Differential Responses of Soybean and Sorghum ... - ACS Publications

Subscriber access provided by University of Sussex Library

Article

Differential responses of soybean and sorghum growth, nitrogen uptake and microbial metabolism in the rhizosphere to cattle manure application: A rhizobox study Qingnan Chu, Zhimin Sha, Takuji Nakamura, Norikuni Oka, Mitsuru Osaki, and Toshihiro Watanabe J. Agric. Food Chem., Just Accepted Manuscript • DOI: 10.1021/acs.jafc.6b03046 • Publication Date (Web): 11 Oct 2016 Downloaded from http://pubs.acs.org on October 20, 2016

Just Accepted “Just Accepted” manuscripts have been peer-reviewed and accepted for publication. They are posted online prior to technical editing, formatting for publication and author proofing. The American Chemical Society provides “Just Accepted” as a free service to the research community to expedite the dissemination of scientific material as soon as possible after acceptance. “Just Accepted” manuscripts appear in full in PDF format accompanied by an HTML abstract. “Just Accepted” manuscripts have been fully peer reviewed, but should not be considered the official version of record. They are accessible to all readers and citable by the Digital Object Identifier (DOI®). “Just Accepted” is an optional service offered to authors. Therefore, the “Just Accepted” Web site may not include all articles that will be published in the journal. After a manuscript is technically edited and formatted, it will be removed from the “Just Accepted” Web site and published as an ASAP article. Note that technical editing may introduce minor changes to the manuscript text and/or graphics which could affect content, and all legal disclaimers and ethical guidelines that apply to the journal pertain. ACS cannot be held responsible for errors or consequences arising from the use of information contained in these “Just Accepted” manuscripts.

Journal of Agricultural and Food Chemistry is published by the American Chemical Society. 1155 Sixteenth Street N.W., Washington, DC 20036 Published by American Chemical Society. Copyright © American Chemical Society. However, no copyright claim is made to original U.S. Government works, or works produced by employees of any Commonwealth realm Crown government in the course of their duties.

Page 1 of 38

Journal of Agricultural and Food Chemistry

Differential responses of soybean and sorghum growth, nitrogen uptake and microbial metabolism in the rhizosphere to cattle manure application: A rhizobox study Qingnan Chu,† Zhimin Sha,‡ Takuji Nakamura,§Norikuni Oka,§Mitsuru Osaki,† Toshihiro Watanabe*,† Author information †

Graduate School of Agriculture, Hokkaido University, Sapporo 060-8589, Japan

‡

School of Agriculture and Biology, Shanghai Jiaotong University, 200240, Shanghai,

China §

NARO Hokkaido Agricultural Research Center, Sapporo 062-8555, Japan

Information for corresponding author *Toshihiro Watanabe, Graduate School of Agriculture, Hokkaido University, Sapporo 060-8589, Japan Tel: +81 11 706 2498; Fax: +81 11 706 2498 E-mail: [email protected]

1

ACS Paragon Plus Environment


Page 2 of 38

1

Abstract

2

In this study, we determined the capacity of soybean (Glycine max L. Merr. cv.

3

Hoyoharuka) and sorghum (Sorghum bicolor L. Moench. cv. Hybrid Sorgo) to utilize

4

different forms of nitrogen (N) in a rhizobox system. Seedlings were grown for 35

5

days without N, or with 130 mg N kg-1 soil as ammonium sulfate or farmyard cattle

6

manure. The soil fractions at different distances from the root were sliced millimeter

7

by millimeter in the rhizobox system. We assessed the distribution of different forms

8

of N and microbial metabolism in different soil fractions in the rhizosphere. There are

9

no treatment-dependent changes in biomass production in the roots and shoots of

10

soybeans, however, the ammonium and manure treatment yielded 1.30 and 1.40 times

11

higher shoot biomass of sorghum than the control. Moreover, the depletion of

12

inorganic N and total amino acids (TAA) in the rhizosphere was largely undetectable

13

at various distances from the soybean roots, regardless of the treatments employed.

14

The addition of ammonium sulfate resulted in a decrease in the nitrate concentration

15

gradient as the distance decreased from the sorghum roots. The addition of manure to

16

the soil increased the N content in the sorghum shoots, 1.57 times higher than the

17

control; this increase was negatively correlated with the concentrations of TAA in the

18

soil of the root compartment. In addition, the application of manure simultaneously

19

induced TAA depletion (i.e. the TAA concentration in root compartment was 1.48

20

times higher than that in bulk soil) and greater microbial activity and diversity in the

21

sorghum rhizosphere, where higher microbial consumption of asparagine, glutamic

2


Page 3 of 38


22

acid, and phenylalanine were also observed near the roots. Our results are first to

23

present the evidence that sorghum may possess a high capacity for taking up amino

24

acids as a consequence of organic matter application and microbial metabolism.

25

Keywords: amino acid, microbial metabolism, nitrogen, rhizobox, rhizosphere,

26

sorghum

27

Introduction

28

N is indispensible for sustaining human activities through its role in the production of

29

food, animal feed, and synthetic chemicals.1 Depending on food demand, the

30

projected consumption of N fertilizers is likely to increase from 105 Mt in 2010 to

31

180 Mt by 2050.2 Thus, the effective utilization of organic materials as N source for

32

crop production is important for the development of sustainable agriculture. The

33

actual pool sizes of organic forms of N can be large in agricultural soils,3 but higher

34

plants generally cannot efficiently assimilate the abundant sources of N bound up in

35

the soil.4 Fully comprehending the different capacities of crops to assimilate organic

36

N is important for advancing crop productivity and sustainable agriculture.

37

It has been well documented that the responses to organic matter in terms of growth

38

and N uptake differ among plant species.5-9 Some plants possess the ability to acquire

39

and metabolize organic N (ON) efficiently in the form of amino acids,10-11

40

peptides,12-13 and proteins.6 Once applied to the soil, organic matters undergo stepwise

41

mineralization, including proteolysis, ammonification, and nitrification, where

3



Page 4 of 38

42

proteinaceous compounds are converted into amino acids, ammonium (NH4+), and

43

nitrate (NO3–) by soil bacteria and fungi. Among the primary microbial extracellular

44

proteases in soils, alkaline metalloprotease, neutral metalloprotease, and serine

45

protease are encoded by the apr, npr and sub genes, respectively.14-15 The interactions

46

between plant roots and microbes in the rhizosphere with applied organic fertilizer

47

have been investigated extensively because of their importance for nutrient

48

management and soil carbon (C) dynamics.16-20 Microorganisms are less limited by

49

the ON pool than plants because they are able to release enzymes to utilize

50

macromolecular soil organic N.21 Most plant roots provide soil microbes with C

51

nutrition, while the microorganisms can decompose soil ON and make N available to

52

the roots.18,

53

increase the rates of N mineralization and bacterial numbers, as well as change the

54

microbial-root interactions to accelerate the flux of N from organic sources into the

55

plant-available N pool.18-19 However, the microbial factors that affect its capability to

56

use ON, as well as the forms utilized have not been well elucidated.

57

Among the cereal crops, the sorghum has been demonstrated to have the capacity to

58

utilize more ON than maize and pearl millet in soil or hydroponic growing

59

conditions.5,

60

capability to absorb proteinaceous compounds but that amino acids may be a more

61

readily utilizable source of N.5 In a previous study, we showed that soybean do not

62

efficiently utilize ON,8 and the N uptake by soybean did not increase significantly

22

The microbial responses to the addition of organic matter likely

23-24

Okamoto and Okada 2004 suggested that sorghum have the

4


Page 5 of 38


63

with the farmyard cattle manure application.25 Consequently, soybeans can be used as

64

a comparative plant in studies of ON utilization compared with sorghum. Moreover,

65

the capacity of symbiotic nitrogen fixation by soybean and the difference of sink N

66

demand between C3 plant (soybean) and C4 plant (sorghum) may cause contrasting N

67

utilization. Therefore, in the current study, we aim to investigate the difference of N

68

uptake and utilization between soybean and sorghum receiving ON application, which

69

is quite important for nitrogen management in the agricultural production.

70

Previously, Youssef and Chino (1988) established a rhizobox system to study the

71

distribution of nutrients in the different soil fractions of the rhizosphere,26 and this

72

rhizobox system has been used extensively to map the distribution of N in different

73

rhizospheric soil fractions.18-19,

74

distribution of N and microbial metabolism has not been determined in a rhizobox

75

study. Therefore, we designed a rhizobox experiment to examine the distribution of

76

inorganic N, amino acids, and the microbial metabolism in the rhizospheric soil of

77

soybean and sorghum after the addition of inorganic and organic N fertilizers. We

78

analyzed the interactions between plant N uptake, the microbial utilization of organic

79

compounds, and the distribution of inorganic N and amino acids in different

80

rhizospheric soil fractions.

81

Materials and Methods

82

Experimental soil Soil classified as Haplic Fluvisols according to the US Soil

27

However, the spatial relationship between the

5



Page 6 of 38

83

Taxonomy Classification was collected from agricultural fields at Hokkaido

84

University. The basic properties of the experimental soil were: pH 6.05, total nitrogen

85

4.5 g kg–1, ammonium-N 4.05 mg kg–1, nitrate-N 13.4 mg kg–1, available P (Truog-P)

86

194 mg kg–1, exchangeable K 0.36 cmol kg–1, cation exchange capacity 37.5 cmol kg-1,

87

and base saturation 30%.

88

Experimental design All experiments were performed in a greenhouse. Rhizobox

89

culture chambers (20 × 10 × 15 cm, Fig. 1) were used to separate the root zone soil

90

from the rhizospheric and bulk soils away from the root zone, according to the

91

method described by Youssef and Chino (1988) and Li et al. (2007).26-27 The rhizobox

92

contained two types of compartments: a root compartment (2 × 10 × 15 cm, Fig. 1),

93

and two rhizosphere compartments on each side of the root compartment (Fig. 1). The

94

root compartment was comprised of a perspex frame and nylon cloth (pore radius
14 mm

128

(L) outward from the nylon mesh of root compartment (Fig. 1). The soil collected in

129

the middle root compartment was defined as RC. Any soil at distances more than 14

130

mm from the RC was regarded as the bulk soil (BS). Two soil slices obtained at the

131

same distance from the root plane on either side were mixed together. After slicing the

132

soil samples were sieved rapidly through a 2-mm sieve. Several fresh soil samples

133

were then stored at 4 °C for Biolog EcoplateTM analysis, whereas the other samples

134

were frozen rapidly with liquid nitrogen, stored at -80 °C, and then lyophilized before

135

analyzing the TAA and inorganic N concentrations.

136

Inorganic N and TAA determination in plant and soil samples To analyze the N

137

contents of plant samples, the samples were digested with (H2SO4 [98%] -H2O2) and

138

the N concentrations were determined using the Kjeldahl method (Page et al. 1982). 28

139

The concentrations of inorganic N, TAA, and available mineral elements were

140

analyzed in the soil samples. The inorganic N (NH4+-N and NO3–-N) was extracted

141

from the soil by shaking with 2 M KCl. 9 The soil (1g) was shaken with the 2 M KCl

142

(5 ml) in 10 ml volume polycarbonate tubes on an end-to-end shaker (200 rpm, 30

143

min). The soil suspension was centrifuged (15,000g, 15min, 4°C) and the supernatant

144

was collected. The inorganic N (NH4+-N and NO3–-N) in the supernatant was

145

determined by colorimetric assays at 630 nm and 538 nm, as described by Page

8


Page 9 of 38


146

(1982). The TAA was extracted from soil by shaking with 6 M HCl and the

147

supernatant collection was the same as inorganic N. The concentrations of TAA were

148

determined in 1:5 (w/v) soil (1 g) /HCl extracts with a colorimetric assay at 570 nm

149

using the ninhydrin method, as described by Jones (2002).29

150

Community level physiological profile (CLPP) analysis CLPPs were determined

151

using Biolog EcoplatesTM (Biolog Inc., CA, USA). Each EcoplateTM consisted of

152

triplicate combinations of 32 wells containing 31 different carbon compounds

153

(carbohydrates, carboxylic acids, amino acids, amines, polymers, and miscellaneous),

154

and a control (water) in each replicate. Tetrazolium dye was added to each substrate

155

and liberated via the microbial breakdown of individual carbon compounds. Soil

156

suspensions (1 g soil in 10 mL of 0.85% sterile saline solution) were shaken and then

157

pre-inoculated for 24 h before inoculation to allow microbial utilization of any soluble

158

organic compounds present in the soil. Hundred-fold dilutions were prepared and

159

150-µL aliquots of the dilutions were added to each well of a Biolog Ecoplate. The

160

plates were incubated at 25°C and the color development in each well was recorded as

161

the optical density at 595 nm using a plate reader at regular 12-h intervals (from 24 to

162

120 h). The plates were incubated in the dark at 25°C between measurements.

163

Normalization was performed against the blank well (control) for each replicate and

164

the average well color development (AWCD) was determined by calculating the mean

165

of each well’s absorbance value (Abs) at 595 nm for every reading time. The Shannon

166

diversity index (DQ = – Σpilog2pi, where pi is the ratio of the Abs of a particular well

9



Page 10 of 38

167

relative to the sum of the Abs of all the microplate wells, and DQ represents the

168

microbial diversity) was calculated by treating the Abs for each well as equivalent to

169

the abundance of individuals in each species.30

170

Statistical analysis All statistical analyses were performed using SPSS version 18.0

171

(SPSS Inc. Chicago, IL, USA). One-way analysis of variance (ANOVA) with was

172

used to evaluate the results at P

Differential Responses of Soybean and Sorghum ... - ACS Publications

Recommend Documents