Kim Larson: Soybean Inoculation

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




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




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




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




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




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]

Documents

Kim Larson: Soybean Inoculation