Kim Larson
River Valley Extension District
Crop Production Agent
[1]
Introduction
Soybean acreage has been expanding in Kansas Increasing number of questions dealing with nodulation problems in
soybeans
Problems associated with
- land not previously planted to soybeans
- out of soybean production for 10 years
- Conservation Reserve Program
[2]
Soybean production distribution, 2009
(USDA)
Soybean production distribution, 2012
(USDA)
Research Objectives
Improve consistency of soybean production, especially on "new" soybean ground.
Compare inoculant products using single and double rates and combinations on fields with varying soybean history.
Determine if there is a negative interaction between inoculant products and common seed treatments.
Discover the influence of inoculated seed storage conditions before planting on the rhizobia’s ability to successfully nodulate soybean roots.
[3]
A Comparison of Inoculant Product
Treatments in Various Soybean
Production Scenarios
[4]
Introduction
Bacterial strain and carrier formulation influence the field
performance and survivability of the bacteria (Albareda et al., 2008).
Nodulation of soybean through inoculation of the bacteria in
B. japonicum - free soil has a significant role in establishing
naturalized bacterial population in the soil for subsequent
years (Kuykendall et al., 1982).
Although bacteria may persist for numerous years, it may still
prove economically beneficial to inoculate fields with soybean
history as increased yields have been obtained (Schulz and Thelen, 2008).
[5]
Objective
Compare inoculant products using single and double
rates and product combinations on fields with varying
soybean history.
[6]
Locations Eleven field experiments were conducted at seven locations in
2011 and 2012. Three of the experiments were on sites that had
soybeans present in recent rotation
Manhattan, soybeans in regular rotation
Scandia – irrigated, soybeans in regular rotation
Belleville, no soybeans ~15 yrs in 2011, ~17 yrs in 2012
Osage, no soybean ~40 yrs in
2011, ~30 yrs in 2012
White City, no soybeans ~40 years
Phillipsburg, no soybean history
[7]
Methods
Treatments:
Raw seed
Standard inoculation products:
Advanced Biological Marketing Excalibre™ and ExcalibreSA™
Becker Underwood Vault® HP and Rhizo-Stick®
Novozymes Optimize® and Soil Implant+/ Cell Tech Granular
TerraMax Maximize
Treatments designed for land not previously in soybean
Urea fertilizer nitrogen application near V4: 60 and 120 lb/a on non-
inoculated plots
[8]
Nitrogen fertilized plot
Methods
Nodulation Analysis:
10 plants dug per plot
Nodule count per plant
Nodule dry mass
Visual rating on a 0 to 5 scale (0 = no nodulation, 5 = several, large nodules at taproot)
Plant Analysis:
Plant top and root dry mass at V4
Plant nitrogen content at V4
Seed characteristics
Yield
[9]
Root nodule analysis Root washer
Field Observations
[10]
Untreated NZ-Treatment Untreated NZ-Treatment
Results:
History Group- Recent Soybean Rotation
[11]
Inoculant treatment Nodule count Nodule rating Yield Seed nitrogen
nodule plant-1 ------bu a-1------- --------g kg-1-------
Check 14 a 2.6 a 49 d 55.5 abc
ABM- Excalibre 12.7 a 2.3 a 56 ab 54.9 bcde
ABM- ExcalibreSA 14.2 a 2.3 a 54 abcd 55.3 abcd
BU- Vault 12.2 a 2.2 a 57 a 55.1 bcde
NZ- Optimize 12.6 a 2.2 a 56 a 55 bcde
TM- Maximize 13.7 a 2.7 a 57 abcd 56.1 a
BU- Vault 2X 11.3 a 2.7 a 58 ab 54.8 bcde
BU- Rhizo Stick 2X 14.3 a 2.8 a 53 abcd 54.9 cde
NZ- Optimize 2X 14.4 a 2.8 a 55 abc 55.5 abc
TM- Maximize 2X 20.1 a 2.9 a 50 abcd 54.2 e
BU- Vault/Rhizo Stick 14.3 a 2.8 a 56 ab 55.4 abc
NZ- Optimize/Cell Tech Granular 16.3 a 2.5 a 52 abcd 55.6 ab
Results: History Group- out of Soybean ≥15 years
[12]
Inoculant treatment Nodule count Nodule rating Yield Seed nitrogen
----nodule plant-1---- -----bu a-1----- -----g kg-1-----
Check 2.0 fg 1.0 f 29 abc 53.9 d
ABM- Excalibre 1.2 g 0.7 f 30 abc 54.4 cd
ABM- ExcalibreSA 1.2 g 0.8 f 31 abc 55.2 abc
BU- Vault 4.8 de 1.7 d 27 c 53.6 d
NZ- Optimize 7.6 abc 2.1 abc 29 abc 54.5 abcd
TM- Maximize 1.8 fg 1.2 ef 26 c 54.3 cd
BU- Rhizo Stick 2X 5.9 bcd 1.8 cd 27 c 55.5 ab
BU- Vault 2X 5.4 cd 1.9 bcd 30 abc 53.6 d
NZ- Optimize 2X 8.5 ab 2.3 ab 28 bc 55.8 a
TM- Maximize 2X 3.0 ef 1.6 de 26 c 55.0 abc
BU- Vault/Rhizo Stick 4.3 de 1.6 d 27 c 54.7 abcd
NZ- Optimize/Cell Tech Granular 11.5 a 2.4 a 32 ab 54.4 bcd
Results: Nodule Count Contrasts
Experimental Locations
2011 2011 2011 2012 2012 2012
Inoculant treatment Belleville Osage White City Belleville Phillips-C Phillips-S
----------------------------------nodules plant-1 --------------------------------
Single rates 7.9 a 7.7 a 3.4 a 10.2 b 1.9 a 4.4 b
2X rates 15.7 a 7.0 a 4.6 a 13.7 a 1.8 a 5.2 a
Single rates 7.9 b 7.7 a 3.4 b 10.2 b 1.9 b 4.4 b
Product combinations 11.2 a 10.5 a 3.9 a 18.5 a 5.2 a 10.2 a
Check 3.6 b 5.7 a 0.0 b - - - - - -
Single rates 7.9 a 7.7 a 3.4 a 10.2 - 1.9 - 4.4 -
[13]
Nodule and Vegetative Plant Results
Summary
There was no benefit of inoculant treatments on nodule
performance on soybean ground in recent soybean rotation.
Ground that had been out of soybean for an extended period of
time generally had a positive nodulation response to inoculant
treatments.
The company formulation tended to be more important than
increased rates or combinations of the products.
The liquid and in-furrow product combination from the highest
performing company had improved nodulation performance at five
out of ten environments.
[14]
Seed Characteristics and Yield Results Summary
Treatment differences in nodulation did not transfer to end of season yield or seed characteristics.
Seed size, nitrogen content, and test weight had few significant responses to inoculant treatments.
Highest N fertilizer rate (120 lb/a) had a positive response over the check for seed size at Phillips-S. Seed nitrogen content also was higher with this N rate compared to the check at Phillips-C and White City.
[15]
Conclusions
Only environments out of soybean production for a minimum of 15 years displayed a treatment difference on nodulation.
The inoculant source company had a greater impact on nodulation performance rather than increased rates or product combinations.
On an individual site basis, there was a significant positive response at five of the ten research environments to the highest performing company’s liquid and in-furrow inoculant combination treatment.
Treatment differences in nodulation did not transfer to end of season yield or seed characteristics.
[16]
Soybean Inoculant and Seed Treatment
Interactions
[17]
Introduction
Several studies have found a negative yield response of bacteria to
seed applied fungicide treatments (Schulz and Thelen, 2008; Hiltbold et al.,
1980; Campo et al., 2009).
The amount of viable B. japonicum on treated seeds decreases with
time but also varies with fungicide product (Revellin et al., 1993).
Inoculant labels included a listing of compatible seed treatments that
the inoculant can be added to after seed treatment.
[18]
Objective
Determine if a negative interaction exists
between inoculant products and common seed
treatments.
[19]
Methods Seed Treatments:
Untreated
ApronMaxx® RFC™
ApronMaxx® RFC™ Cruiser™
ApronMaxx® RFC™ Cruiser™ Avicta®
ApronMaxx® RFC™ Poncho® /VOTiVO®
Inoculant:
Untreated
Advanced Biological Marketing ExcalibreSA™
Becker Underwood Vault® HP
Novozymes Optimize®
TerraMax Maximize
[20]
Results: Nodule Counts
[21]
Results: Nodule Dry Mass
[22]
Results: Nodule Visual Rating
[23]
Summary of Results
Seed treatments did not negatively affect nodule counts, dry
mass, or visual ratings. The only significant main effect was
due to environment.
Plant dry mass and whole above ground plant nitrogen also
were unaffected by seed treatment.
Inoculant product affected seed size, nitrogen content, and
yield rather than seed treatment at several sites.
[24]
NS
NS
NS
NS
NS
0
5
10
15
20
25
Belleville 2011
Belleville 2012
Manhattan 2011
Manhattan 2012
Phillipsburg 2012
White City 2011
No
du
les
pe
r p
lan
t
None
ApronMaxx RFC
ApronMaxx RFC Cruiser
ApronMaxx RFC Cruiser Avicta
ApronMaxx RFC Poncho/Votivo
NS
NS
NS
NS
NS
-0.01
0.00
0.01
0.02
0.03
0.04
0.05
0.06
0.07
0.08
Belleville 2011
Belleville 2012
Manhattan 2011
Manhattan 2012
Phillipsburg 2012
White City 2011
No
du
le D
ry M
ass
(g p
er
pla
nt)
None
ApronMaxx RFC
ApronMaxx RFC Cruiser
ApronMaxx RFC Cruiser Avicta
ApronMaxx RFC Poncho/Votivo
NS
NS
NS
NS
NS
NS
0.0
0.5
1.0
1.5
2.0
2.5
3.0
3.5
4.0
4.5
Belleville2011
Belleville2012
Manhattan2011
Manhattan2012
Phillipsburg2012
White City2011
No
du
le R
atin
g
None
ApronMaxx RFC
ApronMaxx RFC Cruiser
ApronMaxx RFC Cruiser Avicta
ApronMaxx RFC Poncho/Votivo
Conclusions
Seed treatment formulations did not significantly affect soybean nodulation or yield.
Due to growing conditions, it was not unexpected to not find a positive response or benefit of seed treatments.
These results imply that seed treatments are not associated with problems that have been observed on “new” soybean ground with no naturalized Bradyrhizobium japonicum populations.
[25]
NS
NS
NS
NS
NS
0
5
10
15
20
25
Belleville 2011
Belleville 2012
Manhattan 2011
Manhattan 2012
Phillipsburg 2012
White City 2011
No
du
les
pe
r p
lan
t
None
ApronMaxx RFC
ApronMaxx RFC Cruiser
ApronMaxx RFC Cruiser Avicta
ApronMaxx RFC Poncho/Votivo
NS
NS
NS
NS
NS
-0.01
0.00
0.01
0.02
0.03
0.04
0.05
0.06
0.07
0.08
Belleville 2011
Belleville 2012
Manhattan 2011
Manhattan 2012
Phillipsburg 2012
White City 2011
No
du
le D
ry M
ass
(g p
er
pla
nt)
None
ApronMaxx RFC
ApronMaxx RFC Cruiser
ApronMaxx RFC Cruiser Avicta
ApronMaxx RFC Poncho/Votivo
NS
NS
NS
NS
NS
NS
0.0
0.5
1.0
1.5
2.0
2.5
3.0
3.5
4.0
4.5
Belleville2011
Belleville2012
Manhattan2011
Manhattan2012
Phillipsburg2012
White City2011
No
du
le R
atin
g
None
ApronMaxx RFC
ApronMaxx RFC Cruiser
ApronMaxx RFC Cruiser Avicta
ApronMaxx RFC Poncho/Votivo
Inoculated Seed Storage Effect on
Soybean Nodulation
[26]
Introduction
The viability of bradyrhizobium on the seed decreases with time and increasing temperature (Penna et al., 2011).
Inoculated seed storage should preferably be below 20°C (68° F) (Penna et al., 2011).
The number of viable bacteria that are retained on the seed after inoculation decreases rapidly within 30 days of seed storage when stored at 25°C (77°F) (Albareda et al., 2008).
Heat and lack of water causes desiccation of rhizobial cells, resulting in cell death. Cell death caused by these circumstances has been shown to follow a negative linear relationship over time (Mary et al.,1985).
[27]
Objective
Determine the influence of inoculated seed
storage conditions before planting on the
rhizobia’s ability to successfully nodulate
soybean roots.
[28]
Methods Experimental Set-up:
Randomized split-plot factorial design
Four replications
Storage Treatments of Inoculated Seed:
Control – no storage
Storage temperatures:
15°C; 25°C; 35°C; 40°C
59°F; 77°F; 95°F; 104°F
Storage conditions:
Humid; desiccant
Storage length:
4, 12, 24, 48, 168, 336 hours
[29]
CS215 temp and humidity probe set-up
Seed inoculation materials
Results and Conclusions
Nodulation performance did not match expected outcomes
of storage treatment effect on inoculated soybean seeds.
Based on previous knowledge, higher temperatures, longer
storage, and desiccant conditions should have resulted in
reduced cell viability.
Non-inoculated seeds nodulated; therefore the results were
not likely influenced by storage treatment.
This study must be repeated using care that there is no
outside bacterial contamination that could affect the results
before any conclusions based on treatment are made.
[30]
General Summary Inoculation according to company protocol and cool storage of inoculated seed before planting achieved successful nodulation in all of the study environments.
In situations where there was no soybean history, there were fewer numbers of nodules per plant than in environments where there had been soybeans grown in the past.
The inoculant product applied impacts the nodulation performance but not yield.
Seed treatments that are compatible with bacterial inoculants, applied to the seed before inoculation, do not have negative impacts on achieving successful nodulation.
[31]
Acknowledgements
This work was supported by the Kansas Soybean Commission.
Additional thanks goes to:
Major Professor: Dr. Kraig Roozeboom.
Graduate Committee Members: Dr. Charles Rice and Dr. Doug Shoup.
Department of Agronomy facility and staff.
Field cooperators: Doug Shoup, Randall Nelson, Keith Jansen, Phillip Goodyear, Matt Van Allen, and Tony Imm.
Fellow graduate students: Josh Jennings and Bryson Haverkamp.
The graduate and undergraduate students in our cropping systems group.
[32]
Questions?
[33]