Source: WASHINGTON STATE UNIVERSITY submitted to
FRAME: FUNGICIDE RESISTANCE ASSESSMENT, MITIGATION AND EXTENSION NETWORK FOR WINE, TABLE, AND RAISIN GRAPES
Sponsoring Institution
National Institute of Food and Agriculture
Project Status
EXTENDED
Funding Source
Reporting Frequency
Annual
Accession No.
1016359
Grant No.
2018-51181-28364
Project No.
WNP04379
Proposal No.
2018-03375
Multistate No.
(N/A)
Program Code
SCRI
Project Start Date
Sep 1, 2018
Project End Date
Aug 31, 2022
Grant Year
2019
Project Director
Moyer, M. M.
Recipient Organization
WASHINGTON STATE UNIVERSITY
240 FRENCH ADMINISTRATION BLDG
PULLMAN,WA 99164-0001
Performing Department
Agricultural Research Center
Non Technical Summary
In specialty crops, product quality is king. What happens when routine production tactics that influence quality, such as disease management, stop working? This situation has become reality in grape (wine, table, raisin) production in the USA, where control failures of powdery mildew (Erysiphe necator) have recently occurred. Many factors can influence the effectiveness of a disease management program, but fungicide resistance is the most feared. There is currently no effective system to monitor or predict fungicide resistance; it is usually identified after a management failure. Our proposed research and extension efforts will empower growers with data and predictive tools on the potential for fungicide resistance development that will be coupled with improved approaches to managing and mitigating resistance development, allowing them to design and implement fungicide resistance stewardship programs. We will develop rapid monitoring technology and the implementation programs for diagnostic labs. Our systems-approach will enable all audiences (e.g., growers, consultants, extension, chemical manufacturers and resellers) to engage in developing stewardship programs to protect at-risk fungicides. The integration of this project's objectives address: Where fungicide resistance is currently; Improve how we detect and monitor fungicide resistance and improve application efficiency; Predict where and when fungicide resistance will arise; and Develop strategies that help growers, educators, and manufactures answer what they need to do to mitigate resistance development and manage resistance that has already developed. Importantly, this Fungicide Resistance Assessment, Mitigation and Extension Network (FRAME Network) will develop tools that can be applied to other specialty crops facing fungicide resistance challenges.
Animal Health Component
0%
Research Effort Categories
Basic
45%
Applied
45%
Developmental
10%
Classification

Knowledge Area (KA)Subject of Investigation (SOI)Field of Science (FOS)Percent
2124020106060%
2121139106040%
Goals / Objectives
In specialty crops, product quality is king. What happens when routine production tactics that influence quality, such as disease management, stop working? This situation has become reality in grape (wine, table, raisin) production in the USA, where control failures of powdery mildew (Erysiphe necator) have recently occurred. Many factors can influence the effectiveness of a disease management program, but fungicide resistance is the most feared. There is currently no effective system to monitor or predict fungicide resistance; it is usually identified after a management failure. Our proposed research and extension efforts will empower growers with data and predictive tools on the potential for fungicide resistance development that will be coupled with improved approaches to managing and mitigating resistance development, allowing them to design and implement fungicide resistance stewardship programs. We will develop rapid monitoring technology and the implementation programs for diagnostic labs. Our systems-approach will enable all audiences (e.g., growers, consultants, extension, chemical manufacturers and resellers) to engage in developing stewardship programs to protect at-risk fungicides. The integration of this project's objectives address: Where fungicide resistance is currently; Improve how we detect and monitor fungicide resistance and improve application efficiency; Predict where and when fungicide resistance will arise; and Develop strategies that help growers, educators, and manufactures answer what they need to do to mitigate resistance development and manage resistance that has already developed. Importantly, this Fungicide Resistance Assessment, Mitigation and Extension Network (FRAME Network) will develop tools that can be applied to other specialty crops facing fungicide resistance challenges.Objective 1: Extension and OutreachObj. 1 - Activity 1: Pre- and post-project surveys to track short-term learning, knowledge, and skills changes in grape growers across the United StatesObj. 1 - Activity 2: In-the-field demonstrations of management recommendations.Obj. 1 - Activity 3: Develop a "University Service Center" business plan for fungicide resistance testing.Obj. 1 - Activity 4: Develop workshops and other classic extension tools; create and engage with enhanced local and regional industry groups.Objective 2: Group DecisionsObj. 2 - Activity 1: Conduct interviews to identify the externalities negotiating parties are experiencing (i.e., how their pesticide use affects their neighbors' production).Obj. 2 - Activity 2: Use game theory to develop compensating mechanisms that induce cooperation and aid fungicide stewardship by inducing cooperative behavior among growers, crop consultants and chemical companies.Objective 3: Detection and MonitoringSub-Objective 3.1 - Develop Molecular Markers to Improve the Speed of Resistance Detection.Obj. 3.1 - Activity 1: Development of PCR based diagnostic assays.Obj. 3.1- Activity 2: Developing isothermal diagnostic tools for in-field detection of fungicide resistance.Obj. 3.1 - Activity 3: Target-site gene sequencing to diagnose DMI, QoI, and SDHI resistance.Sub-Objective 3.2 - Monitoring for Fungicide Resistance.Obj. 3.2 - Activity 1: Assess the sampling density and pattern needed to estimate proportion of E. necator with fungicide resistance.Obj. 3.2 - Activity 2: Phenotype for fungicide sensitivity.Sub-Objective 3.3 - Understanding the Basis of Resistance.Obj. 3.3 - Activity 1: Elucidate mechanisms of resistance development by experimentally evolving populations with resistance to SDHI and AzN fungicides.Obj. 3.3 - Activity 2: Create reference genome of E. necator for comparative genomic and transcriptomic approaches to identifying fungicide resistance mechanisms.Obj. 3.3 - Activity 3: Identification of genomic markers associated with fungicide resistance through comparative genome and transcriptome analysis.Objective 4: Predicting Resistance SpreadSub-Objective 4.1: Conduct seasonal monitoring and recurring collection of isolates from selected grower vineyards for AzN, DMI, QoI and SDHI resistance and genetic diversity.Obj. 4.1 - Activity 1: Sample collections and phenotyping.Obj. 4.1 - Activity 2: Genome sequencing and allelic diversity analyses.Sub-Objective 4.2 - Dispersion Tracking and Population Prediction.Obj. 4.2 - Activity 1.1: Mapping resistant isolate spread risk.Obj. 4.2 - Activity 1.2: System validation and evaluation.Sub-Objective 4.3 - Fungicide Record Evaluation.
Project Methods
Objective 1: Extension and OutreachHypotheses: (1) Ready-to-use extension and outreach information will improve implementation and use of recommendations; and (2) Providing in-field examples of resistance mitigation techniques will improve grower adoption of proposed management recommendationsEffortsNational survey to develop baseline knowledge; change in knowledge.In-the-field demonstrations of management recommendations.Create and engage with enhanced local industry groups.Develop train-the-trainer workshops and classic extension products on fungicide resistance management.Develop a "Service Center" business plan for resistance testing.EvaluationMitigation efficacy determined by comparing resistance frequency and tracking crop loss in vineyards using modified programs.National survey in yrs 1 and 4 to determine changes in stakeholder knowledge/behavior.Tracking downloads and web hits for Extension presentations, curricula.MetricsShort-Term: Extension curricula improve grower knowledge on resistance fungicide management knowledge.Mid-Term: Wide-spread adoption of routine resistance testing.Long-Term: Reduction in economic loss due to fungicide resistance; reduction in overall pesticide use as growers stop using "rescue" sprays.Objective 2: Group DecisionsHypotheses: (1) Conflicting interests can be solved through negotiation between growers and chemical companies and; (2) Providing a compensating mechanism will induce cooperation among agents involved in the negotiation.EffortsInterviews to identify the source of conflict among the negotiating parties.Game theory simulation to develop compensating mechanisms that induce cooperation.Policy evaluations.EvaluationDiscussion with growers about the result of the simulation and identification of scenarios that represent their current negotiation problem.MetricsShort-Term: Identification of the source of conflict during the bargaining process.Mid-Term: Characterization and simulation of the bargaining process among growers.Long-Term: Policies are developed that promote cooperation towards resistance management.Objective 3: Detection and MonitoringHypotheses: (3.1) Field-level diagnostics based on nucleic acid amplification will be suitable for detecting presence of fungicide resistance. (3.2) Rapid sampling protocols can be used in place of intensive inoculum monitoring to accurately assess fungicide resistance presence and make management decisions. (3.3) Direct sequencing of genes or transcripts encoding fungicide resistance targets will allow for detection of low-frequency resistant genotypes before practical resistance is observed.Efforts3.1 - Developing Molecular Markers to Improve the Speed of Resistance DetectionDevelopment of PCR based diagnostic assays.Developing isothermal diagnostic tools for in-field detection of fungicide resistance.Target-site gene sequencing to diagnose DMI, QoI, and SDHI resistance.3.2 - Monitoring for Fungicide Resistance.Assess the sampling density and pattern needed to estimate proportion of E. necator with fungicide resistance.Phenotype for fungicide sensitivity.Collect spatial and temporal data on fungicide resistance for Objective 4.3.3 - Understanding the Basis of Resistance.Elucidate mechanisms of resistance development by experimentally evolving populations with resistance to SDHIs and AzN.Create reference genome of E. necator for comparative genomic and transcriptomic approaches to identifying fungicide resistance mechanisms.Identification of genomic markers associated with fungicide resistance through comparative genome and transcriptome analysis.EvaluationSampling methods must meet growers' expectations for ease of collection while balancing sensitivity of detection.Surveys will be conducted that assess grower probability of using different collection methods.Adoption of diagnostic assays by private labs and crop consultants.Implementation of in-field assays with growers and crop consultants.MetricsShort-Term: New tools become available to help diagnostic labs identify potential fungicide resistance before field failures occur.Mid-Term: Identify diversity of mutations associated with FRAC 3, 7 and 13 fungicide resistance for new diagnostic assay developmentLong-Term: Fungicide resistance monitoring becomes routine practices in vineyard management plans.Objective 4: Predicting Resistance SpreadHypotheses: (4.1) Sub-regions, or even vineyards, will have different genotypes and fungicide resistance allele frequencies. (4.2) Highly-resolved biophysical models for air turbulence and vineyard microclimate can be used to significantly improve the prediction of regional spread and risk of fungicide resistant E. necator. (4.3) Fungicide resistance development can be predicted by analysis of historical use.Efforts4.1 - Seasonal Monitoring and Recurring Collection of IsolatesEngage with growers to Identify vineyards for recurring isolate collection.Collect isolates for DNA extractions; monospore isolates for DNA sequencing.Phenotype subset of isolates for resistance to FRAC chemistries 3,7,11,13.Collect and analyze next generation sequencing data4.2 - Dispersion Tracking and Population PredictionConstruct FRAME dispersion risk system.Quantify FRAME dispersion risk system uncertainty (and reduce if possible).4.3 - Fungicide Record EvaluationSolicit historical fungicide programs from growers to determine use pattern influence on resistance.EvaluationValidated genetic features are used to develop diagnostic detection tools.Uncertainty of risk maps is bounded and of reasonable magnitude.Grower adoption rate of fungicide use histories in developing immediate and long-term fungicide plans.MetricsShort-Term: Identification of key features from historical fungicide use that may lead to development of resistance; use of that knowledge to improve spray program design.Mid-Term: Use of population models (genetic, dispersion) to predict where fungicide resistant isolates may spread. Understand how fungicide resistant-isolates spread, and how to predict that spread.Long-Term: Policies of sales and distribution of at-risk fungicides are based on risk-data and follow multi-year approaches to mitigate practical resistance development.

Progress 09/01/20 to 08/31/21

Outputs
Target Audience:The target audience includes grape growers, vineyards managers, vineyard interns, vineyard technicians, vineyard laborers, professional crop consultants, Extension professionals, plant health diagnostic clinics, chemical manufacturing companies, chemical distribution companies, industry organizations, undergraduate students, graduate students, and fellow researchers in the fields of fungicide resistance and pathogen dissemination. We have delivered results of the research through formal seminars, regional grower group meetings, Extension newsletters, trade publications and personal interactions with industry members. Peer-networking by growers across growing regions has supported dissemination of project results and increased uptake of recommended mitigation practices including in-field testing. Outreach presentations have been conducted in 7 states across the country. FRAME-specific fungicide spray program full-day virtual workshops (half-day of presentations, half-day of synchronous activities) have been provided in 12 states. Changes/Problems:We have not experienced major changes, except for unprecedented challenges associated with COVID-19 travel and workspace restrictions. This has impacted some members of our team more than others; however, given we are a smaller project, we were able to adopt and shift some of the necessary workload between groups and have been able to keep largely on-time for most project efforts. In some ways, COVID-19 has actually helped teams, such as Extension, due to the forced wide-spread adoption of distance learning, that have allowed us to reach regional grower groups that would normally be difficult to reach due to costs assocaited with time and travel. What opportunities for training and professional development has the project provided?Members of the Extension team have conducted multiple workshops and training sessions across the country since June 2020 (the end of the previous reporting timeline). This includes one-on-one training sessions for growers in multiple states, 4 FRAME-specific fungicide resistance educational virtual workshops, 23 grower presentations, and a poster at a virtual grower meeting. In-person meetings could not be held due to COVID-19 gathering and travel restrictions. There are multiple undergraduate and graduate students who are currently being trained on this project (7 graduate students, and several undergraduate (5) lab rotations). These students have participated in developing newsletter articles, a popular press article, and several peer reviewed papers (2 published, 1 submitted). Graduate students have also presented posters at the American Phytopathological Society in August 2020 and 2021 and given presentations at four separate scientific conferences. There are also 3 postdoctoral research associates who have participated in developing a peer reviewed paper, delivered 3 presentations and 2 posters at grower and scientific meetings. How have the results been disseminated to communities of interest?We have delivered 31 presentations, 4 virtual FRAME fungicide workshops, 1 popular press article, a blog post, 4 peer reviewed articles, and attended 6 scientific conferences. Our team also developed an interactive dashboard of FRAC 11 fungicide resistance results that was updated in real time during the 2021 growing season. Unfortunately, direct interactions and conference participation were limited to virtual methods due to COVID-19. In addition, when growers or consultants submit samples to one or more of our various participating laboratories for fungicide resistance genetic testing, they receive customized letters that include their sample results, along with resistance mitigation action plans. We also routinely post content to our project website at: framenetworks.wsu.edu, as well as our social media accounts (Twitter - @FRAMEnetworks, Facebook -https://www.facebook.com/FRAMEnetworks). What do you plan to do during the next reporting period to accomplish the goals?Objective 1: Extension and Outreach Obj. 1 - Activity 1: Pre- and post-project surveys to track short-term learning, knowledge, and skills changes in grape growers across the United States.Plan of action for year 4 - Begin developing post-project survey questions in fall of 2021 for dissemination in 2022. Obj. 1 - Activity 2: In-the-field demonstrations of management recommendations. Plan of action for year 4 - In-field demonstrations will not occur in year 4 due to timing of the project relative to the growing season. Obj. 1 - Activity 3: Develop a "University Service Center" business plan for fungicide resistance testing. Plan of action for year 4 - Complete compilation of protocols and business plans based on the experiences from the 2020-2021 WSU IAREC lab setup with protocols and budgets from two external labs. Obj. 1 - Activity 4:Develop workshops and other classic extension tools; create and engage with enhanced local and regional industry groups.Plan of action for year 4 - Will continue to host regional workshops, write additional extension articles, and podcasts. Will update and expand online educational content to address the current challenges of in-person training. Maintain the mapping tool to report current and historical survey results in real-time for all participating growing regions. Expand extension efforts to further reach constituents across the country. Objective 2: Group Decisions Obj. 2 - Activity 1:Conduct interviews to identify the externalities negotiating parties are experiencing (i.e., how their pesticide use affects their neighbors' production).Plan of action for year 4 - We are in the process of completing drafting the paper for this activity for submission in 2022. Obj. 2 - Activity 2: Use game theory to develop compensating mechanisms that induce cooperation and aid fungicide stewardship by inducing cooperative behavior among growers, crop consultants and chemical companies. Plan of action for year 4 - We are currently working on a signaling game that characterizes the lack of information that some growers face about the severity of fungicide resistance. We consider a context in which two grape growers have already decided to cooperate (reduce their fungicide usage), however, one of them has more information about the severity of fungicide resistance. We aim to evaluate their strategic behavior and identify under which cases growers will tend to deviate from cooperation (i.e., increase their fungicide usage) Objective 3: Detection and Monitoring Sub-Objective 3.1 - Develop Molecular Markers to Improve the Speed of Resistance Detection. Obj. 3.1 - Activity 1:Development of PCR based diagnostic assays.Plan of action for year 4 - This activity has been completed. Obj. 3.1 - Activity 2: Developing isothermal diagnostic tools for in-field detection of fungicide resistance. Plan of action for year 4 - Finish analysis of results from the separate labs and complete the paper for submission in Winter 2021. Obj. 3.1 - Activity 3:Target-site gene sequencing to diagnose DMI, QoI, and SDHI resistance.Plan of action for year 4 - Continue to monitor the incidence and dynamics of mutations in FRAC 7 - associated genes in powdery mildew from CA, OR, WA, and MI vineyards. Our toxicity assays have shown that next to target-site mutations, there is an additional mechanism in E. necator that mediates tolerance to SDHIs. We will complete comparative transcriptomic analysis to determine these alternative mechanisms. Sub-Objective 3.2 - Monitoring for Fungicide Resistance. Obj. 3.2 - Activity 1:Assess the sampling density and pattern needed to estimate proportion of E. necator with fungicide resistance.Plan of action for year 4 - In CA, commercial vineyards will be monitored for fungicide resistance by surveying powdery mildew colonies and through a network of fixed, rotorod spore traps. In OR, we are monitoring E. necator and fungicide resistance by deploying 25 spore traps in the region. In OR, MI, GA, and WA, vineyards will be intensively sampled using leaf, glove, and isolate sampling to examine which method is the efficient for allowing an accurate grower decision. This sampling will partially be in collaboration with a large consulting group to cover more vineyards on the west coast. Obj. 3.2 - Activity 2:Phenotype for fungicide sensitivity.Plan of action for year 4 - We will repeat the limited introduction of FRAC 3 and 11 products back into spray programs in cooperating vineyards to assess the viability for product use and maintenance of field level control of grape powdery mildew. We will continue to increase the number of live cultures tested for our new expanded fungicide sensitivity profile. Sub-Objective 3.3 - Understanding the Basis of Resistance. Obj. 3.3 - Activity 1: Elucidate mechanisms of resistance development by experimentally evolving populations with resistance to SDHI and AzN fungicides. Plan of action for year 4 - See Obj. 3.3 Activity 3. Obj. 3.3 - Activity 2:Create reference genome ofE. necatorfor comparative genomic and transcriptomic approaches to identifying fungicide resistance mechanisms.Plan of action for year 4 - Paper was published. Activity is complete. Obj. 3.3 - Activity 3: Identification of genomic markers associated with fungicide resistance through comparative genome and transcriptome analysis. Plan of action for year 4 -We will further in building genomic and transcriptomic resources for E. necator to understand the molecular basis of fungicide resistance in this pathogen. We will also continue to monitor for SDHI target-site mutations in populations of E. necator from different US states. Objective 4: Predicting Resistance Spread Sub-Objective 4.1:Conduct seasonal monitoring and recurring collection of isolates from selected grower vineyards for AzN, DMI, QoI and SDHI resistance and genetic diversity. Obj. 4.1 - Activity 1:Sample collections and phenotyping. Plan of action for year 4 - New postdoctoral researcher and technician will build amplicon libraries for pooled powdery mildew samples and submit them for sequencing. Sequenced samples will be analyzed for genetic variability. Obj. 4.1 - Activity 2: Genome sequencing and allelic diversity analyses. Plan of action for year 4 - Primers and probes will be finalized with the completion of the genome sequences in Spring 2021. Sub-Objective 4.2 - Dispersion Tracking and Population Prediction. Obj. 4.2 - Activity 1:Mapping resistant isolate spread risk. Plan of action for year 4 - Code cleanup has been completed. Currently in the process of drafting papers for peer-reviewed publication. Obj. 4.2 - Activity 2: System validation and evaluation. Plan of action for year 4 - the implementation of the QES dispersion modeling system with a focus on real world application of dispersion with full topography and flow fields linked to weather. We will also continue to work on efficiency of QES to make its interface, execution, and analysis smooth. This will enable to project to move from engaging researchers to engaging practitioners. Sub-Objective 4.3 - Fungicide Record Evaluation. Plan of action for year 4 - We will conclude record collections in Fall 2021 and will begin data analysis. A paper will be drafted for publication in 2022.

Impacts
What was accomplished under these goals? Overall- The third annual project meeting was held virtually 23 - 24 Feb, 2021. Participants included all project PIs and Key Personnel, most of the Stakeholder Advisory group, as well as participating technicians and graduate students. Objective Teams have and continue to host regular web conferences from monthly to quarterly (depending on nature of the objective). Objective 1 - We have learned how growers relate to information about fungicide stewardship. We have developed improved extension curriculum for optimized information uptake and adoption, and have a solid resource set on our website for regular grower interaction. Objective 2 - We have learned what drives grower's willingness to cooperate, and will be able to prescribe policy recommendations providing a different approach to subsidies or penalizations. Objective 3 - We have rapid sampling and detection tools that provide near real-time results for grape growers. Objective 4 - We have learned how powdery mildew can spread in a vineyard, and observed general fungicide application practices. Objective 1: Extension and Outreach Obj. 1 - Act. 1: Pre- and post-project surveys. Accomplishments - Published presurvey results in AJEV. Obj. 1 - Act. 2: In-the-field demonstrations.Accomplishments - Five field trials have been initiated in GA (three in 2020 [completed] and two in 2021 [in process]) to review management methods for control of powdery mildew in the presence of FRAC 3 and 11Erysiphe necator resistant populations. All powdery mildew survey data collected to date (2017-2020) were collated and a Dashboard was designed to display data by year, region and grape category (raisin, table, wine and juice). Obj. 1 - Act. 3: Develop a "University Service Center" business plan.Accomplishments - Consulted with local growers and adjusted the testing facility procedures to decrease the amount of time between sample receipt and result dispersion to growers. Obj. 1 - Act. 4:Develop workshops and other classic extension tools. Accomplishments - Delivered an additional 36 FRAME presentations with a cumulative attendance of approximately 2,400 individuals. The audiences included both the industry and the scientific community, with 23 of the 31 presentations targeted to industry. Our FRAME fungicide spray program workshop was reformatted to an asynchronous virtual format containing a series of 5 videos and a half-day interactive design section. This workshop was presented 4 times with attendees from 12 states (IA, IN, MA, MD, MI, MN, ND, NJ, OH, PA, WA, and WI). Additionally, there have been 5 student/technician posters, 1 blog post, 1 trade article, 4 newsletters, and 8 videos since June 2020. Objective 2: Group Decisions Obj. 2 - Act. 1:Conduct interviews to identify the externalities negotiating parties are experiencing.Accomplishments - We finished collecting data from two surveys: grape growers and crop consultants. The paper based on the grower survey is under revision in Journal of Wine Economics (submitted 05/20/2021) and are developing a research paper focused on the strategic behavior of grape growers when their choices of fungicide levels generate fungicide resistance which is under review in the European Review of Agricultural Economics. Objective 3: Detection and Monitoring Sub-Objective 3.1 - Develop Molecular Markers to Improve the Speed of Resistance Detection. Obj. 3.1 - Act.1:Development of PCR based diagnostic assays.Accomplishments - The PCR diagnostics are now in full-use for real-time diagnostics of grower-submitted samples. Obj. 3.1 - Act.2: Developing isothermal diagnostic tools.Accomplishments - A draft of a paper for peer-reviewed publication is being prepared for submission in Winter 2021. We are not pursuing this diagnostic tool further, per the recommendations of our stakeholder advisory group. Obj. 3.1 - Act. 3:Target-site gene sequencing to diagnose DMI, QoI, and SDHI resistance.Accomplishments - Continued expanding the collection of resistant isolates, including isolates resistant to multiple FRAC group fungicides. Sub-Objective 3.2 - Monitoring for Fungicide Resistance. Obj. 3.2 - Act. 1:Assess the sampling density and pattern needed to estimate proportion of E. necator with fungicide resistance.Accomplishments - Spore capture devices (CA, OR) and glove-swabs (GA, MI, OR, WA) were used to monitor inoculum occurrence and incidence of FRAC 11 resistance. Processed more than 5213 swabs samples for FRAC 11 and FRAC 3 genetic markers in OR, and 300 in WA. Obj. 3.2 - Act. 2:Phenotype for fungicide sensitivity.Accomplishments - We collaborated with 3 OR growers that quit using FRAC 3 and 11 fungicides to see if thesefungicides could be rotated back into their program as single applications and still achieve successful field control. We continued to bioassay individual isolates to confirm molecular resting and identify new resistant phenotypes. We expanded the profile of tested materials to include multiple chemistries in the FRAC 3, 7, 9, 11, 13, 50 and U06 groups. Sub-Objective 3.3 - Understanding the Basis of Resistance. Obj. 3.3 - Act. 2:Create reference genome ofE. necator.Accomplishments - Gene annotation of the assembled E. necator genome was completed, with an estimated completeness of 97%. Obj. 3.3 - Act. 3: Identification of genomic markers associated with fungicide resistance.Accomplishments - Functional annotations of major gene categories involved in drug resistance were performed. Further identification marker examinationrevealed that the genotype of EnFRAME01 is similar to east coast genotypes, rather than the other west coastisolates. Transcriptome of EnFRAME01 at six time points during infection has been sequenced and found3,000 genes whose expression varied significantly through time. Among four isolates of E. necator included in the analyses, no allelic variation was observed in cytb apart from the G143A mutation associated with resistance to QoIs. Objective 4: Predicting Resistance Spread Sub-Objective 4.1:Conduct seasonal monitoring and recurring collection of isolates from selected grower vineyards for AzN, DMI, QoI and SDHI resistance and genetic diversity. Obj. 4.1 - Act. 1:Sample collections and phenotyping.Accomplishments - Collected more than 1300 samples representing fields from CA, WA, OR, and GA using gloves and trap plants and tested for QoI resistance. A subset of samples with sufficiently high DNA were selected for sequencing, and several products tested for improving DNA quality prior to sequencing. A postdoctoral researcher was hired and a new technician was hired (both will start September 2021). Obj. 4.1 - Act. 2: Genome sequencing and allelic diversity analyses.Accomplishments - Genome sequence was published and amplicon based sequencing probes (for ~1400 genes) were developed. Sub-Objective 4.2 - Dispersion Tracking and Population Prediction. Obj. 4.2 - Act.1:Mapping resistant isolate spread risk.Accomplishments - The QES GPU based flow solver, the real-time dispersion and spore tracking system, development was completed thoughdynamic coupling. The model was used in initial studies to examine how spore trap placement impacts which vines are sampled in the field. We also developed a powdery mildew 3D disease spread model that incorporates key biophysical components that contribute to spore movement. The model was used in initial studies on the effectiveness of different disease scouting methodologies (e.g., visual identification, impaction traps). Obj. 4.2 - Act. 2: System validation and evaluation.Accomplishments - Models are currently being verified and two publications are being drafted which will be submitted early in 2022. Sub-Objective 4.3 - Fungicide Record Evaluation.Accomplishments - Expanded collected records to 55 CA, 2 GA, 5 MI, 1 OH, 67 OR and 19 WA vineyards for a total of 152 participating vineyards. Additionally, preliminary analyses have begun.

Publications

  • Type: Journal Articles Status: Published Year Published: 2020 Citation: Warres, B., Breeden, S., Winkles, J., Severns, P., Brannen, P., Rogers, D., Covington, R., Mahaffee, W., and Neill, T. 2020. Fungicide comparisons for powdery mildew management in a fungicide-resistant Erisiphe necator population, 2019. Plant Dis. Mgmt. Rep.14:PF014.
  • Type: Journal Articles Status: Published Year Published: 2020 Citation: Warres, B., Breeden, S., Winkles, J., Severns, P., Brannen, P., Rogers, D., Covington, R., Mahaffee, W., and Neill, T. 2020. Efficacy of sulfur and myclobutanil combinations for control of grapevine powdery mildew in Georgia, 2019.Plant Dis. Mgmt. Rep.14:PF015.
  • Type: Journal Articles Status: Published Year Published: 2021 Citation: Warres, B., Johnson, K., Busher, K., Cameron, C., Brannen, P., Rogers, D., Covington, R., Mahaffee, W., and Neil, T. 2021. Fungicide comparisons for powdery mildew management in a fungicide-resistant Erysiphe necator population, 2020.Plant Dis. Mgmt. Rep.15:PF012
  • Type: Journal Articles Status: Published Year Published: 2021 Citation: Warres, B., Johnson, K., Busher, K., Cameron, C., Brannen, P., Rogers, D., Covington, R., Mahaffee, W., and Neil, T. 2021. Assessment of interactions between sulfur and DMI fungicides to control powdery mildew in the presence of a DMI-resistant Erysiphe necator population in Georgia, 2020.Plant Dis. Mgmt. Rep.15:PF013
  • Type: Journal Articles Status: Published Year Published: 2021 Citation: Oliver, C., Cooper, M., Lewis-Ivey, M., Brannen, P., Miles, T., Mahaffee, W., and Moyer, M.2021. Assessing the United States Grape Industrys Understanding of Fungicide Resistance Mitigation Practices Am. J. Enol. Vitic.72: 181-193. https://doi.org/10.5344/ajev.2021.20062
  • Type: Journal Articles Status: Published Year Published: 2021 Citation: Bozorgmehr, B., Willemsen, P., Gibbs, J., Stoll, R., Kim, J., and Pardyjak, E. 2021. Utilizing dynamic parallelism in CUDA to accelerate a 3D red-black successive over relaxation wind-field solverEnvironmental Modelling & Software: with Environment Data News.137
  • Type: Journal Articles Status: Published Year Published: 2021 Citation: Warres, B., Johnson, K., Busher, K., Cameron, C., Brannen, P., Rogers, D., Covington, R., Mahaffee, W., and Neil, T. 2021. Efficacy of DMI (FRAC 3) fungicides for control of powdery mildew in a DMI-tolerant Erysiphe necator population, 2020. Plant Dis. Mgmt. Rep.15:PF014
  • Type: Journal Articles Status: Published Year Published: 2021 Citation: Zaccaron, A., De Souza, J., and Stergiopoulos, I. 2021 The mitochondrial genome of the grape powdery mildew pathogen Erysiphe necator is intron rich and exhibits a distinct gene organization. Scientific Reports. 11:13924
  • Type: Other Status: Published Year Published: 2021 Citation: Oliver, C. and Moyer, M. 2021. FRAME Network: Fungicide Resistance in Wine Grapes. WineVit. Virtually. (Poster)
  • Type: Other Status: Published Year Published: 2021 Citation: Newbold, C., and Mahaffee, W. 2021. Phenotypic variability among Erysiphe necator isolates. American Phytopathological Society. Virtually. (Research On Demand)
  • Type: Other Status: Published Year Published: 2021 Citation: Lowder, S., and Mahaffee, W. 2021. Reduction of the G143A mutation in Erysiphe necator overwintering chasmothecia. American Phytopathological Society. Virtually. (Research On Demand)
  • Type: Other Status: Published Year Published: 2021 Citation: Sharma, N., and Miles, T. 2021. Assessment of fungicide resistant Plasmopara viticola populations in vineyards of the Great Lakes region. American Phytopathological Society. Virtually. (Research On Demand)
  • Type: Other Status: Published Year Published: 2021 Citation: Yang, H., and Miles, T. 2021. Grape powdery mildew spray programs affect non-target fungi populations differentially in California vineyards. American Phytopathological Society. Virtually. (Research On Demand)
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Mahaffee, W. 2020. Finding Needles in Haystacks: Inoculum Monitoring as a Decision Aid. Western Region IPM Center IPM hour Webinar series. http://youtu.be/ewG-Fv5gS4o.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Moyer, M. 2020. A Casual Conversation on Grape Powdery Mildew. Thirsty Thursday Westover Viticulture Grower Meeting. Virtually.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Mahaffe, W. 2020. Improved Scouting Methods for Powdery Mildew in Vineyards. Sustainable Ag Expo. Virtually.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Mahaffe, W. 2020. Rollers, Wakes and Vortices and Other Considerations in Locating Samplers. APS Webinar. Virtually.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Warres, B., and Brannen, P. 2020. Efficacy of Fungicides for Grape Powdery Mildew Control in the Presence of Fungicide-Resistant Erysiphe necator Populations in Georgia. Cumberland-Shenandoah Fruit Workers Conference. Virtually.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2021 Citation: Brannen P. 2021. Fungicide Resistance Management for Powdery and Downy Mildews. Maryland Grape and Wine Industry Annual Conference. Virtually.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2021 Citation: Mahaffe, W. 2021. Updates on fungicide resistance monitoring. LIVE technical meeting. Virtually.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2021 Citation: Moyer, M. 2021. FRAME-ing Grape Powdery Mildew Management for the West Coast. Syngenta Crop Consultant Summit. Virtually.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2021 Citation: Lowder, S. 2021. Fits like a glove: A new method to monitor grape powdery mildew and FRAC 11 fungicide resistance. Orchard and Vineyard Supply Grower Meeting. Virtually.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2021 Citation: Mahaffe, W. 2021. New approaches for improving disease scouting. Coastal Viticulture Consultants. Virtually.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Pardini, C. 2020. Facing Fungicide Resistance in Grape Production: A Game Theoretic Approach. AERE at SEA Virtual Meeting. Virtually.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2021 Citation: Warres, B., and Brannen, P. 2021. Fungicide resistance survey of Georgia wine grapes and fungicide efficacy in the presence of a resistant Erysiphe necator population. Georgia Association of Plant Pathologists meeting. Virtually.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2021 Citation: Warres, B., and Brannen, P. 2021. Survey of QoI fungicide resistance in Erysiphe necator populations in Georgia. Southern Fruit Workers meeting. Virtually.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2021 Citation: Brannen, P. 2021. Downy and Powdery mildew resistance management update. Georgia Wine Producers Conference. Virtually.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2021 Citation: Espinola-Arredondo, A. 2021. Fungicide Resistance and Misinformation: A Game Theoretic Approach. Montreal Workshop on Resource and Environmental Economics. Virtually.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2021 Citation: Moyer, M. 2021. Alternative Approaches for Powdery Mildew Management: Fungicide Resistsance Update. Washington Advancements in Viticulture and Enology (WAVE). Virtually.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2021 Citation: Aoun N. 2021. Monitoring for SDHI resistance in field isolates of Erysiphe necator. Oregon Wine Research Institute Spring Webinar. Virtually.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2021 Citation: Lowder, S. 2021. Fits like a glove: Monitoring powdery mildew and what that can tell us about QoI resistance. Oregon Wine Research Institute Spring Webinar. Virtually.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2021 Citation: Lowder, S., Neill, T., Miles, T., Moyer, M., Ding, S., Oliver, C., Butler, C., Walgenbach, P., and Mahaffee, W. 2021. Spatial and temporal trends in G143A caused QoI resistance in Erysiphe necator across the growing season in the Western US. American Pathological Society - Pacific Division. Virtually.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2021 Citation: Newbold, C., and Mahaffee, W. 2021. Fitness difference in Quinilone outside inhibitor fungicide resistant Erysiphe necator. American Pathological Society - Pacific Division. Virtually.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2021 Citation: Renault, M., Pardyjak, E., and Stoll, R. 2021. Fast-Response, High-Resolution Flow Solver for Vegetation Canopies. 34th Conference on Agricultural and Forest Meteorology/5th Conference on Biogeosciences. Virtually.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2021 Citation: Ulmer, L., Miller, M., Bailey, B., Mahaffee, W., Pardyjak, E., and Stoll., R. 2021. A Semi-Empirical Wind-Resolving Model for Predicting Transport Processes in Row-Organized Agricultural Canopies. 34th Conference on Agricultural and Forest Meteorology/5th Conference on Biogeosciences. Virtually.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2021 Citation: Lowder, S. 2021. Early Powdery Mildew Detection and Fungicide Resistance. Orchard and Vineyard Supply Grower Meeting. Virtually.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2021 Citation: DeLong, J., Naegele, R., Saito, S., Wang, F., and Xiao, C. 2021. Grape powdery mildew spray programs affect non-target fungi populations differentially in California vineyards. American Phytopathological Society. Virtually.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2021 Citation: Oliver, C. 2021. Using Historical Fungicide Records to Evaluate Use Patterns. American Phytopathological Society. Virtually.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2021 Citation: Mahaffe, W. 2021. Mission Impossible? Inoculum monitoring as a decision aide. University of Maryland. Virtually.
  • Type: Other Status: Published Year Published: 2021 Citation: Lowder, S., and Warres, B. 2021. Early-season scouting for grape powdery mildew. Good Fruit Grower. https://www.goodfruit.com/early-season-scouting-for-grape-powdery-mildew/ (trade article)
  • Type: Conference Papers and Presentations Status: Published Year Published: 2021 Citation: Deford, L., Stoll, R., and Pardyjak, E. 2021. Development of a Low-Cost Aspirated Temperature Measurement Radiation Shield. 34th Conference on Agricultural and Forest Meteorology/5th Conference on Biogeosciences. Virtually.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2021 Citation: Oliver, C. 2021. Understanding Fungicide Use Patterns in Vineyards through Historical Use Records. American Society for Enology and Viticulture. Virtually.
  • Type: Other Status: Published Year Published: 2021 Citation: Oliver, C.2021. New FRAME networks Resources for Growers. Washington State University Viticulture and Enology Extension News - Spring 2021. http://s3-us-west-2.amazonaws.com/sites.cahnrs.wsu.edu/wp-content/uploads/sites/66/2021/04/15161804/2021-Spring-VEEN-FINAL.pdf (newsletter)
  • Type: Other Status: Published Year Published: 2021 Citation: Lowder, S., and Mahaffee, W. 2021. Hands on information: Updates on rapid sampling techniques for grape powdery mildew. Oregon Wine Research Institute Technical Newsletter - Spring 2021. (newsletter)


Progress 09/01/19 to 08/31/20

Outputs
Target Audience:The target audience for this project is broad and includes: grape growers, vineyards managers, vineyard interns, vineyard technicians, vineyard laborers, professional crop consultants, Extension professionals, plant health diagnostic clinics, chemical manufacturing companies, chemical distribution companies, industry organizations, undergraduate students, graduate students, and fellow researchers in the fields of fungicide resistance and pathogen dissemination. We are reaching these audiences through one-on-one recommendations and interactions, newsletters, trade publications, regional industry meetings, and academic conferences. Outreach presentations have been conducted in 10 states across the country. FRAME-specific fungicide spray program full day workshops have been provided in 4 states on both coasts. Rapid powdery mildew sampling kits to screen for fungicide resistance have been made available to growers and consultants, along with kits designed to assist growers in determining spray coverage. Kits for fungicide resistance molecular screening assay validation have been created and disseminated to participating labs in 3 states. The FRAME scientific team has also been regularly presenting through scientific venues, including routine publication of research results, along with presentations and posters at scientific conferences such as the American Phytopathological Society annual meeting. Changes/Problems:We continued to have personnel challenges associated with slow hiring processes through the USDA, but many of these were alleviated by the end of Year 2. However, at that time, challenges related to University hiring (freezing of new hires due to financial challenges brought about by COVID-19), arose. While we continue to make excellent progress despite these hiring challenges, they have impacted our ability to spend in salary and wage allocations. Actual field activities (which is a signficant component of this project) were slowed due to COVID-19, but not stopped, so work efforts have proceeded at a similar-to-normal pace. Project Meetings (both routine and annual) will likely move forward on an exclusively digital basis until a more consistent national return-to-work approach has been acheived. The Extension team will also be re-considering education and outreach efforts, to determine how to best deliver those in durable forms that don't require face-to-face gatherings (where those gatherings are restricted). Many stakeholder winter meetings have already gone to ditigal platforms, which will likely provide a positive opporutnity to "interact" with more groups than was possible when travel was required. What opportunities for training and professional development has the project provided?Members of the Extension team have conducted multiple workshops and training sessions across the country since June 2019 (the end of the previous reporting timeline). This includes 2 GA extension agent trainings, one-on-one training sessions for growers in multiple states, 3 FRAME-Specific fungicide resistance educational workshops, 41 grower presentations, and multiple posters at grower meetings. We have also hosted Round-table discussion meetings in California, 3 wine grape spray workshops, and multiple open houses and field days in late summer / early fall of 2019. There are multiple undergraduate and graduate students who are currently being trained on this project (6 graduate students, and several undergraduate lab rotations). These students have participated in developing newsletter articles and are also currently working on a popular press article. They have also given departmental seminars (2), as well as participated in the FRAME resistance training workshops as either participants or working directly with growers on the activities. Graduate students have also presented posters at the American Phytopathological Society in August 2019 and August 2020. How have the results been disseminated to communities of interest?We have delivered 50 presentations, 4 FRAME fungicide workshops, 2 original videos (one on fungicide resistance, one on spore trap assembly), blog posts, email blasts, and direct interactions. In addition, when growers or consultants submit samples to one or more of our various participating laboratories for fungicide resistance genetic testing, they receive customized letters that include their sample results, along with resistance mitigation action plans. We also routinely post content to our project website at: framenetworks.wsu.edu, as well as our social media accounts (Twitter - @FRAMEnetworks, Facebook - https://www.facebook.com/FRAMEnetworks). What do you plan to do during the next reporting period to accomplish the goals?Objective 1: Extension and Outreach Obj. 1 - Act. 1: Pre- and post-project surveys to track short-term learning, knowledge, and skills changes in grape growers across the United States.Plan- Submit pre-project survey for publication; begin developing post-project survey questions. Obj. 1 - Act. 2: In-the-field demonstrations of management recommendations. Plan- Consider alternative means of field demonstrations. Obj. 1 - Act. 3: Develop a "University Service Center" business plan for fungicide resistance testing. Plan- Begin compilation of protocols and business plans based on the experiences from the 2020 WSU IAREC lab setup with protocols and budgets from two external labs. Obj. 1 - Act. 4:Develop workshops and other classic extension tools; create and engage with enhanced local and regional industry groups.Plan- Continue regional workshops (digitally), write extension articles, and podcasts. Develop accessible mapping tool to report current and historical survey results for all participating growing regions. Objective 2: Group Decisions Obj. 2 - Act. 1:Conduct interviews to identify the externalities negotiating parties are experiencing.Plan- All surveys have been distributed. Our objectives for the Fall 2020 are: (1) analyze data, (2) literature review and (3) prepare a paper. Obj. 2 - Act. 2: Use game theory to develop compensating mechanisms that induce cooperation and aid fungicide stewardship by inducing cooperative behavior among growers, crop consultants and chemical companies. Plan- Currently working on signaling game that characterizes the lack of information growers face about the severity of fungicide resistance. We consider a context in which two grapes growers have already decided to cooperate, however, one of them has more information. We aim to evaluate their strategic behavior and identify under which cases growers will deviate from cooperation. Objective 3: Detection and Monitoring Sub-Obj.3.1 - Develop Molecular Markers to Improve the Speed of Resistance Detection. Obj. 3.1 - Act. 1:Development of PCR based diagnostic assays.Plan- Complete FRAC 11 detection manuscript and submit for peer reviewed publication. Continue to optimize isothermal assays for sensitivity and specificity and conduct technology transfer experiments with other partner laboratories. Obj. 3.1 - Act. 2: Developing isothermal diagnostic tools for in-field detection of fungicide resistance. Plan- Finish analysis of results from the separate labs and complete the paper for submission in Spring 2021. Obj. 3.1 - Activity 3:Target-site gene sequencing to diagnose DMI, QoI, and SDHI resistance.Plan- Continue monitoring incidence and dynamics of mutations in the FRAC 7 - genes in powdery mildew from CA, OR, WA, and MI vineyards. Determine the effect of mutations tolerance in E. necator. Develop diagnostic assays for some of the mutations. We will continue to perform target-site gene sequencing as this is currently more cost effective and reduces the response time needed for obtaining accurate results. A multiplex PCR assay is being developed to allow us to amplify 3 SDH genes in a single PCR reaction. Isolates will be collected from vineyards in which the mutations were identified and tested for tolerance to SDHIs using leaf-disc toxicity assays. For mutations that increase the tolerance of the fungus against SDHIs, TaqMan and other PCR-based diagnostic assays will be developed and tested for specificity and sensitivity. High resolution melting analysis will be pursued to simultaneously detect multiple mutations. Sub-Obj. 3.2 - Monitoring for Fungicide Resistance. Obj. 3.2 - Act. 1:Assess the sampling density and pattern needed to estimate proportion of E. necator with fungicide resistance.Plan- In CA, vineyards will be monitored for fungicide resistance by surveying powdery mildew. In OR, monitoring of E. necator and fungicide resistance will be done by deploying 25 spore traps. In OR, MI, GA, and WA, vineyards will be sampled using leaf, glove, and isolate sampling for method validation. Sampling will collaborate with a large consulting group to cover more vineyards on the west coast. Obj. 3.2 - Act. 2:Phenotype for fungicide sensitivity.Plan- We will collaborate with growers that quit using FRAC 3 and 11 fungicides in 2016 no longer have detectable FRAC 11 resistance populations to see if these fungicides can be re-introduced. We continue to bioassay individual isolates to confirm molecular testing and identify new resistant phenotypes. We will optimize rapid testing technique and increase the number of live cultures tested for fungicide sensitivity. Sub-Obj. 3.3 - Understanding the Basis of Resistance. Obj. 3.3 - Act. 1: Elucidate mechanisms of resistance development by experimentally evolving populations with resistance to SDHI and AzN fungicides. Plan- See Obj. 3.3 Act. 3. Obj. 3.3 - Act. 2:Create reference genome ofE. necatorfor comparative genomic and transcriptomic approaches to identifying fungicide resistance mechanisms.Plan- Annotations of E. necator genome will be completed utilizing PacBio's Isoform sequencing method to capture full-length transcripts. Using the methods developed during 2020, 5-6 more isolates will be sequenced to provide additional genetic references to capture the variation between sensitive and resistant isolates of E. necator. Obj. 3.3 - Act. 3: Identification of genomic markers associated with fungicide resistance through comparative genome and transcriptome analysis. Plan- The new genome assembly of E. necator will be used to search for sources of SDHI resistance. This will be primarily done through (i) identification of differentially expressed genes (DEGs) resulting from exposure to different SDHIs compounds and doses, and (ii) characterization of genomic structural variants (SVs). RNAseq reads will be mapped to our reference E. necator genome assembly with HISAT2 and DEGs will be identified with DESeq2. To identify SVs, 3 to 5 additional E. necator strains showing different patterns of SDHI resistance will be sequenced and assembled at near chromosome level. Sequencing will be mainly performed at 100x depth coverage on the PacBio Sequel II system. In addition, Illumina data will be generated to correct single base errors and small INDELs in the assemblies. The genomes assembled with Canu and whole-genome alignments with NUCmer. Isolates created will be saved for use 1) in future fitness costs experiments, 2) validation of molecular, and 3) as a correlative tool to see if resistant isolates belong to specific sub populations of E. necator. Objective 4: Predicting Resistance Spread Sub-Obj. 4.1:Conduct seasonal monitoring and recurring collection of isolates from selected grower vineyards for AzN, DMI, QoI and SDHI resistance and genetic diversity. Obj. 4.1 - Act. 1:Sample collections and phenotyping. Plan- Collection of 2020 field samples from glove swabs and trap plants will be completed. Pooled isolates will be barcoded and submitted for sequencing. Obj. 4.1 - Act. 2: Genome sequencing and allelic diversity analyses. Plan- Primers and probes will be finalized with the completion of the genome sequences in Spring 2021. Sub-Obj. 4.2 - Dispersion Tracking and Population Prediction. Obj. 4.2 - Act. 1:Mapping resistant isolate spread risk. Plan- Wrap up the code clean-up and final code notation in Winter 2020. Obj. 4.2 - Act. 2: System validation and evaluation. Plan- Will use the validated and accelerated dispersion modeling system in an inverse mode with both idealized and realistic vineyard geometries and distributions of E. necator. Will enable us to understand how data collected by FRAME objectives links to the distribution of E. necator. Sub-Obj. 4.3 - Fungicide Record Evaluation. Plan- Will continue to collect fungicide spray records from participating states.

Impacts
What was accomplished under these goals? Overall- Project meeting held 11 - 12 Mar, 2020, in Napa, CA. Participants included: PIs and Key Personnel, Stakeholder Advisory group, and several technicians and graduate students. Objective Teams host regular (weekly to quarterly) web conferences. Objective 1: Extension and Outreach Obj. 1 - Act. 1: Pre- and post-project surveys to track short-term learning, knowledge, and skills changes in grape growers across the United States.Accomplishments - Completed data analysis and drafted paper for publication. Obj. 1 - Act. 2: In-the-field demonstrations of management recommendations. Accomplishments - Due to COVID-19, in-field demonstrations were reduced. A FRAC 3 and 11 fungicide trial was established in GA and sentinel blocks for disease progress and observation were established in OH. In WA, monitoring fungicide trials for FRAC 11 resistance continues. Obj. 1 - Act 3: Develop a "University Service Center" business plan for fungicide resistance testing. Accomplishments - Began collection of budgets, and "dry run" testing facility establishment (purchasing supplies, assay validation, etc.). Obj. 1 - Activity 4:Develop workshops and other classic extension tools; create and engage with enhanced local and regional industry groups.Accomplishments - Delivered 50 FRAME presentations since Jun 2019 (Jun-Aug 2019 not included in previous report). Cumulative attendance of 2,800+ individuals. Audiences included: industry and scientific community, with 42 of the 50 presentations targeted to industry. FRAME fungicide spray program workshop was presented in 4 states (WA, OR, CA, and OH). There were 12 student/tech posters, 3 blog posts, 3 trade articles, 6 newsletters, 2 videos, and a podcast since June 2019. Objective 2: Group Decisions Obj. 2 - Act. 1:Conduct interviews to identify the externalities negotiating parties are experiencing.Accomplishments - Finished data collection from 3 surveys. Currently developing a paper that focuses on the survey results. Developed a second paper titled "Facing Fungicide Resistance in Grape Production: A Game Theoretic Approach." This provides insight into behavior of grape growers when their choices generate a negative intertemporal production externality in the form of fungicide resistance. When growers encounter this externality, the noncooperative fungicide level is higher than the socially optimal level. We examine a compensation mechanism to ameliorate fungicide resistance and find that it induces the socially optimal level; however, misinformation about severity of the fungicide resistance generates distortions. Information available to growers about fungicide resistance is essential for its mitigation with the proposed compensation mechanism. If the misinformed grower considers fungicide resistance to be relatively mild, then the misinformed grower has the compensating role. Objective 3: Detection and Monitoring Sub-Obj. 3.1 - Develop Molecular Markers to Improve the Speed of Resistance Detection. Obj. 3.1 - Act. 1:Development of PCR based diagnostic assays. Accomplishments - A manuscript was published on FRAC 11 molecular diagnostics. Obj. 3.1 - Act. 2: Developing isothermal diagnostic tools for in-field detection of fungicide resistance. Accomplishments - New rapid isothermal tool (LAMP) was developed for FRAC 11 resistance detection. Assay was validated in external labs. During the annual meeting, stakeholders advised that new diagnostics should focus traditional assays. Obj. 3.1 - Act. 3:Target-site gene sequencing to diagnose DMI, QoI, and SDHI resistance. Accomplishments - Collected samples from vineyards with FRAC 7 and 13 resistance to establish living lab cultures. Sub-Obj. 3.2 - Monitoring for Fungicide Resistance. Obj. 3.2 - Act. 1:Assess the sampling density and pattern needed to estimate proportion of E. necator with fungicide resistance. Accomplishments - Repeated collections in CA, GA, MI, OR, and WA. Demonstrated that gloves swab sampling was more sensitive and cost effective than visual scouting. Also was as effective as spore samplers for monitoring for disease. Processed >4000 swabs samples for FRAC 11 and 3 resistance markers. MI processed ~700 samples from multiple locations. Obj. 3.2 - Act. 2:Phenotype for fungicide sensitivity.Accomplishments - Identified pathogen isolates resistant to FRAC 7 and 13 fungicides. Found >14 isolates with FRAC 7 resistance and 4 with FRAC 13 resistance. Identified 100+ isolates with FRAC 11resistance and 48 with FRAC 3 tolerance. Sub-Objective 3.3 - Understanding the Basis of Resistance. Obj. 3.3 - Act. 1: Elucidate mechanisms of resistance development by experimentally evolving populations with resistance to SDHI and AzN fungicides. Accomplishments - Naturally-occurring isolates were found, so this activity is not needed. Obj. 3.3 - Act. 2:Create reference genome ofE. necatorfor comparative genomic and transcriptomic approaches to identifying fungicide resistance mechanisms. Accomplishments - Further refined large-scale culturing of E. necator. Produced enough DNA for sequencing of 2 more E. necator isolates. Obtained a chromosome-scale genome assembly for E. necator. Among the assembled scaffolds, 11 are complete chromosomes that contain telomeric repeats at both ends. This new genome is a considerable improvement over the 2014 assembly - now at 7,570 predicted genes with 97% completeness. Developed transcriptomes of 6 E. necator isolates to refine the genome annotation. Obj. 3.3 - Act. 3: Identification of genomic markers associated with fungicide resistance through comparative genome and transcriptome analysis. Accomplishments - Transcriptomics studies are underway for determining FRAC 13 resistance. In total this project has collected over 1600 Gbp of RNAseq data. These experiments investigate resistant and sensitive isolates to quinoxyfen (FRAC 13) at various timepoints and conditions, in order to develop a resource for understanding the expression regulation of E. necator. For FRAC 7, we continued USA monitoring and nd identified new SDHI target-site mutations. Target-site mutations were identified in 9 of 12 states, with the majority of strains with such mutations originating from CA and MI. The frequency of FRAC 7 target-site mutations almost doubled in 2019 as compared to 2018, although there are were no reports of reduced field efficacy. Objective 4: Predicting Resistance Spread Sub-Obj. 4.1:Conduct seasonal monitoring and recurring collection of isolates from selected grower vineyards for AzN, DMI, QoI and SDHI resistance and genetic diversity. Obj. 4.1 - Act. 1:Sample collections and phenotyping. Accomplishments - Field collections continued. Probes developed for specific powdery mildew genes to facilitate exome-based pooled sequencing. Obj. 4.1 - Act. 2: Genome sequencing and allelic diversity analyses. Accomplishments - Genome sequencing is still underway. Preliminary identification of primers and probes has begun with the genome annotations provided from Obj. 3. Sub-Obj. 4.2 - Dispersion Tracking and Population Prediction. Obj. 4.2 - Act. 1:Mapping resistant isolate spread risk. Accomplishments - Modernized dispersion modeling numerical code base, enabling the use of graphics processing unit (GPU) technology. This has resulted in a greater than 100x reduction in model execution time for our E. necator spore dispersion model. Obj. 4.2 - Act. 2: System validation and evaluation. Accomplishments - Finalized the physics of vineyard dispersion model by performing a rigorous validation against existing vineyard dispersion data. Journal articles for both advancements will be submitted for publication soon. Sub-Obj 4.3 - Fungicide Record Evaluation. Accomplishments - Collected records from 75 CA, 2 GA, 1 OH, 3 MI, and 13 WA vineyards with a history of QoI resistance testing. Records needed extensive quality control.

Publications

  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Oliver C. 2019. Fungicide Stewardship - Best Practices. FRAME Extension Meeting. Salem, OR
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Wong, A. 2019. Powdery Mildew Biology. FRAME Extension Meeting. Salem, OR
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Lowder S. 2019. Rapid sampling techniques to monitor fungicide resistant grape powdery mildew. Mendicino County Grape IPM Conference. Hopland, CA
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Miles T. 2020. Detecting and managing Plant pathogens in Michigan vineyards. Orchard and Vineyard Show. Traverse City, MI
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Warres, B., and Breeden, S. 2020. Wine Grape Powdery and Downy Mildew Resistance Management. Georgia Wine Growers Association. Braselton, GA
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Miles T. 2020. Detecting and managing Plant pathogens in Michigan vineyards. Southwest Michigan Horticultural Days. Benton Harbor, MI
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Mahaffee W. 2020. Recognizing, monitoring, and mitigating fungicide resistance. D&M Chem Inc. Annual Grower Meeting. Moxee, WA
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Lowder S. 2020. Powdery Mildew Management: Monitoring Fungicide Resistance. Oregon Wine Symposium. Portland, OR
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Stergiopoulos I. 2020. Fungicide Resistance in Grape Powdery Mildew Across the Western US; What Do We Know So Far. Current Wine and Winegrape Research Course, UC Davis Extension . Davis, CA
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Lewis-Ivey, M. 2020. Best practices for utilizing fungicides to prevent fungicide resistance development. Ohio State University, Pesticide Safety Education Program. Akron, OH
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Zaccaron A. 2020. A chromosome-level assembly of the grape powdery mildew fungus Erysiphe necator. 2019-2020 Host-Microbe Interactions Meeting. Davis, CA
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Lewis-Ivey, M. 2020. Fungicide Stewardship  Best Practices. FRAME Extension Meeting. Dublin, OH
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Moyer M. 2020. Powdery Mildew Biology and Management . FRAME Extension Meeting. Dublin, OH
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Moyer M. 2020. Best Sprayer Practices. FRAME Extension Meeting. Dublin, OH
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Mahaffee W. 2020. Needles in haystacks: Inoculum monitoring as a decision aid. University of Florida, Dept. Plant Pathology. Gainesville, FL
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Moyer M. 2020. The Past, Present and Future of Grape Pest Management. Ohio Grape Growers Conference. Dublin, OH
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Brannen P. 2020. Fungicide and Insecticide Spray Program Workshop for Extension Agents. Georgia Extension Service. Blairsville, GA
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Moyer M. 2020. Powdery Mildew Biology and Management . FRAME Extension Meeting. Prosser, WA
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Oliver C. 2020. Fungicide Stewardship and Application Practices. FRAME Extension Meeting. Prosser, WA
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Moyer M. 2020. Powdery Mildew: FRAMing the Reality of Fungicide Resistance. Business Enology Viticulture (B.E.V.) New York Annual Meeting. Rochester, NY
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Brannen P. 2020. West Georgia Wine Grape Spray Program Workshop. Carrol County Extension Service. Carrollton, GA
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Brannen P. 2020. Northwest Georgia Wine Grape Spray Program Workshop. Lumpkin County Extension Service. Dahlonega, GA
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Lewis-Ivey, M. 2020. QoI resistance in Ohio vineyards and management strategies to mitigate resistance development and the spread of resistance between vineyards. Ohio Winter Grape School. Madison, OH
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Miles T. 2020. Fungicide resistance research and mitigation strategies for grape powdery mildew . FRAME Extension Meeting. Napa, CA
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Moyer M. 2020. What makes for a challenging powdery mildew season? Environmental and cultural factors. FRAME Extension Meeting. Napa, CA
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Oliver C. 2020. Decision-support tools for fungicide resistance management. FRAME Extension Meeting. Napa, CA
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Brannen P. 2020. North Georgia Wine Grape Spray Program Workshop. Fannin County Extension Service. Ellijay, GA
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Sharma, N. 2020. Prebloom meeting on early season vineyard management and the importance of fungicide resistance. MSU Prebloom series. MI, (ZOOM)
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Moyer M. 2020. Grape Disease Management  Powdery Mildew & Botrytis Bunch . Wine Island Growers Association. British Columbia (ZOOM)
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Moyer M. 2020. Fungicide Resistance and the Acronyms. Pennsylvania Wine and Grape Team. Pennsylvania (ZOOM)
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Pardini, C. 2020. Facing fungicide resistance in grape production: a game theoretic approach. AAEA Virtual Meeting. Virtually
  • Type: Journal Articles Status: Awaiting Publication Year Published: 2020 Citation: Miles, T.D., Neill, T.M., Colle, M., Warneke, B, Robinson, G., Sterigiopoulus, I. and Mahaffee, W.F.(2020). Allele-specific detection methods for QoI fungicide resistant Erysiphe necator in vineyards. Plant Disease. https://doi.org/10.1094/PDIS-11-19-2395-RE In press
  • Type: Other Status: Published Year Published: 2019 Citation: Oliver C. 2019. FRAME Network: Fungicide Resistance in Wine Grapes. Washington State Grape Society. Grandview, WA (poster)
  • Type: Other Status: Published Year Published: 2019 Citation: Alzohairy, S., Sharma, N., Colle, M., Gillett, J., Sysak, R., Miles, T. 2019. 2019 update on fungicide resistance in Michigan vineyards. Great Lakes Fruit and Vegetable Expo. Grand Rapids, MI (poster)
  • Type: Other Status: Published Year Published: 2020 Citation: Oliver C. 2020. FRAME Network: Fungicide Resistance in Wine Grapes. Washington Association of Winegrape Growers. Kennewick, WA (poster)
  • Type: Other Status: Published Year Published: 2020 Citation: Oliver C. 2020. Grower Knowledge and Perception of Fungicide Resistance in US Vineyards. American Phytopathological Society. Virtually (poster)
  • Type: Other Status: Published Year Published: 2020 Citation: Warres, B., Neill, T., Mahaffee, W., and Brannen, P. 2020. Field efficacy of fungicides for management of grape powdery mildew in the presence of fungicide - resistant populations of Erysiphe necator in Georgia. American Phytopathological Society. Virtually (poster)
  • Type: Other Status: Published Year Published: 2020 Citation: Wong, A., and Mahaffee, W. 2020. A High Throughput Bioassay to CharacterizeErysiphe necator Tolerance to Multiple Fungicide Classes. American Phytopathological Society. Virtually (poster)
  • Type: Other Status: Published Year Published: 2020 Citation: Newbold, C., and Mahaffee, W. 2020. Phenotypic variability in the grape powdery mildew pathogen, Erysiphe necator. American Phytopathological Society. Virtually (poster)
  • Type: Other Status: Published Year Published: 2020 Citation: Lowder, S., and Mahaffee, W. 2020. Fits like a glove: Rapid, cost-effective sampling techniques to monitor Erysiphe necator. American Phytopathological Society. Virtually (poster)
  • Type: Other Status: Published Year Published: 2019 Citation: Moyer M., M. Cooper, P. Brannen, W. Mahaffee, M. Lewis-Ivey, T. Miles, and C. Oliver. 2019. Good to Know: Why Some Seasons are Worse for Powdery Mildew. Good Fruit Grower . https://www.goodfruit.com/why-some-seasons-are-worse-for-powdery-mildew/ (trade magazine)
  • Type: Other Status: Published Year Published: 2020 Citation: Miles T. 2020. Fungicide resistance a growing concern: Michigan grape growers must take steps to manage powdery mildew, botrytis bunch rot. Good Fruit Grower . https://www.goodfruit.com/fungicide-resistance-a-growing-concern/ (trade magazine)
  • Type: Other Status: Published Year Published: 2020 Citation: Lewis-Ivey, M. 2020. Managing Fungicide Resistance in the Vineyard. American Vineyard. https://americanvineyardmagazine.com/managing-fungicide-resistance-in-the-vineyard/ (trade magazine)
  • Type: Other Status: Published Year Published: 2020 Citation: Lewis-Ivey, M.2020. Managing Fungicide Resistance in the Vineyard. Ohio Grape Newsletter . https://ohiograpeweb.cfaes.ohio-state.edu/sites/grapeweb/files/imce/pdf_newsletters/March%202020%20OGEN.pdf (newsletter)
  • Type: Other Status: Published Year Published: 2020 Citation: Warres, B., and Brannen, P. 2020. Widespread QoI resistance and emergence of DMI resistance in powdery mildew in Georgia. Integrated Pest Management Newsletter. https://site.extension.uga.edu/ipm/2020/04/08/widespread-qoi-resistance-and-emergence-of-dmi-resistance-in-powdery-mildew-in-georgia/ (newsletter)
  • Type: Other Status: Published Year Published: 2020 Citation: Warres, B., and Brannen, P. 2020. Widespread QoI resistance and emergence of DMI resistance in powdery mildew in Georgia. Small Fruit News. https://smallfruits.org/2020/04/widespread-qoi-resistance-and-emergence-of-dmi-resistance-in-powdery-mildew-in-georgia/ (newsletter)
  • Type: Other Status: Published Year Published: 2020 Citation: Sharma, N., and Miles, T.2020. Grapevine powdery mildew fungicide resistance survey. MSU Extension News. https://www.canr.msu.edu/news/grapevine-powdery-mildew-fungicide-resistance-survey (newsletter)
  • Type: Other Status: Published Year Published: 2020 Citation: Tekip, A., and Miles, T.2020. Detecting emerging pesticide resistance in grapes. MSU AgBio Research. https://www.canr.msu.edu/news/detecting-emerging-pesticide-resistance-in-grapes (newsletter)
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Brannen P. 2019. Update on fungicide resistance in downy and powdery mildew diseases of grape in Georgia. Southeastern Professional Fruitworkers Conference. Tifton, GA
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Moyer M. 2019. The Past, Present, and Future of Viticulture Research and Extension at Washington State University. Washington State University - IAREC Centennial Celebration. Prosser, WA
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Stergiopoulos I. 2019. New developments in managing powdery mildew resistance. 8th Annual Vineyards & Wineries Continuing Education Class Series. Napa, CA
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Mahaffee W. 2019. Finding Needles in haystacks: Disease monitoring and risk assessment. National Grape Research Alliance. Sacramento, CA
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Mahaffee W. 2019. Needles in haystacks; Monitoring and mitigating fungicide resistance. University of California - Davis, Department Plant Pathology. Davis, CA
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Mahaffee W. 2019. Tracking fungicide resistant grape powdery mildew. Northwest Small Fruit Center Annual Conference. Ferndale, WA
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Stergiopoulos I. 2019. Fungicide Resistance in grape powdery mildew across the Western US; What do we know so far?. FPS Grape Advisory Committee Meeting. Davis, CA
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Miles T. 2019. Fungicide Resistance management in grapes: powdery mildew and botrytis. Great Lakes Fruit and Vegetable Expo. Grand Rapids, MI
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Lowder S. 2019. Grape Powdery Mildew Management: Mildew's and Mildon'ts. FRAME Extension Meeting. Salem, OR
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Moyer M. 2019. Best Sprayer Practices. FRAME Extension Meeting. Salem, OR


Progress 09/01/18 to 08/31/19

Outputs
Target Audience:The target audience for this project is broad and includes: grape growers, vineyards managers, vineyard interns, vineyard technicians, vineyard laborers, professional crop consultants, Extension professionals, plant health diagnostic clinics, chemical manufacturing companies, chemical distribution companies, industry organizations, undergraduate students, graduate students, and fellow researchers in the fields of viticulture, integrated pest management, plant pathology, fungicide resistance and disease epidemiology. We are reaching these audiences through surveys, on-farm and in-meeting demonstrations, workshops, and industry and extension publications (trade magazines, email blasts, social media posts, websites, and peer-reviewed extension publications).Rapid powdery mildew-sampling kits to screen for fungicide resistance have been made available to growers and consultants for the 2019 growing season. Sprayer coverage kits have been developed (in English and Spanish) to begin with initial training on what on-farm practices can lead to the selection of resistance, and how to correct for those actions. Nationwide surveys have been deployed to assess the general knowledge of growers, managers, and consultants on the presence of fungicide resistance; the responses from these surveys will be used to tailor and develop future extension and outreach efforts to target these specific audiences in the different geographical and cultural regions of the United States. Changes/Problems:We currently have no major changes within the scope of this project after the first 9 months. We did have a few delays in project hiring (postdoctoral scientists and graduate students). These were related to institutional delays in funding dispersement and account set-ups (most universities will not allow hiring of staff until accounts are fully established, which can take months after initial fund receival). The government shutdown over Dec 2018 - Feb 2019 significantly impacted two of our CoPIs during prime hiring time for students and postdocs related to project activities. We have since overcome these staffing delays, and have hired, or are in the process of hiring, the remaining budgeted staff and students associated with the project. What opportunities for training and professional development has the project provided?We (Extension team) delivered a full-day fungicide program design workshop, with 65 participants in Washington state, which provided additional opportunities for FRAME-associated postdocs and graduate students to participate and delivered pre-designed content, providing opportunities in public speaking and group-work with industry members. Postdocs and students have also engaged growers in OR and WA with specific demonstration workshops on how to properly scout for, and collect, grape powdery mildew. In addition, a YouTube instructional video was also made by a graduate student. Graduate students have also presented several posters and small research-based presentations (including short presentations at our annual project meeting) between September 2018 and May 2019. In addition to our postdoctoral research associates and graduate students, this project is providing training for1 undergraduate student, currently. How have the results been disseminated to communities of interest?Members of the Extension team (including graduate students) have delivered 35 FRAME-related presentations in multiple states since September 2018 with total cumulative attendance of over 1600 individuals. The audience type included both the industry and scientific community, with 31 of the 35 presentations targeted to industry. We have also presented several posters at regional grape conferences, and have had multiple articles shared via our university news feeds and trade magazines such as the Good Fruit Grower. We have also launched our project website at: framenetworks.wsu.edu, which will provide a holding ground for our durable outreach products. Social media accounts, such as Twitter and Facebook pages, have been established to help with the dispersal of materials. Additionally, regional grower targeted products such as blog posts, extension bulletins, and newsletters have been published in CA, GA, OR, MI, and WA. All articles and podcasts are featured on our FRAME project website. What do you plan to do during the next reporting period to accomplish the goals?Objective 1: Extension and Outreach Obj. 1 - Activity 1: Pre- and post-project surveys to track short-term learning, knowledge, and skills changes in grape growers across the United States. Plan of action - We plan on publishing our pre-project survey results in a peer-reviewed journal, highlighting key areas for targeting teaching and engagement. Obj. 1 - Activity 2: In-the-field demonstrations of management recommendations. Plan of action - Coordinated field demonstrations will begin in summer 2019, focusing on fungicide program design that incorporate rotation with non-risk products. After national surveying of the extent of fungicide resistance in 2019, these trials will be expanded in summer 2020 to reflect regional approaches to disease management. Obj. 1 - Activity 3: Develop a "University Service Center" business plan for fungicide resistance testing. Plan of action - No activity until years 3 and 4. Obj. 1 - Activity 4:Develop workshops and other classic extension tools; create and engage with enhanced local and regional industry groups.Plan of action - We will continue to host regional workshops relating to basic powdery mildew management. We plan on writing additional extension articles on the basis of fungicide resistance and will work towards developing short videos and podcasts on basic fungicide resistance education. Objective 2: Group Decisions Obj. 2 - Activity 1:Conduct interviews to identify the externalities negotiating parties are experiencing (i.e., how their pesticide use affects their neighbors' production).Plan of action - All surveys have been distributed. Our objectives for the fall semester are: (1) analyze data, (2) literature review and (3) prepare a paper. Obj. 2 - Activity 2: Use game theory to develop compensating mechanisms that induce cooperation and aid fungicide stewardship by inducing cooperative behavior among growers, crop consultants and chemical companies. Plan of action - No intended activity until after year 2. Objective 3: Detection and Monitoring Sub-Objective 3.1-Develop Molecular Markers to Improve the Speed of Resistance Detection. Obj. 3.1 - Activity 1:Development of PCR based diagnostic assays.Plan of action - Complete FRAC 11 detection manuscript and submit for peer reviewed publication. For SDH mutations that increase the most tolerance of the fungus against SDHIs, TaqMan and other PCR-based diagnostic assays will be developed and tested for specificity and sensitivity. High resolution melting analysis will also be pursued as a method to simultaneously detect multiple mutations in the SDH genes. Obj. 3.1- Activity 2: Developing isothermal diagnostic tools for in-field detection of fungicide resistance. Plan of action -Continue to optimize isothermal assays for sensitivity and specificity and conduct technology transfer experiments with other partner laboratories. Obj. 3.1 - Activity 3:Target-site gene sequencing to diagnose DMI, QoI, and SDHI resistance.Plan of action - We will continue to monitor the incidence and dynamics of SDH gene mutations bytarget-site gene sequencing in collected samples from multiple states,and determine the effect of these mutations on SDHI tolerance.A multiplex PCR assay is being developed that will allow us to amplify the three SDH genes in a single PCR reaction and subsequently sequence them with gene-specific primers. Sub-Objective 3.2-Monitoring for Fungicide Resistance. Obj. 3.2 - Activity 1:Assess the sampling density and pattern needed to estimate proportion of E. necator with fungicide resistance.Plan of action - Field collections are currently under way for the Spring 2019 growing season. In CA, commercial vineyards will be monitored for fungicide resistance by surveying powdery mildew colonies and through a network of fixed, rotorod spore traps. In OR, we are monitoring for E. necator and fungicide resistance by deploying 25 spore traps in the region. In OR, MI, GA, and WA, vineyards will be intensively sampled using leaf, glove, and isolate sampling to examine which method is the efficient for allowing an accurate grower decision. This sampling will be in collaboration with a large consulting group to cover more vineyards on the west coast. Obj. 3.2 - Activity 2:Phenotype for fungicide sensitivity.Plan of action - We will collaborate with 3 growers that quit using DMI and QoI fungicides in 2016 and have lower or no detectable QoI resistance populations to see if QoIs and DMI fungicides can be rotated back into their program. We will continue to bioassay individual isolates to confirm molecular resting and identify new resistant phenotypes. Sub-Objective 3.3-Understanding the Basis of Resistance. Obj. 3.3 - Activity 1: Elucidate mechanisms of resistance development by experimentally evolving populations with resistance to SDHI and AzN fungicides. Plan of action - Populations will start to be developed in year two, with activity focus occuring in year 3 and 4. Obj. 3.3 - Activity 2:Create reference genome ofE. necatorfor comparative genomic and transcriptomic approaches to identifying fungicide resistance mechanisms.Plan of action - A combined assembly will be generated using two draft assemblies produced from DNA long-reads and Hi-C data. The final assembly will be polished and completeness will be assessed using a minimum of five standard metrics, including N50 and L50 measures. If necessary, additional long-read sequences will be pursued by Nanopore sequencing, using theUC Davis genome center. After full assembly, transcripts will be annotated during the winter of 2019, finishing the process and publishing the genome in early 2020. Obj. 3.3 - Activity 3: Identification of genomic markers associated with fungicide resistance through comparative genome and transcriptome analysis.Plan of action - We are currently in the process of hiring another graduate student to continue work on FRAC 7 (SDHI) fungicide resistance. Objective 4: Predicting Resistance Spread Sub-Objective 4.1:Conduct seasonal monitoring and recurring collection of isolates from selected grower vineyards for AzN, DMI, QoI and SDHI resistance and genetic diversity. Obj. 4.1 - Activity 1:Sample collections and phenotyping. Plan of action - A postdoctoral research candidate has been identified to work on the project starting in July, and federal clearance paperwork has been initiated. Sampling from each location will continue through the end of the grape growing season in each location. Obj. 4.1 - Activity 2: Genome sequencing and allelic diversity analyses. Plan of action - Initial population sequencing will begin in year 2, with activity focus in years 3 and 4. Sub-Objective 4.2- Dispersion Tracking and Population Prediction. Obj. 4.2 - Activity 1.1:Mapping resistant isolate spread risk. Plan of action - A postdoctoral research candidate has been identified to work on the project and a formal offer has been extended to the candidate. Construction of model domains for each FRAME test vineyard will commence throughout the summer and downscaled weather forecasts in both a reanalysis and forecast model will be performed and archived. Obj. 4.2 - Activity 1.2: System validation and evaluation. Plan of action - Major activity not planned until after year 2. Sub-Objective 4.3-Fungicide Record Evaluation. Plan of action - Collection of fungicide records will begin during the spring and will continue through the fall 2019. Data analysis of records will begin in winter 2019, with preliminary models comparing use patterns to time of noticeable control failure and resistance detection developed by end of year 2. Model refinement and validation in years 3 and 4.

Impacts
What was accomplished under these goals? At the time of reporting, we are currently only 9 months into the project, and just starting the first in-field season. Not all project goals and objectives were slated for the first year. Updates for what was completed / in progress for Year 1 are below: Overall accomplishments - The first annual project meeting for FRAME was held on 27 Feb - 1 Mar, 2019, in Prosser WA. Participants included all project PIs and Key Personnel,Stakeholder Advisory group, technicians and students. Objective Teams hosted regular web conferences from weekly to quarterly, depending on nature of the objective). The project website (https://framenetworks.wsu.edu/), was developed. Objective 1: Extension and Outreach Obj. 1 - Activity 1: Pre- and post-project surveys to track short-term learning, knowledge, and skills changes in grape growers across the United States.Year 1 accomplishments - Completed our nation-wide pre-project survey, designed to gather regional baseline understanding of fungicide resistance among grape growers. Data analysis to begin in late summer 2019. Obj. 1 - Activity 2: In-the-field demonstrations of management recommendations. This is just starting for 2019. Obj. 1 - Activity 3: Develop a "University Service Center" business plan for fungicide resistance testing. Not scheduled until years 3 and 4. Obj. 1 - Activity 4:Develop workshops and other classic extension tools; create and engage with enhanced local and regional industry groups.Year 1 accomplishments - Members of the Extension team delivered 35 FRAME-related presentations since Sept. 2018 to a cumulative 1600 individuals. The audience type included both the industry and scientific community, with 31 of the 35 presentations targeted to industry. There were several student posters, a pod cast, and several feature articles on the project between September 2018 and May 2019. All articles and podcasts are featured on the FRAME website. Social media accounts have been established to assist in information "push". Objective 2: Group Decisions Obj. 2 - Activity 1:Conduct interviews to identify the externalities negotiating parties are experiencing (i.e., how their pesticide use affects their neighbors' production).Year 1 accomplishments -We developed three surveys which were distributed (Growers Survey, Crop Consultant Survey) and May (Manufacturers survey). The Growers Survey aims to identify the level of knowledge about fungicide resistance among grape growers and their willingness to adjust their own practices if it meant a regional benefit in mitigate crop loss due to fungicide resistance. The Crop Consultant Survey seeks to determine the level of knowledge about fungicide resistance among crop consultant and their willingness to adjust their sales and program designs if it meant a regional improvement in management of fungicide resistance. The Manufacturers Survey will evaluate what changes in production or marketing manufacturers would deem appropriate in light of potential product efficacy loss due to resistance. Obj. 2 - Activity 2: Use game theory to develop compensating mechanisms that induce cooperation and aid fungicide stewardship by inducing cooperative behavior among growers, crop consultants and chemical companies. Not scheduled until after year 2. Objective 3: Detection and Monitoring Sub-Objective 3.1-Develop Molecular Markers to Improve the Speed of Resistance Detection. Obj. 3.1 - Activity 1:Development of PCR-based diagnostic assays.Year 1 accomplishments - An improved rapid sampling technique was developed that reduced sampling kit manufacturing time by 80% and the time to collect sample by 60%, with no effect on sample processing time or sensitivity. We refined the y136F qPCR assay for detecting Y136F mutation associated with FRAC 3 resistance, including correlating Cyp51 copy number in relation to genome copy number (and subsequent expression of FRAC 3 resistance phenotype). Obj. 3.1- Activity 2: Developing isothermal diagnostic tools for in-field detection of fungicide resistance. Year 1 accomplishments - We developed an isothermal molecular diagnostic marker for FRAC 11 resistance, which should allow for field detection of fungicide resistance. Obj. 3.1 - Activity 3:Target-site gene sequencing to diagnose DMI, QoI, and SDHI resistance.Year 1 accomplishments - Genetic libraries were developed to study FRAC 13 resistance and to examine the transcriptome profile of FRAC 13 sensitive and resistant isolates. Libraries are actively being created for examination of SDHI resistance profiles to screen for consistant mutation prevalence and relavence to field resistance. Sub-Objective 3.2-Monitoring for Fungicide Resistance. Obj. 3.2 - Activity 1:Assess the sampling density and pattern needed to estimate proportion of E. necator with fungicide resistance.Year 1 accomplishments - We standardized sampling protocols across collaborators in CA, GA, MI, OR, and WA; it included three experimental sampling proceedures and research plot design. Obj. 3.2 - Activity 2:Phenotype for fungicide sensitivity.Year 1 accomplishments - A germination bioassay for FRAC 3, 9, 11, 13, 50 (formally U8) and U6 fungicides was develop that improved throughput by >400%. Sub-Objective 3.3-Understanding the Basis of Resistance. Obj. 3.3 - Activity 1: Elucidate mechanisms of resistance development by experimentally evolving populations with resistance to SDHI and AzN fungicides. Just starting this field season. Obj. 3.3 - Activity 2:Create reference genome ofE. necatorfor comparative genomic and transcriptomic approaches to identifying fungicide resistance mechanisms.Year 1 accomplishments - We developed an improved procedure for growing E. necator to increase DNA concentrations for Hi-C or PacBio sequencing. We have produced over 20mg of E. necator E1-101 DNA for sequencing, meeting Objective target 9 months ahead of schedule. A current draft of a genome is under construction using 3 separate methods for validation and consensus comparison. Obj. 3.3 - Activity 3: Identification of genomic markers associated with fungicide resistance through comparative genome and transcriptome analysis. Not slated year 2. Objective 4: Predicting Resistance Spread Sub-Objective 4.1:Conduct seasonal monitoring and recurring collection of isolates from selected grower vineyards for AzN, DMI, QoI and SDHI resistance and genetic diversity. Obj. 4.1 - Activity 1:Sample collections and phenotyping. Year 1 accomplishments - Selection of field sampling locations and physical sample collectionsfor the 2019 season (first full growing season) are underway. Obj. 4.1 - Activity 2: Genome sequencing and allelic diversity analyses. Not slated until after year 2. Sub-Objective 4.2- Dispersion Tracking and Population Prediction. Obj. 4.2 - Activity 1.1:Mapping resistant isolate spread risk. Year 1 accomplishments - A model for airborne isolate spread at the vineyard scale was developed and validated prior to the project's start. Further tests downscaling forecasted regional weather to the vineyard scale (~1 km) where performed. These initial tests downscaled forecasts from the North American Mesoscale Forecast System (NAM) at 12 km to 1 km using the Weather Research and Forecasting (WRF) model. Obj. 4.2 - Activity 1.2: System validation and evaluation. Not slated until after year 2. Sub-Objective 4.3-Fungicide Record Evaluation. Year 1 accomplishments - We began soliciting fungicide records from participating growers at the start of the 2019 growing season (April).

Publications

  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Miles, T. (2019). Dormant and early season management including information about "softer chemicals". presented at MI Statewide Pre-Season Kickoff meeting, Benton Harbor, MI.
  • Type: Websites Status: Published Year Published: 2018 Citation: https://framenetworks.wsu.edu/
  • Type: Other Status: Published Year Published: 2019 Citation: Lowder, S. (2019). Rapid sampling techniques to monitor group 11 fungicide resistant grape powdery mildew. Poster presentation to grower groups, Corvallis, OR.
  • Type: Other Status: Published Year Published: 2019 Citation: Newbold, C. (2019). Succinate dehydrogenase inhibitor resistance in isolates of Erysiphe necator from the Western U.S. Poster presentation to grower groups, Corvallis, OR.
  • Type: Other Status: Published Year Published: 2019 Citation: Wong, A. (2019). A High Throughput Bioassay to Rapidly Characterize Erysiphe necator Resistance to Demethylation Inhibitor Fungicides. Poster presentation to grower groups, Corvallis, OR.
  • Type: Other Status: Published Year Published: 2019 Citation: Neill, T. (2019). An International Mildew Management Problem  fungicide resistance from coast to coast. Poster presentation to grower groups, Corvallis, OR.
  • Type: Conference Papers and Presentations Status: Submitted Year Published: 2019 Citation: Colle., M. Neill, T., Mahaffee, W., Miles, T. (2019). Development of a rapid isothermal assays to detect QoI resistance in Erysiphe necator the causal agent of grape powdery mildew. Phytopathology Abstr.
  • Type: Conference Papers and Presentations Status: Submitted Year Published: 2019 Citation: Sharma, N. Neill, T., Mahaffee, W., Miles, T.D. (2019). Development of a rapid isothermal assays to detect QoI resistance in Erysiphe necator the causal agent of grape powdery mildew. Phytopathology Abstr.
  • Type: Journal Articles Status: Other Year Published: 2020 Citation: Colle, M., Neill, T., Warnke, B., Mahaffee, W., Miles, T.D. (201X). Detection of Erysiphe necator fungicide-resistant alleles in leaf and air samples using novel molecular diagnostic techniques. (in preparation, submitting to Plant Disease).
  • Type: Other Status: Published Year Published: 2019 Citation: Hansen, M., and M. Moyer. 2019. Basic Training for Combating Mildew. Wine Business Monthly. March 2019: 84  91. Online: https://www.winebusiness.com/wbm/
  • Type: Other Status: Published Year Published: 2019 Citation: Oliver, C. 2019. Avoiding Selection of Fungicide Resistance. WSU Viticulture and Enology Extension News - Spring 2019. Online: http://wine.wsu.edu/extension/viticulture-enology-news-veen/
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Miles, T. (2019). Grape Disease Management and Program Directions. presented at Southwest Horticulture Days, Benton Harbor, MI.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Miles, T. (2019). How Fungicide Resistance Occurs, and Using Resistance Management Concepts in Mildew Management Programs. presented at FRAME Extension Meeting, Prosser, WA.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Miles, T. (2019). Vineyard Integrated Pest Management: Diseases. presented at MI Statewide MSU Grape School, Benton Harbor, MI.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Mahaffee, W. (2018). Fungicide Resistance in grape production. presented to BASF representatives, Raleigh, NC.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Mahaffee, W. (2018). Fungicide resistance, selection, and timing. presented at the Sustainable Ag Expo, San Luis Obispo, CA.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Mahaffee, W. (2018). Fungicide stewardship. presented to PCA representatives, Woodland, CA.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Mahaffee, W. (2018). Grape powdery mildew management in the era of fungicide resistance. presented to the Growest PCA conference, Woodland, CA.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Mahaffee, W. (2019). What the mildew?. presented at the Lodi Grape Growers Conference, Lodi, CA.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Mahaffee, W. (2019). Fungicide resistance, selection, and timing. presented to Nutrien Crop Consultants, McMinnville, OR.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Mahaffee, W. (2019). Fungicide resistance, selection, and timing. presented to North-Coast CAPCA, Santa Rosa, CA.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Robinson, G. (2019). Monitoring of QoI and SDHI Resistance in Grape Powdery Mildew through Target-Site Mutation Detection  A Critical Update. presented to UC Davis Extension via Current Wine and Winegrape Research Course, Davis, CA.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Stergipopoulos, I. (2019). An update on SDHI resistance monitoring and the (mt) genome of Erysiphe necator. presented to FRAME research team, Prosser, WA.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Moyer, M. (2018). Realizing the Role of People in Grape Disease Management. presented to WSU Dept of Plant Pathology, Prosser, WA.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Moyer, M. (2018). Grape Powdery Mildew Update. presented to Washington State Grape Society, Grandview, WA.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Moyer, M. (2019). Monitoring and Mapping Grape Powdery Mildew Fungicide Resistance and Crown Gall Incidence in Washington Vineyards. presented to Washington State Grape and Wine Research Program, Prosser, WA.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Moyer, M. (2019). Fungicide Management. presented to Washington Winegrowers Association, Kennewick, WA.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Moyer, M. (2019). Managing Mildew and Fungicide Resistance: Back to Basics. presented to Gallo Grape Growers - South Valley, Fresno, CA.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Moyer, M. (2019). Managing Mildew and Fungicide Resistance: Back to Basics. presented to Gallo Grape Growers - North Valley, Lodi, CA.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Moyer, M. (2019). Designing a Mildew Spray Program. presented during FRAME extension meeting, Prosser, WA.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Moyer, M. (2019). Managing Crown Gall and Powdery Mildew Diseases in the Columbia Gorge. presented to WAVEx (Washington State Wine Commission), Dalles, OR.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Moyer, M. (2019). Powdery Mildew Management in Concord - An Update. presented to National Grape Cooperative Growers, Grandview, WA.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Cooper, M. (2018). Powdery Mildew Fungicide Resistance Update. presented at UCCE Lake/Mendocino IPM Conference, Hopland, CA.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Cooper, M. (2018). Strategies for managing fungicide resistance. presented to Napa Regional Grower Group, Oakville, CA.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Cooper, M. (2019). Monitoring fungicide resistance with a spore trap array in Napa County. presented to Napa Regional Grower Group, Oakville, CA.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Brannen, P. (2018). Grape Post Harvest Round Table. presented to Georgia Wine Grape Producers, White, GA.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Brannen, P. (2018). Fungicide Sensitivity Profiling of Grapevine Downy Mildew in Georgia. presented at Southeastern Professional Fruit Workers Conference, Manchester, TN.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Brannen, P. (2019). Powdery and Downy Mildew Resistance Update. presented to Georgia Wine Growers Association, Braselton, GA.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Brannen, P. (2019). Powdery and Downy Mildew Resistance Updates. presented to Georgia Wine Grape IPM Meeting, Elijay, GA.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Lowder, S. (2019). Rapid methods for monitoring fungicide resistance in grape powdery mildew. presented at Raisin and Table Grape Growers annual Conference, Visalia, CA.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Lowder, S., Neill, T., Moyer, M., Miles, T., Stergiopoulous, I., and Mahaffee, W. (2019). Monitoring fungicide resistant grape powdery mildew and the use of micronized sulfur. California Table Grape Commission Conference, Visalia, CA.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Miles, T. (2018). Fungicide resistance and viruses commonly found in Michigan. presented at Great Lakes Fruit and Vegetable Expo, Grand Rapids, MI.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Miles, T. (2019). Detecting viruses and fungicide resistant pathogens in MI vineyards. presented at Orchard and Vineyard Show, Traverse City, MI.