Location: Floral and Nursery Plants Research Unit
Project Number: 1230-22000-032-13
Start Date: Sep 01, 2013
End Date: Aug 31, 2018
1. Conduct a broad comparative analysis of detection methods for R3bv2. Following listing of R3bv2 as a Select Agent in 2002, a diverse array of methods has been developed that promise rapid, sensitive detection of this pathogen subgroup. Many involve novel technologies while others are modifications of existing PCR or immunological detection techniques. However, to date these assorted methods have not been directly compared. The speed, sensitivity, accuracy, and cost of multiple published detection protocols will be quantified on a set of coded samples from geranium and potato tissue containing known numbers of R. solanacearum cells, either R3bv2 or an endemic North American strain. The resulting publication will serve as an external validation and practical reference for clinicians, industry and agency scientists seeking an optimal detection strategy for this Select Agent. 2. Conduct post-genomic studies to identify pathogen traits enabling formation of latent infections and infections in temperate conditions. The supposedly unique cold tolerance of R3bv2 strains is the basis of their Select Agent designation, and the ability of R. solanacearum in general to form latent (asymptomatic) infections contributes to the challenge of identifying infected plants. Understanding the fundamental biology underlying these traits is therefore of considerable interest. A comparative transcriptomic analysis of genes expressed at cool and warm temperatures in R3bv2 and tropical R. solanacearum strains identified several candidate cool tolerance genes. Mutagenesis, gene swaps, and biochemical analyses will be tested to determine the biological functions of these genes’ products. In parallel, it is proposed to use a similar strategy to identify bacterial genes differentially expressed in R3bv2 strains during latent and full-blown bacterial wilt disease. 3. Develop transgenic bacteriocin-producing geranium lines with immunity to R. solanacearum R3bv2. Many bacteria produce bacteriocins, which are extracellular proteins that specifically target and kill closely related bacteria. Several non-Select Agent strains of R. solanacearum produce diverse bacteriocins that can specifically inhibit or kill R3bv2 strains have been identified by ARS scientists. These bacteriocins, named solanacins, may explain why R3bv2 strains compete poorly with other R. solanacearum strains in the field. This project will characterize the range, activity, and stability of several solanacins and identify the genes that encode their synthesis. The long-term goal is to modify these genes for expression in roots and vascular tissue of geranium plants, potentially generating lines that are completely immune to R3bv2. This would be accomplished using an Agrobacterium-based plant transformation plasmid encoding anti-R3bv2 solanacins that industry scientists could introduce into their proprietary genetic lines. Input from ornamental industry representatives regarding the potential usefulness of genetically modified (GM) ornamental plants to geranium producers will be sought.