Comparative secretome analyses of toxigenic and atoxigenic Rathayibacter species

Authors: Tancos, Matthew; McMahon, Michael - Mike; Garrett, Wesley; Luster, Douglas - Doug; Rogers, Elizabeth

Submitted to: Phytopathology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 1/25/2021
Publication Date: 1/26/2021
Citation: Tancos, M.A., Mcmahon, M.B., Garrett, W.M., Luster, D.G., Rogers, E.E. 2021. Comparative secretome analyses of toxigenic and atoxigenic Rathayibacter species. Phytopathology. 111:1530-1540. https://doi.org/10.1094/PHYTO-11-20-0495-R.
DOI: https://doi.org/10.1094/PHYTO-11-20-0495-R

Interpretive Summary: The Rathayibacter genera comprises a unique, understudied group of plant-associated bacteria that include several nematode-transmitted bacterial pathogens of grasses and cereals. Rathayibacter toxicus is the most notorious species due to its ability to make neurotoxins in the seedheads of forage grasses, which causes neurological damage in grazing livestock termed annual ryegrass toxicity. Rathayibacter toxicus is currently found only in Australia and is listed as a Plant Pathogen Select Agent by the U.S. Department of Agriculture – Animal and Plant Health Inspection Service. Due to these complex, nematode-dependent disease cycles and Rathayibacter species being difficult to genetically manipulate, we currently lack a clear understanding of how Rathayibacter causes disease. Therefore, we performed the first analysis of secreted and bacterial surface-associated proteins for three agronomically-damaging Rathayibacter species. This comparative analysis of secreted proteins led to the identification of unique proteins that appear to be essential for attachment to nematodes and the ability to cause disease. Interestingly, the recently identified secreted proteins have more similarities to proteins associated with bacterial animal pathogens than true plant pathogens; thereby, suggesting a reevaluation of this nematode-transmitted bacterium. Moreover, the identification of highly abundant and unique secreted proteins will help in the development of protein-based diagnostic assays. Quick, accurate, and reliable diagnostic assays are essential for the rapid identification of these pathogens due to the increased occurrence of R. toxicus-like poisonings outside of Australia, which suggests they may be more widespread than originally thought.

Technical Abstract: Phytopathogenic Rathayibacter species are unique bacterial plant pathogens as they are obligately vectored by plant parasitic Anguinid nematodes to the developing seedheads of forage grasses and cereals. This understudied group of plant-associated Actinomycetes includes the neurotoxigenic Plant Pathogen Select Agent, R. toxicus, which causes annual ryegrass toxicity in grazing livestock. The complex Rathayibacter disease cycle requires intimate interactions with the nematode cuticle and plant hosts, which warrants an increased understanding of the secretory and surface-associated proteins that mediate these diverse eukaryotic interactions. Here we present the first comparative secretome analysis for this complex, nematode vectored Rathayibacter genera that compares three agronomically damaging Rathayibacter species, R. toxicus, R. iranicus, and R. tritici. The exoproteomic comparison identified 1,423 unique proteins between the three species using LC-MS/MS. Of the uniquely identified proteins, 94 homologous proteins were conserved between the three Rathayibacter exoproteomes and comprised between 43.4 – 58.6% of total protein abundance. Comparative analyses revealed both conserved and uniquely expressed extracellular proteins, which, interestingly, had more similarities to extracellular proteins commonly associated with bacterial animal pathogens than classical plant pathogens. This comparative exoproteome analysis will facilitate the characterization of proteins essential for vector and host colonization and assist in the development of diagnostic targets.