New Approaches to Study the Cyst Nematode/Sudden Death Syndrome Interaction in Soybean

1. 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 • Heteroderaglycines • 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 • Meloidogynehapla (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 • Fusariumvirguliforme (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., 1989- greenhouse • McLean and Lawrence, 1993 - greenhouse • Rupe et al., 1991; field trials • Hershman et al., 1990, field trials • Xing and Westphal, 2006 - microplots • Additive • Gao et al., 2006 - 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

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.

14. Experimental Design • 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

16. Inoculum 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

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.