status; and iii) the effect of elevation (site) and maturity group on the proportion of nitrogen derived from N2 fixation. The mean soil/air temperatures (C) during the experimental period were 24/23, 23/20, and 20/18 at 320, 660, and 1050 m elevations, respectively.
In a second experiment, inoculated and uninoculated soybean and common bean were grown with either 9, 120 or 900 kg N ha-1 at elevations of 320 and 1050 m. The objectives were i) to evaluate the growth responses of G. max and P. vulgaris to cool and warm temperatures under varying levels of nitrogen nutrition; and ii) to contrast the nitrogen assimilation patterns of G. max and P. vulgaris. The mean soil/air temperatures (C) during this experiment were 25/23 and 21/19 at 320 and 1050 m elevations, respectively.
In the first experiment, differences in soil, temperature, and soybean growth between sites had little effect on the pattern of nodule occupancy by B. japonicum strains USDA 110, 136b and 138 inoculated in a mixture. USDA 110 occupied 81% of nodules and plant growth parameters were unrelated to differences in nodule occupancy by the strains. Average dry matter and nitrogen accumulation at the highest site were only 48 and 41% respectively of that obtained at the lowest site. Nitrogen fixation contributed 80, 66, and 97% of total plant nitrogen at the lowest, intermediate and highest sites, respectively. Although total nitrogen assimilation between varieties differed in some cases by 400%, the proportions of nitrogen derived from fixation were similar within a site. Soybean maturity was delayed with increasing elevation mainly due to extended vegetative growth periods averaging 23 and 64% longer at the 660 and 1050 m sites compared to the lowest site. The effect of elevation on the duration of the seed-fill phase was minimal, but the rate of seed dry matter accumulation (kg ha-1 d-1) decreased with increasing elevation. Non-nodulating isolines matured earlier than their nodulating counterparts due to nitrogen insufficiency.
In the second experiment, the number of days to flowering and maturity for both soybean and common bean were delayed at 1050 m compared to the 320 m elevation; low temperature had a relatively greater effect on soybean than common bean. Average crop growth rate (kg biomass ha-1 d-1) decreased by 26% at the 1050 m site compared to the 320 m site, but each legume attained similar dry weights among the sites due to prolonged duration of growth at the 1050 m site. Grain yield of soybean was 30% lower at the 1050 m site due to reduced harvest index. In contrast to soybean, the grain yield of common bean was similar between elevations. Application of 900 kg N ha-1 increased the grain yield an average of 22% primarily by increasing the crop growth rate and total dry matter production. Common bean accumulated 46% more dry weight than soybean during vegetative growth due to a higher crop growth rate and a longer vegetative duration than soybean. Average crop growth rate for the entire growth period for the two legumes was similar; the total dry matter of soybean was 9% greater than common bean because of longer crop duration. Soybean had increased leaf production (leaf area and leaf weight ratio) and substantially reduced unit leaf rate (g m-2 d-1) at the high elevation compared to common bean. Nitrogen application increased the leaf area and crop growth rate of both legumes. With 900 kg N ha-1, N assimilation by both legumes increased by 25% compared to treatments receiving 9 kg N ha-1. With 900 kg N ha-1, N assimilation by soybean at the 320 m site was 320 kg N ha-1 which was 67% higher than the maximum N assimilation by common bean. Total N of soybean at the 1050 m site was 31% less than that at the 320 m site; total N of common bean did not differ between the two sites. Because crop duration of both species was greatly extended at the high, cool site, total N assimilation was reduced far less than the rate of N assimilation. Nitrogen assimilation was more rapid during the early reproductive phase of soybean while for common bean N assimilation was relatively uniform throughout growth. Nitrogen fixation by soybean followed a pattern similar to that of N assimilation; the period of maximum N assimilation coincided with the period of peak N2 fixation. Common bean relied relatively more on mineral N due to greater fertilizer N use efficiency than soybean, especially during the vegetative phase.
The results of the two experiments indicated that: i) rhizobial strains superior in competitiveness may be identified for a broad range of host genotypes and environments; ii) the proportion of nitrogen derived from symbiotic nitrogen fixation by soybean was approximately constant at a site and therefore the contribution of the symbiosis to a crop's nitrogen requirement may be predicted after characterizing soil nitrogen availability at a site; iii) low temperature prolonged the vegetative and total growth of soybean and common bean; iv) maturity of soybean was dependent on the nitrogen nutritional status of the crop and nitrogen stress hastened maturity; v) extended growth duration and increased leaf production were two primary responses which compensated for reduced rate of growth of soybean and common bean at the higher elevation; vi) growth and yield of common bean were more stable across a wider range of temperatures than was the case for soybean; vii) maximum N assimilation by soybean and common bean does not occur if there is significant N2 fixation; and viii) nitrogen derived from N2 fixation by common bean probably can only be maximized by genetic manipulation of its N assimilation characteristics.
APPENDIX
The following tables are part of the experiment described in Chapters 5 and 6. They were deemed to be not of sufficient importance to be incorporated into the body of the dissertation.
Table A-1. Effect of elevation and N level on nodule dry weight at end bloom (R2/R3).
__________________________________________________________
N applied1 Soybean Common bean
Uninoculated Inoculated
__________________________________________________________
kg N ha-1 - - - - - - - g m-2 - - - - - - -
320m elevation
3 8.6 4.6 6.9
40 6.7 2.7 5.7
300 3.3 0.5 0.5
1050m elevation
3 3.9 0.6 3.8
40 2.6 0.8 2.4
300 1.4 0.6 1.0
_________________________________________________________
1 N applied at planting.
Table
A-2. Days to flowering (R1) and
physiological maturity (R7) of the non-nodulating isoline of
soybean grown at two elevations with three levels of
mineral N.
____________________________________________________
Nitrogen
applied1 R1 R7
____________________________________________________
kg N ha-1 - - - -days- - - - -
320 m elevation
9 28 74
120 28 79
900 28 82
1050 m elevation
9 43 110
120 43 118
900 43 120
___________________________________________________
1 Total N applied was split into equal applications
at planting, end bloom, and pod fill.
Table A-3. Effect of N level on the harvest index
of legumes grown at two elevations.
___________________________________________________
N applied1 Soybean Common bean
Nod Non-nod
___________________________________________________
kg N ha-1 320m elevation
9 0.47 0.34 0.51
120 0.48 0.43 0.47
900 0.50 0.50 0.50
1050m elevation
9 0.36 0.32 0.45
120 0.38 0.36 0.45
900 0.33 0.28 0.45
__________________________________________________
1 Total N applied was split into equal applications
at planting, end bloom, and pod fill.
Table
A-4. Grain yield of the non-nodulating
isoline of
of mineral N.
______________________________________________________
Elevation (m)
N applied1 320 m 1050 m
______________________________________________________
kg N ha-1 - - - kg ha-1 - - -
9 1316 2030
120 2387 2464
900 4163 2074
______________________________________________________
1 Total N applied was split into equal applications at
planting, end bloom and pod fill.
Table
A-5. Total N assimilation by the
non-nodulating isoline of
three levels of mineral N.
____________________________________________________
Elevation (m)
N applied1 320 m 1050 m
____________________________________________________
kg N ha-1 - - - kg ha-1 - - -
9 84 165
120 152 203
900 299 232
____________________________________________________
1 Total N applied was split into equal applications
at planting, end bloom and pod fill.
Table A-6. Effect of N level on the leaf area index (LAI) and leaf weight ratio (LWR) of legumes at different growth stages.
____________________________________________________________
N applied1 End bloom2 Pod fill3 Maturity4
LAI LWR LAI LWR LAI LWR
____________________________________________________________
kg N ha-1 - - - - - - - kg ha-1 - - - - - - - - - -
9 3.1 0.49 3.8 0.26 1.9 0.11
120 3.4 0.47 4.2 0.26 2.0 0.11
900 3.6 0.47 4.2 0.28 1.7 0.09
____________________________________________________________
1 Total N applied was split into equal applications at
planting, end bloom, and pod fill.
2 End bloom (R2/R3) harvest was 35 and 51 days after
planting (DAP) for soybean and 41 and 55 DAP for common
bean at 320 and 1050m sites, respectively.
3 Pod fill (R5/R6) harvest was at 58 and 76 DAP for soybean
and 62 and 77 DAP for common bean at 320 and 1050m sites,
respectively.
4 Maturity harvest was at physiological maturity (R7).
Table A-7. Effect of N level on the unit leaf rate of legumes at different growth stages.
____________________________________________________________
N applied1 End bloom2 Pod fill3 Maturity4
____________________________________________________________
kg N ha-1 - - - - - - - g m-2 d-1 - - - - - - -
9 2.72 3.55 2.73
120 2.77 3.38 2.97
900 2.76 3.31 3.60
____________________________________________________________
1 Total N applied was split into equal applications at
planting, end bloom, and pod fill.
2 End bloom (R2/R3) harvest was 35 and 51 days after
planting (DAP) for soybean and 41 and 55 DAP for common
bean at 320 and 1050m sites, respectively.
3 Pod fill (R5/R6) harvest was at 58 and 76 DAP for soybean
and 62 and 77 DAP for common bean at 320 and 1050m sites,
respectively.
4 Maturity harvest was at physiological maturity (R7).
Table A-8. Fertilizer use efficiency of legumes grown
at two elevations with three N levels.
_____________________________________________________
N applied1 Soybean Common bean
320m 1050m 320m 1050m
_____________________________________________________
kg N ha-1 - - - - - - % - - - - - - -
9 26 47 25 48
120 37 49 31 49
900 21 16 15 13
_____________________________________________________
1 Total N applied was split into equal applications
at planting, end bloom, and pod fill.
Table A-9. KCl-extractable soil N after harvest of legumes grown at two elevations with three N levels.
____________________________________________________________
N applied1 Soybean Common bean
320m 1050m 320m 1050m
____________________________________________________________
kg N ha-1 - - - - - g N kg-1 soil - - - - -
9 0.015 0.016 0.016 0.016
120 0.015 0.015 0.013 0.024
900 0.150 0.200 0.179 0.251
____________________________________________________________
1 Total N applied was split into equal applications at
planting, end bloom, and pod fill.
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