1a. Objectives (from AD-416)
The primary objective of the project is to investigate the ecology, population biology and pathogenic mechanisms of plant pathogens as they interaction with cool season grain legumes that include pea, chickpea and lentil. The project will focus on pathogens, such as species of Ascochyta, Erysiphe, Sclerotinia, and Stemphylium, with economical significance to US production of cool season grain legumes. The second objective of the project is to develop management practices for controlling or mitigating the adverse effect of pathogens on crop production.
1b. Approach (from AD-416)
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 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 methods for DNA isolation and quantification. Microsatellite alleles of isolates will be determined using PCR, and haplotypes (multi locus genotype or combination of microsatellite alleles) will be determined for each isolate and used to examine genetic relatedness among isolates. 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 will be detected and quantified using high performance liquid chromatography. Unknown compounds will be identified using nuclear magnetic resonance spectroscopy and mass spectrometry. 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. 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, function of the mutated genes, and patterns of gene expression. 5. To identify resistant sources 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 scales and resistance will be rated and evaluated in repeated experiments.
3. Progress Report
Asochyta blight is a serious disease of cool season grain legumes and Asochyta blight of chickpea is particularly devastating in all chickpea production areas. The disease is caused by the fungal pathogen Ascochyta rabiei. The pathogen has two mating types and goes through a sexual cycle producing ascospores during cool and moist winter months if both mating types are available. In the United States, the sexual cycle was documented in the Pacific Northwest. A signature feature of sexual reproduction is even distribution of both mating types. Research was conducted to determine the mating type distribution of the pathogen in the Northern tier states. It was found that the two mating types are distributed in a 1:1 ratio, suggesting that sexual reproduction is occurring in the northern Greater Plains. Sexual reproduction allows for genetic recombination and generating new genotypes, which gives greater potential for responding to changing agricultural practices such as the use of new resistance cultivars and application of fungicides. Ascochyta rabiei also produces phytotoxin solanopyrones which may aid pathogenesis. The genes coding for the production of the toxins were identified in Ascochyta rabiei. Identification of the genes will allow us to conduct mutagenesis studies in order to gain insight into the role of the toxins in pathogenesis. Understanding the pathogenic mechanisms will allow us to investigate and develop appropriate management strategies for this globally important disease of chickpea. The lead scientist monitors the research progress by regular communications through meetings and e-mails. Lead scientist and cooperator meet regularly to discuss and review research results and progress at the cooperator’s facility at Washington State University. Discussion regarding results at various regional and national meetings of growers or professional and trade associations were also conducted between the cooperator and the lead scientist.