A SYSTEMS APPROACH TO UNDERSTANDING COMBINED BIOTIC AND ABIOTIC STRESS RESPONSES IN PLANTS

• Sponsoring Institution: National Institute of Food and Agriculture
• Project Status: COMPLETE
• Funding Source: HATCH
• Reporting Frequency: Annual
• Accession No.: 1009898
• Project Start Date: Jul 1, 2016
• Project End Date: Jun 30, 2021
• Program Code: [(N/A)]- (N/A)

Recipient Organization
COLORADO STATE UNIVERSITY
(N/A)
FORT COLLINS,CO 80523

Performing Department
Agricultural Biology

Non Technical Summary
Many plant diseases are predicted to intensify as environmental temperatures increase, and many widely used single gene sources of disease resistance (R genes) are less effective at high temperatures. While the phenomenon of temperature-induced susceptibility is well documented, the mechanisms are not known. For example, high temperatures are conducive to bacterial blight (BB) disease of rice caused by Xanthomonas oryzae pv. oryzae, and most R genes are less effective at controlling disease at high relative to low temperature regimes. We will investigate the mechanisms behind increased disease at high temperatures. Preliminary data suggest that high temperatures cause an increased expression of genes that mediate disease susceptibility and decreased expression of genes that mediate disease resistance, and that these changes might contribute to increased BB disease. In the proposed studies, the impact of high temperatures on rice gene expression during X. oryzae infection will be determined. Transcriptome analyses and phenotypic assays will be used to ask if, over time, high temperatures induce disease susceptibility genes, suppress defense pathways important to R gene-mediated resistance, alter primary metabolism to enhance disease, and/or activate abiotic stress response genes. Second, the transcriptome and phenotypic data will be integrated using a co-expression network analysis to identify common components and master regulators of high temperature and pathogen stress responses. Future steps will be to validate the function of predicted genes at the network nodes.

Animal Health Component

10%

Research Effort Categories

Basic

90%

Applied

10%

Developmental

(N/A)

Classification

Knowledge Area (KA)	Subject of Investigation (SOI)	Field of Science (FOS)	Percent
203	1530	1060	100%

Knowledge Area
203 - Plant Biological Efficiency and Abiotic Stresses Affecting Plants;

Subject Of Investigation
1530 - Rice;

Field Of Science
1060 - Biology (whole systems);

Keywords

climate change

bacterial blight

rice

abiotic stress

resistance

biotic stress

Goals / Objectives
Increasing temperatures associated with climate change impact plants in many ways, including how plants respond to pathogens. Many plant diseases intensify as environmental temperatures increase, and many widely used single gene sources of disease resistance (R genes) are less effective at high temperatures. While the phenomenon of temperature-induced susceptibility is well documented, the mechanisms are not known. For example, high temperatures are conducive to bacterial blight (BB) disease of rice caused by Xanthomonas oryzae pv. oryzae, and most R genes are less effective at controlling disease at high relative to low temperature regimes. The increase in disease susceptibility and reduced efficacy of most R genes at high temperatures provide an excellent model to study the impacts of climate change-associated temperature changes on plant-bacterial interactions in an important crop plant.Our goal is to understand the mechanisms behind increased disease at high temperatures. We hypothesize that warmer temperatures increase disease by affecting the regulation of host factors involved in susceptibility in ways not exclusively associated with altered R gene function. Our preliminary data suggest that high temperatures cause an increased expression of genes that mediate disease susceptibility and decreased expression of genes that mediate disease resistance, and that these changes might contribute to increased BB disease. In the proposed studies, the impact of high temperatures on rice gene expression during X. oryzae infection will be determined. Transcriptome analyses and phenotypic assays will be used to ask if, over time, high temperatures induce disease susceptibility genes, suppress defense pathways important to R gene-mediated resistance (also called effector triggered immunity, ETI), alter primary metabolism to enhance disease, and/or activate abiotic stress response genes. Second, the transcriptome and phenotypic data will be integrated using a co-expression network analysis to identify common components and master regulators of high temperature and pathogen stress responses. Finally, expression of key genes at network nodes will be validated using a broader sampling. Our specific objectives are to:Obj 1. Determine if high temperatures enhance plant disease by early and/or increased expression of susceptibility genes and/or suppression of genes that function in resistance (ETI).Obj 2. Identify the common components and master regulators of high temperature and pathogen stress responses.Obj 3. Confirm that the target-node-genes exhibit altered expression at high temperature, ETI, susceptibility, and combined biotic-abiotic stress responses.

Project Methods
Obj 1. Determine if high temperatures enhance plant disease by early and/or increased expression of susceptibility genes and/or suppression of genes that function in resistance (ETI).Preliminary evidence from transcript profiling at 24 hpi with Xoo suggests that many plant responses to combined temperature and pathogen stresses are different from responses to the individual stresses. We will profile changes in the rice transcriptome over time after exposure to individual or combined stresses to identify specific genes whose expression is altered. We will compare responses in rice grown at high (35°C day:29°C night) and low (29°C day:21°C night) temperature regimes as described (23). Transcript changes will be assessed using RNA sequencing (RNASeq) (29). Transcriptome responses will be monitored in uninoculated rice, during disease responses with Xoo, and during resistance responses governed by two R genes (Xa10 and Xa7). The efficacy of these two genes is oppositely affected by temperature, enabling us to study the effects of temperature on ETI (23). We use rice lines near-isogenic for Xa7 and Xa10, and a single X. oryzae strain with/without cosmid-borne copies of TAL effector genes avrXa7 or avrXa10. Lesion lengths and bacterial numbers in planta will provide a quantitative measure of disease (enhanced vs 'normal' disease vs resistance) (30, 31). We predict changes in genes known to be associated with ETS and ETI, but also anticipate that the combined stresses may affect very diverse plant responses, including genes involved in hormone production/signaling and heat stress, justifying a comprehensive profiling approach. Obj 2. Identify the common components and master regulators of high temperature and pathogen stress responses.Plants respond to multiple concurrent stresses through mostly non-overlapping transcriptional networks (32, 33). Integration of transcriptome data, phenotypic analyses, and information on rice protein-protein interactions will allow the identification of expression networks that are similarly and/or differently regulated by temperature and pathogen interactions.To understand the molecular responses associated with each stress, as well as their combined contribution, we will develop an integrated stress response network that will manifest coexpressed gene clusters specific to each type of biotic and abiotic stress. The rationale of using coexpression networks is that genes that are highly correlated together are more likely to be involved in a common biological process. Currently, we have downloaded expression data from 27 experiments on pathogen interactions with rice from GEO, representing a total of 457 rice Affymetrix microarrays. These samples will be analyzed for quality, and datasets with more than 10 samples and at least two replicates each will be integrated in a normalized correlation protocol to build a genome wide coexpression network. This biotic stress coexpression network will be used to identify coexpression modules conserved in both the stresses and specific to each type to derive the rice ecosystem response coexpression network.Co-expression analyses of the regulated genes identified in our RNA-Seq experiments will be used to identify transcriptional responses and pathways that are specific to each stress, as well as those common to both processes (34). We will use the integrated stress network for stress-disease interaction analysis to allow identification of functional gene modules, which often manifest themselves as dense subnetworks (35), and to identify genes that together are regulated similarly or are coordinated to act together in a biological process (e.g. disease response). The network analysis will reveal dense clusters of genes or functional modules associated with the experimental "parameters" of interest (R-gene genotypes, pathogen strain, temperature, and time points) that are found associated from multiple experiments, and need not be differentially expressed under the specific parameters of the experiments conducted, yet have important roles in the biological processes. To identify potential regulatory pathways and biological processes that cannot be described on the basis of rice gene annotation alone, we will derive active subnetworks of genes for each experimental parameter combination (e.g. specific genotype/condition) using the ActiveModules algorithm (36), to identify gene clusters that are regulated similarly in specific processes. Protein-protein interaction data from RiceNet (37), Plant-Pathogen Interactome (38) and the Rice Interactome Network (39), will be probed to identify other genes/proteins that interact with the genes identified in our co-expression analyses, to increase the potential of identifying suitable candidates for further analysis and validation.Dissection of the individual and combined responses will be instrumental to address whether the changes in plant susceptibility at higher temperatures are derived from a general change in plant physiology, pathogen virulence, or as expected, a combination of both. Further, these analyses are likely to identify genes that contribute to both biotic and abiotic stress, likely candidates for conferring general stress tolerance in plants.Obj. 3. Confirm that the target-node-genes exhibit altered expression at high temperature, ETI, susceptibility, and combined biotic-abiotic stress responses.To validate expression patterns for predicted nodes in abiotic/biotic stress interactions, we will examine expression of predicted node genes using a quantitative RT-PCR approach. The gene expression of marker genes related to the target-node-genes will be assayed in tissues derived from different scenarios of treatments, e.g., high temperature vs. low temperature, with relevant strains of Xoo. The protocols of the disease assay will be as described (23). Overall, Obj 3 will confirm expression of the response networks and key nodes in those networks that are related to temperature and pathogen stress responses, individually and combined. Genes identified in these analyses will be likely candidates for the generation of stable edited non-transgenic lines and/or marker-assisted selection for increased biotic/abiotic tolerance in future breeding projects. Future studies (beyond this project) of a select number of confirmed target-node-genes can then proceed using a CRISPR/Cas9 approach to provide evidence for which of the target-node-genes play positive or negative roles in biotic (ETI or ETS) or abiotic (temperature) stresses, or, importantly, interactions of the two.

Progress 07/01/16 to 06/30/21

Outputs
Target Audience: Researchers: molecular plant-stress interactions Breeders: identify markers for improving plants for resistance to combined stresses Early career scientists: train in collaborative work to solve international problems. Changes/Problems:The COVID pandemic impacted our progress significantly, due to (1) restricted access to the laboratory in early 2021, (2) restricted travel to interact with collaborators, and (3) inability to hold any of the planned international workshops. However, despite these interruptions, we moved the project forward through publication and successful grant applications. In doing so, we have added Early Career Scientists to the project, expanding their training in collaborative work to solve international problems. What opportunities for training and professional development has the project provided?Overall, the project provided training opportunities for a PhD and an MS graduate student, two undergraduate students, and a post doctoral fellow. How have the results been disseminated to communities of interest?We have shared the conceptual idea of using conserved stress-response CRE/CRM as markers for genome-wide breeding through a Perspectives Review in New Phytologist. We are also working with plant breeders to validate the approach. What do you plan to do during the next reporting period to accomplish the goals?Importantly, we received funding from FFAR to collaborate with plant breeders and pathologists at CIAT in Colombia to (1) improve our bioinformatic pipeline to identify CRE and CRM associated with heat and resistance to a broad spectrum of pathogens, (2) develop molecular markers for tracking the incorporation of heat tolerance and broad-spectrum disease resistance into rice, and (3) accelerate genome-wide breeding for heat and disease tolerance aided by conserved CRE/CRM markers.

Impacts
What was accomplished under these goals? Higher temperatures associated with climate change are predicted to increase disease risk for many crop plant species. While the importance and implications of these predictions are accepted, the mechanisms for why plants are more susceptible to disease under increased temperatures are not understood. Our approaches to understand host (rice) responses that are influenced by temperature during interactions with an important bacterial pathogen will provide new insights into how plants have adapted to temperature stresses in the environment. Through this work, we hope to identify target points, potentially breeding markers, to guide future remediation efforts, by predicting ideal gene promoter composition for climate-change resilient, disease resistant rice. Thus, the proposed research has impacts that extend well beyond rice. Tolerance in crop plants to stresses, such as heat and disease, involves changes in expression patterns of large numbers of genes. These changes are controlled by short sequences in the promoters of the genes, specifically cis-regulatory elements (CRE) or combinations of CRE that are organized as modules (called cis-regulatory modules or CRM). We demonstrated that variation in CRE and CRM content and organization in promoters of disease defense response genes is linked to remarkable expression differences within and among rice varieties. Through a meta-analysis strategy that exploited gene expression data generated by multiple studies investigating rice responses against diverse pathogens, we identified CRE and CRM that are enriched in stress induced genes relative to the rest of the genome. We also demonstrated that accumulation of broad spectrum defense response (BSDR) genes containing active CRM leads to decreased susceptibility against diverse pathogens in the field. These findings indicate that CRM could serve as genetic markers to select for active genes in breeding programs. These findings are relevant because they point not only to the existence of common regulatory systems that can be exploited to develop cultivars with durable BSDR, but also they may represent an evolutionary strategy of plants to cope with simultaneous stresses. Indeed, our analysis of rice responses to multiple simultaneous stresses (e.g. pathogen and heat stress) demonstrated the existence of common regulatory pathways based on activation of a core set of genes. All of these findings support our hypothesis that that optimization of stress-responsive CRE/CRM content can be used to improve plant tolerance to stresses, in general, including enhanced tolerance to heat and disease stress. In other words, that heat and disease tolerance in crop plants can be simultaneously improved through the development, testing and application of conserved molecular markers based on CRE/CRM for breeding. In addition to revealing the association of specific CRE/CRM with stress-responsive promoters, we produced a preliminary bioinformatic pipeline to identify these associations more broadly. This publicly available pipeline can be used by others.

Publications

• Type: Journal Articles Status: Published Year Published: 2021 Citation: Tonnessen BW, AM Bossa-Castro, F Martin, JE Leach. 2021. Intergenic spaces: A new frontier to improving plant health. New Phytol 232:1540-1548. doi: 10.1111/nph.17706.
• Type: Journal Articles Status: Published Year Published: 2021 Citation: Echeverri-Rico J, E Petro, PA Fory, GM Mosquera, JM Lang, JE Leach, JD Lobaton, G Garces, R Perafan, N Amezquita, S Toro, B Mora, JB Cuasquer, J Ramierez-Villegas, MC Rebolledo, EA Torres. 2021. Understanding the complexity of disease-climate interactions for rice bacterial panicle blight under tropical conditions. PLoS ONE 16(5):e0252061. doi: 10.1371/journal.pone.0252061.
• Type: Journal Articles Status: Published Year Published: 2021 Citation: Diddi, N, L Lai, BP Brookbank, S Hussain, E Nambara, C Todd, M Nourimand, B Taran, D Song, L Holbrook, K Doshi, M Loewen, EK Luna, J Shipp, JE Leach, SJ Robinson, SR Abrams. 2021. 3-(Phenyl alkynyl) analogs of abscisic acid: Synthesis and biological activity of potent ABA antagonists. Organic & Biomolecular Chemistry 19: 2978-2985. 10.1039/D1OB00166C
• Type: Journal Articles Status: Published Year Published: 2020 Citation: Cohen S, EK Luna, JM Lang, J Ziegle, C Chang, JE Leach, M Le-Saux, P Portier, R Koebnik, JM Jacobs. 2020. High-quality complete genome resource of Xanthomonas hyacinthi generated via long-read sequencing. Plant Disease doi.org/10.1094/PDIS-11-19-2393-A
• Type: Journal Articles Status: Published Year Published: 2021 Citation: Trivedi P, C Mattupalli, K Eversole, JE Leach*. 2021. Tansley Review: Enabling sustainable agriculture through understanding and enhancement of microbiomes. New Phytologist 230: 2129-2147. DOI: 10.1111/nph.17319

Progress 10/01/19 to 09/30/20

Outputs
Target Audience: Scientific Community of agricultural and biological scientists Graduate and undergraduate students Plant breeders Plant disease management specialists Scientific Community of agricultural and biological scientists Graduate and undergraduate students Plant breeders Plant disease management specialists Changes/Problems:COVID has had a clear impact on progress. The student on this project was unable to accomplish her goals related to the project. In addition, we were not able to hold a second workshop due to the pandemic. What opportunities for training and professional development has the project provided?A new MS student, Jennifer Shipp, is working on a master's degree, studying the role of temperature in plant disease and resistance. She will develop skills in plant pathology, molecular biology, and plant physiology. The COVID pandemic has severely restricted Jennifer's progress on this project, but we are hopeful that with the advent of vaccines, she will be able to resume work on this full time. The first Advanced Crop Improvement short course provided training and personal development opportunities to 29 participants from 13 countries. We could not hold the course in 2020, due to COVID. We are submitting proposals to continue the course, with new sources of funding. How have the results been disseminated to communities of interest?Yes, in presentations at scientific meetings and universities, newsletters, podcasts, and in two papers published. What do you plan to do during the next reporting period to accomplish the goals?In the first four years, we identified several rice genes and pathways as candidates for involvement in susceptibility to bacterial blight disease at high temperature, and the enhanced activity of one resistance gene Xa7. We are currently testing some of these experimentally in rice plants in growth chamber experiments (Obj 3). We are testing the roles of different hormone responses in the enhanced activity of resistance at high temperatures. We also submitted a proposal to FFAR with the goal of understanding how rice responds to high temperature generally. We will submit a new proposal in 2020 to continue the ACI course at AfricaRice in spring 2021.

Impacts
What was accomplished under these goals? Rice generally up-regulates hormone-responsive genes during stress responses (Cohen et al 2018; 2019), most notably genes in the abscisic acid, jasmonic acid and salicylic acid pathways. In particular, we have been following the up-regulation of genes in the abscisic acid (ABA) biosynthetic pathway as well as ABA-regulated genes in rice under simultaneous Xanthomonas oryzae (Xo) infection and high temperature stress. We are investigating the hormonal responses during resistant (with the Xa7 bacterial blight resistance gene) and susceptible interactions at elevated and normal temperatures. The Xa7 gene in rice increases resistance to Xo at high temperatures, and unlike other known resistance genes. Transcriptomics analysis shows that ABA biosynthesis and signaling are down-regulated in inoculated Xa7 plants, suggesting that Xa7 preserves pathogen defense by minimizing responses to heat stress. Downstream response to salicylic acid (SA), a hormone involved in rice immunity to Xo, was also down-regulated in Xo interactions with Xa7. Exogenous treatment with ABA enhanced Xa7 activity (resistance), indicating its complex role during this interaction. We are investigating the roles of ABA and SA during simultaneous Xo infection and high temperature stress through treatment of rice with a novel ABA antagonist. We demonstrated that the ABA antagonist blocks ABA-induced inhibition of germination in rice seeds. Understanding rice hormone signaling during simultaneous stresses will improve plant breeding strategies to address environmental conditions currently restricting rice production. Following the success of the 2019 shortcourse, "Advanced Crop Improvement: Meeting Challenges for Food Security" (ACI), held at AfricaRice, we are preparing to submit new versions of the course as the Broader Impacts components of different NSF proposals. The goal of the course remains the training of a new generation of scientists who understand and are able to rationally discuss challenging science issues related to international food security, and who can effectively communicate science to a broad and diverse audience. In new iterations of the course, we are focusing on the agricultural microbiome and on the role that micobiomes play in protecting plants from stresses, including abiotic stresses. Our goal is that these opportunities will provide participants with an appreciation for the complexity of adopting new technologies in the developed and developing worlds. Ultimately, the participants will see how various approaches to crop health intimately links to food security, the national and international politics of food and agriculture, and science communication.

Publications

• Type: Journal Articles Status: Published Year Published: 2020 Citation: Cohen S, JE Leach. 2020. High temperature-induced plant disease susceptibility: more than the sum of its parts. Curr Opin Plant Biol. 2020;S1369-5266(20)30028-5. 10.1016/j.pbi.2020.02.008.
• Type: Journal Articles Status: Published Year Published: 2020 Citation: Trivedi P, JE Leach, SG Tringe, T Sa, BK Singh. 2020. Plant-microbiome interactions. Nat Rev Microbiol 18L607-621. doi: 10.1038/s41579-020-0412-1.
• Type: Conference Papers and Presentations Status: Other Year Published: 2020 Citation: Leach, JE. 2020. Pursuing durable, broad-spectrum disease resistance in plants. Noel T. Keen Invited Speaker. Dec 11, UC Riverside, Riverside, CA (Virtual due to COVID).
• Type: Conference Papers and Presentations Status: Other Year Published: 2020 Citation: Leach, JE. 2020. Complex traits for broad-spectrum, durable disease resistance in rice. Invited webinar speaker Taking MPMI Discoveries to the Field. Dec 2, (Virtual due to COVID)
• Type: Conference Papers and Presentations Status: Other Year Published: 2020 Citation: Leach, JE. 2020. Building broad-spectrum disease resistance in rice. Invited Symposium speaker. Korean Society of Plant Pathology Annual Meetings. Oct 14, (Virtual due to COVID)
• Type: Conference Papers and Presentations Status: Other Year Published: 2020 Citation: Leach, JE. 2020. Impacts of high temperatures on disease and resistance to bacterial blight disease of rice. Symposium speaker, Plant Health 2020, APS Annual Meeting. Aug 10, (Virtual due to COVID)
• Type: Conference Papers and Presentations Status: Other Year Published: 2020 Citation: Leach, JE. 2020. Challenges for Agriculture: Impacts of climate change on plant disease. Invited speaker, 40th Philippine Academy of Science & Engineering APAMS meeting. Jul 26, (Virtual due to COVID)
• Type: Conference Papers and Presentations Status: Other Year Published: 2020 Citation: Leach, JE. 2020. Building broad-spectrum disease resistance in rice. Invited Seminar Speaker. Feb 19, Donald Danforth Plant Science Center, St Louis, MO.
• Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Shipp, Jennifer, EK Luna, SP Cohen, S Abrams, L Lai, JE Leach. 2020. Why does heat make rice more susceptible to bacterial blight disease? Poster presentation, Plant Health 2020, APS Annual Meeting. Aug 10-14, (Virtual due to COVID)

Progress 10/01/18 to 09/30/19

Outputs
Target Audience:1. Scientific Community of agricultural and biological scientists 2. Graduate and undergraduate students 3. Plant breeders 4. Plant disease management specialists ? Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?A large part of this project was conducted by a Ph.D. student, Stephen Cohen, who graduated in 2019. Through this project, Stephen developed skills in computational biology, plant pathology, molecular biology, and plant physiology. Stephen received the APS I.E. Melhus Graduate Student Symposium Award Recipient 2019 to present a talk at the Plant Health 2019 meetings, and was also awarded aUSDA NIFA Education and Workforce Development Postdoctoral Fellowship in 2019. The Advanced Crop Improvement short course provided training and personal development opportunities to 29 participants from 13 countries. How have the results been disseminated to communities of interest?Yes, in presentations at scientific meetings, podcasts, and in two papers published. ? What do you plan to do during the next reporting period to accomplish the goals?In the first two years, we identified several rice genes and pathways as candidates for involvement in susceptibility to bacterial blight disease at high temperature. We are currently testing some of these experimentally in rice plants in growth chamber experiments (Obj 3). We will submit a new proposal in 2020 to continue the ACI course at AfricaRice in spring 2021. ?

Impacts
What was accomplished under these goals? To identify the core stress response in rice, we performed a meta-analysis of publicly available rice transcriptome data (Cohen et al. 2019). Our results confirm our previous report (Cohen et al., 2018) and reports of others that rice universally down-regulates photosynthesis in response to both abiotic and biotic stress. Rice also generally up-regulates hormone-responsive genes during stress response, most notably genes in the abscisic acid, jasmonic acid and salicylic acid pathways. We identified several promoter motifs that are likely involved in stress-responsive regulatory mechanisms in rice. With this work, we also developed a list of candidate genes to study for improving rice stress tolerance in light of environmental stresses. Our work also serves as a proof of concept to show that meta-analysis of diverse transcriptome data is a valid approach to develop robust hypotheses for how plants respond to stress. In lab experimental validation of some of the pathways is now in progress. A new shortcourse, "Advanced Crop Improvement: Meeting Challenges for Food Security" (ACI) is a collaboration between AfricaRice, Colorado State University, Cornell University and IRD (France). The goal of the course was to train a new generation of plant scientists who understand and are able to rationally discuss challenging science issues related to international food security, and who can effectively communicate science to a broad and diverse audience. The ACI course showcased real-world issues faced in crop production and delved into state-of-the-art crop improvement methods and the sometimes controversial issues associated with their use. Participants experienced diverse cropping systems, and interviewed international agricultural scientists as well as consumers, growers, and marketers to produce podcasts (https://sciencecolorado.podbean.com/). These opportunities provided participants with an appreciation for the complexity of adopting new technologies in the developed and developing worlds. Ultimately, the participants saw how the science of crop improvement intimately links to food security, the national and international politics of food and agriculture, and science communication.

Publications

• Type: Journal Articles Status: Published Year Published: 2019 Citation: Cohen SP, JE Leach. 2019. Abiotic and biotic stress induce a core transcriptome response in rice. Scientific Reports 9:6273. doi: 10.1038/s41598-019-42731-8.
• Type: Journal Articles Status: Published Year Published: 2019 Citation: Tonnessen, BW, A Bossa-Castro, R Mauleon, N Alexandrov, JE Leach. 2019. Shared cis-regulatory architecture across defense response genes predicts broad spectrum quantitative resistance in rice. Scientific Reports 9:1536 DOI: 10.1038/s41598-018-38195-x
• Type: Conference Papers and Presentations Status: Other Year Published: 2019 Citation: Leach, JE. 2019 Interactions in the Phytobiome: Understanding the System to Manage Plant Health; Plenary speaker, Annual Meetings of the Society of Nematology, July 13-19, Raleigh, NC
• Type: Conference Papers and Presentations Status: Other Year Published: 2019 Citation: Leach, JE. 2019. Tipping the Balance: Long-Lasting Disease Resistance to Multiple Pathogens, 2019 Agropolis Louis Malassis International Prize for Food and Agriculture --Distinguished Scientist Award, May 23, Montpellier, France
• Type: Conference Papers and Presentations Status: Other Year Published: 2019 Citation: Leach, JE. 2019. Pieces of the Phytobiome: Environmental Influences on Plant Health, Invited speaker, 3rd Ag & Climate Conference, Mar 24-26, Budapest, Hungary
• Type: Conference Papers and Presentations Status: Other Year Published: 2019 Citation: Leach, JE. 2019. Interactions in the phytobiome: Understanding the system to manage plant health. Phytobiomes and plant health: from basics to applications Jan 23-25, Thessaloniki, Greece

Progress 10/01/17 to 09/30/18

Outputs
Target Audience:1. Scientific Community of agricultural and biological scientists 2. Plant breeders 3. Plant disease management specialists Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?This project is being conducted by a Ph.D. student, Stephen Cohen. Stephen is being mentored by Leach and Argueso. Through this project, Stephen is building skills in computational biology, plant pathology, molecular biology, and plant physiology. Stephen attended two conferences in 2018 [International Congress on Plant Pathology in Boston (7/2018) and the Xanthomonas Genomics Workshop in Halle, Germany (7/2018)]. He was invited as a speaker at the International Congress on Plant Pathology based on his expertise in the area of combined stresses. Stephen also was awarded a Sustainability Leadership Fellowship at CSU. The Rice: Research to Production short course provided training and personal development opportunities to 29 participants. How have the results been disseminated to communities of interest?Yes, in presentations at scientific meetings, and in two papers published. What do you plan to do during the next reporting period to accomplish the goals?In the first two years, we identified several rice genes and pathways as candidates for involvement in susceptibility to bacterial blight disease at high temperature. We are currently testing some of these experimentally in rice plants in growth chamber experiments (Obj 3). The Rice: Research to Production short course will serve as a template for a new 2 week short course that has been proposed to occur at the AfricaRice international center in Senegal, Africa. We have been raising funding for the course, which is planned for fall 2019 in Senegal.

Impacts
What was accomplished under these goals? In this report, we focus on progress made towards Objective 2 and 4. Both abiotic and biotic stresses cause major yield losses to crops. Breeding tolerance for a single stress (e.g. drought, salinity, pathogen, etc.), or a single stress type (e.g. abiotic or biotic) may be risky because plants respond uniquely to different or simultaneous stresses, and increasing tolerance to one stress may be at the expense of tolerance to another. With climate change, more extreme weather events are occurring, increasing the likelihood that plants experience multiple stresses in the field, including additional pressure from plant diseases. There is, therefore, a need to understand the similarities and differences among stress response pathways to best optimize targeted crop improvement. To identify overlapping pathways in stress responses, we used a meta-analysis of publicly available rice transcriptome data, to explore the rice core stress response. Our results confirm our previous report (Cohen et al., 2018) and reports of others that rice universally down-regulates photosynthesis in response to both abiotic and biotic stress. Rice also generally up-regulates hormone-responsive genes during stress response, most notably genes in the abscisic acid, jasmonic acid and salicylic acid pathways. We identified several promoter motifs that are likely involved in stress-responsive regulatory mechanisms in rice. With this work, we also developed a list of candidate genes to study for improving rice stress tolerance in light of environmental stresses. Our work also serves as a proof of concept to show that meta-analysis of diverse transcriptome data is a valid approach to develop robust hypotheses for how plants respond to stress. In lab experimental validation of some of the pathways is now in progress. The Rice: Research to Production 3 week short course was held at the International Rice Research Institute (IRRI) in August of 2017, the courses' 9th year. Leach led the coordination of the course, in collaboration with IRRI scientists. The goal of the course is to create a globally aware, well-networked core of innovative researchers ready to address international issues. In 2017, there were 29 registered participants (15 male and 14 female) from 14 countries: Australia (1), Bangladesh (2) Brazil (2) China (1), Fiji (1), India (7), Indonesia (1), Kenya (1), Mozambique (1), Philippines (2), Sri Lanka (1), Togo (1), Uganda (1) and USA (10).

Publications

• Type: Journal Articles Status: Published Year Published: 2018 Citation: Cohen, SP, JM Jacobs, JE Leach. 2018. In planta bacterial transcriptomics predict plant disease outcomes. Trends Plant Sci. 23(9): 751-753.
• Type: Journal Articles Status: Published Year Published: 2018 Citation: Cohen SP, JM Jacobs, JE Leach. 2018. Spotlight: In planta bacterial transcriptomics predict plant disease outcomes. Trends in Plant Science. 23:751-753 doi: 10.1016/j.tplants.2018.06.008
• Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Cohen SP, Liu H, Verdier VM, Leach JE. 2018. Rice hormone response is involved in the temperature-dependent function of Xa7-mediated bacterial blight resistance. International Congress on Plant Pathology 2018, Boston, MA. https://apsjournals.apsnet.org/doi/pdf/10.1094/PHYTO-108-10-S1.240 [Invited Talk]

Progress 10/01/16 to 09/30/17

Outputs
Target Audience:1. Plant scientists 2. Plant breeders 3. Plant Disease Management Specialists 4. 30 Student/Post doc participants in the Rice: Research to Production Course Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?This effort was conducted by a graduate student, with assistance from an undergraduate student. How have the results been disseminated to communities of interest?Yes, this work was published (Cohen et al., 2017) and has been presented as talks or posters at scientific meetings. What do you plan to do during the next reporting period to accomplish the goals?We are now working on experiments to define the role of ABA in simultaneous abiotic stress with plant disease and resistance.

Impacts
What was accomplished under these goals? Obj 1. To better understand host plant responses during simultaneous heat and pathogen stress, we conducted a transcriptomics experiment for rice plants (cultivar IRBB61) containing Xa7, a bacterial blight disease resistance (R) gene, that were infected with Xanthomonas oryzae, the bacterial blight pathogen of rice, during high temperature stress. Xa7-mediated resistance is unusual relative to resistance mediated by other R genes in that it functions better at high temperatures. Using RNA-Seq technology, we identified 8,499 differentially expressed genes as temperature responsive in rice cultivar IRBB61 experiencing susceptible and resistant interactions across three time points. Notably, genes in the plant hormone abscisic acid biosynthesis and response pathways were up-regulated by high temperature in both mock-treated plants and plants experiencing a susceptible interaction and were suppressed by high temperature in plants exhibiting Xa7-mediated resistance. Genes responsive to salicylic acid, an important plant hormone for disease resistance, were down-regulated by high temperature during both the susceptible and resistant interactions, suggesting that enhanced Xa7-mediated resistance at high temperature is not dependent on salicylic acid signaling. A DNA sequence motif similar to known abscisic acid-responsive cis-regulatory elements was identified in the promoter region upstream of genes up-regulated in susceptible but down-regulated in resistant interactions. The results of our study suggest that the plant hormone abscisic acid is an important node for cross-talk between plant transcriptional response pathways to high temperature stress and pathogen attack. Genes in this pathway represent an important focus for future study to determine how plants evolved to deal with simultaneous abiotic and biotic stresses.

Publications

• Type: Journal Articles Status: Published Year Published: 2017 Citation: Cohen, S, H Liu, C Argueso, A Pereira, C Vera Cruz, V Verdier, JE Leach*. 2017. RNA-Seq analysis reveals insight into enhanced rice Xa7-mediated bacterial blight resistance at high temperature. PLoS One 12(11):e0187625. doi: 10.1371/journal.pone.0187625.
• Type: Books Status: Published Year Published: 2017 Citation: Annual Review of Phytopathology. 2017. J.E. Leach, S. Lindow. Volume 55. Annual Reviews, Palo Alto, CA
• Type: Journal Articles Status: Published Year Published: 2017 Citation: Leach JE*, LR Triplett, C Argueso, P Trivedi. 2017. Communication in the Phytobiome. Cell 169:587-596 DOI: 10.1016/j.cell.2017.04.025
• Type: Journal Articles Status: Published Year Published: 2017 Citation: Michelmore, RW, G Coaker, R Bart, GA Beattie, A Bent, T Bruce, D Cameron, J Dangl, S Dinesh-Kumar, R Edwards, S Eves-van den Akker, W Gassmann, J Greenberg, R Harrison, P He, J Harvey, A Huffaker, S Hulbert, R Innes, JD Jones, I Kaloshian, S Kamoun, F Katagiri, JE Leach, W Ma, J McDowell, J Medford, B Meyers, R Nelson, RP Oliver, Y Qi, D Saunders, M Shaw, P Subudhi, L Torrance, BM Tyler, J Walsh. 2017. Foundational and translational research opportunities to improve plant health. Mol Plant Microbe Interact. doi: 10.1094/MPMI-01-17-0010-CR
• Type: Journal Articles Status: Published Year Published: 2017 Citation: Busby PE, C Soman, MR Wagner, ML Friesen, J Kremer, A Bennett, M Morsy, JA Eisen, JE Leach, J Dangl. 2017. Research priorities for harnessing plant microbiomes in sustainable agriculture. PLoS Biology. 15(3): e2001793. doi: 10.1371/journal.pbio.2001793

Progress 07/01/16 to 09/30/16

Outputs
Target Audience: Scientific community Plant Breeders Plant Disease Management Specialists Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?This project is being conducted by a Ph.D. student, Stephen Cohen. Stephen is being mentored by Leach and Argueso. Through this project, Stephen is building skills in plant pathology, molecular biology, and plant physiology. Based on an abstract he submitted, he was selected for an symposium presentation at the Keystone Symposium. Phytobiomes: From Microbiomes to Ecosystems held fromNov 8-12in Santa Fe, NM. How have the results been disseminated to communities of interest?Yes, to date in presentations at scientific meetings. We are preparing a manuscript on Stephen Cohen's work now. What do you plan to do during the next reporting period to accomplish the goals?In the first year, we identified several rice genes as candidate positive regulators of susceptibility to bacterial blight disease at high temperature. In the next year, we will investigate the role of these candidate genes in bacterial blight progression at high temperature.

Impacts
What was accomplished under these goals? Crop diseases are often more severe at high temperatures, which could pose a serious risk to global food security in light of climate variability. Because the mechanisms underlying this phenomenon are not well understood, this problem highlights the importance of studying the phytobiome for crop improvement in a changing environment. Bacterial blight of rice, caused by Xanthomonas oryzae pv. oryzae, is able to overcome many mechanisms of host immunity during concurrent infection and high temperature stress. In this first year, to study this phenomenon, we focused on Objective 1, i.e., to Determine if high temperatures enhance plant disease by early and/or increased expression of susceptibility genes and/or suppression of genes that function in resistance (ETI). We conducted a transcriptomics experiment using rice with a resistance mechanism, Xa7-mediated resistance, that is more effective at high temperatures. Differential gene expression analysis revealed that resistant plants underwent drastic transcriptional changes during simultaneous high temperature and disease stress. When compared to the susceptible controls, resistant plants downregulated several genetic responses to abiotic stress, suggesting that resistant plants prioritize resistance to disease stress over abiotic stress, while susceptible plants do the opposite. Hormone balance was also perturbed due to simultaneous stresses, with resistant plants exhibiting reduced abscisic acid signaling at high temperature. Several rice genes were identified as candidate positive regulators of susceptibility to bacterial blight at high temperature. We are now investigating the role of these candidate genes in bacterial blight progression at high temperature.

Publications

• Type: Conference Papers and Presentations Status: Other Year Published: 2016 Citation: Cohen S, C Argueso, H Liu, V Verdier, J Leach. 2016. Transcriptomic analysis reveals key genetic responses involved in the rice response to simultaneous abiotic (high temperature) and biotic (bacterial blight) stresses. Keystone Symposium. Phytobiomes: From Microbiomes to Ecosystems. Nov 8-12, Santa Fe, NM. [Invited Oral Presentation]
• Type: Journal Articles Status: Published Year Published: 2016 Citation: Triplett LR, S Cohen, C Heffelfinger, C Tekete, V Verdier, CL Schmidt, A Huerta, S Dellaporta, AJ Bogdanove, JE Leach. 2016. A resistance locus in the American heirloom rice variety Carolina Gold Select is triggered by diverse TAL effectors and is effective against African strains of Xanthomonas oryzae pv. oryzicola. Plant J 87(5):472-83. doi: 10.1111/tpj.13212.