Suppressing Plant Diseases with VermicompostA. Jack Cornell University 2008 Suppressing Plant...

A. Jack Cornell University 2008

Suppressing Plant Diseases with VermicompostResearch reports from Cornell University

Allison L. H. JackDr. Eric B. Nelson’s research groupNC State Vermicomposting Workshop

May 20, 2008


Outline

• Broader context of disease suppression•

Impacts on root microbial communities‒

Soil microbiology primer (new tools)

• Impacts on plant diseases•

Non-aerated compost tea (preliminary)


We shall not cease from explorationAnd the end of all our exploringWill be to arrive where we started

And know the place for the first time-

T. S. Eliot

1. Background

Traditional farmingGreen revolution ‒Industrial agriculture


Paradigm shift in agriculture

Soil properties

physical

chemical

biological

Green Revolution Agriculture post WWII industrial nations

Era of Agroecology10,000 B.C. ‒

now

Dr. David R. BouldinCornell University

“We were wrong”


Science only answers the questions it asks!

Dr. Helda

Morales, EcoSur

Chiapas, MX


Brief history of disease suppression research

•

Late 1800s: suppressive soils documented [Huber & Schneider 1982]

•

1959: Biological nature of suppression documented [Menzies

1959]

•

1930s ‒

1940s: Link made between composts and soil health [Howard 1942]

•

1970s -

1980s: Extensive work done on suppressive composts [Hoitink

& Kuter

1986, Weltzein

1989]


Terminology

•

Specific suppression‒

Single microorganism is responsible for observed suppression

•

General suppression‒

Multiple microorganisms are responsible

‒

Mechanism is generally assumed to be competition for nutrients


Disease suppressive composts

•

Single organism vs. pathogens‒

Antibiosis (antibiotic production)

‒

Parasitism‒

Competition for nutrients

‒

Induced Systemic Resistance (ISR)

•

Communities vs. pathogens‒

Our understanding is shockingly limited!


Example: Pythium spp. (damping off)

Post-emergence damping off

[www.ipmimages.org]


vegetative hyphae

sporangium

germinating sporangium

zoosporangium zoospores

antheridium

oogonium

oogonium oospore

Germinatingoospore

asexual

sexual

direct

indirect

DISEASE

[modified from Matthews 1931]

Dr. Velma D. MatthewsUniversity of North Carolina(1904-1958)

P. aphanidermatum

A. JackCornell University 2008


Antibiosis

Root surfaceBacillus subtilis

Zwittermicin

A (antibiotic)

[Shang et al. 1999]

Pythiumzoospore


Competition for nutrients

Seed exudates

Cucumber seed

Linoleicacid

Pythiumsporangium

[van Dijk

and Nelson 2000]

Enterobacter cloacae

Linoleicacid

Pythiumsporangium


Parasitism

www.nysaes.cornell.edu/ent/biocontrol/pathogens/trichoderma


Induced Systemic Resistance (ISR)

Pseudomonas corrugata Pythium

sporangium

[Chen et al. 2000]


Biocontrol vs. compost

•

Biocontrol agents are expensive to develop and not always effective in the field

•

Composts provide a wide variety of microbes for the plant to “chose”

from at

a much lower cost, BUT composts are not always effective either


Common approach for studying disease suppression

1. Run a bioassay to document suppression

2. Run a battery of microbiological tests on the compost

3. Attempt to correlate specific measurements with suppression


No clear pattern!Factor SOrganic matter decomposition [Nanjappa

et al. 2001, Boehm et al.

1993, Stone et al. 2001, Grunwald

et al. 2000]

?

High microbial biomass, activity [Van Os & van Ginkel

2001,

Lumsden

et al. 1987]

Y

Cultural bacteria, total:active

ratios, and molecular fingerprinting [Postma

et al. 2000,

Kowalchuck

et al. 2003,

Scheuerell

& Mahaffee

2005]

Y,N

Ability to metabolize fatty acids [McKellar & Nelson 2003] YActinomycetes

[Postma

et al. 2000, McKellar & Nelson 2003] YPseudomonas spp. [Postma

et al. 2000, Lumsden

et al. 1987] Y,NBacillus spp. [Postma

et al. 2000] N


Alternative approaches

[McKellar & Nelson 2003]

Suppressive compost Conducive compost

Entire seed colonizing community removed, then used as seed treatment

Individual isolates used as seed treatment #

of is

olat

es

S C

# of

isolat

es

S C


The whole does NOT equal the sum of the parts!

“Emergent properties”& further complications...


One gram of soil = 1,000,000 species of bacteria

1,000,000,000 individuals

Microbes we can culture ‒

0.1 to 10%

www.csb.yale.edu

K. Loeffler

Microbes we can measure with DNA ‒

80% ?

www.dees.dri.edu

Microbes we know are there ‒

100%(microscopy)


2. Impacts of Organic Transplant Media on Root Bacterial

Communities

Anu

Rangarajan, Allison Jack, Thanwalee

Sooksa-Nguan, Janice Thies

Department of Crop and Soil Sciences

Department of Horticulture

Funded by the Toward Sustainability Foundation


Potting media amendmentsSUN Industry standard: peat-based potting mix

with turkey litter compost & blood mealBASE Negative control: 70:30 (v:v) mixture of

sphagnum peat moss and vermiculiteTC Composted dairy manure solids (20%v/v)

RT SolutionsVC Vermicomposted dairy manure solids (20% v/v)

RT SolutionsSM1 Sesame meal (1% v/v)SM2.5 Sesame meal (2.5% v/v)AM5 Alfalfa meal (5%)


Earthworms farm microbes The ‘external rumen’

for manure dwellers

microbes Organicmatter

INPUT

castOUTPUT

Decomposed more available nutrients

[Swift 1979, Brown et al. 2000]

http://www.earthworms.ca/pics/earthworm.gif


‘Sleeping Beauty’

Paradox

[Lavelle et al. 1995, Brown et al. 2000]


[Brown et al. 2000, Hartenstein et al. 1981, Sampedro

et al. 2006]

Mixing and grinding

Water secreted

Intestinal mucus secreted

Water and mucusreabsorbed

Earthworm gutEisenia fetida ‒ 2.5 hr

food

casting

Viable microbes

Size of microbial community

Species composition


Are bacterial communities different in compost and

vermicompost?


Molecular techniques

•

Allow us to study microbes without cultivating them

•

Field of environmental microbiology is rapidly expanding because of these techniques


Molecular clock

•

Essential genes evolve VERY slowly‒

ribosomal RNA genes

•

Small changes in sequence indicates a different species

•

We can extract DNA, amplify these genes, and use them in different “fingerprinting”

techniques


Polymerase Chain Reaction

Tool used to amplify specific fragments of DNA for use in fingerprinting techniques


Denaturing Gradient Gel Electrophoresis (DGGE)

Increasing Denaturant C

oncentration

Separation of DNA fragments of same size based on the DNA melting properties

Low Tm

Higher Tm

R. Kantety

45%

55%


Bacterial communities: Potting mediaDGGE2

100

908070605040

DGGE2

TCTC

TC

VC

VC

VC

A

A

A

SM

SM

SM

AM

AM

AM

B

B

B

60 70 80 90 100

5040

% Similarity

TC

VC

SUN

SM

AM

BASE

45% 55%

A. Jack Cornell University 2008 http://rdp8.cme.msu.edu/html/t-rflp_jul02.html

Terminal Fragment Length Polymorphism (T-RFLP)

A. Jack Cornell University 2008 http://www.softgenetics.com/images/rawdata.jpg

Why you’re happy you’re not a scientist ☺


Are bacterial communities different in compost and

vermicompost?


IPCA1

IPCA

2

1.00.50.0-0.5-1.0-1.5

1.0

0.5

0.0

-0.5

-1.0

-1.5

TYPE OF ORGANIC MATTER

POTTING MEDIA

Plant basedamendments

Manure basedcompost amendments

TC & VC

SUN & BASE TCVCBASESUNSM1AM5SM25A. Jack

Cornell University 2008


Will tomato plants select for different bacterial rhizosphere communities based on the

potting media used?


IPCA1

IPCA

2

1.51.00.50.0-0.5-1.0

1.0

0.5

0.0

-0.5

-1.0

-1.5

TOMATO RHIZOSPHERE - JUNE

TCVCBASESUNSM1AM5SM25


VC

TC

SUN & BASEPlant basedamendments

AM

SM

TYPE OF ORGANIC MATTERA. JackCornell University 2008


Root sampling

JuneGreenhouse

Original plugJuly

August

Organic system = NOTHING added after rye/vetch cover crop plow down


Will treatment differences in root bacterial communities

persist once the seedlings are transplanted in the field?


TOMATO RHIZOSPHERE - JULY

IPCA1

IPCA

2

1.00.50.0-0.5-1.0

2.0

1.5

1.0

0.5

0.0

-0.5


TYP

E O

F O

RG

AN

IC M

ATT

ER


Plant based amendments

VC & TC

SUN &BASE

AM

SM25

SM1



IPCA1

IPCA

2

1.00.50.0-0.5-1.0-1.5-2.0

1.5

1.0

0.5

0.0

-0.5

-1.0

TIME


COMPILED DATA – ENTIRE SEASON

Potting media

June

July

AugustPlant based amendments

Manure basedcompostamendments

P

C

P

C


Conclusions

•

VC had good germination, highest transplant dry weight and significantly higher early yield

•

Tiny amounts of potting media amendments can affect root microbial communities for much of the life of the plant‒

How does this impact plant health?


3. Mechanisms of disease suppression

Allison Jack, Eric NelsonDepartment of Plant Pathology

and Plant Microbe Biology


The Spermosphere

Pythiumzoospore

cucumber seed

Seed exudates

[Nelson 2004, 2006]A. JackCornell University 2008


General questions•

Can vermicompost suppress Pythium damping off in cucumber?

•

How are zoospores prevented from infecting seeds sown in suppressive composts?

Goal•

Understand how it works, so we can develop consistently suppressive vermicomposts


Bioassay set upHeigh

t of w

ater

colum

n co

ntro

ls m

oistur

e in fu

nnels

85 cm

= -8

.5 kPa

mat

ricpo

tent

ial



Sterile glass fiber filterSoil or Soil/compost mixtureCucumber seeds

Soil or Soil/compost mixture

A. Jack Cornell University 2008A. Jack

Cornell University 2008


General questions

•

Can vermicomposted dairy manure suppress damping off in cucumber?

•

YES•

Is this suppression due to a biological factor?

A. Jack Cornell University 2008 Treatment*inoculation interaction term p < 0.001

cAver

age dise

ase ra

ting

Disease rating 5 = healthy, 0 = dead

Soil VC1 VC2 Sterile VC2


General questions

•

Can vermicomposted dairy manure suppress damping off in cucumber?

•

YES•

Is this suppression due to a biological factor?

•

YES


Release Infection

Homing

Attachment/encystment

Germination

150 um sec-1

[Nelson 2006]


General questions

•

How are zoospores prevented from infecting seeds sown in suppressive composts?‒

We know zoospores reach seed within 24 hours, what interaction is taking place in this critical time period?


24 hr incubationin seed exudate

24 hr. seed germination in:Sterile soilCDVCCDTCSterile water & filter paper

Seeds rinsed in sterile waterMicrobes remain

Seeds removed, Exudate filtered to remove cells

Microbially modified seed exudate exudate

Zoospore suspension

Agarose spotted with different types of modified seed exudate Encysted zoospores are counted after 30 minutes.

Zoospore attraction

assay

[Modified from Heungens

& Parke 2001]


Zoospores can’t find the seed!

Soil VermicompostSeed exudate from seeds sown in:


-0.2

0

0.2

0.4

0.6

0.8

1

1.2

1.4

1.6

1.8

2

V T W S H

Prop

ortio

n of

enc

yste

d zo

ospo

res r

elat

ive to

wat

er con

trol

3 reps on 3 days, 9 total replicates of the experiment

Vermicompost Compost Water Soil No seed


General questions

•

How are zoospores prevented from infecting seeds sown in suppressive composts? ‒

Seed colonizing microbes are changing the exudate so that the zoospores are no longer attracted to it

‒

Differences between VC & TC aren’t clear•

Are the microbes making a toxin or degrading an important signal?


Filtered Seed

Exudate

Control exudate

Filtered Seed

Exudate

Treatmentexudate

1. Conclude inhibitory substance Low to none High Low

2. Conclude signal missing Low to none High High

Control exudateNo exudate

Treatment exudate +

Controlexudate

Number of zoospores

Mechanism?


Future directions

•

Measure precise chemical changes•

Use this information to back track and find out which microbes are responsible for disrupting zoospore swimming

•

Develop tools to screen composts for these microbes‒

Are they reliable indicators of suppression?


Non-aerated VC tea

0

1

2

3

4

5

6

7

8

9

1:5 1:10 1:15 1:20 1:25 1:30 1:35 1:40 1:45 1:50 1:55 1:60

Ratio of VC to water (m:m)

pHDO ppmEC mS cm^-1


Non-aerated VC tea

0.0E+00

5.0E+05

1.0E+06

1.5E+06

2.0E+06

2.5E+06

3.0E+06

1:5 1:10 1:15 1:20 1:25 1:30 1:35 1:40 1:45 1:50 1:55 1:60

Ratio of VC to water (m:m)

Bac

teria

CFU

mL-

1


Potential use of vermicompost as a substitute for synthetic inputs to horticulture and nursery production

•

New York Farm Viability Institute funding‒

Eric Nelson, P.I. (PPPMB) & me ☺•

Use understanding of mechanism to develop predictive tools, study disease suppression in the field

‒

Anu

Rangarajan

(HORT)•

On-farm field trials (greenhouse/field)•

strawberry•

nursery industry•

vegetable farms‒

Chuck Nicholson (AEM)•

Economic analysis‒

Jean Bonhotal

(CSS-CWMI)

•

Outreach and educational workshops‒

Tom Herlihy

(RTS)

•

Industry collaborator, makers of “Worm Power”


New website coming soon!•

www.plantpath.cornell.edu/Labs/ENelson/Vermicompost.html

Workshop on vermicompost use (understand your markets!)

Summer 2009

[email protected]

http://www.plantpath.cornell.edu/Labs/ENelson/Vermicompost.html


AcknowledgementsNelson Lab past and present:Mary Ann KarpSofia WindstamMei-Hsing ChenSarah ArnoldMegan AckermanCristina McGuireHillary DavisMy committee:Eric Nelson (PPPMB)Anthony Hay (MICRO)Anu Rangarajan (HORT)Kathie Hodge (PPPMB)Scott Peters (EDUC)

Financial support:Department of Plant Pathology and Plant Microbe Biology

USDA BARD

Knight Institute for Writing in the Disciplines

NY State Foundation for Science, Technology and Innovation - Center for Advanced Technology (with RT Solutions)

USDA Small Business Innovation Research(with RT Solutions)

Organic Farming Research Foundation

Organic Crop Improvement Association

Andrew W. Mellon Fellowship

New York Farm Viability Institute

The “Worm Guy”Tom Herlihy – RT Solutions“Boo Boo”Steffen Jack

A. Jack Cornell University 2008Vermicompost screener at RTS’ facility

Building growth chamber apparatus for bioassays

Thanks!!!

http://www.earthworms.ca/pics/earthworm.gif

Suppressing Plant Diseases with VermicompostA. Jack Cornell University 2008 Suppressing Plant...

Documents

Transcript of Suppressing Plant Diseases with VermicompostA. Jack Cornell University 2008 Suppressing Plant...