Location: Crops Pathology and Genetics Research
Project Number: 2032-22000-017-000-D
Project Type: In-House Appropriated
Start Date: Apr 25, 2022
End Date: Apr 24, 2027
Objective:
Objective 1: Characterize and incorporate resistance to disease into woody perennial crops.
Sub-objective 1.A: Characterize and field test crown gall (CG) resistant rootstock genotypes for walnut production systems.
Sub-objective 1.B: Identify, develop, characterize, and field test CG resistant rootstock genotypes for almond production systems.
Sub-objective 1.C: Identify, characterize, and field test Phytophthora resistant rootstock genotypes for walnut production systems.
Sub-objective 1.D: Identify, develop, characterize, and field test Phytophthora resistant rootstock genotypes for almond production systems.
Sub-objective 1.E: Develop Phytophthora resistant almond and walnut rootstocks using RNA interference (RNAi).
Sub-objective 1.F: Characterize the genetics behind phosphite-induced resistance against Phytophthora in walnut.
Sub-objective 1.G: Identify sources of recessive alleles in walnut germplasm that confer resistance to Cherry leaf roll virus (CLRV).
Objective 2: Characterize soil/phytomicrobiome communities and targeted phytopathogens to understand their impact on plant and soil health, enhance pathogen diagnostics, and develop optimal disease management strategies for woody perennial crops.
Sub-objective 2.A: Characterize potential genes linked to CG and Phytophthora crown and root rot resistance/tolerance in walnut and the in planta gene expression of A. tumefaciens and P. pini.
Sub-objective 2.B: Determine antagonistic interactions between A. tumefaciens strains isolated from walnut orchards.
Sub-objective 2.C: Characterize populations of Phytophthora in almond and walnut orchards and surface sources of irrigation water.
Sub-objective 2.D: Characterize metatranscriptomic and metagenomic profiles of walnut rootstock to which Paradox canker was graft transmitted versus healthy walnut rootstock.
Sub-objective 2.E: Develop improved primers for detection of Grapevine fanleaf virus (GFLV) and Grapevine leafroll-associated virus 4 (GLRaV-4).
Objective 3: Develop novel and sustainable biologically-based management strategies to control targeted pathogens and replant disorders.
Sub-objective 3.A: Examine potential nutritional and microbial contributions of ASD treatment components to PRD induction and management in almond.
Sub-objective 3.B: Determine which volatile organic compounds (VOCs) are produced in ASD-treated soils as a function of carbon source.
Sub-objective 3.C: Develop sentinel grapevine genotypes for early detection of red blotch virus in vineyards.
Approach:
Objective 1 1.A: Inoculate walnut rootstocks Agrobacterium tumefaciens and rate crown gall (CG) disease symptoms. Map genetic loci mediating CG resistance. 1.B: Inoculate almond with A. tumefaciens and rate CG severity in greenhouse trials. Evaluate field performance of selected resistant rootstocks. 1.C: Inoculate walnut rootstocks with P. cinnamomi and rate symptoms. Analyze resistance phenotypes and genotyping data to resolve quantitative trait loci. 1.D: Rate experimental and commercial almond rootstocks for resistance to Phytophthora in orchard trials. 1.E: Rate disease symptoms in walnut lines carrying host-induced gene silencing (HIGS) contructs inoculated with Phytophthora. Extract RNA from inoculated and non-inoculated walnut lines and perform RNA-Seq to identify microRNA produced from HIGs vectors. 1.F: Using an in vitro disease assay system, generate and analyze transcriptomes of phosphite-treated walnut inoculated with Phytophthora and controls treated with water treatment and mock-inoculated. 1.G: Graft uninfected walnut trees onto Cherry leaf roll virus-infected ‘Chandler’ trees and monitor for virus infection. Objective 2 2.A: Extract RNA from resistant and susceptible walnut rootstocks inoculated with A. tumefaciens or P. pini. Identify genes mediating CG or Phytophthora resistance in walnut and transcriptional changes in pathogens during infection. 2.B. Assess antagonistic interactions of A. tumefaciens strains with different opine types. Measure growth inhibition zones of A. tumefaciens strains exposed to another strain. Perform Tn5 mutagenesis and tests for phenotype restoration to identify genes involved in antagonism. 2.C: Isolate Phytophthora from almond and walnut orchards with Phytophthora-associated diseases and identify isolates using genotype-by-sequencing. Sample surface water for DNA extraction and sequencing of ITS genes and isolation and identification Phytophthora. Relate Phytophthora populations in orchards and surface sources of irrigation water. 2.D: Extract DNA and RNA from walnut tissue from graft experiments of Paradox canker disease to compare healthy control and infected tissues to identify potential causal agents. 2.E: Using an existing virome database and additional sequences derived from RNA-seq of 576 Vitis vinifera accessions, design with Primer Express 3 and test diagnostic primers and TaqMan probes for Grapevine fanleaft virus and Grapevine leafroll-associated virus 4. Objective 3 3.A: Establish field trial of anaerobic soil disinfestation (ASD) with rice bran- and almond hull and shell-based treatments. Plant almond trees and apply fertilizer treatments of with differing amounts of nitrogen and phosphorus. Profile soil and root microbiomes, soil physicochemistry, nematode populations, and measure tree growth. 3.B: Establish ASD mesocoms with different carbon source treatments. Collect volatiles for GC-MS analysis and soils for microbiome and metabolomic profiling. 3.C: For grapevine, construct synthetic silent expression construct that will result in loss of color upon infection of Grapevine red blotch virus and evaluate performance in greenhouse and field trials.
