1. Southern Illinois University
New Approaches to Study the Cyst Nematode/Sudden Death Syndrome Interaction in Soybean
Michael Schmidt
Jason P. Bond
Ahmad Fakhoury
Southern Illinois University

2. Distribution of SCN and SDS
SCN SCN & SDS

3. Soybean Cyst Nematode Heterodera glycines Many HG types classified
Resistance is qualitative/quantitative

4. Root Knot Nematode Two species of concern in Illinois
Meloidogyne incognita (southern root-knot)
Has over 500 different hosts including corn
Meloidogyne hapla (northern root-knot)
More limited host range, corn is a non host
Both species are important in soybean, crops, orchards, vineyards, etc.

5. Sudden Death Syndrome Fusarium virguliforme (Fv).
Soilborne, root rotting fungus that colonizes tap root and crown.
Produces a toxin causing foliar symptoms (scorch).

6. Components of SDS Resistance
Foliar symptoms - resistance to toxin effects, measured as DX. DX=DS*DI/9. DS= disease severity valued on a 1-9 scale. DI=disease incidence as a percentage of plants in plot having disease.
Root infection/colonization - resistance to the fungal pathogen

7. SCN Interactions with SDS
Synergistic –
Roy et al., greenhouse
McLean and Lawrence, greenhouse
Rupe et al., 1991; field trials
Hershman et al., 1990, field trials
Xing and Westphal, microplots
Additive
Gao et al., greenhouse

8. Limitations - SCN/SDS Research
Field studies were/are conducted in fields with SCN
SCN densities – generally the resolution at field level.
Providing an environment suitable for expression of SDS.
Difficult y in quantifying Fv in soil and roots.
Need for the right compliment of soybean isolines contrasting for SCN and SDS resistance.

9. New Tools to Study the Nematode/Fv/Soybean Interaction
Develop a transformation system for Fv
Generate fluorescing strains of the fungus – GFP = Green fluorescing protein.
Identify GFP phenotypes differing in aggressiveness toward soybean.
To study the infection & colonization
To screen for resistance
Producing tools
Transformation system
Tagged
Production of tagged mutants

10. Restriction Enzyme Mediated Integration (REMI)
Linearized Plasmid
Fungal DNA
Integration of the Linearized Plasmid
New Phenotypes Developed
Phenotypic Analysis
Isolation & Characterization of Regions Flanking the Integration
Characterizing the disrupted gene(s)

11. GFP Strain in Soybean Roots

12. REMI Results
An aggressive Fv isolate known as Mont1 (Monticello, IL) was chosen as source population for transformation.
Several GFP strains were produced.
An aggressive strain of Fv was identified.
A non-aggressive strain was identified.

13. OBJECTIVE: To use the aggressive and non-aggressive strains to:
Determine the role of SCN and RKN in the ability of Fv to penetrate and colonize soybean roots.
Determine the role of SCN and RKN in the ability of Fv to cause SDS foliar symptoms.
OBJECTIVE: To use the aggressive and non-aggressive strains to:

14. Experimental Design Greenhouse experiment.
Greenhouse experiment.
Five soybean cultivars different in their resistance to SCN, RKN, and SDS.
Cultivar s were challenged with the GFP-expressing aggressive strain and the GFP-expressing non-aggressive strain.
Cultivars were co-challenged with SCN and RKN.
The experiment consisted of 30 treatments replicated 5 times.
Duration of the experiment was 15 days.
Fv root colonization quantified via QPCR.
SDS leaf scorch evaluated on a 1-9 scale.

15. Cultivars Cultivar H. glycines M. incognita F. virguliforme Forrest R
P94M50 S
Spencer
GH3983 LS

16. Inoculum Mock Inoculation Aggressive strain Non-aggressive strain
SCN + aggressive strain
SCN + non-aggressive strain
RKN + aggressive strain
Five cultivars x Six inoculum levels = 30 treatments.
Inoculum: SCN=2300egs/plant, RKN=2400 eggs/plant, Fv=7,700,000spores.

17. Greenhouse Experiment
Waterbath system maintains constant temperature across all experimental units.
Five replications, sample size of 3 plants.
~0.5 liter soil/container.

18. Fungus
Fungus + SCN
Fungus +RKN
Cultivar
H. glycines
M. incognita
F. virguliforme
Forrest R
P94M50 S
Spencer
GH3983 LS

19. Cultivar
H. glycines
M. incognita
F. virguliforme
Forrest R
P94M50 S
Spencer
GH3983 LS

20. Cultivar
H. glycines
M. incognita
F. virguliforme
Forrest R
P94M50 S
Spencer
GH3983 LS

21. Cultivar
H. glycines
M. incognita
F. virguliforme
Forrest R
P94M50 S
Spencer
GH3983 LS

22. Results
Co-inoculation with Fv and either SCN or RKN lead to a significant increase in Fv root colonization and SDS foliar symptoms for all cultivars.
Co-inoculating with the fungus and the nematode can overwhelm plant resistance to SDS under greenhouse conditions.