Project Number: 2090-21000-029-01-S
Project Type: Non-Assistance Cooperative Agreement
Start Date: Sep 13, 2013
End Date: Aug 31, 2018
The main objective is to investigate the etiology, ecology and population biology of fungal and viral pathogens as they interact with cool season grain legumes that include pea, chickpea and lentil. The project will focus on pathogens that frequently occur with economical significance on the grain legumes in the pacific northwest, such as Ascochyta, Sclerotinia, Stemphylium, Rhizoctonia, Pea enation mosaic virus, bean leaf roll virus and other viruses. The other objective is to study the management practices in controlling or mitigating the adverse effect of the pathogens on grain legume production and grain yields.
1. Strains of the pathogens will be collected from infected cool season grain legume plants from various geographic regions, and isolated in pure culture using standard mycological techniques. Isolates will be maintained in cellulose filter paper and in 15% glycerol at -80 C. Additional isolates will be obtained from cooperators from other locations under appropriate USDA permits. 2. To study population structure of the grain legume pathogens, total genomic DNA will be isolated from each isolate using standard DNA isolation procedure and quantified using spectrophotometric method. Microsatellite alleles of the isolates will be determined using PCR, and haplotypes (multi locus genotype or combination of microsatellite alleles) will be determined for each isolate and used in analysis using clustering and Bayesian methods. 3. Some secondary metabolites like toxins of fungal pathogens play important roles in causing diseases. To study secondary metabolites of the fungal pathogens, pathogen cultures will be grown in appropriate culture media. Secreted metabolites from the cultures will be isolated and purified using appropriate solvents and concentrated in rotary evaporator, detected and quantified using high performance liquid chromatography. Unknown compounds will be identified using nuclear magnetic resonance spectroscopy and mass spectrometer. Biological activities of the secondary metabolites will be tested using appropriate bioassays. 4. To increase our mechanistic understanding of pathogenesis of grain legume pathogens, genomic segments related to or responsible for pathogenesis will be identified through generation of non-pathogenic mutants of the grain legume pathogens. Random mutagenesis will be used to generate tagged mutations and mutants will be screened for virulence. Mutants with altered virulence will be further characterized in terms of genetic mutations and function of the mutated genes, and other genomic methods like transcriptomics. 5. For virus diseases, infected tissues of grain legumes, particularly of chickpea plants, will be collected and the presence and identity of viruses in the tissues will be detected with ELISA and PCR techniques. 6. To identify resistance sources to diseases in grain legumes, germplasm lines and cultivars of pea, chickpea and lentil will be planted in the greenhouse. At appropriate growth stages, the test plants will be inoculated with the target pathogens and incubated for disease development at environmental conditions conducive to disease development. Disease severity of the plant genotypes will be scored with appropriate rating scale and resistance will be rated and evaluated in repeated experiments.