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ARS Home » Northeast Area » Frederick, Maryland » Foreign Disease-Weed Science Research » Research » Research Project #431767

Research Project: Molecular Identification, Characterization, and Biology of Foreign and Emerging Viral and Bacterial Plant Pathogens

Location: Foreign Disease-Weed Science Research

Project Number: 8044-22000-044-00-D
Project Type: In-House Appropriated

Start Date: Mar 26, 2017
End Date: Mar 25, 2022

Objective:
Objective 1: Develop broad range nucleic acid, antibody and metabolomics based diagnostics for vectored plant pathogens. [NP303, C1, PS1] 1-A. Develop E-probe Diagnostic Nucleic acid Assay (EDNA) diagnostics for the detection of plant pathogen vectors. 1-B. Develop massively parallel sequencing based diagnostic for the detection of bacterial pathogens in vectors. 1-C. Develop immunodiagnostic reagents for specific and sensitive detection of Rathayibacter toxicus and tunicamycin toxin in plant products. 1-D. Collect and characterize foreign and emerging bacterial plant pathogens. Objective 2: Assess the effects of host metabolism and environmental factors on transmission, biology and evolution of threatening and emerging insect-transmitted plant pathogens. [NP303, C2, PS2C] 2-A. Assess the effects of vernalization on Plum pox virus adaptation to new hosts. 2-B. Determine the effects of Plum pox virus infection on host plant metabolomics. 2-C. Xylella fastidiosa subsp. pauca comparative genomes and proteomes. 2-D. Transmission of Xylella fastidiosa subsp. pauca CoDiRO by glassy-winged sharpshooter. Objective 3: Identify genes and proteins required for infection, toxin production and pathogenicity of foreign bacterial plant pathogens. [NP303, C2, PS2A] 3-A. Control of toxin production in Rathayibacter toxicus. 3-B. Rathayibacter toxicus gall transcriptome and proteome.

Approach:
Metagenomics based detection of pathogens and vectors will utilize E-probe Diagnostic Nucleic acid Assay (EDNA) diagnostics. E-probes will be developed for vectors and vectored bacterial pathogens, and tested on metagenomes from controlled simulated insect traps, then extended to test assay success on real world samples. Immunoassays for Rathayibacter toxicus will be developed by the identification of soluble, high abundance, extracellular and/or secreted pathogen proteins as potential diagnostic targets followed by production of monoclonal antibodies. Obtain cultures of target bacteria from major international collections, foreign collaborators, and by traveling abroad. Accessions will be cloned, checked for authenticity using biochemical tests and added to the FDWSRU International Collection of Phytopathogenic Bacteria. Effects of vernalization on Plum pox virus (PPV) biology will be assessed using parallel lines of PPV in peaches, one undergoing artificial vernalization and the other without undergoing vernalization. PPV effects on the metabolome of peaches will be assessed using standard methods, testing PPV positive symptomatic and non-symptomatic trees and comparing the results to metabolomic profiles from healthy and Prunus necrotic ringspot infected trees. The genomes of multiple Xylella fastidosa subsp. pauca isolates will be sequenced, and comparative genomics will be used to assess potential host range and pathogenicity factors. The transmission of the olive strain will be tested using glassy-winged sharpshooter biotypes from the U.S. The genes responsible for toxin production in Rathyaibacter toxicus will be confirmed by gene knockouts, and the regulation and control of these genes will be studied using transcriptomics and proteomics.