Location: Emerging Pests and Pathogens Research
2022 Annual Report
Objectives
Objective 1: Characterize the genomes of emerging and persistent bacterial plant pathogens, including Pectobacterium and Dickeya species, to identify pathogenicity and virulence factors.
Objective 2: Functionally characterize key metabolic and virulence pathways that contribute to pathogenesis in emerging and persistent bacterial pathogens of potato and tomato.
Sub-Objective 2.1: Characterize bacterial regulators that contribute to virulence.
Sub-Objective 2.2: Characterize the roles of bacterial genes involved in calcium precipitation.
Sub-Objective 2.3: Identify genes involved in host-pathogen interactions.
Objective 3: Develop and test strategies that target pathogen biology or host interactions for control of emerging and persistent bacterial plant pathogens.
Sub-Objective 3.1: Test anti-virulence (AV) approaches for inhibiting bacterial virulence and plant disease.
Sub-Objective 3.2: Identify novel inhibitors that target bacterial genes involved in calcium precipitation.
Sub-Objective 3.3: Identify and characterize antisense RNA molecules that target metabolic or virulence factors of bacterial pathogens.
Objective 4: Develop datasets and computational tools to facilitate the development and refinement of genomes, genome annotations, and other data sets for type strains and field isolates of select bacterial plant pathogens [NP303, C2, PS2A].
Sub-Objective 4.1: Develop deep proteogenomic data sets to guide the annotation of poorly characterized type strains and field isolates of select strains of bacterial plant pathogens and other plant-associated bacteria.
Approach
Bacterial plant pathogens cause significant economic losses by reducing crop yields and value or by degrading post harvest handling and storage qualities. High value, vegetable, fruit and nursery crops, are particularly vulnerable because diseases reduce productivity and value by diminishing appearance. The threat of newly emerging plant pathogens has increased due to the combined influence of globalization and climate change, which serve to introduce and alter pathogen range and disease dynamics. As such, research is needed to develop novel control strategies that enable growers to quickly and effectively respond to emerging and persistent bacterial plant pathogens.
Our proposed studies will use state of the art high-throughput genomic and molecular methods to understand how bacteria infect and cause plant disease and how this information can be directed toward the development of novel methods to manage bacterial plant pathogens of agricultural importance. Specifically, we will focus our efforts on bacterial pathogens of solanaceous crops, such as bacterial speck of tomato caused by Pseudomonas syringae pv. tomato and blackleg disease of potatoes caused by a disease complex that includes Pectobacterium spp. and Dickeya spp. We expected to discover novel conceptual information regarding microbial adaptations that facilitate plant associations and disease. This information will guide new and environmentally sound management strategies that target features of the pathogen's biology or host interactions, specifically virulence factors. Our proposed studies are expected to result in new and innovative approaches for managing plant pathogens and ultimately increase plant health and production.
Progress Report
This is the final report for project 8062-21000-042-000D, which terminated in February 2022. All planned experiments were completed prior to the start of FY2022.
Substantial progress was made over the 5 years of the project. Over the course of the five-year project ending in February 2022, we provided diagnostic analysis for potato diseases. We used these samples to investigate the genetic relationships between bacterial strains from a wide range of locations. Results showed that there are two groups of bacteria causing soft rot and blackleg disease of potato. In one group, the bacteria were all genetically identical even though they were found at many geographically isolated farms, suggesting the bacteria came from a common source, most likely in contaminated seed potatoes. The other group were much more diverse, which is consistent with the bacteria coming from many different places, most likely at the farms where the disease outbreaks occurred. We worked with scientists at the U.S. Potato Genebank in Sturgeon Bay, Wisconsin, to screen tubers of wild potato relatives for resistance to bacterial soft rot disease. We found potato plants that have natural resistance, which can be incorporated into cultivated potato. This type of natural resistance is the most environmentally sound and sustainable method to control plant diseases, which should translate into increased food security and favorably affect the economics of potato production in the U.S. We also developed methods and performed experiments to study the interactions between soft rot bacteria and susceptible and tolerant potatoes. Using this information we characterized the plant defense responses involved in these interactions and identified natural plant products that could represent promising sources of antimicrobials. The studies advance our understanding of the molecular interactions between potato plants and soft rot pathogens and provide information to help in the development of disease management strategies and disease-resistance potatoes. We identified new pathogenic bacterial species causing disease on potato and onion in New York, Oregon, Washington and Hebei Provence, China. Efforts were also spent on identification of some new and undescribed diseases of important floriculture crops. Using molecular methods, we sequenced and assembled the genomes of bacteria pathogenic to nursery crops. The results increased our understanding of bacterial pathogens invading leaves and vascular systems of flower crops and potatoes.
Our studies also identified key bacterial regulatory networks and functions that are critical to plant disease progression. We discovered that some plant pathogenic bacteria make themselves invisible to plants at an early stage of the infection. These bacteria do this by turning off production of molecules that plants use to detect an infection and produce an immune response. We constructed a high-density barcoded Transposon library in several plant pathogenic bacteria, namely Dickeya species. These libraries allowed for the identification of fitness contributions to growth. These experiments also revealed the importance of five novel transcriptional regulators to fitness of Dickeya in potato tubers. These discoveries have helped us get closer to understanding the strategies bacteria use to cause disease and is foundational knowledge that can be used to guide development of new strategies to prevent plant diseases caused by bacteria. Additionally, the valuable new libraries are being used to discover genes required for Dickeya’s broad host-range lifestyle, identify common virulence strategies used by related phytopathogenic Dickeya strains, and more generally, investigate the role of diverse genes required for necrotrophic colonization of host plants and post-harvest diseases.
Global transcript profiling (RNA-Seq) was conducted to investigate the early interactions between Dickeya and potato plants. We found that when potatoes are challenged with Dickeya, small, naturally occurring anti-microbial molecules such as terpenes and latex proteins demonstrate changes in expression compared to unchallenged potato plants, indicating that these plant molecules might confer potato tolerance or susceptibility to Dickeya. Looking at expression of bacterial genes, we saw that Dickeya expresses many genes that code for proteins related to chemotaxis when in planta. Overall, using molecular and functional genomic methods we have been able to decipher the mechanisms involved in bacterial pathogenesis, specifically to understand how bacterial plant pathogens gauge and respond to host signals and nutrients to modulate growth, virulence, and cause disease as well as gain a greater understanding of bacterial-plant interactions.
We identified a mechanism of antimicrobial resistance in bacterial soft rot pathogens. We found that bacterial strains of Dickeya and Pectobacterium sp. are sensitive to the antimicrobial potassium tetraborate. However, the bacterial strains become resistant to the antimicrobial. The bacterial genes responsible for the antimicrobial resistance were identified. The study advances our understanding of antimicrobial resistance in bacteria and provides information on the potential risks associated with using potassium tetraborate as chemical control for bacterial soft rot pathogens.
Over the past several years of this project, we optimized our PCR-based diagnostics for important viruses and bacteria using the DNA/RNA extracted from the Flinders Technology Associates (FTA) cards for determining disease incidence estimates in dormant tubers for potato seed certification. We provided seed certification with protocols for determining disease incidence estimates using dormant tuber testing and helped with the implementation of the new procedures. The protocol provides the growers and seed buyers with swifter and more accurate potato seed health data.
Accomplishments
Review Publications
Georgoulis, S., Shalvarjian, K., Helmann, T.C., Hamilton, C., Carlson, H., Deutschbauer, A., Lowe-Power, T. 2021. Genome-wide identification of tomato xylem sap fitness factors for plant-pathogenic Ralstonia. mSystems. 6(6):e01229-21. https://doi.org/10.1128/mSystems.01229-21.
Ma, X., Brazil, J., Rivedal, H.M., Perry, K.L., Frost, K., Swingle, B.M. 2022. First report of Pectobacterium versatile causing potato soft rot in Oregon and Washington. Plant Disease. https://apsjournals.apsnet.org/doi/epdf/10.1094/PDIS-08-21-1635-PDN.
Ma, X., Lofton, L., Bamberg, J.B., Swingle, B.M. 2022. Identification of resistance to Dickeya dianthicola soft rot in Solanum microdontum. American Journal of Potato Research. https://doi.org/10.1007/s12230-021-09859-8.
Helmann, T.C., Filiatrault, M.J., Stodghill, P. 2022. Genome-wide identification of genes important for growth of dickeya dadantii and dickeya dianthicola in potato (solanum tuberosum) tubers. Frontiers in Microbiology. 13:778927. https://doi.org/10.3389/fmicb.2022.778927.
Djami-Tchatchou, A., Li, Z., Stodghill, P., Filiatrault, M.J., Kunkel, B. 2021. Identification of IAA-regulated genes in Pseudomonas syringae pv. tomato strain DC3000. Journal of Bacteriology. https://doi.org/10.1128/JB.00380-21.
