Skip to main content
ARS Home » Northeast Area » Frederick, Maryland » Foreign Disease-Weed Science Research » Research » Publications at this Location » Publication #367356
Research Project: Molecular Identification, Characterization, and Biology of Foreign and Emerging Viral and Bacterial Plant Pathogens

Location: Foreign Disease-Weed Science Research

Title: Immunoreagents for development of a diagnostic assay specific for Rathayibacter toxicus

Author
item Luster, Douglas - Doug
item McMahon, Michael - Mike
item Carter, Melissa
item Sechler, Aaron
item Rogers, Elizabeth
item SCHROEDER, BRENDA - University Of Idaho
item MURRAY, TIMOTHY - Washington State University

Submitted to: Food and Agricultural Immunology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 12/22/2019
Publication Date: 1/11/2020
Citation: Luster, D.G., Mcmahon, M.B., Carter, M.L., Sechler, A.J., Rogers, E.E., Schroeder, B.K., Murray, T.D. 2020. Immunoreagents for development of a diagnostic assay specific for Rathayibacter toxicus. Food and Agricultural Immunology. 31(1):231-242. https://doi.org/10.1080/09540105.2020.1714554.
DOI: https://doi.org/10.1080/09540105.2020.1714554

Interpretive Summary: Rathayibacter toxicus is bacterial plant pathogen causing gummosis of forage grasses, and produces a toxin responsible for recurring livestock poisonings in Australia. Because R. toxicus is an APHIS-listed Plant Pathogen Select Agent, threatening grass seed and livestock production areas in the Pacific Northwest and elsewhere, we are developing antibodies for use in diagnostic assays to prepare for the possible introduction of the pathogen into the U.S. Antibodies were evaluated for sensitivity and specificity against a panel of Rathayibacter spp. and near neighbor genera. The antibodies exhibited high sensitivity and specificity for R. toxicus in assays with cultured bacterial cells. The highest-performing antibodies were identified for further testing with field samples collected in Australia and for eventual development of an immunoassay for bacterial detection in grass samples. Such assays would help to prevent the inadvertent introduction of the pathogen as a contaminant in imported commodities and protect livestock production areas in the U.S. Pacific Northwest.

Technical Abstract: Rathayibacter toxicus is gram-positive bacterium causing gummosis of forage grasses, and produces a corynetoxin responsible for recurring livestock poisonings in Australia. Because R. toxicus is an APHIS-listed Plant Pathogen Select Agent, threatening grass seed and livestock production areas in the Pacific Northwest and elsewhere, we are developing diagnostic assays to prepare for the possible introduction of the pathogen into the U.S. Using proteomic, transcriptomic and comparative genomics data, we identified proteins expressed in R. toxicus but not in other Rathayibacter spp. Potential candidate antigens were selected based upon predicted extracellular localization, antigenicity, and solubility. Recombinant proteins corresponding to extracellular domains were used as immunogens to raise polyclonal and monoclonal antibodies against the two top candidate protein targets. Antibodies were evaluated for sensitivity and specificity in western blot and ELISA format immunoassays against a panel of Rathayibacter spp. and near neighbor genera. One polyclonal and multiple monoclonal antibodies exhibited high sensitivity and specificity for R. toxicus in assays with cultured bacterial cells. The highest-performing immunoreagents were identified for further testing with field samples collected in Australia and for eventual development of an immunoassay for bacterial detection in grass samples.