DIVERSITY AND HOST RESISTANCE TO NOVEL POTATO VIRUS Y ISOLATES IN COLORADO

• Sponsoring Institution: National Institute of Food and Agriculture
• Project Status: ACTIVE
• Funding Source: OTHER GRANTS
• Reporting Frequency: Annual
• Accession No.: 1027480
• Grant No.: 2021-70006-35809
• Cumulative Award Amt.: $138,485.00
• Proposal No.: 2021-04881
• Project Start Date: Sep 1, 2021
• Project End Date: Aug 31, 2025
• Grant Year: 2021
• Program Code: [ARDP]- Applied Research and Development Program

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

Performing Department
Agricultural Biology

Non Technical Summary
This is an ARDP research (single function) project. Potato virus Y (PVY) is the most economically-important potato pathogen that reduces yield, tuber quality, and profit margins. In recent years, the prevalence of recombinant PVY strains has increased. These strains spread quickly and also fail to induce foliar symptoms hampering identification during seed certification. Furthermore, no single test can accurately classify the multitude of emerging PVY strains. While virus-resistant varieties are commercially available, their responses to new strains are not known. Knowledge of virus strains occurring in a given area and host genotypes' response is critical for developing and deploying resistant varieties. In the current study, we propose to use cutting-edge third-generation sequencing to sequence PVY strains in Colorado and determine host genotype responses to these novel strains. This research aims to develop tools to improve virus detection and facilitate the development and release of virus-resistant varieties. Specific objectives of the research are:Characterize the diversity, prevalence, and incidence of PVY strainsIdentify host plant resistance to recombinant PVY strainsDevelop effective methods to provide research-based information regarding PVY detection and disease management to potato producersThe research we propose addresses CPPM focus areas (1) "Plant Protection Tools and Tactics" by identifying resistant germplasm and (3) "enhancing Agricultural Biosecurity" by providing growers information on PVY strain diversity. Integrative tactics will promote the National IPM Roadmap goals of improving cost-benefit analyses when adopting IPM practices, reducing potential health risks, and minimizing pests and related management strategies' adverse environmental effects.

Animal Health Component

20%

Research Effort Categories

Basic

70%

Applied

20%

Developmental

10%

Classification

Knowledge Area (KA)	Subject of Investigation (SOI)	Field of Science (FOS)	Percent
212	1310	1101	75%
202	1310	1081	25%

Knowledge Area
212 - Pathogens and Nematodes Affecting Plants; 202 - Plant Genetic Resources;

Subject Of Investigation
1310 - Potato;

Field Of Science
1101 - Virology; 1081 - Breeding;

Keywords

breeding

detection

diversity

extreme resistance

genetic markers

potato virus y

potatoes

Goals / Objectives
The U.S. potato industry and over 330 growers identify PVY as the number one disease problem affecting the seed potato crop (Stewart Grey, personal communication). The commercial crop will be at risk if the tuber necrotic strains become widespread. The emergence of other tuber necrotic viruses such as PMTV and TRV only exacerbates this problem; these are, however, not the focus of the proposal. Controlling tuber necrotic viruses and PVY remains an ongoing challenge, and current methods are not working effectively. Virus diseases cannot be managed using chemical or biological protectants as is often effective for fungal, bacterial, or nematode pathogens. Chemical control of the vectors by insecticides and fungicides is not effective against PVY.Recent data indicates that nearly 40% of the U.S. seed lots had detectable levels of PVY, and more than 20% of all lots exceed the 2% rejection level prescribed by the State National Harmonization Program. Much of the known virus incidence in the seed can be traced to four factors: (1) virus strains that induce mild or transient foliar symptoms, (2) the emergence of novel or "rare" recombinant strains, (3) release of tolerant cultivars that do not express foliar symptoms, and (4) late-season infections that do not manifest symptoms in the foliage, but a high percentage of tubers in some cultivars can still become infected. All these factors lead to low virus incidence observed and reported from summer field inspections conducted by seed certification programs. Still, high tuber infection levels may exist in the seed, requiring breeding programs to grow out plants for an entire season before they can determine if a resistance response is occurring. A post-harvest test of all seed lots is prohibitively expensive and not done in most states. Knowing which cultivars are most at risk for inaccurate visual summer field evaluations and having reliable diagnostics to determine accurate infection levels would allow limited certification resources to be directed where most efficacious. Seed certification agencies have identified the development of training materials and applications that will assist seed certification inspectors and growers in identifying the various foliar and tuber symptoms induced by the various strains of PVY and other viruses predominant in that region as a priority. This need is addressed in this proposal.Virus resistance in potatoes has not been a priority for U.S. breeding programs until recently. Now it is recognized that virus resistance is a desirable trait, and, out of necessity, it has become an important strategy to manage virus incidence and spread. Although resistance genes to PVY have been identified in wild and cultivated potato, only a few provide broad-spectrum resistance to PVY [25; 38]. However, only a few newly released cultivars have been screened against all predominant PVY strains. None have been screened against emerging novel and "rare" recombinant strains to determine their susceptibility or their symptom expression when infected. Growers and breeders need to know which are most susceptible to viruses, emphasizing those susceptible to tuber necrosis and methods to facilitate the development and release of cultivars with levels of resistance useful in virus disease management. These needs are addressed in this proposal.ObjectivesObjective 1. Characterize the diversity, prevalence, and incidence of PVY strains in Colorado.This research-led objective aims to characterize the genetic diversity of PVY in the San Luis Valley.Objective 2. Identify host plant resistance to recombinant PVY strains in the Colorado potato breeding germplasm.The goal of this research-led objective is to screen selections (a combination of early, intermediate, and late generations) in the Colorado Potato Breeding Program (CPBP) for their response to the predominant recombinant strains of PVY in the Valley (identified as part of objective 1)Objective 3. Develop effective methods to provide research-based information regarding PVY detection and disease management to potato growers.The goal of this extension-led objective is to provide resources and training to stakeholders.

Project Methods
Objective 1. Characterize the diversity, prevalence, and incidence of PVY strains in Colorado.Commercial fields and plant sampling: As part of a current proposal funded by the Colorado Potato Administrative Committee (letter attached), we sample ten potato fields in the San Luis Valley (Fig. 5). For the proposed work, we plan on expanding the number of fields by 10 for a total number of 20 fields. The fields will be sampled to determine the prevalence and incidence of PVY. Systematic samples will be collected by walking the fields in a zig-zag pattern. Samples will be collected from 20 randomly chosen plants per field. Individual samples will consist of six to ten leaves from new growth per potato plant. Samples will be collected during the potato growing seasons of 2021 and 2022. Several years (since 2017) of aphid monitoring data indicate that peak aphid flights in the Valley occur at the end of July and the beginning of August. Therefore, all of our samplings will be performed between the third and fourth week of August. By doing so, we will monitor the in-season spread of various PVY strains in the Valley. The proportion of infected plants will be used as a measure of PVY incidence within fields.Sampling from Post-Harvest Tests: PHTs are usually performed from October to February each year. The PHT location is either in greenhouses or in warmer locations such as Hawaii or Florida. To capture many growers, genotypes (varieties), and lots, samples from the 2021 and 2022 PHTs will be included in our analysis. In this instance, samples will be collected from plants showing symptoms ascribable to virus infection (mainly leaf mosaic, but also leaf distortion, rugosity, yellowing and crinkling). Samples (composed of six to seven leaves/plant) will be collected from all plants showing symptoms. The Potato Certification Service staff will perform a sample collection and send it to the P.I.'s lab at CSU. Based on previous PHTs and the Co-P.I.'s experience, we estimate that between 100 - 120 plants will be sampled each year. Besides, seed inspectors from the Potato Certification Service visit seed fields during the summer. Samples will also be collected from symptomatic plants during these field visits and sent to the P.I.'s lab for further analysis.Simultaneous detection and identification of PVY: Immunocapture-reverse transcription-polymerase chain reaction (IC-RT-PCR) is a sensitive, reproducible, and robust method for detecting and identify RNA viruses such as PVY. The I.C. step provides a simple method to isolate virus particles from plant tissue, thus enabling subsequent RT-PCR amplification for large-scale virus testing and typing. A multiplex format of the PCR has been developed for PVY. It will detect and identify fourteen strains and recombinants of PVY simultaneously. The protocol followed will be as per Chikh-Ali and Karasev, 2015.Identification of novel and "rare" recombinants: The IC-RT-PCR assay targets the most common recombination sites and can potentially mistype "rare" and newly described genotypes. Therefore, samples that show single or mixed infections sustained by different PVY genotypes will be selected and subjected to Illumina sequencing technology to reconstruct the whole genome viral sequences. As controls, samples that show any of the 14 individual PVY genotypes previously detected by the IC-RT-PCR assay will also be included. The Illumina datasets will be processed, and recombination and phylogenetic analyses performed as described in Della Bartola et al., 2020.Objective 2. Identify host resistance sources to recombinant PVY strains in the Colorado potato breeding germplasm.Currently, there are no widely accepted potato cultivars that are resistant to PVY. The Colorado potato breeding germplasm maintained by Dr. David Holm (Co-PI) has selections at various stages of variety development and advanced breeding clones. The markers used to screen for the presence of the genes have been used in breeding programs the world over: (1) RYSC3 marker to detect Ryadg, (2) YES3-3A, and YES3-3B to detect Rysto, and (3) Ry186 markers to detect Rychc. The Co-PI, Dr. Holm, uses this information to decide whether the selections will be advanced further along in the breeding program or abandoned. Similar decisions will be made regarding the advanced breeding clones (Table 1). In the current proposal, we plan to continue screening the selections and advanced breeding clones for the extreme resistance gene markers and further expand the work to include cataloging symptom development as detailed below.Marker-based screening for PVY resistance: Based on the parentage of the crosses, a hundred individual selections and advanced breeding clones will be evaluated or the presence of the three extreme resistance (ER) genes. Plant samples will be collected from field-grown plants in the summer of 2022 and 2023. Genomic DNA will be extracted from leaf samples. The presence of the resistance will be performed to determine the presence/absence of molecular markers for Ryadg, Rysto, and Rychc genes.Greenhouse screening for PVY response: Selections and advanced breeding clones containing the ER genes will be planted in the greenhouse at the SLVRC in the winter of 2022 and 2023. The lines will be evaluated for their response to multiple PVY strains (Four prevalent strains -NTN, -N-Wi, -O, and -N:O) and two unidentified novel and "rare" recombinant strains that are prevalent in the Valley identified from the survey performed in objective 1. Three to five plants for each selection and advanced clones will be grown in the greenhouse. Plants will be mechanically inoculated with different strains of PVY four weeks after planting. Inoculated plants will be monitored for visual symptoms, and ELISA (Enzyme-Linked ImmunoSorbent Assay) will be performed 20 and 40-days post-inoculation to confirm virus presence. As controls, we will also include ten major commercial potato varieties grown in the Valley and monitor symptom development in these lines in response to infection by novel and "rare" recombinants.Objective 3. Develop outreach and education materials that can be disseminated by extension professionals to growers, potato breeders, seed inspectors, and crop consultants.Outreach and education strategies. Providing resources and training for stakeholders is critical to successfully managing PVY. Because the current prevalent strains of PVY cause mild foliar symptoms, it is critical that the breeders, growers, and seed inspectors are aware of the novel and "rare" PVY strains emerging in the Valley. They have to be aware of the symptoms they have to be looking out for in commonly grown cultivars in the Valley. Diverse and numerous multimedia and presentation strategies are planned to reach our most diverse stakeholder audiences. We will maintain a blog/website that will provide up-to-date information regarding the diversity, prevalence, and incidence of PVY strains in the Valley. Besides, a catalog of symptom development (upon infection by novel and "rare" strains) will be posted on the blog/website. We propose targeting potato growers and seed inspectors, and crop advisors, including crop consultants and agricultural industry personnel (seed, chemical, and cooperative employees) with various digital and traditional media. We also propose using traditional indoor meetings, such as winter Extension meetings, and field days such as the one held at the San Luis Valley Research Center. These events can be used to demonstrate the progress of disease symptoms on important potato cultivars. More advanced and hands-on training opportunities will also be made available to advanced clientele and colleagues upon request.

Progress 09/01/23 to 08/31/24

Outputs
Target Audience:Throughout the duration of our project, our primary focus was on two distinct but interconnected target audiences: farmers and growers in the San Luis Valley and fellow scientists working on potato necrotic viruses, especially potato virus Y. These two groups were central to our efforts as they represented the key stakeholders who would directly benefit from the outcomes of our research and knowledge dissemination. 1. Farmers and Growers in the San Luis Valley: Geographic Focus: Farmers in the San Luis Valley, known for its unique environmental conditions and potato cultivation. Crop-Centric: We aimed to provide insights and tools for optimizing potato crop management, virus control, and resource efficiency. Community Engagement: We actively engaged with the local farming community through newsletters and collaborative initiatives. 2. Fellow Scientists Specializing in Potato Necrotic Viruses: Specialized Expertise: We aimed to contribute new knowledge to scientists specializing in potato necrotic viruses. Collaboration: Promoting collaboration and information exchange among scientists in this field was a priority. Data and Resources: We provided valuable data, research findings, and tools to support ongoing research. Knowledge Dissemination: Sharing our research through publications and conferences was central to our engagement with this audience. By targeting these two audiences, we aimed to bridge the gap between scientific research and practical solutions, empowering farmers to enhance potato cultivation in the San Luis Valley and increasing our understanding of an important potato pathogen. Changes/Problems:The major problem we encountered during the project was the data processing phase of high throughput sequence data, particularly in our efforts to identify novel recombinant strains of Potato Virus Y (PVY). Originally, we had planned to utilize existing bioinformatics pipelines to detect and confirm the absence of novel recombinant strains of PVY in our samples as part of Objective 2. However, due to the complexity of the data and the unique characteristics of PVY strains in our study area, we faced significant challenges in achieving conclusive results using the initially selected bioinformatics tools and protocols. As a result of these difficulties, we could not initiate Objective 2, which relied on identifying and characterizing novel PVY strains. As a direct outcome of addressing the challenges encountered in Objective 1, we did succeed in identifying one novel strain of PVY. This novel strain was found to be highly prevalent in the San Luis Valley and exhibited the intriguing characteristic of evading detection when established and published detection protocols were applied. This discovery highlighted the need for further investigation into the novel strain's behavior and its potential impact on potato crops in the region. To adapt to these changes and address the new findings, we have modified our approach. Specifically, we plan to focus on determining the symptom development of this newly identified PVY strain in commonly grown cultivars and advanced breeding lines. These assessments will be conducted under controlled greenhouse conditions, allowing us to gain insights into the strain's behavior and its potential consequences for potato cultivation in the San Luis Valley. This shift in approach is a response to the unexpected challenges encountered during data processing and the importance of investigating the novel PVY strain we have identified. What opportunities for training and professional development has the project provided?The project has created valuable opportunities for training and professional development, benefiting both graduate students and researchers involved. Some of the notable opportunities and achievements include: Graduate Student Training: The project has served as a platform for the training and development of one graduate student, Jennifer Rushton. Jen had the chance to actively participate in various aspects of the research and actively gain hands-on experience in experimental design, data collection, analysis, and interpretation. This training has equipped her with practical skills and research acumen that will be invaluable for her future career. Undergraduate Student Training: The graduate student gained significant experience in mentoring two undergraduate students throughout the duration of the grant. Attendance at Local and National Conferences: One significant opportunity provided by the project was the ability of the graduate student to attend conferences. These conferences are key forums for staying updated on the latest developments in their respective fields, networking with fellow researchers, and gaining exposure to diverse perspectives and research methodologies. Attendance at such conferences has broadened their horizons and expanded their knowledge base. These events have allowed them to disseminate their research findings, receive constructive feedback, and engage with peers and experts in their fields. Presenting at conferences not only enhances their communication skills but also builds their professional reputation and contributes to the wider dissemination of project outcomes. In summary, the project has provided opportunities for training and professional development. One graduate and two undergraduate students have received practical training, gained exposure to international conferences, and presented their research at various national and local conferences. These experiences collectively contribute to their growth as researchers and professionals in their respective fields, ensuring they are well-prepared for future endeavors in academia and beyond. How have the results been disseminated to communities of interest?The project's results have been disseminated to communities of interest through various channels: Scientific Manuscript: A scientific manuscript to showcase the significant findings from this research was recently accepted in the journla Phytopathology and first look version of the manuscript has been published, ensuring that the broader scientific community gains access to our insights. Newsletter Publication: In 2022, we published a newsletter in collaboration with the Colorado Potato Administrative Committee. This newsletter served as a vital communication tool to inform potato growers about the incidence and prevalence of Potato Virus Y (PVY), sharing essential information with a key stakeholder group. Conference Presentations: Our graduate and undergraduate students actively presented the research findings at multiple regional and national conferences. This dissemination strategy allowed us to reach a wider audience of experts and stakeholders, fostering knowledge exchange and discussion surrounding our work. What do you plan to do during the next reporting period to accomplish the goals?Objectives 1 and 3 werecompleted during the previous reporting period. During the next reporting period, we plan to focus on objective 3, i.e. screening of Colorado potato breeding germplasm to the different recombinant strains of PVY that were detected in SLV.

Impacts
What was accomplished under these goals? Under the specified goals, the following accomplishments have been achieved: Objective 1. Characterize the diversity, prevalence, and incidence of PVY strains in Colorado. Genetic Diversity Analysis: Extensive research efforts have been dedicated to characterizing the genetic diversity of Potato Virus Y (PVY) within the San Luis Valley. This involved thorough genetic profiling to identify different strains and their prevalence and incidence in the region. Determined the incidence and prevalence of PVY in the San Luis Valley over two years. Determined the influence of aphid abundance on PVY incidence and strain diversity. Identified the presence of an SNP in a strain of PVY that reduces its detection efficiency in published and routinely used protocols. Identified the presence of several samples with mixed infections, which has the potential to lead to novel recombinant strains. Did not identify a single novel recombinant strain of PVY. Determined that in 2021, in-season PVY incidence was especially high, and a plausible reason could be the presence of Potato Virus S. Objective 2: Identify host plant resistance to recombinant PVY strains in the Colorado potato breeding germplasm. Screening selection: Unfortunately, the outcomes for this goal were not met, given that objective 1 took longer than expected. However, we plan to complete this goal under greenhouse conditions in the next six months. Objective 3: Develop effective methods to provide research-based information regarding PVY detection and disease management to potato growers. Resource Development: Efforts in this objective have focused on creating resources and providing training to stakeholders, primarily potato growers. The aim is to equip them with research-based information on PVY detection and effective disease management strategies. These resources are designed to enhance disease control and crop health within the San Luis Valley. Overall, the accomplishments under these goals reflect a comprehensive approach that spans from understanding the genetic diversity and prevalence of PVY strains to in the future identifying resistant plant selections within the Colorado potato breeding germplasm. Additionally, efforts have been made to translate research findings into practical resources and training for potato growers, ultimately contributing to more effective PVY management and enhanced crop productivity in the region.

Publications

• Type: Peer Reviewed Journal Articles Status: Awaiting Publication Year Published: 2023 Citation: Ms. J. Rushton, Mr. B. Larson, Dr. A. Houser, Dr. W. J. Pitt, Dr. A. O. Charkowski, Dr. M. Chikh-Ali, and Dr. V. J. Nalam. Detection and Characterization of Potato Virus Y (PVY) Strains and Mixed Infections in San Luis Valley, Colorado. PhytoFrontiers" 0 0:ja

Progress 09/01/22 to 08/31/23

Outputs
Target Audience:Throughout the duration of our project, our primary focus was on two distinct but interconnected target audiences: farmers and growers in the San Luis Valley and fellow scientists working on potato necrotic viruses, especially potato virus Y. These two groups were central to our efforts as they represented the key stakeholders who would directly benefit from the outcomes of our research and knowledge dissemination. 1. Farmers and Growers in the San Luis Valley: • Geographic Focus: Farmers in the San Luis Valley, known for its unique environmental conditions and potato cultivation. • Crop-Centric: We aimed to provide insights and tools for optimizing potato crop management, virus control, and resource efficiency. • Community Engagement: We actively engaged with the local farming community through newsletters and collaborative initiatives. 2. Fellow Scientists Specializing in Potato Necrotic Viruses: • Specialized Expertise: We aimed to contribute new knowledge to scientists specializing in potato necrotic viruses. • Collaboration: Promoting collaboration and information exchange among scientists in this field was a priority. • Data and Resources: We provided valuable data, research findings, and tools to support ongoing research. • Knowledge Dissemination: Sharing our research through publications and conferences was central to our engagement with this audience. By targeting these two audiences, we aimed to bridge the gap between scientific research and practical solutions, empowering farmers to enhance potato cultivation in the San Luis Valley and increasing our understanding of an important potato pathogen. Changes/Problems:The major problem we encountered during the project was the data processing phase of high throughput sequence data, particularly in our efforts to identify novel recombinant strains of Potato Virus Y (PVY). Originally, we had planned to utilize existing bioinformatics pipelines to detect and confirm the absence of novel recombinant strains of PVY in our samples as part of Objective 2. However, due to the complexity of the data and the unique characteristics of PVY strains in our study area, we faced significant challenges in achieving conclusive results using the initially selected bioinformatics tools and protocols. As a result of these difficulties, we could not initiate Objective 2, which relied on identifying and characterizing novel PVY strains. As a direct outcome of addressing the challenges encountered in Objective 1, we did succeed in identifying one novel strain of PVY. This novel strain was found to be highly prevalent in the San Luis Valley and exhibited the intriguing characteristic of evading detection when established and published detection protocols were applied. This discovery highlighted the need for further investigation into the novel strain's behavior and its potential impact on potato crops in the region. To adapt to these changes and address the new findings, we have modified our approach. Specifically, we plan to focus on determining the symptom development of this newly identified PVY strain in commonly grown cultivars and advanced breeding lines. These assessments will be conducted under controlled greenhouse conditions, allowing us to gain insights into the strain's behavior and its potential consequences for potato cultivation in the San Luis Valley. This shift in approach is a response to the unexpected challenges encountered during data processing and the importance of investigating the novel PVY strain we have identified. What opportunities for training and professional development has the project provided?The project has created valuable opportunities for training and professional development, benefiting both graduate students and researchers involved. Some of the notable opportunities and achievements include: 1. Graduate Student Training: The project has served as a platform for the training and development of one graduate student, Jennifer Rushton. Jen had the chance to actively participate in various aspects of the research and actively gain hands-on experience in experimental design, data collection, analysis, and interpretation. This training has equipped her with practical skills and research acumen that will be invaluable for her future career. 2. Undergraduate Student Training: The graduate student gained significant experience in mentoring two undergraduate students throughout the duration of the grant. 3. Attendance at Local and National Conferences: One significant opportunity provided by the project was the ability of the graduate student to attend conferences. These conferences are key forums for staying updated on the latest developments in their respective fields, networking with fellow researchers, and gaining exposure to diverse perspectives and research methodologies. Attendance at such conferences has broadened their horizons and expanded their knowledge base. These events have allowed them to disseminate their research findings, receive constructive feedback, and engage with peers and experts in their fields. Presenting at conferences not only enhances their communication skills but also builds their professional reputation and contributes to the wider dissemination of project outcomes. In summary, the project has provided opportunities for training and professional development. One graduate and two undergraduate students have received practical training, gained exposure to international conferences, and presented their research at various national and local conferences. These experiences collectively contribute to their growth as researchers and professionals in their respective fields, ensuring they are well-prepared for future endeavors in academia and beyond. How have the results been disseminated to communities of interest?The project's results have been disseminated to communities of interest through various channels: • Scientific Manuscript: We have submitted a scientific manuscript to showcase the significant findings from this research, ensuring that the broader scientific community gains access to our insights. • Newsletter Publication: In 2022, we published a newsletter in collaboration with the Colorado Potato Administrative Committee. This newsletter served as a vital communication tool to inform potato growers about the incidence and prevalence of Potato Virus Y (PVY), sharing essential information with a key stakeholder group. • Conference Presentations: Our graduate and undergraduate students actively presented the research findings at multiple regional and national conferences. This dissemination strategy allowed us to reach a wider audience of experts and stakeholders, fostering knowledge exchange and discussion surrounding our work. What do you plan to do during the next reporting period to accomplish the goals?In order to accomplish Objective 2, we plan to conduct greenhouse trials.

Impacts
What was accomplished under these goals? Under the specified goals, the following accomplishments have been achieved: Objective 1. Characterize the diversity, prevalence, and incidence of PVY strains in Colorado. Genetic Diversity Analysis: Extensive research efforts have been dedicated to characterizing the genetic diversity of Potato Virus Y (PVY) within the San Luis Valley. This involved thorough genetic profiling to identify different strains and their prevalence and incidence in the region. • Determined the incidence and prevalence of PVY in the San Luis Valley over two years. • Determined the influence of aphid abundance on PVY incidence and strain diversity. • Identified the presence of an SNP in a strain of PVY that reduces its detection efficiency in published and routinely used protocols. • Identified the presence of several samples with mixed infections, which has the potential to lead to novel recombinant strains. • Did not identify a single novel recombinant strain of PVY. • Determined that in 2021, in-season PVY incidence was especially high, and a plausible reason could be the presence of Potato Virus S. Objective 2: Identify host plant resistance to recombinant PVY strains in the Colorado potato breeding germplasm. • Screening selection: Unfortunately, the outcomes for this goal were not met, given that objective 1 took longer than expected. However, we plan to complete this goal under greenhouse conditions in the next six months. Objective 3: Develop effective methods to provide research-based information regarding PVY detection and disease management to potato growers. • Resource Development: Efforts in this objective have focused on creating resources and providing training to stakeholders, primarily potato growers. The aim is to equip them with research-based information on PVY detection and effective disease management strategies. These resources are designed to enhance disease control and crop health within the San Luis Valley. Overall, the accomplishments under these goals reflect a comprehensive approach that spans from understanding the genetic diversity and prevalence of PVY strains to in the future identifying resistant plant selections within the Colorado potato breeding germplasm. Additionally, efforts have been made to translate research findings into practical resources and training for potato growers, ultimately contributing to more effective PVY management and enhanced crop productivity in the region.

Publications

• Type: Journal Articles Status: Submitted Year Published: 2024 Citation: Rushton, J., Larsen, B., Pitt, W.J., Chikh-Ali, M., Charkowski, A. O., Nalam, V.J. Detection and Characterization of Potato Virus Y (PVY) Strains and Mixed Infections in San Luis Valley, Colorado. Submitted to PhytoFrontiers
• Type: Other Status: Published Year Published: 2021 Citation: Rushton, J. and Nalam, V.J., Prevalence of PVY in San Luis Valley. Colorado Potato Administrative Committee newsletter
• Type: Conference Papers and Presentations Status: Accepted Year Published: 2024 Citation: Nalam. V.J., Detection and Characterization of Mixed Infection of Potato Virus Y (PVY) in the San Luis Valley. Southern Rocky Mountain Ag Conference, Monte Vista, February 6 - 7, 2024
• Type: Conference Papers and Presentations Status: Published Year Published: 2023 Citation: Rushton, J. and Nalam, V.J. Potato Virus Y in the San Luis Valley, Colorado. Colorado Fruit and Vegetable Growers Association conference, Denver, Co (February 21  22, 2023)

Progress 09/01/21 to 08/31/22

Outputs
Target Audience:Throughout the duration of our project, our primary focus was on two distinct but interconnected target audiences: farmers and growers in the San Luis Valley and fellow scientists working on potato necrotic viruses, especially potato virus Y. These two groups were central to our efforts as they represented the key stakeholders who would directly benefit from the outcomes of our research and knowledge dissemination. 1. Farmers and Growers in the San Luis Valley: Geographic Focus: Farmers in the San Luis Valley, known for its unique environmental conditions and potato cultivation. Crop-Centric: We aimed to provide insights and tools for optimizing potato crop management, virus control, and resource efficiency. Community Engagement: We actively engaged with the local farming community through newsletters and collaborative initiatives. 2. Fellow Scientists Specializing in Potato Necrotic Viruses: Specialized Expertise: We aimed to contribute new knowledge to scientists specializing in potato necrotic viruses. Collaboration: Promoting collaboration and information exchange among scientists in this field was a priority. Data and Resources: We provided valuable data, research findings, and tools to support ongoing research. Knowledge Dissemination: Sharing our research through publications and conferences was central to our engagement with this audience. By targeting these two audiences, we aimed to bridge the gap between scientific research and practical solutions, empowering farmers to enhance potato cultivation in the San Luis Valley and increasing our understanding of an important potato pathogen. Changes/Problems:The major problem we encountered during the project was the data processing phase of high throughput sequence data, particularly in our efforts to identify novel recombinant strains of Potato Virus Y (PVY). Originally, we had planned to utilize existing bioinformatics pipelines to detect and confirm the absence of novel recombinant strains of PVY in our samples as part of Objective 2. However, due to the complexity of the data and the unique characteristics of PVY strains in our study area, we faced significant challenges in achieving conclusive results using the initially selected bioinformatics tools and protocols. As a result of these difficulties, we could not initiate Objective 2, which relied on identifying and characterizing novel PVY strains. As a direct outcome of addressing the challenges encountered in Objective 1, we did succeed in identifying one novel strain of PVY. This novel strain was found to be highly prevalent in the San Luis Valley and exhibited the intriguing characteristic of evading detection when established and published detection protocols were applied. This discovery highlighted the need for further investigation into the novel strain's behavior and its potential impact on potato crops in the region. To adapt to these changes and address the new findings, we have modified our approach. Specifically, we plan to focus on determining the symptom development of this newly identified PVY strain in commonly grown cultivars and advanced breeding lines. These assessments will be conducted under controlled greenhouse conditions, allowing us to gain insights into the strain's behavior and its potential consequences for potato cultivation in the San Luis Valley. This shift in approach is a response to the unexpected challenges encountered during data processing and the importance of investigating the novel PVY strain we have identified. What opportunities for training and professional development has the project provided?The project has created valuable opportunities for training and professional development, benefiting both graduate students and researchers involved. Some of the notable opportunities and achievements include: Graduate Student Training: The project has served as a platform for the training and development of one graduate student, Jennifer Rushton. Jen had the chance to actively participate in various aspects of the research and actively gain hands-on experience in experimental design, data collection, analysis, and interpretation. This training has equipped her with practical skills and research acumen that will be invaluable for her future career. Undergraduate Student Training: The graduate student gained significant experience in mentoring two undergraduate students throughout the duration of the grant. Attendance at Local and National Conferences: One significant opportunity provided by the project was the ability of the graduate student to attend conferences. These conferences are key forums for staying updated on the latest developments in their respective fields, networking with fellow researchers, and gaining exposure to diverse perspectives and research methodologies. Attendance at such conferences has broadened their horizons and expanded their knowledge base. These events have allowed them to disseminate their research findings, receive constructive feedback, and engage with peers and experts in their fields. Presenting at conferences not only enhances their communication skills but also builds their professional reputation and contributes to the wider dissemination of project outcomes. In summary, the project has provided opportunities for training and professional development. One graduate and two undergraduate students have received practical training, gained exposure to international conferences, and presented their research at various national and local conferences. These experiences collectively contribute to their growth as researchers and professionals in their respective fields, ensuring they are well-prepared for future endeavors in academia and beyond. How have the results been disseminated to communities of interest?The project's results have been disseminated to communities of interest through various channels: Scientific Manuscript: We are currently preparing a scientific manuscript to showcase the significant findings from this research, ensuring that the broader scientific community gains access to our insights. Newsletter Publication: In 2022, we published a newsletter in collaboration with the Colorado Potato Administrative Committee. This newsletter served as a vital communication tool to inform potato growers about the incidence and prevalence of Potato Virus Y (PVY), sharing essential information with a key stakeholder group. Conference Presentations: Our graduate and undergraduate students actively presented the research findings at multiple regional and national conferences. This dissemination strategy allowed us to reach a wider audience of experts and stakeholders, fostering knowledge exchange and discussion surrounding our work. What do you plan to do during the next reporting period to accomplish the goals? Nothing Reported

Impacts
What was accomplished under these goals? Under the specified goals, the following accomplishments have been achieved: Objective 1. Characterize the diversity, prevalence, and incidence of PVY strains in Colorado. Genetic Diversity Analysis: Extensive research efforts have been dedicated to characterizing the genetic diversity of Potato Virus Y (PVY) within the San Luis Valley. This involved thorough genetic profiling to identify different strains and their prevalence and incidence in the region. Determined the incidence and prevalence of PVY in the San Luis Valley over two years. Determined the influence of aphid abundance on PVY incidence and strain diversity. Identified the presence of an SNP in a strain of PVY that reduces its detection efficiency in published and routinely used protocols. Identified the presence of several samples with mixed infections, which has the potential to lead to novel recombinant strains. Did not identify a single novel recombinant strain of PVY. Determined that in 2021, in-season PVY incidence was especially high, and a plausible reason could be the presence of Potato Virus S. Objective 2: Identify host plant resistance to recombinant PVY strains in the Colorado potato breeding germplasm. Screening selection: Unfortunately, the outcomes for this goal were not met, given that objective 1 took longer than expected. However, we plan to complete this goal under greenhouse conditions in the next six months/ Objective 3: Develop effective methods to provide research-based information regarding PVY detection and disease management to potato growers. Resource Development: Efforts in this objective have focused on creating resources and providing training to stakeholders, primarily potato growers. The aim is to equip them with research-based information on PVY detection and effective disease management strategies. These resources are designed to enhance disease control and crop health within the San Luis Valley. Overall, the accomplishments under these goals reflect a comprehensive approach that spans from understanding the genetic diversity and prevalence of PVY strains to in the future identifying resistant plant selections within the Colorado potato breeding germplasm. Additionally, efforts have been made to translate research findings into practical resources and training for potato growers, ultimately contributing to more effective PVY management and enhanced crop productivity in the region.

Publications